This document describes a study that evaluated the infection course in mice induced by L. major parasites in the presence of positively charged liposomes containing CpG oligodeoxynucleotides (CpG ODN). Mice were inoculated with L. major plus various liposome formulations and lesion development and parasite burden were analyzed. The results showed that mice treated with DMPC liposomes containing CpG ODN developed significantly smaller lesions and had lower parasite levels in the spleen and lymph nodes compared to other treatment groups. Additionally, this treatment induced a Th1 immune response profile. Therefore, immune modulation using DMPC liposomes containing CpG ODN may be an effective approach to improve the safety of leishman
Objective: To investigate the changes in the retina due to deltamethrin toxicity and the process in cell inflammation and apoptosis.
Study Design: Sixteen Wistar albino rats were randomly divided into two groups as control (n=8) and deltamethrin (n=8) groups. Saline was given to the control group, and 0.5 mL of 5 mg/kg deltamethrin was given to the deltamethrin group for 14 days each. Blood was collected for biochemical analysis. Retinal tissue was processed for histological examination.
Results: Compared to the control group, MDA levels were high while GSH and CAT levels were low in the deltamethrin group. Histopathological analysis showed spaces between the pigment epithelium, irregularity in the delimiting membrane, degenerated ganglion, cone and bacillus cell, pyknotic nuclei, thinned inner limitation membrane, and thickened vascular wall. The control group showed FAS expression in the pigment layer limiting membranes, in the nuclei of many cone and bacillus cells, and ganglion cells in the control group sections. In the deltamethrin group, FAS expression was observed in the inner and outer limiting membranes of the pigment epithelium, cone and bacillus cells, and ganglion cell nuclei. In the control group, negative NOS expression in the pigment epithelium and outer limiting membranes, internal limitation membrane, and ganglion cells in the cone and bacillus cell nuclei were observed. In the deltamethrin group, NOS expression was positive in the pigment epithelium, cone and bacillus, and ganglion cell nuclei.
Conclusion: We suggest that deltamethrin toxicity induced apoptotic process due to increased inflammation in the retina and may cause visual impairment as a result of neural damage.
Keywords: deltamethrin, FAS, insecticides, NOS, nitric oxide synthase, retina
Objective: To identify interstitial cells of Cajal (ICC) in the common bile duct of Kunming mice.
Study Design: Common bile ducts obtained from the Kunming mice were prepared for immunohistochemical investigations using the c-kit antibody. Immunoelectron microscopy was used to detect the expression of c-kit in the ICC of the common bile duct. Transmission electron microscopy showed ultrastructure of ICC in the murine bile duct. Reverse transcription–polymerase chain reaction (RT-PCR) and western blot were used to confirm the expression of mRNA specific for the c-kit gene and production of c-kit protein in the Kunming mice common bile duct.
Results: Immunohistochemistry revealed that ICC in the murine common bile duct are c-kit positive and the ICC are located in the tela submucosa and the tunica muscularis of the murine common bile duct and do not connect with each other. Immunoelectron microscopy confirmed the expression of Kit by ICC in the murine common bile duct. Transmission electron microscopy showed that ICC in the murine common bile duct have long processes, abundant mitochondria, plenty of smooth endoplasmic reticulum (sER), a lot of lysosomes, and dense bodies. The caveolae of ICC are distinctive. At the same time, RT-PCR indicated that the Kunming mice common bile duct expressed mRNA specific for the c-kit gene, and western blot analysis showed the evidence of production of c-kit protein in the Kunming mice common bile duct.
Conclusion: ICC are found in the Kunming mice common bile duct, which is likely to lead to the development of motility study of the common bile duct.
Keywords: common bile duct; electron microscopy; immuno-electron microscopy; interstitial cells of Cajal; intestines; smooth muscle; tyrosine kinase receptor (c-kit)
Objective(s):
This study aimed to find the effects of silver nanoparticles (Ag-NPs) (40 nm) on skin wound healing in mice Mus musculus when innate immune system has been suppressed.
Materials and Methods:
A group of 50 BALB/c mice of about 8 weeks (weighting 24.2±3.0 g) were randomly divided into two groups: Ag-NPs and control group, each with 25 mice. Once a day at the same time, a volume of 50 microliters from the nanosilver solution (10ppm) was applied to the wound bed in the Ag-NPs group while in the untreated (control) group no nanosilver solution was used but the wound area was washed by a physiological solution. The experiment lasted for 14. Transforming growth factor beta (TGF-β), complement component C3, and two other immune system factors involving in inflammation, namely C-reactive protein (CRP) and rheumatoid factor (RF) in sera of both groups were assessed and then confirmed by complement CH50 level of the blood.
Results:
The results show that wound healing is a complex process involving coordinated interactions between diverse immunological and biological systems and that Ag-NPs significantly accelerated wound healing and reduce scar appearance through suppression of immune system as indicated by decreasing levels of all inflammatory factors measured in this study.
Conclusion:
Exposure of mice to Ag-NPs can result in significant changes in innate immune function at the molecular levels. The study improves our understanding of nanoparticle interaction with components of the immune system and suggests that Ag-NPs have strong anti-inflammatory effects on skin wound healing and reduce scarring.
Nano-adjuvanted polio vaccine: Preparation and characterization of chitosan ...Nanomedicine Journal (NMJ)
Abstract
Objective(s):
It is proposed that particulate antigens could better interact with the antigen presenting cells (APCs). A fast, simple and scalable process for preparation of polymeric nanoparticles (NPs) is coating of charged antigenic particles, like viruses, with oppositely charged polymers. A second coating with a charged polymer could increase the stability and modify the immunomodulatory potentials of NPs.
Materials and Methods:
Negatively charged inactivated polio virus (IPV) was coated with cationic polymers, chitosan (CHT) and trimethylchitosan (TMC) by a simple incubation method. CHT: IPV and TMC: IPV NPs were coated by anionic polymer, sodium alginate (ALG). Physical characteristics and stability of NPs were studied. Cytocompatibility of NPs was checked with MTT assay. DC maturation study was used for evaluation of the NPs potential in interaction with DCs.
Results:
Among the various polymer to antigen ratios tested, the least size and PDI and the highest ZP was seen in TMC: IPV (2:1), CHT: IPV (2:1), ALG: TMC: IPV (2:2:1) and ALG: CHT: IPV (4:2:1). The physical stability of TMC: IPV and CHT: IPV was preserved until 15 days. After an early de-association of some part of coated alginate, ALG: CHT: IPV and ALG: TMC: IPC NPs were stable until the end of study (25th day). No one of the NPs formulations had a negative effect on cell viability. Compared with plain IPV, nanoparticulate IPV formulations failed to increase the expression of CD40 and CD86 markers of DCs.
Conclusion:
NPs prepared with simple and scalable method, had reasonable physical characteristics, stability and cytocompatibility and could be tested in vivo for their immunoadjuvant potential.
Objective: To investigate the changes in the retina due to deltamethrin toxicity and the process in cell inflammation and apoptosis.
Study Design: Sixteen Wistar albino rats were randomly divided into two groups as control (n=8) and deltamethrin (n=8) groups. Saline was given to the control group, and 0.5 mL of 5 mg/kg deltamethrin was given to the deltamethrin group for 14 days each. Blood was collected for biochemical analysis. Retinal tissue was processed for histological examination.
Results: Compared to the control group, MDA levels were high while GSH and CAT levels were low in the deltamethrin group. Histopathological analysis showed spaces between the pigment epithelium, irregularity in the delimiting membrane, degenerated ganglion, cone and bacillus cell, pyknotic nuclei, thinned inner limitation membrane, and thickened vascular wall. The control group showed FAS expression in the pigment layer limiting membranes, in the nuclei of many cone and bacillus cells, and ganglion cells in the control group sections. In the deltamethrin group, FAS expression was observed in the inner and outer limiting membranes of the pigment epithelium, cone and bacillus cells, and ganglion cell nuclei. In the control group, negative NOS expression in the pigment epithelium and outer limiting membranes, internal limitation membrane, and ganglion cells in the cone and bacillus cell nuclei were observed. In the deltamethrin group, NOS expression was positive in the pigment epithelium, cone and bacillus, and ganglion cell nuclei.
Conclusion: We suggest that deltamethrin toxicity induced apoptotic process due to increased inflammation in the retina and may cause visual impairment as a result of neural damage.
Keywords: deltamethrin, FAS, insecticides, NOS, nitric oxide synthase, retina
Objective: To identify interstitial cells of Cajal (ICC) in the common bile duct of Kunming mice.
Study Design: Common bile ducts obtained from the Kunming mice were prepared for immunohistochemical investigations using the c-kit antibody. Immunoelectron microscopy was used to detect the expression of c-kit in the ICC of the common bile duct. Transmission electron microscopy showed ultrastructure of ICC in the murine bile duct. Reverse transcription–polymerase chain reaction (RT-PCR) and western blot were used to confirm the expression of mRNA specific for the c-kit gene and production of c-kit protein in the Kunming mice common bile duct.
Results: Immunohistochemistry revealed that ICC in the murine common bile duct are c-kit positive and the ICC are located in the tela submucosa and the tunica muscularis of the murine common bile duct and do not connect with each other. Immunoelectron microscopy confirmed the expression of Kit by ICC in the murine common bile duct. Transmission electron microscopy showed that ICC in the murine common bile duct have long processes, abundant mitochondria, plenty of smooth endoplasmic reticulum (sER), a lot of lysosomes, and dense bodies. The caveolae of ICC are distinctive. At the same time, RT-PCR indicated that the Kunming mice common bile duct expressed mRNA specific for the c-kit gene, and western blot analysis showed the evidence of production of c-kit protein in the Kunming mice common bile duct.
Conclusion: ICC are found in the Kunming mice common bile duct, which is likely to lead to the development of motility study of the common bile duct.
Keywords: common bile duct; electron microscopy; immuno-electron microscopy; interstitial cells of Cajal; intestines; smooth muscle; tyrosine kinase receptor (c-kit)
Objective(s):
This study aimed to find the effects of silver nanoparticles (Ag-NPs) (40 nm) on skin wound healing in mice Mus musculus when innate immune system has been suppressed.
Materials and Methods:
A group of 50 BALB/c mice of about 8 weeks (weighting 24.2±3.0 g) were randomly divided into two groups: Ag-NPs and control group, each with 25 mice. Once a day at the same time, a volume of 50 microliters from the nanosilver solution (10ppm) was applied to the wound bed in the Ag-NPs group while in the untreated (control) group no nanosilver solution was used but the wound area was washed by a physiological solution. The experiment lasted for 14. Transforming growth factor beta (TGF-β), complement component C3, and two other immune system factors involving in inflammation, namely C-reactive protein (CRP) and rheumatoid factor (RF) in sera of both groups were assessed and then confirmed by complement CH50 level of the blood.
Results:
The results show that wound healing is a complex process involving coordinated interactions between diverse immunological and biological systems and that Ag-NPs significantly accelerated wound healing and reduce scar appearance through suppression of immune system as indicated by decreasing levels of all inflammatory factors measured in this study.
Conclusion:
Exposure of mice to Ag-NPs can result in significant changes in innate immune function at the molecular levels. The study improves our understanding of nanoparticle interaction with components of the immune system and suggests that Ag-NPs have strong anti-inflammatory effects on skin wound healing and reduce scarring.
Nano-adjuvanted polio vaccine: Preparation and characterization of chitosan ...Nanomedicine Journal (NMJ)
Abstract
Objective(s):
It is proposed that particulate antigens could better interact with the antigen presenting cells (APCs). A fast, simple and scalable process for preparation of polymeric nanoparticles (NPs) is coating of charged antigenic particles, like viruses, with oppositely charged polymers. A second coating with a charged polymer could increase the stability and modify the immunomodulatory potentials of NPs.
Materials and Methods:
Negatively charged inactivated polio virus (IPV) was coated with cationic polymers, chitosan (CHT) and trimethylchitosan (TMC) by a simple incubation method. CHT: IPV and TMC: IPV NPs were coated by anionic polymer, sodium alginate (ALG). Physical characteristics and stability of NPs were studied. Cytocompatibility of NPs was checked with MTT assay. DC maturation study was used for evaluation of the NPs potential in interaction with DCs.
Results:
Among the various polymer to antigen ratios tested, the least size and PDI and the highest ZP was seen in TMC: IPV (2:1), CHT: IPV (2:1), ALG: TMC: IPV (2:2:1) and ALG: CHT: IPV (4:2:1). The physical stability of TMC: IPV and CHT: IPV was preserved until 15 days. After an early de-association of some part of coated alginate, ALG: CHT: IPV and ALG: TMC: IPC NPs were stable until the end of study (25th day). No one of the NPs formulations had a negative effect on cell viability. Compared with plain IPV, nanoparticulate IPV formulations failed to increase the expression of CD40 and CD86 markers of DCs.
Conclusion:
NPs prepared with simple and scalable method, had reasonable physical characteristics, stability and cytocompatibility and could be tested in vivo for their immunoadjuvant potential.
ST8 micellar/niosomal vesicular nanoformulation for delivery of naproxen in c...Vahid Erfani-Moghadam
Naproxen (NPX) is a non-steroidal anti-inflammatory drug (NSAID) used against a variety of diseases, including autoimmune disorders and chronic inflammations. However, low water solubility limits its therapeutic efficacy and novel nanoformulations are required to bypass its poor bioavailability to reach its therapeutic effect. The purpose of the study was to investigate the role of the nanoformulation of biocompatible molecules; Squalene (S) and Tween 80 (T8) Micellar/Niosomal Vesicles (ST8MNV) prepared, by thin-film hydration method and their potential as a drug delivery system for NPX. The percentage of encapsulation efficiency was calculated to be 99.5 ± 0.2% for 5% of NPX weight in total ingredients of micellar/niosomal vesicles (w/w). The ST8MNV nanoformulation exhibited a slower rate of NPX release from the drug encapsulated over seven days, suggesting a stable complex of NPX. Finally, cell toxicity assay demonstrated that the half-maximal inhibitory concentrations (IC50) of NPX were drastically reduced by ST8MNV nanoformulation in MCF-7, A549, HeLa, and MDA-MB-231 cancer cell lines. Our data show this micellar/niosomal naproxen nanoformulation is a great candidate for the future in vitro and in vivo studies for potential clinical anti-inflammatory and anticancer applications.
ABSTRACT- Multiple Drug resistance (MDR) tuberculosis timely diagnose is of utmost clinical relevance and needs to be diagnose at initial stages for the proper treatment. The current study was done to detect the several genes for MDR tuberculosis (TB) in clinical isolates by molecular tools. 60 clinical isolates were collected and subjected for AFB smear preparation, Nested PCR (IS6110) for mycobacterium tuberculosis complex detection and MDR TB PCR targeting rpoB, kat G, mab A promoter. 12 came positive for AFB smears, out of which 08 were pulmonary and 04 were extra pulmonary. Nested PCR targeting IS6110 gene was amplified at 123 base pairs with 340 base pairs as IC (internal control) was seen in 25 cases which include 19 pulmonary and 6 extra pulmonary. The Positive TB PCR specimens were subjected for MDRTB PCR Only 06 cases yielded, an amplicon of 315 bp confirming the rpoB gene resistance for resistance for rifampcin drug. In any of the 06 positives none of the other resistance gene other than rpoB was amplified. Targeting multiple genes at once, additional information will be gained from a single test run that otherwise would require several times the reagents and more time to perform. Current study signifies the usage of quick, cost effective, DNA sequences based method for MDR TB detection where disease will be diagnosed earlier and hence treatment would be started at an early stage.
Keywords: Multiple drug resistance, amplicon, Polymerase chain reaction, Nested PCR, Rifampicin.
Objective: To evaluate the antibacterial effects of 4 different cavity disinfectants on Streptococcus mutans, Lactobacillus acidophilus, and Enterococcus faecalis bacteria in different time periods.
Study Design: The antibacterial effects of Cavity Cleanser, Tubulicid Red Label, Chloraxid 2%, and Oxygenated Water cavity disinfectant solutions on E. faecalis (ATCC 29212), S. mutans (ATCC 25175), and L. acidophilus (RSKK 03037) bacterial strains were evaluated by disk diffusion method. In the study where vancomycin antibiogram disc constituted the positive control group, physiological saline solution was used as the negative control group. Standard, sterile, blank antibiogram discs of 5 mm in diameter, in which 15 μL of each material were added, were placed on agar plates at 2.5–3 cm intervals. The inhibition zone diameters formed around the discs that were left to incubate for 24–48 hours at 37°C were measured in millimeters. Statistical analysis of the data was performed using one-way analysis of variance, Kolmogorov-Smirnov, Levene, and Bonferroni tests.
Results: At the end of the study the solutions tested showed a statistically significant antibacterial effect on all bacterial strains used (p<0.05). Cavity Cleanser disinfectant containing 2% chlorhexidine showed the highest antibacterial effect on S. mutans and L. acidophilus, and benzalkonium-containing Tubulicid Red disinfectant on E. faecalis.
Conclusion: The antibacterial effect of all cavity disinfectants used in the study was found to be higher at the end of the 48th hour than at the end of the 24th hour, but there was no statistically significant difference (p>0.05).
Keywords: antibacterial agents; antibacterial effect; cavity disinfectants; chlorhexidine; contamination; dental caries; disinfection; disc diffusion; gram-negative bacteria; gram-positive bacteria
Personalized nanomedicine for the treatment of vascular hypertensionSusanta Kumar Rout
This study includes designing a nanomedical device for the treatment of vascular hypertension in polycystic kidney diseases (PKD) model through cilia targeting.
They generated and compared two different metal and polymer cilia-targeted nanoparticle drug delivery systems (DDS), i.e. gold (Au) and poly-lactic-co-glycolic acid (PLGA) nanoparticles (NPs)
The target is Dopamine-receptor type-5 (DRS) on primary cilia.
The drug-loaded is Fenoldopam (FD).
Ameliorative potentials of a combination of fenugreek and alpha-tocopherol on...Prof. Hesham N. Mustafa
Background: The current study aimed to elucidate the protective role of combined fenugreek and a-tocopherol against cadmium induced histopathological
changes in the testes.
Materials and methods: Thirty adult male albino rats divided into three equal
groups 10 rats each. Group I is the control group. Group II received 5 mg/kg/day
cadmium chloride. Group III received 5 mg/kg/day cadmium chloride and 150
mg/kg/day fenugreek and 100 mg/kg/day of a-tocopherol. The treatment of all
groups was done by oral gavage for 60 consecutive days. The testes were removed
and subjected to histopathological and ultrastructure study.
Results: Rats exposed to cadmium showed severe histopathological changes in
the testicular spermatogenic series, many vacuoles and multinucleated giant cells.
Treatment with fenugreek and a-tocopherol partially improved the morphological
changes, reduced tissue damage and rebuilt of the spermatogonia layer.
Conclusions: Fenugreek and a-tocopherol might represent a promising medicinal
combination to ameliorate the toxic effects of cadmium exposure. (Folia Morphol
2015; 74, 3: 325–334)
Key words: cadmium chloride, fenugreek, a-tocopherol, seminiferous
epithelium, ultrastructure
ST8 micellar/niosomal vesicular nanoformulation for delivery of naproxen in c...Vahid Erfani-Moghadam
Naproxen (NPX) is a non-steroidal anti-inflammatory drug (NSAID) used against a variety of diseases, including autoimmune disorders and chronic inflammations. However, low water solubility limits its therapeutic efficacy and novel nanoformulations are required to bypass its poor bioavailability to reach its therapeutic effect. The purpose of the study was to investigate the role of the nanoformulation of biocompatible molecules; Squalene (S) and Tween 80 (T8) Micellar/Niosomal Vesicles (ST8MNV) prepared, by thin-film hydration method and their potential as a drug delivery system for NPX. The percentage of encapsulation efficiency was calculated to be 99.5 ± 0.2% for 5% of NPX weight in total ingredients of micellar/niosomal vesicles (w/w). The ST8MNV nanoformulation exhibited a slower rate of NPX release from the drug encapsulated over seven days, suggesting a stable complex of NPX. Finally, cell toxicity assay demonstrated that the half-maximal inhibitory concentrations (IC50) of NPX were drastically reduced by ST8MNV nanoformulation in MCF-7, A549, HeLa, and MDA-MB-231 cancer cell lines. Our data show this micellar/niosomal naproxen nanoformulation is a great candidate for the future in vitro and in vivo studies for potential clinical anti-inflammatory and anticancer applications.
ABSTRACT- Multiple Drug resistance (MDR) tuberculosis timely diagnose is of utmost clinical relevance and needs to be diagnose at initial stages for the proper treatment. The current study was done to detect the several genes for MDR tuberculosis (TB) in clinical isolates by molecular tools. 60 clinical isolates were collected and subjected for AFB smear preparation, Nested PCR (IS6110) for mycobacterium tuberculosis complex detection and MDR TB PCR targeting rpoB, kat G, mab A promoter. 12 came positive for AFB smears, out of which 08 were pulmonary and 04 were extra pulmonary. Nested PCR targeting IS6110 gene was amplified at 123 base pairs with 340 base pairs as IC (internal control) was seen in 25 cases which include 19 pulmonary and 6 extra pulmonary. The Positive TB PCR specimens were subjected for MDRTB PCR Only 06 cases yielded, an amplicon of 315 bp confirming the rpoB gene resistance for resistance for rifampcin drug. In any of the 06 positives none of the other resistance gene other than rpoB was amplified. Targeting multiple genes at once, additional information will be gained from a single test run that otherwise would require several times the reagents and more time to perform. Current study signifies the usage of quick, cost effective, DNA sequences based method for MDR TB detection where disease will be diagnosed earlier and hence treatment would be started at an early stage.
Keywords: Multiple drug resistance, amplicon, Polymerase chain reaction, Nested PCR, Rifampicin.
Objective: To evaluate the antibacterial effects of 4 different cavity disinfectants on Streptococcus mutans, Lactobacillus acidophilus, and Enterococcus faecalis bacteria in different time periods.
Study Design: The antibacterial effects of Cavity Cleanser, Tubulicid Red Label, Chloraxid 2%, and Oxygenated Water cavity disinfectant solutions on E. faecalis (ATCC 29212), S. mutans (ATCC 25175), and L. acidophilus (RSKK 03037) bacterial strains were evaluated by disk diffusion method. In the study where vancomycin antibiogram disc constituted the positive control group, physiological saline solution was used as the negative control group. Standard, sterile, blank antibiogram discs of 5 mm in diameter, in which 15 μL of each material were added, were placed on agar plates at 2.5–3 cm intervals. The inhibition zone diameters formed around the discs that were left to incubate for 24–48 hours at 37°C were measured in millimeters. Statistical analysis of the data was performed using one-way analysis of variance, Kolmogorov-Smirnov, Levene, and Bonferroni tests.
Results: At the end of the study the solutions tested showed a statistically significant antibacterial effect on all bacterial strains used (p<0.05). Cavity Cleanser disinfectant containing 2% chlorhexidine showed the highest antibacterial effect on S. mutans and L. acidophilus, and benzalkonium-containing Tubulicid Red disinfectant on E. faecalis.
Conclusion: The antibacterial effect of all cavity disinfectants used in the study was found to be higher at the end of the 48th hour than at the end of the 24th hour, but there was no statistically significant difference (p>0.05).
Keywords: antibacterial agents; antibacterial effect; cavity disinfectants; chlorhexidine; contamination; dental caries; disinfection; disc diffusion; gram-negative bacteria; gram-positive bacteria
Personalized nanomedicine for the treatment of vascular hypertensionSusanta Kumar Rout
This study includes designing a nanomedical device for the treatment of vascular hypertension in polycystic kidney diseases (PKD) model through cilia targeting.
They generated and compared two different metal and polymer cilia-targeted nanoparticle drug delivery systems (DDS), i.e. gold (Au) and poly-lactic-co-glycolic acid (PLGA) nanoparticles (NPs)
The target is Dopamine-receptor type-5 (DRS) on primary cilia.
The drug-loaded is Fenoldopam (FD).
Ameliorative potentials of a combination of fenugreek and alpha-tocopherol on...Prof. Hesham N. Mustafa
Background: The current study aimed to elucidate the protective role of combined fenugreek and a-tocopherol against cadmium induced histopathological
changes in the testes.
Materials and methods: Thirty adult male albino rats divided into three equal
groups 10 rats each. Group I is the control group. Group II received 5 mg/kg/day
cadmium chloride. Group III received 5 mg/kg/day cadmium chloride and 150
mg/kg/day fenugreek and 100 mg/kg/day of a-tocopherol. The treatment of all
groups was done by oral gavage for 60 consecutive days. The testes were removed
and subjected to histopathological and ultrastructure study.
Results: Rats exposed to cadmium showed severe histopathological changes in
the testicular spermatogenic series, many vacuoles and multinucleated giant cells.
Treatment with fenugreek and a-tocopherol partially improved the morphological
changes, reduced tissue damage and rebuilt of the spermatogonia layer.
Conclusions: Fenugreek and a-tocopherol might represent a promising medicinal
combination to ameliorate the toxic effects of cadmium exposure. (Folia Morphol
2015; 74, 3: 325–334)
Key words: cadmium chloride, fenugreek, a-tocopherol, seminiferous
epithelium, ultrastructure
Cutaneous Leishmaniasis in Pakistan.
Cutaneous Leishmaniasis (CL) is a rising epidemic in Pakistan.
Cutaneous leishmaniasis is found in all the four provinces of Pakistan, Punjab, Sindh, Balochistan, KPK.
A comprehensive description of leischmaniasis with its types, transmission, epidemiology, pathogenesis, prevention and control. It also includes details regarding lab diagnosis, disease agent, vector and host.
Las leishmaniasis son un grupo de enfermedades causadas por diferentes parásitos que
pertenecen a la familia Tripanosomatidae, género Leishmania, transmitidas al ser humano por
la picadura de distintas especies de insectos flbótomos.
Estos flbótomos, que son los vectores de la enfermedad, son diferentes según la especie de
Leishmania.
Estas enfermedades se caracterizan por comprometer la piel, mucosas y vísceras. Dicho compromiso dependerá fundamentalmente de la especie de Leishmania, pero también de la respuesta inmune del huésped entre otros factores.
Las leishmaniasis se consideran enfermedades reemergentes, y un problema creciente de salud
pública en el mundo, debido al aumento de la cantidad de afectados como consecuencia de la
mayor exposición de las personas a los vectores de la enfermedad.
Esta mayor exposición a los vectores se produce en el caso de las leishmaniasis cutáneas en
América por cambios en las condiciones del ambiente (deforestación, cambios climáticos), en
el caso de la leishmaniasis visceral urbana en América por tránsito, tráfio y patrones culturales
del manejo de mascotas, y en ambas por migraciones con urbanización rápida y desorganizada, que incluyen defiiencias en el saneamiento ambiental (disposición inadecuada de excretas
y basura) y viviendas precarias, así como tendencias regionales a la tropicalización.
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
All manuscripts are subject to rapid peer review. Those of high quality (not previously published and not under consideration for publication in another journal) will be published without delay.
Abnormal expression of Pygopus 2 correlates with a malignant phenotype in hum...Enrique Moreno Gonzalez
Pygopus 2 (Pygo2) is a Pygo family member and an important component of the Wnt signaling transcriptional complex. Despite this data, no clinical studies investigating Pygo2 expression in lung cancer have yet been reported.
Content Cytotoxicity Studies of Colorectal Carcinoma Cells Using Printed Impe...journalBEEI
Monitoring the effectiveness of drugs on cancer cells is crucial for chemotherapeutics studies. In-vitro cell-based biosensors can be used as an alternative for characteristic studies of cells’ response to drugs. Cell-based sensors provide real-time measurements and require smaller sample volumes compared to conventional T-flask measurement methods. This paper presents a biosensor that detects in real-time, impedance variations of human colon cancer, HCT-116 cells when treated with anti-cancer agent, 5-Fluorouracil (5-FU). Two different extracellular matrix (ECM); polyaniline and gelatin were tested and evaluated in terms of attachment quality. Polyaniline was found to provide the best attachment for HCT-116 cells and was used for cytotoxicity studies. Cytokinetic behavior indicated that 5-FU inhibited HCT-116 cells at IC50 of 6.8 µg/mL. Trypan blue exclusion method for testing cell viability was used to validate the impedance measurements, where the cancer cell concentrations were reduced to ~35% when treated with 2.5 µg/mL, and 50% when treated with 6.8 µg/mL. The results generated by the microfabricated impedance biosensor are comparable to the Trypan blue method since both gave similar cell growth trend. It can be concluded that the impedance biosensor has potential to be used as an alternative method in drug testing applications.
Low expression of N-myc downstream-regulated gene 2 in oesophageal squamous c...Enrique Moreno Gonzalez
It is currently unclear whether a correlation exists between N-myc downstream-regulated gene 2 (NDRG2) expression and oesophageal squamous cell carcinoma (ESCC). The aim of this study was to examine the underlying clinical significance of NDRG2 expression in ESCC patients and to investigate the effects of NDRG2 up-regulation on ESCC cell growth in vitro and in vivo.
ABSTRACT- Coronary artery disease (CAD) is suspected as a leading cause of mortality in developed countries. Due
to cholesterol and fat deposit plaque is forming into the inner walls of the arteries of the heart, which leads to narrowing
of blood vessels of heart and reduce the blood flow rate into heart. Proprotein convertase subtilisin-like kexin type 9
(PCSK9) is one of the candidate gene that regulate lipoprotein retention pathway of CAD development. It is a newly
discovered serine protease that plays a key role in LDL-C homeostasis by mediating LDL receptor (LDLR). The LDL
receptor is breakdown through a post transcriptional mechanism and induces the production of very low-density
lipoprotein in the fasting state. The aim of this study was to investigate the frequency of single nucleotide
polymorphism (SNP) of PCSK9 gene of 155 CAD patients and 102 ages matched healthy controls. Serum lipids
including total cholesterol (TC), triglycerides (TG), HDL, LDL, and VLDL were analyzed. PCR-RFLP analysis was
carried out to genotype regions carrying Eam 1104I restriction site in the PCSK9. Gene considering significant
difference in serum TC, TG, HDL-C, LDL-C and VLDL-C levels (P<0.001, <0.0001) of patients and control samples.
In CAD patients, G allele frequency is less than A allele frequency. G allele is responsible for decreasing the
LDL: HDL ratio which shows evidence in having its protecting effect on the occurrence of CAD in West Bengal Population.
Key-words- CAD, PCSK9, SNP, Eam1104I, Polymorphism, West Bengal population
Paratuberculosis (PTB) remains one of the most obstacles limit animal breeding sector all over the world. The current study aimed to detect the etiology of PTB in tissues of clinically suspected small ruminants using histopathological and real-time polymerase chain reaction (RT-PCR) methods. Clinical examination showed 10 (26.4%) PTB suspected cases out of the total (38) examined animals. The suspected cases were euthanized, necropsied, gross lesions were recorded and tissue samples were collected for histopathological and molecular procedures. Grossly intestinal and mesenteric lymph nodes thickening, corrugations and edematous swellings were recorded. Semi-thin sections of the intestine and mesenteric lymph nodes stained with toluidine blue demonstrated MAP organism inside epithelium cells and macrophages. RT-PCR detected MAP IS900 gene in all suspected cases (100%), thus we recommend using RT-PCR as a rapid sensitive method in the diagnosis of PTB.
Key-words: Paratuberculosis, Mycobacterium, Semi thin sections, Toluidine blue, IS900 gene
In vivo evaluation of mucoadhesive properties of nanoliposomal formulations u...Nanomedicine Journal (NMJ)
Objective(s):
Drug delivery via mucosal routes has been confirmed to be effective in inducing strong immune responses. Liposomes could enhance immune responses and mucoadhesive potentials, make them useful mucosal drug delivery systems. Coating of liposomes by mucoadhesive polymers succeeded in enhancing immune responses. Our studies aim at preparation and characterization of trimethylchitosan-coated nanoliposomes for nasal delivery of a model antigen, tetanus toxoid (TT).
Materials and Methods:
Anionic liposomes were prepared by dehydration-rehydration method with an average size of 100 nm and were coated with 0.01% (w/v) solution of trimethyulchitosan (TMC) with 50±10% of quaternization. Surface properties and zeta potential were evaluated by DLS. Antigen stability and integrity were studied by SDS-PAGE electrophoresis. Nasal clearance rate and mucoadhesive properties of liposomes were studied by gamma scintigraphy method using 99mTc-labelled liposomes.
Results:
The zeta potential of non-coated and TMC-coated liposomes was -40 and +38.8, respectively. Encapsulation rate of tetanus toxoid was 77 ± 5.5%. SDS-PAGE revealed that the antigens remained intact during formulation procedure. Gamma scintigraphy confirmed that both types of liposomes could remain in nasal cavity up to ten folds over the normal residence time for conventional nasal formulations.
Conclusion:
TMC-coated nanoliposomes have several positive potentials including good mucoadhesive properties, preserved integrity of loaded antigen and presence of TMC as a mucoadhesive polymer with innate immunoadjuvant potential which make them suitable for efficient adjuvant/delivery system.
Similar to Evaluation of infection course in mice induced by L. major in presence of positively charged liposomes containing CpG ODN (20)
Evaluation of the effect of crocetin on antitumor activity of doxorubicin enc...Nanomedicine Journal (NMJ)
Objective(s): The current study reports investigation of codelivery by PLGA nanoparticles (NPs) loaded with crocetin (Cro), a natural carotenoid dicarboxylicHYPERLINK “http://en.wikipedia.org/wiki/Carboxylic_acid” acid that is found in the crocus flower, and Doxorubicin (DOX).
Materials and Methods: Double emulsion/solvent evaporation method was used for preparation of PLGA nanoparticles containing Dox and Cro. Characterizations of prepared NPs were investigated by atomic force microscopy (AFM) and dynamic light scattering analysis. In vitro Cytotoxicity of DOX and Cro loaded PLGA NPs (PLGA-DOX-Cro) on MCF-7 cell line was evaluated using MTT test. Flow cytometry experiments were implemented to distinguish cells undergoing apoptosis from those undergoing necrosis. Furthermore the expression of caspase 3 was examined by western blot analysis.
Results: The prepared formulations had size of 150- 300 nm. Furthermore, PLGA-DOX-Cro nanoparticles inhibited MCF-7 tumor cells growth more efficiently than either DOX or Cro alone at the same concentrations, as quantified by MTT assay and flow cytometry. Studies on cellular uptake of DOX-Cro-NPs demonstrated that NPs were effectively taken up by MCF-7 tumor cells.
Conclusion: This study suggested that DOX-Cro-NPs may have promising applications in breast cancer therapy.
Effects of combination of magnesium and zinc oxide nanoparticles and heat on ...Nanomedicine Journal (NMJ)
Objective: The objective of this study was to investigate the antibacterial activities of combination of MgO and ZnO nanoparticles in the presence of heat against Escherichia coli and Staphylococcus aureus.
Materials and Methods:Bacteria were grown on either agar or broth media followed by the addition of ZnO and MgO nanoparticles. Then the combined effect of ZnO and MgO nanoparticles was investigated. Furthermore, the media containing nanoparticles were treated with mild heat and their synergistic antibacterial activity was investigated against E. coli and S. aureus in milk.
Results: The data showed that the nanoparticles used in this study had no effect on the bacteria in the agar medium. However, the results showed that ZnO and MgO nanoparticles resulted in a significant decrease in the number of E. coli (P<0.000) and S. aureus (Pd”0.05) in the broth medium. The combination of nanoparticles and mild heat exhibited a significant decrease in the number of E. coli and S. aureus indicating the synergistic effects of nanoparticles and heat.
Conclusion: Using a combination of mild heat, ZnO and MgO nanoparticles, E. coli and S. aureus can be controlled successfully in the milk. Mild heating plus ZnO and MgO nanoparticles has a synergistic effect which would reduce the need for high temperature and also the concentrations of ZnO and MgO nanoparticles required for pathogen control in minimally processed milk during maintaining.
Preparation and evaluation of electrospun nanofibers containing pectin and ti...Nanomedicine Journal (NMJ)
Objective(s):The aim of this study was to prepare electrospun nanofibers of celecoxib using combination of time-dependent polymers with pectin to achieve a colon-specific drug delivery system for celecoxib.
Materials and Methods:Formulations were produced based on two multilevel 22 full factorial designs. The independent variables were the ratio of drug:time-dependent polymer (X1) and the amount of pectin in formulations (X2). Electrospinning process was used for preparation of nanofibers. The spinning solutions were loaded in 5 mL syringes. The feeding rate was fixed by a syringe pump at 2.0 mL/h and a high voltage supply at range 10-18 kV was applied for electrospinning. Electrospun nanofibers were collected and evaluated by scanning electron microscopy and drug release in the acid and buffer with pH 6.8 with and without pectinase.
Results:Electrospun nanofibers of celecoxib with appropriate morphological properties were produced via electrospinning process. Drug release from electrospun nanofibers was very low in the acidic media; while, drug release in the simulated colonic media was the highest from formulations containing pectin.
Conclusion: Formulation F2 (containing drug:ERS with the ratio of 1:2 and 10% pectin) exhibited acceptable morphological characteristics and protection of drug in the upper GI tract and could be a good candidate as a colonic drug delivery system for celecoxib.
The combined effects of Aloe vera gel and silver nanoparticles on wound heali...Nanomedicine Journal (NMJ)
Objective(s): This study was aimed at investigating the synergy effects of Aloe vera gel and silver nanoparticles on the healing rate of the cutting wounds.
Materials and Methods: In order to determine the concentration of silver nanoparticles in Aloe vera gel, the MBC methods were applied on the most common bacteria infecting wounds, Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa. The cutting wounds with Full-thickness skin were dorsally created on rats; then the rats were divided into 4 groups. The treatments groups included: mixture of Aloe vera gel and silver nanoparticles, Aloe vera gel alone and silver nanoparticles alone in addition to control groups. The treatment was carried out for 2 weeks and the size of the wound closures were measured by an image software analysis.
Results:There was no significant difference (p<0.05) in healing rate between the control and mixture group. However, there were significant differences between the silver nanoparticles and Aloe vera groups using Tukey’s analysis on the 6th, 8th and 10th days.
Conclusion:The Aloe vera gel increased the rate of wound healing whereas the silver nanoparticles had a delay effect; and when they were mixed, it was similar to the average effect of both Aloe vera gel and silver nanoparticles.
Simultaneous loading of 5-florouracil and SPIONs in HSA nanoparticles: Optimi...Nanomedicine Journal (NMJ)
Objective(s): Over the past two decades, considerable interest has been focused on utilizing biocompatible magnetic nanoparticles (MNPs) for biomedical applications. In this study, production of human serum albumin (HSA) nanoparticles using desolvation technique that were simultaneous loaded with high amounts of superparamagnetic iron oxide nanoparticles (SPIONs) and 5-flourouracil (5-FU) was investigated.
Materials and Methods: 5-FU loading (%) and SPIONs entrapment efficiency (%) were optimized using response surface methodology (RSM). The design expert software used to analyse the interactive effects of pH, 5-FU and SPIONs concentrations.
Results:The optimum conditions found to be pH of 8.2, drug concentration of 1.5 mg/ml and SPIONs concentration of 2.79 mg/ml. Under the mentioned optimum conditions, particles with the size of 111.8 nm, zeta potential of -37.1 mV, 5-FU loading of 15.8% and SPIONs entrapment efficiency of 41.1% were obtained. In vitro cumulative release of 5-FU from the nanoparticles was evaluated in phosphate buffer saline (pH 7.4, 37 °C). Results indicated that 85% of the 5-FU released during 95 h, which revealed a sustained release profile. In addition, Vibrating Sample Magnetometer (VSM) analyses confirmed the superparamagnetic properties of magnetic albumin nanoparticles manufactured under the optimum conditions.
Conclusion: According to the findings,SPIONs and 5-FU loaded HAS nanoparticles arepromising for use as novel targeted delivery system due to proper magnetic and drug release behaviours.
Antimicrobial and cytotoxicity effect of silver nanoparticle synthesized by C...Nanomedicine Journal (NMJ)
Objective(s): For the development of reliable, ecofriendly, less expensive process for the synthesis of silver nanoparticles and to evaluate the bactericidal, and cytotoxicity properties of silver nanoparticles synthesized from root extract of Croton bonplandianum, Baill.
Materials and Methods: The synthesis of silver nanoparticles by plant part of Croton bonplandianum was carried out. The formation of nanoparticles was confirmed by Transmission Electron Microscopy (TEM), Scanning Electron Microscopy (SEM), XRD and UV-Vis spectrophotometric analysis. The biochemical properties were assayed by antibacterial study, cytotoxicity assay using cancer cell line.
Results: The formation of silver nanoparticles was confirmed by UV-VIS spectroscopic analysis which showed absorbance peak at 425 nm. X-ray diffraction photograph indicated the face centered cubic structure of the synthesized AgNPs. TEM has displayed the different dimensional images of biogenic silver nanoparticles with particle size distribution ranging from 15-40 nm with an average size of 32 nm. Silver particles are spherical in shape, clustered. The EDX analysis was used to identify the elemental composition of synthesized AgNPs. Antibacterial activity of the synthesized AgNPs against three Gram positive and Gram negative bacteria strains like Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa carried out showed significant zones of inhibition. The cytotoxicity study by AgNPS also showed cytotoxicity on ovarian cancer cell line PA-1 and lung epithelial cancer cell line A549.
Conclusion: The present study confirms that the AgNPs have great promise as antibacterial, and anticancer agent.
Investigation of the effect of different parameters on the phase inversion te...Nanomedicine Journal (NMJ)
Objective(s): Nanoemulsions are a kind of emulsions that can be transparent, translucent (size range 50-200 nm) or “milky” (up to 500 nm). Nanoemulsions are adequatly effective for transfer of active component through skin which facilitate the entrance of the active component . The transparent nature of the system and lack of the thickener and fluidity are among advantages of nanoemulsion.
Materials and Methods: In this study, a nanoemulsion of lemon oil in water was prepared by the phase inversion temperature (PIT) emulsification method in which the tween 40 was used as surfactant. The effect of concentration of NaCl in aqueous phase, pH and weight percent of surfactant and aqueous on the PIT and droplet size were investigated. Results: The results showed that with increasing of concentration of NaCl from 0.05 M to 1 M, PIT decrease from 72 to 50. The average droplet sizes, for 0.1, 0.5 and 1 M of NaCl in 25 ºC are 497.3, 308.1 and 189.9 nm, respectively and the polydispersity indexes are 0.348, 0.334 and 0.307, respectively.
Conclusion: Considering the characteristics of nanoemulsions such as being transparent, endurance of solution and droplet size can provide suitable reaction environment for polymerization process used in making hygienic and medical materials.
Mechanism of oxidative stress involved in the toxicity of ZnO nanoparticles a...Nanomedicine Journal (NMJ)
ZnO NPs (zinc oxide nanoparticles) has generated significant scientific interest as a novel antibacterial and anticancer agent. Since oxidative stress is a critical determinant of ZnO NPs-induced damage, it is necessary to characterize their underlying mode of action. Different structural and physicochemical properties of ZnO NPs such as particle surface, size, shape, crystal structure, chemical position, and presence of metals can lead to changes in biological activities including ROS (reactive oxygen species) production. However, there are some inconsistencies in the literature on the relation between the physicochemical features of ZnO NPs and their plausible oxidative stress mechanism. Herein, the possible oxidative stress mechanism of ZnO NPs was reviewed. This is worthy of further detailed evaluations in order to improve our understanding of vital NPs characteristics governing their toxicity. Therefore, this study focuses on the different reported oxidative stress paradigms induced by ZnO NPs including ROS generated by NPs, oxidative stress due to the NPs-cell interaction, and role of the particle dissolution in the oxidative damage. Also, this study tries to characterize and understand the multiple pathways involved in oxidative stress induced by ZnO NPs. Knowledge about different cellular signaling cascades stimulated by ZnO NPs lead to the better interpretation of the toxic influences induced by the cellular and acellular parameters. Regarding the potential benefits of toxic effects of ZnO NPs, in-depth evaluation of their toxicity mechanism and various effects of these nanoparticles would facilitate their implementation for biomedical applications.
Combined effects of PEGylation and particle size on uptake of PLGA particles ...Nanomedicine Journal (NMJ)
Abstract
Objective:
At the present study, relationship between phagocytosis of PLGA particles and combined effects of particle size and surface PEGylation was investigated.
Materials and Methods:
Microspheres and nanospheres (3500 nm and 700 nm) were prepared from three types of PLGA polymers (non-PEGylated and PEGylation percents of 9% and 15%). These particles were prepared by solvent evaporation method. All particles were labeled with FITC-Albumin. Interaction of particles with J744.A.1 mouse macrophage cells, was evaluated in the absence or presence of 7% of the serum by flowcytometry method.
Results:
The study revealed more phagocytosis of nanospheres. In the presence of the serum, PEGylated particles were phagocytosed less than non-PEGylated particles. For nanospheres, this difference was significant (P<0/05) and their uptake was affected by PEGylation degree. In the case of microsphere formulation, PEGylation did not affect the cell uptake. In the serum-free medium, the bigger particles had more cell uptake rate than smaller ones but the cell uptake rate was not influenced by PEGylation.
Conclusion:
The results indicated that in nanosized particles both size and PEgylation degree could affect the phagocytosis, but in micron sized particles just size, and not the PEGylation degree, could affect this.
Synthesis of silver nanoparticles and its synergistic effects in combination ...Nanomedicine Journal (NMJ)
Abstract
Objectives:
Biofilms are communities of bacteria attached to surfaces through an external polymeric substances matrix. In the meantime, Acinetobacterbaumannii is the predominant species related to nosocomial infections. In the present study, the effect of silver nanoparticles alone and in combination with biocides and imipenem against planktonic and biofilms of A. baumannii was assessed.
Materials and Methods:
Minimum inhibitory concentrations (MICs) of 75 planktonic isolates of A. baumannii were determined by using the microdilution method as described via clinical and laboratory standards institute (CLSI). Among all strains, 10 isolates which formed strong biofilms were selected and exposed to silver nanoparticles alone and in combination with imipenem, bismuth ethandithiol (BisEDT) and bismuth propanedithiol (BisPDT) to determine minimum biofilm inhibitory concentrations (MBIC). Subsequently, minimum biofilm eradication concentrations (MBECs) of silver nanoparticles alone and in combination with imipenem against mature biofilm of the isolates were evaluated.
Results:
Results showed that 29.3% of isolates were susceptible to silver nanoparticles and could inhibit the growth and eradicate biofilms produced by the isolates. For this reason, ∑FIC, ∑FBIC and ∑FBEC ≤ 0.05 were reported which shows synergism between silver nanoparticles and imipenem against not only planktonic cells but also inhibition and eradication of biofilms. The results of ∑FBIC >2 indicated to antagonistic impacts between silver nanoparticles and BisEDT/BisPDT against biofilms.
Conclusion:
It can be concluded that silver nanoparticles alone can inhibit biofilm formation but in combination with imipenem are more effective against A. baumannii in planktonic and biofilm forms.
Abstract
Objective(s):
Zinc oxide nanoparticles (ZNP) are increasingly used in sunscreens, biosensors, food additives and pigments. In this study the effects of ZNP on liver of rats was investigated.
Materials and Methods:
Experimental groups received 5, 50 and 300 mg/kg ZNP respectively for 14 days. Control group received only distilled water. ALT, AST and ALP were considered as biomarkers to indicate hepatotoxicity. Lipid peroxidation (MDA), SOD and GPx were detected for assessment of oxidative stress in liver tissue. Histological studies and TUNEL assay were also done.
Results:
Plasma concentration of zinc (Zn) was significantly increased in 5 mg/kg ZNP-treated rats. Liver concentration of Zn was significantly increased in the 300 mg/kg ZNP-treated animals. Weight of liver was markedly increased in both 5 and 300 mg/kg doses of ZNP. ZNP at the doses of 5 mg/kg induced a significant increase in oxidative stress through the increase in MDA content and a significant decrease in SOD and GPx enzymes activity in the liver tissue. Administration of ZNP at 5 mg/kg induced a significant elevation in plasma AST, ALT and ALP. Histological studies showed that treatment with 5 mg/kg of ZNP caused hepatocytes swelling, which was accompanied by congestion of RBC and accumulation of inflammatory cells. Apoptotic index was also significantly increased in this group. ZNP at the dose of 300 mg/kg had poor hepatotoxicity effect.
Conclusion:
It is concluded that lower doses of ZNP has more hepatotoxic effects on rats, and recommended to use it with caution if there is a hepatological problem.
Synthesis of graphene oxide-TiO2 nanocomposite as an adsorbent for the enrich...Nanomedicine Journal (NMJ)
Abstract
Objective(s):
In our study, graphene oxide-TiO2 nanocomposite (GO/TiO2) was prepared and used for the enrichment of rutin from real samples for the first time.
Materials and Methods:
The synthesized GO/TiO2 was characterized by X-ray diffraction, scanning electron microscopy, and FT-IR spectra. The enrichment process is fast and highly efficient. The factors including contact time, pH, and amount of GO/TiO2 affecting the adsorption process were studied.
Results:
The maximum adsorption capacity for ciprofloxacin was calculated to be 59.5 mg/g according to the Langmuir adsorption isotherm. The method yielded a linear calibration curve in the concentration ranges from 15 to 200 μg/L for the rutin with regression coefficients (r2) of 0.9990. The limits of detection (LODs, S/N=3) and limits of quantification (LOQs, S/N=10) were found to be 8 μg/Land 28 μg/L, respectively. Both the intra-day and inter-day precisions (RSDs) were < 10% .
Conclusion:
The developed approach offered wide linear range, and good reproducibility. Owing to the diverse structures and unique characteristic, GO/TiO2 possesses great potential in the enrichment and analysis of trace rutin in real aqueous samples.
Preparation and evaluation of vitamin A nanosuspension as a novel ocular drug...Nanomedicine Journal (NMJ)
Abstract
Objective(s):
The aim of this study was to prepare a nanosuspension formulation as a new vehicle for the improvement of the ocular delivery of vitamin A.
Material and Methods:
Formulations were designed based on full factorial design. A high pressure homogenization technique was used to produce nanosuspensions. Fifteen formulations were prepared by the use of different combinations of surfactants Tween 80, benzalkonium chloride and Pluronic and evaluated for pH, particle size, entrapment efficiency, differential scanning calorimetry (DSC), stability and drug release. Also, Draize test was used to evaluate the irritation of rabbit eye by formulations.
Results:
All formulations showed a small mean size that is well suited for ocular application. Also it was observed that the particle size decreased with increase in the amount of surfactant. Drug entrapment increased with increasing amount of surfactant. It was shown that initial and final drug release can be controlled by the ratio and the total amount of surfactants, respectively.
Conclusion:
It was concluded that the use of Tween 80 and Pluronic in the formualtions with a proper ratio does not show eye irritation and could be useful to achieve a suitable nanosuspension of vitamin A as a novel ocular delivery system.
A comparative study about toxicity of CdSe quantum dots on reproductive syste...Nanomedicine Journal (NMJ)
Abstract
Objective(s):
Medicinal benefits of quantum dots have been proved in recent years but there is little known about their toxicity especially in vivo toxicity. In order to use quantum dots in medical applications, studies ontheir in vivo toxicity is important.
Materials and Methods:
CdSe:ZnS quantum dots were injected in 10, 20, and 40 mg/kg doses to male mice10 days later, mice were sacrificed and five micron slides were prepared structural and optical properties of quantum dots were evaluated using XRD.
Results:
Histological studies of testis tissue showed high toxic effect of CdSe:ZnS in 40 mg/kg group. Histological studies of epididymis did not show any effect of quantum dots in terms of morphology and tube structure. Mean concentration of LH and testosterone and testis weight showed considerable changes in mice injected with 40 mg/kg dose of CdSe:ZnS compared to control group. However, FSH and body weight did not show any difference with control group.
Conclusion:
Although it has been reported that CdSe is highly protected from the environment by its shell, but this study showed high toxicity for CdSe:ZnS when it is used in vivo which could be suggested that shell could contribute to increased toxicity of quantum dots. Considering lack of any previous study on this subject, our study could potentially be used as an basis for further extensive studies investigating the effects of quantum dots toxicity on development of male sexual system.
Functionalization of carbon nanotubes and its application in nanomedicine: A ...Nanomedicine Journal (NMJ)
Abstract
This review focuses on the latest developments in applications of carbon nanotubes (CNTs) in medicine. A brief history of CNTs and a general introduction to the field are presented.
Then, surface modification of CNTs that makes them ideal for use in medical applications is highlighted. Examples of common applications, including cell penetration, drug delivery, gene delivery and imaging, are given. At the same time, there are concerns about their possible adverse effects on human health, since there is evidence that exposure to CNTs induces toxic effects in experimental models. However, CNTs are not a single substance but a growing family of different materials possibly eliciting different biological responses. As a consequence, the hazards associated with the exposure of humans to the different forms of CNTs may be different. Understanding the structure–toxicity relationships would help towards the assessment of the risk related to these materials. Finally, toxicity of CNTs, are discussed. This review article overviews the most recent applications of CNTs in Nanomedicine, covering the period from 1991 to early 2015.
The role of surface charge of ISCOMATRIX nanoparticles on the type of immune ...Nanomedicine Journal (NMJ)
Abstract
Objective(s):
ISCOMATRIX vaccines have now been shown to induce strong antigen-specific cellular or humoral immune responses to a broad range of antigens of viral, bacterial, parasite or tumor. In the present study, we investigated the role of ISCOMATRIX charge in induction of a Th1 type of immune response and protection against Leishmania major infection in BALB/c mice.
Materials and Methods:
Positively and negatively charged ISCOMATRIX were prepared. BALB/C mice were immunized subcutaneously, three times with 2-week intervals, with different ISCOMATRIX formulations. Soluble Leishmania antigens (SLA) were mixed with ISCOMATRIX right before injection. The extent of protection and type of immune response were studied in different groups of mice.
Results:
The group of mice immunized with negatively charged ISCOMATRIX showed smaller footpad swelling upon challenge with L. major and the highest IgG2a production compared with positively charged one. The mice immunized with positively charged ISCOMATRIX showed the lowest splenic parasite burden compared to the other groups. Cytokine assay results indicated that the highest level of IFN- γ and IL-4 secretion was observed in the splenocytes of mice immunized with negatively charged ISCOMATRIX as compared to other groups.
Conclusion:
The results indicated that ISCOMATRIX formulations generate an immune response with mixed Th1/Th2 response that was not protective against challenge against L. major.
Abstract
In the last decade, developments in nanotechnology have provided a new field in medicine called “Nanomedicine”. Nanomedicine has provided new tools for photodynamic therapy. Quantum dots (QDs) are approximately spherical nanoparticles that have attracted broad attention and have been used in nanomedicine applications. QDs have high molar extinction coefficients and photoluminescence quantum yield, narrow emission spectra, broad absorption, large effective stokes shifts. QDs are more photostable and resistant to metabolic degradation. These photosensitizing properties can be used as photosensitizers for Photodynamic Therapy (PDT). PDT has been recommended for its unique characteristic, such as low side effect and more efficiency. Therefore, nanomedicine leads a promising future for targeted therapy in cancer tumor. Furthermore, QDs have recently been applied in PDT, which will be addressed in this review letter. Also this review letter evaluates key aspects of nano-particulate design and engineering, including the advantage of the nanometer scale size range, biological behavior, and safety profile.
Preparation of protein-loaded PLGA-PVP blend nanoparticles by nanoprecipitati...Nanomedicine Journal (NMJ)
Abstract
Objective(s):
Despite of wide range applications of polymeric nanoparticles in protein delivery, there are some problems for the field of protein entrapment, initial burst and controlled release profile.
Materials and Methods:
In this study, we investigated the influence of some changes in PLGA nanoparticles formulation to improve the initial and controlled release profile. Selected parameters were: pluronic F127, polysorbate 80 as surfactant, pH of inner aqueous phase, L/G ratio of PLGA polymer, volume of inner aqueous phase and addition of polyvinylpyrrolidone as an excipient. FITC-HSA was used as a model hydrophilic drug. The nanoparticles were prepared by nanoprecipitation.
Results:
Initial release of FITC-HSA from PLGA-tween 80 nanoparticles (opt-4, 61%) was faster than control (PLGA-pluronic) after 2.30 h of incubation. Results showed that decrease in pH of inner aqueous phase to pI of protein can decrease IBR but the release profile of protein is the same as control. Release profile with three phases including a) initial burst b) plateau and c) final release phase was observed when we changed volume of inner aqueous phase and L/G ratio in formulation. Co-entrapment of HSA with PVP and pluronic reduced the IBR and controlled release profile in opt-19. Encapsulation efficiency was more than 97% and nanoparticles size and zeta potentials were mono-modal and -18.99 mV, respectively.
Conclusion:
In this research, we optimized a process for preparation of PLGA-PVP-pluronic nanoparticles of diameter less than 300 nm using nanoprecipitation method. This formulation showed a decreased initial burst and long lasting controlled release profile for FITC-HSA as a model drug for proteins.
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A Strategic Approach: GenAI in EducationPeter Windle
Artificial Intelligence (AI) technologies such as Generative AI, Image Generators and Large Language Models have had a dramatic impact on teaching, learning and assessment over the past 18 months. The most immediate threat AI posed was to Academic Integrity with Higher Education Institutes (HEIs) focusing their efforts on combating the use of GenAI in assessment. Guidelines were developed for staff and students, policies put in place too. Innovative educators have forged paths in the use of Generative AI for teaching, learning and assessments leading to pockets of transformation springing up across HEIs, often with little or no top-down guidance, support or direction.
This Gasta posits a strategic approach to integrating AI into HEIs to prepare staff, students and the curriculum for an evolving world and workplace. We will highlight the advantages of working with these technologies beyond the realm of teaching, learning and assessment by considering prompt engineering skills, industry impact, curriculum changes, and the need for staff upskilling. In contrast, not engaging strategically with Generative AI poses risks, including falling behind peers, missed opportunities and failing to ensure our graduates remain employable. The rapid evolution of AI technologies necessitates a proactive and strategic approach if we are to remain relevant.
2024.06.01 Introducing a competency framework for languag learning materials ...Sandy Millin
http://sandymillin.wordpress.com/iateflwebinar2024
Published classroom materials form the basis of syllabuses, drive teacher professional development, and have a potentially huge influence on learners, teachers and education systems. All teachers also create their own materials, whether a few sentences on a blackboard, a highly-structured fully-realised online course, or anything in between. Despite this, the knowledge and skills needed to create effective language learning materials are rarely part of teacher training, and are mostly learnt by trial and error.
Knowledge and skills frameworks, generally called competency frameworks, for ELT teachers, trainers and managers have existed for a few years now. However, until I created one for my MA dissertation, there wasn’t one drawing together what we need to know and do to be able to effectively produce language learning materials.
This webinar will introduce you to my framework, highlighting the key competencies I identified from my research. It will also show how anybody involved in language teaching (any language, not just English!), teacher training, managing schools or developing language learning materials can benefit from using the framework.
Unit 8 - Information and Communication Technology (Paper I).pdfThiyagu K
This slides describes the basic concepts of ICT, basics of Email, Emerging Technology and Digital Initiatives in Education. This presentations aligns with the UGC Paper I syllabus.
Macroeconomics- Movie Location
This will be used as part of your Personal Professional Portfolio once graded.
Objective:
Prepare a presentation or a paper using research, basic comparative analysis, data organization and application of economic information. You will make an informed assessment of an economic climate outside of the United States to accomplish an entertainment industry objective.
Honest Reviews of Tim Han LMA Course Program.pptxtimhan337
Personal development courses are widely available today, with each one promising life-changing outcomes. Tim Han’s Life Mastery Achievers (LMA) Course has drawn a lot of interest. In addition to offering my frank assessment of Success Insider’s LMA Course, this piece examines the course’s effects via a variety of Tim Han LMA course reviews and Success Insider comments.
Francesca Gottschalk - How can education support child empowerment.pptxEduSkills OECD
Francesca Gottschalk from the OECD’s Centre for Educational Research and Innovation presents at the Ask an Expert Webinar: How can education support child empowerment?
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Introduction to AI for Nonprofits with Tapp NetworkTechSoup
Dive into the world of AI! Experts Jon Hill and Tareq Monaur will guide you through AI's role in enhancing nonprofit websites and basic marketing strategies, making it easy to understand and apply.
Evaluation of infection course in mice induced by L. major in presence of positively charged liposomes containing CpG ODN
1. 28 Nanomed J, Vol. 1, No. 1, Autumn 2013
Received: May. 23, 2013; Accepted: Jul. 12 , 2013
Vol. 1, No. 1, Autumn 2013, page 28-37
Received: Apr. 22, 2014; Accepted: Jul. 12, 2014
Vol. 1, No. 5, Autumn 2014, page 298-301
Online ISSN 2322-5904
http://nmj.mums.ac.ir
Original Research
Evaluation of infection course in mice induced by L. major in presence of
positively charged liposomes containing CpG ODN
Hesamoddin Hoseinjani1
, Mahmoud Reza Jaafari1,2
, Ali Khamesipour3
, Azam Abbasi1
, Zahra
Saberi1
, Ali Badiee1*
1
Nanotechnology Research Center, School of Pharmacy, Mashhad University of Medical Science, Mashhad, Iran
2
Biotechnology Research Center, School of Pharmacy, Mashhad University of Medical Science, Mashhad, Iran
3
Center for Research and Training in Skin Diseases and Leprosy, Tehran University of Medical Sciences,
Tehran, Iran
Abstract
Objective(s): An inoculation of virulent Leishmania major is known as leishmanization (LZ)
which is proven to be the most effective control measure against Cutaneous Leishmaniasis
(CL). However, using LZ is restricted due to various side effects such as uncontrolled lesion
development.
Materials and Methods: In the present research, the efficacy of cationic nanoliposomes
containing CpG oligodeoxynucleotides (CpG ODN) as an improved adjuvant delivery system
was studied to diminish the lesion development and infection course of L. major after
inoculation into the mice. BALB/c mice were inoculated subcutaneously (SC) with L. major
plus empty DSPC, DSPC (CpG ODN), DSPC (Non CpG ODN), empty DMPC, DMPC (CpG
ODN), DMPC (Non CpG ODN) or HEPES buffer.
Results: The results showed that group of mice received DMPC (CpG ODN) nanoliposomes
developed a significantly smaller lesion and showed minimum number of L. major in the
spleen and draining lymph nodes. In addition, using DMPC (CpG ODN) liposomes resulted
in a Th1 type of immune response with a preponderance of IgG2a isotype which is concurrent
with the production of DMPC (CpG) induced IFN-γ in the spleen of the mice.
Conclusion: Taken together, the results suggested that immune modulation using DMPC
(CpG ODN) nanoliposomes might be a practical approach to improve the safety of LZ.
Keywords: CpG ODN, DMPC (CpG ODN), Nanoliposomes, Immune response,
Leishmanization, L. major
*Corresponding author: Ali Badiee, School of Pharmacy, Mashhad University of Medical Sciences, Mashhad, Iran.
Tel: +98 511 8823255, Email: badieea@mums.ac.ir
A.B. and M.R.J. equally designed research
2. Cationic liposomes in lesion development by L. major
Nanomed J, Vol. 1, No. 1, Autumn 2013 29
Introduction
Leishmanization (LZ) is the practice of
inducing infection by injecting live virulent
parasites in an aesthetically acceptable site of
the body in healthy individuals. Despite of
limited efficacy results seen in clinical trials
using first generation Leishmania vaccine
consists of killed Leishmania with or without
adjuvant, inoculation of live L. major showed
to be highly efficient tool to control CL. LZ
has been used for centuries in various parts of
Asian countries to protect against further
infection especially multiple injections in the
face. Mass LZ was used in Iran during the
1980s and the results showed that LZ is highly
effective. Due to various reasons such as
development of uncontrolled lesion, LZ
practice was stopped in different countries
except in Uzbekistan (1).
The ability of CpG ODN to induce both innate
and adaptive cellular immune responses makes
it a prospective prophylactic and therapeutic
vaccine adjuvant for diseases requiring cellular
immunity (2, 3). Unmethylated CpG motifs
present at high frequency in bacterial but not
vertebrate DNA are recognized by Toll-like
receptor 9 expressed in B cells and
plasmacytoid dendritic cells (pDCs) (4, 5). A
potential method to increase ODN uptake by
the cells of the immune system involves
liposome encapsulation of ODN (6, 7) as we
also showed previously against leishmaniasis
(8, 9). Moreover, the efficacy of
coadminstration of live L. major with lipo-
some–protamine–DNA nanoparticle (LPD)
containing immunostimulatory CpG oligodeo-
xynucleotides (CpG ODN) which is an
improved adjuvant delivery system was
examined to check Leishmania pathology and
immune response generated (10). The results
suggested that immune modulation using LPD
nanoparticles might be a practical approach to
improve the safety of inoculation of live L.
major.Cationic liposomes are used as an
adjuvant delivery system for CpG ODN in the
current study. Cationic liposomes are
particularly attractive due to their favorable
characteristics such as biodegradability,
minimal toxicity, and relative ease of large-
scale production and simplicity of use (11).
These particles target antigens for endocytosis
by APCs more efficiently as the cationic
entities interact with the negatively charged
molecules on the surface of APCs (12).
In addition, there have been reports of the use
of cationic liposomes for the generation of
CD8+
T cell response, which requires antigen
presentation in the context of the MHC class I
pathway (13).
They have also been successfully used as
delivery systems for nucleic acids in gene
therapy (14).
In the present study, cationic nanoliposomes
containing CpG ODN was used instead of
LPD nanoparticle, according to their simplicity
during formulation and stability issues, to
explore the possibility to induce a milder
lesion and infection after live Leishmania
parasites inoculation in susceptible BALB/c
mice.
Materials and Methods
Animals, parasites and CpG ODNs
Female BALB/c mice 4-6 weeks old were
purchased from Pasteur Institute (Tehran,
Iran). All mice were maintained in animal
house of Pharmaceutical Research Center and
fed with tap water and standard laboratory diet
(Khorasan Javane Co., Mashhad, Iran).
Animals were housed in a colony room 12/12h
light/dark cycle at 21˚ C with free access to
food and water. Animal experiments were
carried out according to Mashhad University
of Medical Sciences, Ethical Committee Acts.
The Leishmania major strain (MRHO/I-
R/75/ER) used in this experiment is the one
which has been used for experimental
Leishmania vaccine and Leishmanin test in
Iran (15-17).
The CpG ODNs used in this study was a
Nuclease-resistant phosphorothioate-modified
sequence 1826 containing two CpG motifs
(underlined: 5’-TCC ATG ACG TTC CTG
ACG TT -3’) with known immunostimulatory
effects on murine immune response (2, 18)
which was provided by Microsynth (Micro-
3. Hoseinjani H, et al.
30 Nanomed J, Vol. 1, No. 1, Autumn 2013
synth, Balgach, Switzerland).
Preparation of cationic nanoliposomes
Liposomes containing DMPC or DSPC
phospholipids, DOTAP and cholesterol were
prepared using lipid film method. Briefly, lipid
film consisting of dimyristoyl phosphatidyl
cholin (DMPC) or distearyl phosphatidyl
cholin (DSPC) (24µmol/ml; Avanti Polar
lipids, USA), 1, 2-dioleoyl-3-trimethylam-
monium propane (DOTAP) (Avanti Polar
lipids, USA) and cholesterol (Avanti Polar
lipids, USA) (3:1:1 molar ratio) were prepared
in a glass vial by evaporating the chloroform:
methanol (2:1, v/v) solution under rotary
evaporation (Heidolph, Germany). The lipid
film was then hydrated and dispersed by
adding required amount of HEPES buffer with
sucrose 10%. The resulting multilamellar
vesicles (MLVs) were sonicated in a bath-type
sonicator (Decon, England) at 50˚C for 15 min
followed by extrusion (Avestin, Canada)
through 400, 100 nm membrane filter to form
100 nm SUVs (small unilamellar vesicles).
CpG ODNs (200µg/ml) or Non CpG ODNs
(200µg/ml) was then added by drop wise
addition to cationic nanoliposomes as it was
swirling gently.
Size distribution and zeta potential analysis of
nanoliposomes
Liposome and LPD nanoparticles size dis-
tribution and zeta potential were measured
using a Zetasizer (Nano-zs, Malvern, UK).
Particle sizes were reported as the means ±
standard deviation and poly dispersity index
(PDI) (n=3). Zeta potentials were reported as
the means ± zeta deviation (n=3).
Inoculation of BALB/c mice
Different groups of mice, 10 mice per group,
were subcutaneously (SC) inoculated in the
left hind footpad (LF) with either of the
following formulations plus parasites (P)
suspension (1 × 106
promastigotes of L. major
harvested at stationary phase in 25µl
buffer/mouse): HEPES buffer (25µl/mouse),
empty DSPC (474.1 µg DSPC, 139.6 µg
DOTAP, 77.2 µg cholesterol/25 µl/mouse),
empty DMPC (406.7 µg DMPC, 139.6 µg
DOTAP, 77.2 µg cholesterol/25 µl/mouse),
DSPC (CpG ODN) (10 µg CpG ODN, 474.1
µg DSPC, 139.6 µg DOTAP, 77.2 µg
cholesterol/25 µl/mouse), DMPC (CpG ODN)
(10 µg CpG ODN, 406.7 µg DMPC, 139.6 µg
DOTAP, 77.2 µg cholesterol/25 µl/mouse),
DSPC (Non CpG ODN) (10 µg Non CpG
ODN, 474.1 µg DSPC, 139.6 µg DOTAP,
77.2 µg cholesterol/25 µl/mouse) or DMPC
(Non CpG ODN) (10µg Non CpG ODN,
406.7 µg DMPC, 139.6 µg DOTAP, 77.2 µg
cholesterol/25 µl/mouse). The progress of
infection was followed by weekly measure-
ment the thickness of the footpad using a
metric caliper (Mitutoyo Measuring Instru-
ments, Japan).
Quantitative parasite burden
The number of viable L. major parasites was
enumerated in the spleen and subiliac lymph
nodes of the mice using limiting dilution assay
method (19). Briefly, the mice were sacrificed
at week 14 post inoculation; the spleens and
lymph nodes were aseptically removed and
homogenized in RPMI 1640 supplemented
with 10% v/v heat inactivated FCS (Eurobio,
Scandinavie), 2 mM glutamine, 100 U/mL of
penicillin and 100 µg/mL of streptomycin
sulfate (RPMI-FCS). The homogenates were
diluted with the same media in 8-serial 10-fold
dilutions and then placed in each well of flat-
bottom 96-well microtiter plates (Nunc)
containing solid layer of rabbit blood agar in
triplicate and kept at 25 ˚C for 7-10 days. The
number of viable parasites per spleen or lymph
node was calculated by ELIDA software, a
statistical method for limiting dilution assay
(20).
Antibody isotype assay
Blood samples were collected from mice at
week 6 post inoculation and serum samples
were analyzed for specific anti-SLA IgG, IgG1
and IgG2a antibodies using ELISA method
(Nunc, Denmark) according to the manu-
facturer’s instructions. Briefly, 96-well micro-
titer plates were coated with 50 µl of SLA (0.5
µg/ml) overnight at 4 °C. Plates were washed
4. Cationic liposomes in lesion development by L. major
Nanomed J, Vol. 1, No. 1, Autumn 2013 31
and blocked for 1 h at 37 °C using 200 µL per
well of 1% bovine serum albumin (BSA) in
PBS-Tween. Serum samples were diluted to
1:200 with PBS-Tween and applied to the
plates. The plates were then treated with HRP-
rabbit anti-mouse IgG isotype according to the
manufacturer’s instructions (Zymed Laborato-
ries Inc., USA). Optical density was deter-
mined at 450 nm using 630 nm as the
reference wavelength.
In vitro spleen cell response
Three mice in each group were sacrificed at
week 14 post inoculation. The spleens were
aseptically removed and a single-cell
suspension was obtained by homogenization
of the tissue. Mononuclear cells were isolated
by Ficoll-Hypaque (Biogene, Iran) density
centrifugation method from spleen cell
suspension. The cells were washed and
resuspended in complete medium (RPMI
1640-FCS) and seeded at 2 × 106
cells/ml in
96-well flat-bottom plates (Nunc, Denmark).
The spleen cells were then stimulated in vitro
in the presence of SLA (5 µg/ml), SLA (10
µg/ml), Concanavalin A (ConA) (2.5 µg/ml),
or medium alone and incubated at 37˚ C with 5
% CO2. Supernatants were collected at 72h of
culture and the concentration of IFN-γ and IL-
4 were assayed using ELISA method
according to the manufacturer’s instructions
(MabTech, Sweden).
Statistical analysis
One-way ANOVA statistical test was used to
analyze the data. In the case of significant F
value, Tukey–Kramer multiple comparisons
test was carried out as a post-test to compare
the means in different groups of mice.
Differences were considered significant when
P < 0.05.
Results
Physical properties of nanoliposomes
Liposomes used in this study were morpho-
logically unilamellar vesicles with mean diam-
eters of 77.6 ± 16.1, 143.2 ± 15.4, 161.5 ±
17.4, 212.9 ± 20.7, 205.6 ± 24.4 and 209.1 ±
19.2 nm (n=3) and the zeta potential of 71.2 ±
21.4, 54.4 ± 13.2, 69.5 ± 11, 48 ± 5.32, 75.7 ±
8.06 and 47.8 ± 3.74 mV, as calculated by
particle size analyzer for empty DSPC, empty
DMPC, DSPC (CpG ODN), DMPC (CpG
ODN), DSPC (Non CpG ODN) and DMPC
(Non CpG ODN), respectively.
Footpad thickness post inoculation
In order to establish the efficacy of cationic
nanoliposomes at the site of L. major
inoculation, the evolution of lesions was
monitored by weekly measurement of footpad
thickness (Figure 1). The group of mice
received DMPC (CpG ODN) along with L.
major showed significantly (P<0.001) the
smallest lesion size compared to untreated
control group (HEPES) throughout the study
period. While the group of mice received
empty DMPC or DMPC (Non CpG ODN)
showed no significant difference compare with
those inoculated with HEPES (P>0.05).
However, there was a significant difference in
lesion size in mice immunized with DSPC
(CpG ODN) and those received HEPES buffer
during 8 weeks post injection (P<0.001).
Moreover, the results of footpad measuring
during 14 weeks showed that there was no
significant difference between mice treated
with DMPC (CpG ODN) and those received
DSPC (CpG ODN) (P>0.05). The results of
footpad measuring showed that there was a
significant difference between mice treated
with empty DSPC and those received HEPES
buffer (P<0.001). However, no significant
difference between the group of mice
immunized with empty DSPC and those
received DSPC (CpG ODN) was observed
during 14 weeks post injection (P>0.05).
Although the group of mice received empty
DSPC showed no significant difference
compare with those inoculated with DMPC
(CpG ODN) during 11 weeks post injection
(P>0.05), later the difference became sign-
5. Hoseinjani H, et al.
32 Nanomed J, Vol. 1, No. 1, Autumn 2013
Figure 1. Footpad swelling in BALB/c mice inoculated subcutaneously (SC) in the left footpad with empty DSPC, empty
DMPC, DSPC (CpG ODN), DMPC (CpG ODN), DSPC (Non CpG ODN), DMPC (Non CpG ODN) or HEPES in
combination with 106
L. major promastigotes. The values represent the mean lesion diameters ± standard error mean (SEM)
(n=10). (***), P<0.001 indicates that the values of mice received DMPC (CpG ODN) or DSPC (CpG ODN) nanoliposomes
are significantly different from those received HEPES.
ificant until the end of measuring period
(P<0.01). The group of mice immunized with
DSPC (Non CpG ODN) showed significant
lower footpad thickness compare with those
inoculated with HEPES buffer during 3 weeks
post injection (P<0.001) but later the differe-
nce became insignificant until the week 8 of
measuring period (P<0.01). The measurement
of footpad swelling was stopped in some
control groups (Figure 1) at week 8 or 9
because the mice had lost their footpads so that
the measurement was not accurate.
Parasite burden
The number of viable L. major parasites was
determined in the subiliac lymph nodes and
spleens of different groups of mice at week 14
post inoculation (Figure 2A and B). As noted,
the group of mice inoculated with DMPC
(CpG ODN) or DSPC (CpG ODN) showed
almost no parasites in the lymph nodes and
spleens compared with the control group
received HEPES (P<0.001). Mice immunized
with empty DSPC, empty DMPC, DSPC (Non
CpG ODN) or DMPC (Non CpG ODN)
showed significantly lower parasite burden
than those received HEPES buffer (P<0.001).
Although, the parasite number in these groups
were higher than the groups of mice
immunized with DMPC (CpG ODN) or DSPC
(CpG ODN), but the differences were not
significant (except for empty DMPC group in
which the significant higher number of
parasites were observed in spleen (P<0.001)
and lymph nodes (P<0.05) comared with
DMPC (CpG ODN) or DSPC (CpG ODN)
groups).
Antibody response
To assess the type of immune response
generated in different groups of mice, serum
levels of anti-SLA specific IgG, IgG1 or
IgG2a antibodies were evaluated at week 6
post infections. As shown in Figure 3A, B and
C, the sera of mice immunized with DSPC
(CpG ODN), DMPC (CpG ODN) and DSPC
(Non CpG ODN) showed a significantly
(P<0.001, P<0.05 and P<0.01 respectively)
lower level of IgG total antibody titer com-
pared with the group received HEPES.
Moreover, the level of IgG1 antibody in the
above groups of mice was significantly
(P<0.001) lower than the control group
received HEPES.
The highest level of IgG1 was observed in sera
of mice received HEPES buffer compared to
other groups. In case of IgG2a, the sera of
mice immunized with DMPC (CpG ODN)
showed no significant difference with the
group received HEPES. Interestingly, the
highest ratio of IgG2a/IgG1 antibodies was
seen in the sera of mice immunized with
DMPC (CpG ODN) com-pared with all other
groups.
6. Cationic liposomes in lesion development by L. major
Nanomed J, Vol. 1, No. 1, Autumn 2013 33
Figure 2. Parasite burden in (A) local draining lymph
nodes and in (B) spleens of BALB/c mice inoculated
subcutaneously (SC) in the left footpad with empty DSPC,
empty DMPC, DSPC (CpG ODN), DMPC (CpG ODN),
DSPC (Non CpG ODN), DMPC (Non CpG ODN) or
HEPES in combination with 106
L. major. A limiting
dilution analysis was performed at week 14 post infection
on the cells isolated from three mice (n=3) and cultured in
the RPMI-FCS for 10 days at 25ºC in serial 8-fold
dilutions. The wells were assessed microscopically for L.
major growth, and the number of viable parasite was
determined by ELIDA software. The values represent
mean ± SD. (***), P<0.001 indicate that the values of mice
received DMPC (CpG ODN) or DSPC (CpG ODN)
nanoliposomes are significantly different from those
received HEPES.
The level of IgG2a antibody in the sera of
mice received DSPC (CpG ODN) or DSPC
(Non CpG ODN) was significantly lower than
those received HEPES (P<0.001). In terms of
the group of mice received empty DSPC or
empty DMPC, the results showed no
significant differences in the level of IgG or
IgG1 antibodies compared with HEPES.
In the case of IgG2a, the sera of mice imm-
unized with empty DSPC showed sign-
ificantly lower level compared with the group
received HEPES (P<0.05), however no sig-
nificant difference between the group of mice
immunized with empty DMPC with those
received HEPES was observed (P>0.05).
In vitro cytokine production by splenocytes
The supernatant of cultured splenocytes
restimulated in vitro at week 14 post inoc-
ulations with SLA to analyze the level of IFN-
Figure 3. The levels of anti-SLA specific IgG1(A), IgG2a
(B) or IgG (C) antibodies based on mean absorbance in the
sera of BALB/c mice inoculated subcutaneously (SC) in
the left footpad with empty DSPC, empty DMPC, DSPC
(CpG ODN), DMPC (CpG ODN), DSPC (Non CpG
ODN), DMPC (Non CpG ODN) or HEPES in
combination with 106
L. major promastigotes. The assays
were performed in triplicate with 200, 2000, 20000 or
200000-fold diluted serum samples. Values represent the
mean ± standard deviation (SD). (*), P<0.05 and (***),
P<0.001 indicate that the values of mice received DMPC
(CpG ODN) or DSPC (CpG ODN) nanoliposomes are
significantly different from the group received HEPES.
γ and IL-4, cytokine markers of Th1 and Th2
immune responses, respectively. As shown in
figure 4A and B, even though DMPC (CpG
ODN) or DSPC (CpG ODN) inoculated mice
indicated a statistically significant reduction in
the level of IL-4 comparing PBS inoculated
mice (P<0.001). Regarding the group of mice
immunized with DMPC (CpG ODN) the level
of IL-4 was comparable to that of DSPC (CpG
ODN) (P>0.05), and also a significant high
7. Hoseinjani H, et al.
34 Nanomed J, Vol. 1, No. 1, Autumn 2013
amount of IFN-γ over the cells compared to
DSPC (CpG ODN) (P<0.001). Although the
group of mice immunized with empty DSPC
showed significant lower level of IL-4
compared with those inoculated with DSPC
(CpG ODN) (P<0.05), the level of IFN-γ was
significantly lower than that of DSPC (CpG
ODN) (P<0.001). In terms of empty DMPC
inoculated mice, although they indicated
comparable level of IFN-γ to DMPC (CpG
ODN) treated ones (P>0.05), they induced a
significantly enhanced level of IL-4 (P<0.01),
too.
Discussion
Induction of protection against future lesion
following recovery from CL was used as a
rationale to inoculate children with materials
from active lesions to induce a lesion, which is
then known as “leishmanization” (LZ) (1). LZ
showed to be the most effective control
measure against CL; However, LZ was stop-
ped due to several reasons such as develop-
ment of large troublesome lesions in 2-3% of
inoculated persons (21). According to the
nature of immune response to the live parasite
vaccines, the current study is mainly focused
on using cationic nanoliposomes containing
CpG ODN adjuvant to explore the possibility
of inducing a Th1 type of immune response
and reduce pathology of L. major infection in
BALB/c mice. Inoculation of mice with
virulent L. major promastigotes mixed with
DMPC (CpG ODN) or DSPC (CpG ODN)
cationic nano-liposomes showed to
induce a significantly smaller lesion size
compared with all other groups. Besides the
kinetic of lesion development, the results also
showed differ-rences in the immune response
generated in group of mice received L. major
mixed with DMPC (CpG ODN) versus the
group of mice received L. major in the other
groups.
As expected, this moderate dermal pathology
was correlated with a substantial
reduction in parasite burden in the spleen and
subiliac lymph nodes of immunized mice as
shown previously (8, 22). Immunostimulatory
activity of CpG motifs was also supported by
Figure 4. Splenic cell response in BALB/c mice
inoculated subcutaneously (SC) in the left footpad with
empty DSPC, empty DMPC, DSPC (CpG ODN), DMPC
(CpG ODN), DSPC (Non CpG ODN), DMPC (Non CpG
ODN) or HEPES in combination with 106
L. major
promastigotes. At week 14 post inoculation, the spleens
were removed and the splenocytes were stimulated in vitro
with either SLA (5 µg/ml), SLA (10 µg/ml), Concanavalin
A (2.5 µg/ml) or medium alone. Amount of IFN-γ (A) and
IL-4 (B) were assessed by ELISA method in supernatants
collected at 72 h of in vitro incubation. Values represent
mean concentration of triplicate assays ± SD. (*), P<0.05,
(**), P<0.01 and (***), P<0.001 indicate that the values of
mice received DMPC (CpG ODN) or DSPC (CpG ODN)
nanoliposomes are significantly different from those
received HEPES.
the observation that applying CpG sequences
at the site of live vaccination with L. major in
C57BL/6 mice enhances primary immunity
and thereby moderates the pathology
associated with L. major infection (23).
Moreover, coadminstration of CpG ODN with
L. major in susceptible BALB/c mice also
induced a significantly smaller lesion size and
a lower death rate compared with the group
which received L. major parasites alone (24).
Inoculation of mice with virulent L. major
promastigotes mixed with LPD (CpG)
nanoparticles also induced a significantly
8. Cationic liposomes in lesion development by L. major
Nanomed J, Vol. 1, No. 1, Autumn 2013 35
smaller lesion size compared with those
received L. major alone.
In the current study, cationic nanoliposomes
containing a neutral phospholipid (DMPC or
DSPC), DOTAP and cholesterol were used as
a delivery system for CpG ODN. DOTAP as a
cationic lipid was used in liposome form-
ulation based on two main purposes. The first
one was to prepare positively charged lipo-
somes which interact more efficiently with the
negatively charged molecules such as CpG
ODNs so increasing the entrapment of CpG
ODN into the liposomes, and the second one
was using the intrinsic adjuvanticity of this
synthetic quaternary ammonium compound
(25).
Liposomes by themselves act as delivery
system, but to further increase the
adjuvanticity of liposomes, immune-
stimulatory adjuvants should be incorporated
into the lipid structures (26). The stronger
immune response is seen when CpG is
encapsulated in an appropriate delivery system
(9). Liposomes protect CpG ODNs from
nuclease activity and hamper the distribution
of CpG ODNs to tissue, consequently
increasing the CpG ODNs half-life (27). At the
APCs level, CpG ODNs through TLR9,
augment activation and maturation of DC as
well as the induction of proinflammatory
cytokines (28). Thus, the endogenous prod-
uction of IL-12, IL-18, and other soluble
mediators from activated DC induced by CpG
ODNs are likely to result in a more
physiologic cognate interaction between the
DC and T cell, resulting in both a qualitatively
and quantitatively different type of CD4+
and
CD8+
T cell response (22). The current results
elucidated the effect of the presence of both
cationic lipid and CpG for full immune-
stimulation activity required for counteracting
the drawbacks of L. major inoculation. As
compared with control groups, DMPC (CpG
ODN) demonstrated a significant control on
the progression of the growth of the footpad
and desired immune response.
To develop a leishmanization protocol with a
mild lesion, co-administration of live parasites
with neutral liposomes containing CpG ODN
was studied before (23, 24).
Interestingly, the presence of CpG ODN
decreased lesion size and there was no
significant difference between mice receiving
CpG ODN in liposomal form and those
receiving CpG ODN in soluble form,
particularly when DSPC was used as neutral
lipid. However, it was not true for the
liposomes consisting DMPC (Fig. 1). It might
be due to the transition temperature of lipid
used in liposome formulation (i.e. DSPC). As
DSPC has a very high transition temperature
(Tm 55 ºC) and produces a very rigid and
stable bilayer structure in liposome
formulation which resists releasing a suitable
amount of CpG ODN for interaction with
TLR9 receptor located in the phagosomes (24)
while DMPC has low Tm (23 ºC) was not true
for. Previously, when we used 1,2-dioleoyl-3-
trimethylammoniumpropane (DOTAP) (Tm
∼0 ºC) in LPD nanoparticles, the results
showed a significant difference in protection
compared with CpG ODN in soluble (10). The
difference may be correlated with the
availability of the free form of CpG ODN in
phagosomes due to the very low Tm of the
lipid component. On the other hand, the
nanoparticles were destabilized more easily in
phagosomes and as a result CpG ODN is
available for interaction with the receptors.
The level of both IFN-γ and IL-4 cytokines
was high in HEPES received group due to the
inoculation of parasite alone without any
adjuvants, whereas mice inoculated with
DMPC (CpG ODN) or DSPC (CpG ODN)
plus parasites significantly suppressed the IL-4
production. Although DMPC (CpG ODN)
inoculated mice indicated a comparable level
of IL-4 compared to DSPC (CpG ODN)
treated ones (P>0.05), but they induced a
significantly enhanced level of IFN-γ
(P<0.001). The highest level of IFN-γ was
detected in mice treated with HEPES that it
might not be due to the induction of Th1
response, because of the high amount of IL-4,
IgG1, and IgG titer observed in this group.
Moreover, generalization of infection to the
spleen was detected by parasite burden
experiment. The same trend was reported
when LPD nanoparticles was used in
combination with L. major (10). In group of
9. Hoseinjani H, et al.
36 Nanomed J, Vol. 1, No. 1, Autumn 2013
mice received DMPC (CpG ODN), though the
level of IFN- γ was lower than those received
HEPES, the significant lower IL-4 production
and the highest IgG2a/IgG1 ratio was also
seen. Altogether, the results of cytokine and
Ab production analysis show that DMPC
(CpG ODN) nanoliposomes prevent to induce
a Th2 type of response compared to other
groups.
It is proposed that inoculation of live parasites
into BALB/c mice induces a specific IgG
response in the absence of adjuvant, with a
preponderance of the IgG1 isotype (a marker
of Th2 response) (29).
In this study, DMPC (CpG ODN)
nanoliposomes enhanced the production of
IgG2a and particularly IgG2a/ IgG1 ratio, as a
hallmark of Th1 type of immune response
which is essential for counteracting
intracellular pathogens (6).Taken together, the
current results suggested that immune
modulation using DMPC (CpG ODN) cationic
nanoliposomes might be a practical approach
to improve the safety of inoculation of live L.
major and can be utilized instead of LPD
nanoparticles.
Conclusion
In conclusion, the current results suggested
that immune modulation using DMPC (CpG
ODN) cationic nanoliposomes might be a
practical approach to improve the safety of
inoculation of live L. major and can be utilized
instead of LPD nanoparticles.
There are limited studies using live
Leishmania parasites concurrently with
adjuvants, thus, further studies are needed to
explore the detailed role of co-administration
of cationic nanoliposomes with live
Leishmania.
Acknowledgments
The financial support of the Nanotechnology
Research Center and Biotechnology Research
Center, Mashhad University of Medical Sciences
are gratefully acknowledged.
This study was part of Pharm. D. dissertation of
HA that was done in Nanotechnology Research
Center, MUMS, Iran.
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