1. The study examined the effects of repeated restraint stress on Peyer's patches (PPs), which are lymphoid tissues in the intestine that play a key role in immune responses.
2. It found that repeated restraint stress did not modify the morphological structure of PPs, but it did reduce the total number of lymphocytes, CD8+ T cells, B cells, and plasma cells in PPs.
3. Specifically, restraint stress reduced the number of IgA-producing plasma cells in the dome region of PPs, where these cells are usually most numerous. Since IgA protects against intestinal infections, repeated stress may increase susceptibility to pathogens.
Increase your Understanding of the Pathogenesis of Gluten Spectrum DisordersCell Science Systems
Recently, researchers at Harvard University, Alessio Fasano et. al., and the National Institutes of Health (laboratories of immunology and cellular and molecular biology), reported real-time microscopic observations of gluten-induced neutrophil activation.
According to authors, " To what extent neutrophil function adds to, or protects against, gluten intolerance is currently under vigorous investigation."
This presentation will shed light on this question. It will also review the Fasano study and examine the role of neutrophil function in multiple disease conditions, as well as explore how neutrophil function may also play a dual role in protecting the body from the untoward effects of dietary and environmental agents.
This document discusses the use of animal models in biomedical research. It describes how animals are used to study human disease and test potential treatments in order to advance human health without risking harm to people. Common animal models mentioned include mice, rats, dogs, primates, and rabbits, which can provide insights into conditions like heart disease, cancer, neurological disorders, and infectious diseases. However, the use of animals in research also raises ethical issues, as it often involves invasive procedures that can cause pain and distress. Large numbers of animals are required for activities like vaccine production and drug testing, but only a small percentage of potential treatments ultimately succeed.
This document provides an overview of animal models used in periodontal research. It discusses the definition and history of animal models, the need for animal models in periodontal research given limitations of human studies, and various categories and classifications of animal models. The document then examines specific animal models used in periodontal research, including rats, mice, and hamsters, describing their anatomy, how periodontal disease presents in each, and advantages and limitations of each model.
Kinsenoside isolated from Anoectochilus formosanus suppresses LPS-Stimulated ...Cây thuốc Việt
In the present study, we reported that kinsenoside, a major component of Anoectochilus formosanus, inhibited inflammatory reactions in mouse peritoneal lavage macrophages and protects mice from endotoxin shock. In LPSstimulated mouse peritoneal lavage macrophages, kinsenoside inhibited the inflammatory mediators, such as nitric oxide, TNF-!, IL-1", monocyte chemoattractant protein 1, and macrophage migration inhibitory factor production. Furthermore, kinsenoside decreased the formation of a nuclear factor .BYDNA complex and nuclear p65 and p50 protein levels. Kinsenoside inhibited nuclear factor .B translocation through both I.B!-dependent and -independent pathway. In contrast, it stimulated anti-inflammatory cytokine IL-10 generation and enhanced the mRNA expression of IL-10 and suppressor of cytokine signaling 3 in the same cells induced by LPS. In an animal model, both pretreatment and posttreatment of kinsenoside increased the survival rate of ICR mice challenged by LPS (80 mg/kg, i.p.). Pretreatment with kinsenoside decreased serum levels of TNF-!, IL-1", IL-10, monocyte chemoattractant protein 1, and migration inhibitory factor at 1 h after sublethal dose of LPS (40 mg/kg, i.p.) in mice. In contrast, kinsenoside enhanced serum IL-10 level at 24 h after LPS injection in mice. In conclusion, kinsenoside inhibited the production of inflammatory mediators and enhanced antiinflammatory cytokine generation. Therefore, kinsenoside can alleviate acute inflammatory hazards.
A study on the toxic effect of different doses of Diclofenac sodium on the de...Prof. Hesham N. Mustafa
The toxic effects of different doses of diclofenac sodium (DS) on the kidney on the postnatal period (0-7 days) by morphometrical and immunohistochemical methods were investigated. For this purpose, 15 female adult wistar albino rats were used and divided into 5 main groups. Group Ia served as normal control, physiologic group Ib received normal saline, group II received low dose (3.9 mg/kg), group III received medium dose (9 mg/kg) and group IV received high dose (18 mg/kg). Male offspring’s from 0-7 days after birth were used in this study. On the 8th day of postnatal life, all animals were anesthetized. Then, the kidney samples were analyzed. Haematoxylin and eosin staining showed degeneration and necrosis, apparent atrophy of the glomeruli, mononuclear cell infiltration, congested vessels, increased fibrous tissue and distortion of the proximal convoluted tubules with interruption of the brush margin of the DS treated group. Increased level of Caspase-3 and upregulation of TNF-α with different doses of DS. In light of our findings, DS may lead to adverse effects that are dose-dependent in the prenatal subjected kidney to this drug.
Keywords : Diclofenac sodium; Proximal convoluted tubules; Apoptosis; Cyclooxygenase.
Animal models are important tools in toxicological and biomedical research. Regulations aim to ensure animal welfare while enabling scientific progress. Key points of regulations include:
- Licensing of research facilities and oversight by ethics committees.
- Focus on replacing, reducing and refining animal use (3Rs principle).
- Standards for humane care and treatment of research animals.
- Requirements vary by country but most have adopted versions of the 3Rs and facility licensing with inspections. Self-regulation is common but some places have more direct legal oversight.
This study analyzed diagnostic records from the Taiwan National Laboratory Animal Center from 2004 to 2007 to assess the health status of rodent colonies in Taiwan. The key findings were:
1) Demand for pathogen diagnostic services increased steadily each year over the study period, indicating growing awareness of animal health issues.
2) Many mouse colonies tested positive for pathogens such as mouse parvovirus, mouse hepatitis virus, Theiler murine encephalomyelitis virus, and Mycoplasma pulmonis.
3) Nearly 40% of rat colonies tested positive for Mycoplasma pulmonis and rat parvovirus, and some also contained viruses like Kilham rat virus or intestinal parasites.
4
Increase your Understanding of the Pathogenesis of Gluten Spectrum DisordersCell Science Systems
Recently, researchers at Harvard University, Alessio Fasano et. al., and the National Institutes of Health (laboratories of immunology and cellular and molecular biology), reported real-time microscopic observations of gluten-induced neutrophil activation.
According to authors, " To what extent neutrophil function adds to, or protects against, gluten intolerance is currently under vigorous investigation."
This presentation will shed light on this question. It will also review the Fasano study and examine the role of neutrophil function in multiple disease conditions, as well as explore how neutrophil function may also play a dual role in protecting the body from the untoward effects of dietary and environmental agents.
This document discusses the use of animal models in biomedical research. It describes how animals are used to study human disease and test potential treatments in order to advance human health without risking harm to people. Common animal models mentioned include mice, rats, dogs, primates, and rabbits, which can provide insights into conditions like heart disease, cancer, neurological disorders, and infectious diseases. However, the use of animals in research also raises ethical issues, as it often involves invasive procedures that can cause pain and distress. Large numbers of animals are required for activities like vaccine production and drug testing, but only a small percentage of potential treatments ultimately succeed.
This document provides an overview of animal models used in periodontal research. It discusses the definition and history of animal models, the need for animal models in periodontal research given limitations of human studies, and various categories and classifications of animal models. The document then examines specific animal models used in periodontal research, including rats, mice, and hamsters, describing their anatomy, how periodontal disease presents in each, and advantages and limitations of each model.
Kinsenoside isolated from Anoectochilus formosanus suppresses LPS-Stimulated ...Cây thuốc Việt
In the present study, we reported that kinsenoside, a major component of Anoectochilus formosanus, inhibited inflammatory reactions in mouse peritoneal lavage macrophages and protects mice from endotoxin shock. In LPSstimulated mouse peritoneal lavage macrophages, kinsenoside inhibited the inflammatory mediators, such as nitric oxide, TNF-!, IL-1", monocyte chemoattractant protein 1, and macrophage migration inhibitory factor production. Furthermore, kinsenoside decreased the formation of a nuclear factor .BYDNA complex and nuclear p65 and p50 protein levels. Kinsenoside inhibited nuclear factor .B translocation through both I.B!-dependent and -independent pathway. In contrast, it stimulated anti-inflammatory cytokine IL-10 generation and enhanced the mRNA expression of IL-10 and suppressor of cytokine signaling 3 in the same cells induced by LPS. In an animal model, both pretreatment and posttreatment of kinsenoside increased the survival rate of ICR mice challenged by LPS (80 mg/kg, i.p.). Pretreatment with kinsenoside decreased serum levels of TNF-!, IL-1", IL-10, monocyte chemoattractant protein 1, and migration inhibitory factor at 1 h after sublethal dose of LPS (40 mg/kg, i.p.) in mice. In contrast, kinsenoside enhanced serum IL-10 level at 24 h after LPS injection in mice. In conclusion, kinsenoside inhibited the production of inflammatory mediators and enhanced antiinflammatory cytokine generation. Therefore, kinsenoside can alleviate acute inflammatory hazards.
A study on the toxic effect of different doses of Diclofenac sodium on the de...Prof. Hesham N. Mustafa
The toxic effects of different doses of diclofenac sodium (DS) on the kidney on the postnatal period (0-7 days) by morphometrical and immunohistochemical methods were investigated. For this purpose, 15 female adult wistar albino rats were used and divided into 5 main groups. Group Ia served as normal control, physiologic group Ib received normal saline, group II received low dose (3.9 mg/kg), group III received medium dose (9 mg/kg) and group IV received high dose (18 mg/kg). Male offspring’s from 0-7 days after birth were used in this study. On the 8th day of postnatal life, all animals were anesthetized. Then, the kidney samples were analyzed. Haematoxylin and eosin staining showed degeneration and necrosis, apparent atrophy of the glomeruli, mononuclear cell infiltration, congested vessels, increased fibrous tissue and distortion of the proximal convoluted tubules with interruption of the brush margin of the DS treated group. Increased level of Caspase-3 and upregulation of TNF-α with different doses of DS. In light of our findings, DS may lead to adverse effects that are dose-dependent in the prenatal subjected kidney to this drug.
Keywords : Diclofenac sodium; Proximal convoluted tubules; Apoptosis; Cyclooxygenase.
Animal models are important tools in toxicological and biomedical research. Regulations aim to ensure animal welfare while enabling scientific progress. Key points of regulations include:
- Licensing of research facilities and oversight by ethics committees.
- Focus on replacing, reducing and refining animal use (3Rs principle).
- Standards for humane care and treatment of research animals.
- Requirements vary by country but most have adopted versions of the 3Rs and facility licensing with inspections. Self-regulation is common but some places have more direct legal oversight.
This study analyzed diagnostic records from the Taiwan National Laboratory Animal Center from 2004 to 2007 to assess the health status of rodent colonies in Taiwan. The key findings were:
1) Demand for pathogen diagnostic services increased steadily each year over the study period, indicating growing awareness of animal health issues.
2) Many mouse colonies tested positive for pathogens such as mouse parvovirus, mouse hepatitis virus, Theiler murine encephalomyelitis virus, and Mycoplasma pulmonis.
3) Nearly 40% of rat colonies tested positive for Mycoplasma pulmonis and rat parvovirus, and some also contained viruses like Kilham rat virus or intestinal parasites.
4
This study established a modified immunohistochemistry method using murine antiserum as the primary antibody to detect mouse hepatitis virus (MHV) and Mycoplasma pulmonis antigens in formalin-fixed tissue sections. Using this method, MHV antigen was detected in the liver, stomach, caecum, colon and spleen of infected mice. Mycoplasma pulmonis antigen was demonstrated on the luminal surface of bronchioles in infected rats. This technique provides a useful method for diagnosing MHV and M. pulmonis infections when commercial antibodies are unavailable or serological diagnosis is not possible due to immunosuppression.
This document discusses various animal models used for research including invertebrate models like Drosophila and C. elegans, rodent models, rabbit models, and large animal models. These models are used to study processes like genetics, development, and disease due to their similarities to humans. Drosophila and C. elegans have been important for discoveries in development and genetics. Rodent models are widely used due to their similarities to humans and short lifespans. Larger animal models are needed for pre-clinical research due to closer mimicry of human physiology. A variety of animal models at different sizes are essential for advancing biomedical research.
This document summarizes a study that investigated the expression of melatonin receptor proteins (MT1 and MT2) in the thyroid gland of male mice at different ages, and measured levels of thyroxine (T4) and thyrotropin (TSH) hormones. The study found that aged mice had decreased levels of serum T4 but unchanged levels of serum TSH compared to young mice. Expression of both MT1 and MT2 melatonin receptor proteins in the thyroid gland decreased with age. This suggests that aging modulates the responsiveness of the thyroid gland to melatonin through changes in melatonin receptor expression, which may contribute to declining thyroid function in older organisms.
Journal of Feline Medicine and Surgery (2012). Congress AbstractsEmily Wallinder
The document summarizes several clinical/research abstracts that were accepted for presentation at the inaugural poster session of the International Society of Feline Medicine Congress in 2012. Specifically, it provides brief summaries of 3 abstracts:
1) A case report of a cat with feline gastrointestinal eosinophilic sclerosing fibrolasia associated with zygomycete fungi.
2) A study finding that the renal toxin binder AST-120 was well tolerated and accepted by cats when mixed with food, even at high doses.
3) A clinical trial finding that probiotic therapy reduced relapse risk in cats treated with ronidazole for Tritrichomonas foetus-associated diarrhea.
1) The study examined the ability of Lactobacillus plantarum MON03 (LP), Tunisian montmorillonite clay (TM), and their composite to remove the mycotoxin zearalenone (ZEA) in vitro and prevent its immunotoxic effects in mice.
2) Results showed that both TM and the LP+TM composite highly adsorbed ZEA (87.2% and 94.2% respectively) after 24 hours, while LP alone removed 78%. The amount of ZEA adsorbed was highest for the LP+TM composite.
3) In mice exposed to ZEA, treatment with LP, TM, or their composite prevented the immunotoxic effects of ZEA
This document describes a microarray analysis comparing gene expression profiles in the large intestine, small intestine, liver, and spleen of mice with different gut microbiota colonization models: specific pathogen-free mice, germ-free mice colonized at birth, and germ-free mice colonized at 5 weeks of age. The analysis found hundreds of differentially expressed genes in each tissue and colonization model. Gene set enrichment analysis identified overrepresented gene ontology categories related to immune system development and antigen presentation in intestines of mice colonized at birth, and metabolic processes in intestines of specific pathogen-free mice. Analysis of signaling pathways found prominent changes in toll-like receptor and type 1 interferon signaling pathways in intestines of mice
The study characterized the Campylobacter jejuni IAL 2383 strain isolated from humans in Brazil. They found that the strain harbored important virulence genes and expressed major virulence factor transcripts. It grew better at 41°C than 37°C, indicating ability to colonize avian hosts. The strain was sensitive to most antibiotics tested and could serve as an experimental model for interactions with host cells and acquisition of antibiotic resistance.
1) The study evaluated the antidepressant, anxiolytic, anticonvulsant, and sedative effects of standardized extracts of flavonoids from Byrsonima crassifolia in mice.
2) The methanolic extract produced a significant antidepressant effect in the forced swimming test at a dose of 500 mg/kg but did not show anxiolytic, sedative, or anticonvulsant effects.
3) Although the main compound in the extract was identified as quercetin 3-O-xyloside, flavonoids such as rutin, quercetin, and hesperidin may be involved in the antidepressant effects.
- Oral insulin capsules developed by Oramed Pharmaceuticals were found to successfully lower blood sugar levels in human clinical trials. The company is now conducting a larger 90-day study to further evaluate the capsules' effects on HbA1c and their potential as a safer, more convenient alternative to injected insulin.
- Scientists at the John Innes Centre identified the last missing genes in the Madagascar periwinkle plant that allow it to produce the important cancer-fighting alkaloids vinblastine and vincristine. Understanding these genes could help increase sustainable production of these drugs through plant or synthetic biology techniques.
- Researchers at multiple institutions reported developments that could help advance cancer treatment, including
The document summarizes a study that tested a novel antifungal drug (Drug A) in a murine model of invasive pulmonary aspergillosis. Mice were infected with Aspergillus fumigatus and then received various doses of Drug A or a positive control, Posaconazole. The mice were divided into groups for assessing fungal burden or survival. Higher doses of Drug A and Posaconazole reduced fungal counts in the lungs, showing the drug's antifungal activity. However, Drug A unexpectedly increased mouse mortality compared to controls, suggesting it may be toxic. The results point to an experimental error requiring the study to be repeated.
This document discusses the history and types of animal experimentation. It notes that Aristotle and Erasistratus were among the first to use living animals in experiments. It outlines the types of animal research including basic research, applied research, toxicology testing, and xenotransplantation. Common animal models used are rats, mice, rabbits, and guinea pigs. The document also discusses the principles of replacement, reduction and refinement of animal experiments and the ethical requirements for conducting such research.
Experiment modelling of Auto-immune diseasesPratik Parikh
The document discusses experimental animal models of autoimmune diseases. It describes how animal models can help understand the pathogenesis and potential treatment of human autoimmune conditions since their immune systems share many similarities to humans. Some commonly used spontaneous animal models include the obese strain chicken for Hashimoto's thyroiditis and the non-obese diabetic mouse for type 1 diabetes. Induced models include the experimental autoimmune encephalomyelitis model in mice and rats for multiple sclerosis. Proper animal models are selected based on their relevance to human diseases and characteristics like ease of handling and rapid reproduction.
The study investigated the in vitro anti-inflammatory activity of an aqueous leaf extract of Vitex negundo. Rat peritoneal cells and erythrocytes were used to study the effects. The extract inhibited nitric oxide production by rat peritoneal cells in a dose-dependent manner and also stabilized erythrocyte membranes, as shown by dose-dependent inhibition of heat-induced hemolysis. Higher concentrations of the extract were cytotoxic to rat peritoneal cells, while lower concentrations showed no cytotoxicity. The results suggest that the extract's anti-inflammatory activity is due to inhibition of nitric oxide production by immune cells and membrane stabilizing effects.
This document summarizes a proteomic analysis of the excretory/secretory (ES) proteins of Toxoplasma gondii, the causative agent of toxoplasmosis. 34 proteins were identified from the ES proteins of the RH strain of T. gondii using LC/MS/MS analysis and spectral counting methods. The three most abundant proteins identified were GRA1, GRA7, and GRA2. A variety of other proteins were also identified, including micronemal proteins, rhoptry proteins, and dense granule proteins, which play important roles in T. gondii host cell invasion and survival within host cells. This analysis provides new insights into the ES proteome of T
Screening of immunomodulatory activity of Sphaeranthus indicus Linn. whole plantiosrjce
This document summarizes a study that evaluated the immunomodulatory activity of the methanolic extract of Sphaeranthus indicus Linn. (MESI) whole plant in rats. The study assessed the effects of MESI at doses of 100, 200, and 400 mg/kg on humoral immunity (antibody titers, plaque forming cells), cellular immunity (delayed type hypersensitivity, T-cell populations), and myelosuppression. MESI showed significant increases in circulating antibody titers, plaque forming cells, delayed type hypersensitivity responses, and T-cell populations compared to control, indicating immunostimulatory effects. The results suggest that Sphaeranthus indicus has potential as
Screening models for testing of immunological factorsKundlik Rathod
This document discusses screening models for testing immunological factors. It describes both in vitro and in vivo methods. The in vitro methods covered include inhibition of histamine release from mast cells, neutrophil locomotion and chemotaxis assays, and using cell lines like THP-1 monocytes. The in vivo methods covered are using murine models to study humoral antibody response, assessing delayed type hypersensitivity reaction, and measuring macrophage phagocytosis using the carbon clearance test. The goal of these screening models is to test substances that can modulate the immune system.
The document is a poem describing the various microhabitats within the human body that are suitable for commensal microbes to colonize, including pores, arm pits, and scalp. It welcomes microbes to build colonies and supplies them warmth, moisture, and nutrients, on the condition they do not cause issues like acne or athlete's foot. The poem acknowledges microbes as guests in the body on this new year's day.
Animal models like inbred mouse strains and techniques like adoptive transfer are useful for immunology research by reducing genetic variability. Inbred strains are genetically identical due to inbreeding, allowing the study of immune responses without interference from individual genetic differences. Adoptive transfer involves transferring lymphocytes from a donor mouse into an irradiated recipient mouse to study the immune response in isolation. SCID mice that lack mature immune cells accept grafts from other species, enabling the study of human immune system development within a mouse model through the use of SCID-human mice implanted with human tissues.
This study investigated the effects of L-alpha-glycerylphosphorylcholine (GPC), a deacylated derivative of phosphatidylcholine (PC), in a rat model of small intestinal ischemia-reperfusion injury. The results showed that:
1) Intestinal ischemia-reperfusion increased oxidative stress markers, microcirculatory dysfunction, and liver ATP depletion.
2) Both pre-treatment and post-treatment with GPC significantly reduced oxidative stress markers, protected microcirculation, and alleviated hepatic ATP depletion caused by ischemia-reperfusion.
3) GPC therapies were effective in attenuating the inflammatory response to ischemia-reperfusion injury, providing indirect
This document summarizes key advances in neurogastroenterology and motility research from 2011. Three main points are:
1) Studies showed that gut microbes and nutrients can affect mood and food intake through the vagus nerve and endocannabinoid signaling. Stress was also found to exacerbate visceral pain through changes in primary afferent neurons and spinal glia.
2) Two studies provided evidence that enteric glia can generate new neurons in the gut after injury, indicating they may serve as neuronal precursors.
3) Research found that neuronal serotonin protects the enteric nervous system and regulates motility and inflammation, while mucosal serotonin contributes to visceral pain. A new
Explore the cell's role in mediating adverse reactions 7 c09Paul Thiessen
This document discusses the role of neutrophils and macrophages in mediating various physiological and pathological processes. It summarizes several scientific studies that found:
1) Neutrophils can be recruited by substances like gliadin and mediate local inflammatory responses in tissues like the intestine.
2) Neutrophils and macrophages produce reactive molecules that can damage cells and tissues, and their activation levels correlate with conditions like infertility and acute coronary syndrome.
3) Chronic activation of the innate immune system by these cells may underlie metabolic syndrome by stimulating inflammation and hormonal changes.
4) Oxidative stress can increase blood levels of modified lipids implicated in atherosclerosis, coinciding with increased neutrophil counts.
This study established a modified immunohistochemistry method using murine antiserum as the primary antibody to detect mouse hepatitis virus (MHV) and Mycoplasma pulmonis antigens in formalin-fixed tissue sections. Using this method, MHV antigen was detected in the liver, stomach, caecum, colon and spleen of infected mice. Mycoplasma pulmonis antigen was demonstrated on the luminal surface of bronchioles in infected rats. This technique provides a useful method for diagnosing MHV and M. pulmonis infections when commercial antibodies are unavailable or serological diagnosis is not possible due to immunosuppression.
This document discusses various animal models used for research including invertebrate models like Drosophila and C. elegans, rodent models, rabbit models, and large animal models. These models are used to study processes like genetics, development, and disease due to their similarities to humans. Drosophila and C. elegans have been important for discoveries in development and genetics. Rodent models are widely used due to their similarities to humans and short lifespans. Larger animal models are needed for pre-clinical research due to closer mimicry of human physiology. A variety of animal models at different sizes are essential for advancing biomedical research.
This document summarizes a study that investigated the expression of melatonin receptor proteins (MT1 and MT2) in the thyroid gland of male mice at different ages, and measured levels of thyroxine (T4) and thyrotropin (TSH) hormones. The study found that aged mice had decreased levels of serum T4 but unchanged levels of serum TSH compared to young mice. Expression of both MT1 and MT2 melatonin receptor proteins in the thyroid gland decreased with age. This suggests that aging modulates the responsiveness of the thyroid gland to melatonin through changes in melatonin receptor expression, which may contribute to declining thyroid function in older organisms.
Journal of Feline Medicine and Surgery (2012). Congress AbstractsEmily Wallinder
The document summarizes several clinical/research abstracts that were accepted for presentation at the inaugural poster session of the International Society of Feline Medicine Congress in 2012. Specifically, it provides brief summaries of 3 abstracts:
1) A case report of a cat with feline gastrointestinal eosinophilic sclerosing fibrolasia associated with zygomycete fungi.
2) A study finding that the renal toxin binder AST-120 was well tolerated and accepted by cats when mixed with food, even at high doses.
3) A clinical trial finding that probiotic therapy reduced relapse risk in cats treated with ronidazole for Tritrichomonas foetus-associated diarrhea.
1) The study examined the ability of Lactobacillus plantarum MON03 (LP), Tunisian montmorillonite clay (TM), and their composite to remove the mycotoxin zearalenone (ZEA) in vitro and prevent its immunotoxic effects in mice.
2) Results showed that both TM and the LP+TM composite highly adsorbed ZEA (87.2% and 94.2% respectively) after 24 hours, while LP alone removed 78%. The amount of ZEA adsorbed was highest for the LP+TM composite.
3) In mice exposed to ZEA, treatment with LP, TM, or their composite prevented the immunotoxic effects of ZEA
This document describes a microarray analysis comparing gene expression profiles in the large intestine, small intestine, liver, and spleen of mice with different gut microbiota colonization models: specific pathogen-free mice, germ-free mice colonized at birth, and germ-free mice colonized at 5 weeks of age. The analysis found hundreds of differentially expressed genes in each tissue and colonization model. Gene set enrichment analysis identified overrepresented gene ontology categories related to immune system development and antigen presentation in intestines of mice colonized at birth, and metabolic processes in intestines of specific pathogen-free mice. Analysis of signaling pathways found prominent changes in toll-like receptor and type 1 interferon signaling pathways in intestines of mice
The study characterized the Campylobacter jejuni IAL 2383 strain isolated from humans in Brazil. They found that the strain harbored important virulence genes and expressed major virulence factor transcripts. It grew better at 41°C than 37°C, indicating ability to colonize avian hosts. The strain was sensitive to most antibiotics tested and could serve as an experimental model for interactions with host cells and acquisition of antibiotic resistance.
1) The study evaluated the antidepressant, anxiolytic, anticonvulsant, and sedative effects of standardized extracts of flavonoids from Byrsonima crassifolia in mice.
2) The methanolic extract produced a significant antidepressant effect in the forced swimming test at a dose of 500 mg/kg but did not show anxiolytic, sedative, or anticonvulsant effects.
3) Although the main compound in the extract was identified as quercetin 3-O-xyloside, flavonoids such as rutin, quercetin, and hesperidin may be involved in the antidepressant effects.
- Oral insulin capsules developed by Oramed Pharmaceuticals were found to successfully lower blood sugar levels in human clinical trials. The company is now conducting a larger 90-day study to further evaluate the capsules' effects on HbA1c and their potential as a safer, more convenient alternative to injected insulin.
- Scientists at the John Innes Centre identified the last missing genes in the Madagascar periwinkle plant that allow it to produce the important cancer-fighting alkaloids vinblastine and vincristine. Understanding these genes could help increase sustainable production of these drugs through plant or synthetic biology techniques.
- Researchers at multiple institutions reported developments that could help advance cancer treatment, including
The document summarizes a study that tested a novel antifungal drug (Drug A) in a murine model of invasive pulmonary aspergillosis. Mice were infected with Aspergillus fumigatus and then received various doses of Drug A or a positive control, Posaconazole. The mice were divided into groups for assessing fungal burden or survival. Higher doses of Drug A and Posaconazole reduced fungal counts in the lungs, showing the drug's antifungal activity. However, Drug A unexpectedly increased mouse mortality compared to controls, suggesting it may be toxic. The results point to an experimental error requiring the study to be repeated.
This document discusses the history and types of animal experimentation. It notes that Aristotle and Erasistratus were among the first to use living animals in experiments. It outlines the types of animal research including basic research, applied research, toxicology testing, and xenotransplantation. Common animal models used are rats, mice, rabbits, and guinea pigs. The document also discusses the principles of replacement, reduction and refinement of animal experiments and the ethical requirements for conducting such research.
Experiment modelling of Auto-immune diseasesPratik Parikh
The document discusses experimental animal models of autoimmune diseases. It describes how animal models can help understand the pathogenesis and potential treatment of human autoimmune conditions since their immune systems share many similarities to humans. Some commonly used spontaneous animal models include the obese strain chicken for Hashimoto's thyroiditis and the non-obese diabetic mouse for type 1 diabetes. Induced models include the experimental autoimmune encephalomyelitis model in mice and rats for multiple sclerosis. Proper animal models are selected based on their relevance to human diseases and characteristics like ease of handling and rapid reproduction.
The study investigated the in vitro anti-inflammatory activity of an aqueous leaf extract of Vitex negundo. Rat peritoneal cells and erythrocytes were used to study the effects. The extract inhibited nitric oxide production by rat peritoneal cells in a dose-dependent manner and also stabilized erythrocyte membranes, as shown by dose-dependent inhibition of heat-induced hemolysis. Higher concentrations of the extract were cytotoxic to rat peritoneal cells, while lower concentrations showed no cytotoxicity. The results suggest that the extract's anti-inflammatory activity is due to inhibition of nitric oxide production by immune cells and membrane stabilizing effects.
This document summarizes a proteomic analysis of the excretory/secretory (ES) proteins of Toxoplasma gondii, the causative agent of toxoplasmosis. 34 proteins were identified from the ES proteins of the RH strain of T. gondii using LC/MS/MS analysis and spectral counting methods. The three most abundant proteins identified were GRA1, GRA7, and GRA2. A variety of other proteins were also identified, including micronemal proteins, rhoptry proteins, and dense granule proteins, which play important roles in T. gondii host cell invasion and survival within host cells. This analysis provides new insights into the ES proteome of T
Screening of immunomodulatory activity of Sphaeranthus indicus Linn. whole plantiosrjce
This document summarizes a study that evaluated the immunomodulatory activity of the methanolic extract of Sphaeranthus indicus Linn. (MESI) whole plant in rats. The study assessed the effects of MESI at doses of 100, 200, and 400 mg/kg on humoral immunity (antibody titers, plaque forming cells), cellular immunity (delayed type hypersensitivity, T-cell populations), and myelosuppression. MESI showed significant increases in circulating antibody titers, plaque forming cells, delayed type hypersensitivity responses, and T-cell populations compared to control, indicating immunostimulatory effects. The results suggest that Sphaeranthus indicus has potential as
Screening models for testing of immunological factorsKundlik Rathod
This document discusses screening models for testing immunological factors. It describes both in vitro and in vivo methods. The in vitro methods covered include inhibition of histamine release from mast cells, neutrophil locomotion and chemotaxis assays, and using cell lines like THP-1 monocytes. The in vivo methods covered are using murine models to study humoral antibody response, assessing delayed type hypersensitivity reaction, and measuring macrophage phagocytosis using the carbon clearance test. The goal of these screening models is to test substances that can modulate the immune system.
The document is a poem describing the various microhabitats within the human body that are suitable for commensal microbes to colonize, including pores, arm pits, and scalp. It welcomes microbes to build colonies and supplies them warmth, moisture, and nutrients, on the condition they do not cause issues like acne or athlete's foot. The poem acknowledges microbes as guests in the body on this new year's day.
Animal models like inbred mouse strains and techniques like adoptive transfer are useful for immunology research by reducing genetic variability. Inbred strains are genetically identical due to inbreeding, allowing the study of immune responses without interference from individual genetic differences. Adoptive transfer involves transferring lymphocytes from a donor mouse into an irradiated recipient mouse to study the immune response in isolation. SCID mice that lack mature immune cells accept grafts from other species, enabling the study of human immune system development within a mouse model through the use of SCID-human mice implanted with human tissues.
This study investigated the effects of L-alpha-glycerylphosphorylcholine (GPC), a deacylated derivative of phosphatidylcholine (PC), in a rat model of small intestinal ischemia-reperfusion injury. The results showed that:
1) Intestinal ischemia-reperfusion increased oxidative stress markers, microcirculatory dysfunction, and liver ATP depletion.
2) Both pre-treatment and post-treatment with GPC significantly reduced oxidative stress markers, protected microcirculation, and alleviated hepatic ATP depletion caused by ischemia-reperfusion.
3) GPC therapies were effective in attenuating the inflammatory response to ischemia-reperfusion injury, providing indirect
This document summarizes key advances in neurogastroenterology and motility research from 2011. Three main points are:
1) Studies showed that gut microbes and nutrients can affect mood and food intake through the vagus nerve and endocannabinoid signaling. Stress was also found to exacerbate visceral pain through changes in primary afferent neurons and spinal glia.
2) Two studies provided evidence that enteric glia can generate new neurons in the gut after injury, indicating they may serve as neuronal precursors.
3) Research found that neuronal serotonin protects the enteric nervous system and regulates motility and inflammation, while mucosal serotonin contributes to visceral pain. A new
Explore the cell's role in mediating adverse reactions 7 c09Paul Thiessen
This document discusses the role of neutrophils and macrophages in mediating various physiological and pathological processes. It summarizes several scientific studies that found:
1) Neutrophils can be recruited by substances like gliadin and mediate local inflammatory responses in tissues like the intestine.
2) Neutrophils and macrophages produce reactive molecules that can damage cells and tissues, and their activation levels correlate with conditions like infertility and acute coronary syndrome.
3) Chronic activation of the innate immune system by these cells may underlie metabolic syndrome by stimulating inflammation and hormonal changes.
4) Oxidative stress can increase blood levels of modified lipids implicated in atherosclerosis, coinciding with increased neutrophil counts.
1) The study investigated the effects of gasdermin D on pyroptosis in a mouse model of sepsis-induced acute kidney injury.
2) The results showed that gasdermin D expression was increased in mice with sepsis-induced acute kidney injury and promoted inflammation and pyroptosis in kidney cells.
3) Downregulating gasdermin D decreased inflammation and pyroptosis, and the NLRP3 inflammasome was identified as an important target of gasdermin D in mediating inflammation during sepsis-induced acute kidney injury.
The current slide focuses on different screening models for neurodegenerative diseases along with a brief description of the diseases where the slides are to the points and brief with detailed evaluation.
This study investigated the role of autophagy on human retinal pigment epithelial (RPE) cell viability and apoptosis under oxidative stress. RPE cells were divided into control, H2O2, and H2O2+3-MA groups. H2O2 treatment activated autophagy and increased apoptosis while decreasing cell viability. Inhibition of autophagy with 3-MA decreased apoptosis. The results suggest that autophagy is involved in H2O2-induced apoptosis of RPE cells under oxidative stress conditions.
Morphological and functional state of immune organs in rats with experimental...QUESTJOURNAL
This study examined the morphological and functional changes in the immune organs (thymus and spleen) of rats with experimentally induced type 1 diabetes mellitus (DM-1). Rats with DM-1 showed initial pathological changes in these organs, including increased medulla in the thymus and increased white pulp in the spleen, indicating early inflammatory and degenerative processes. Treatment with the phytopreparation BNO 10.30 was found to help restore cell structure in the thymus and spleen by stimulating immune function. The results demonstrate the importance of early detection of immune organ changes in patients with type 1 diabetes.
The current study investigated the immunomodulatory
potential of ethyl acetate soluble supernatant of
Lactobacillus casei (LC-EAS) in vitro. The effect of
LC-EAS on nitric oxide release was analyzed in RAW
264.7 cells, wherein, an inhibition in nitric oxide production
through suppression of inducible nitric oxide synthase
mRNA expression was observed. Evaluation of LC-EAS
on LPS-induced peripheral blood mononuclear cells
showed a down-regulation in TNF-a and IL-6 genes and an
upregulation of IL-10. An inhibition in the protein
expression of NF-kB, ERK1/2 and STAT3 phosphorylation
confirms the immunomodulatory potential of LC-EAS. The
effect of LC-EAS on in vitro intestinal epithelial cells was
investigated using HT-29 human colon adenocarcinoma
cancer cells. LC-EAS exhibited an inhibition of NF-jB and
ERK1/2 phosphorylation, whereas STAT3 phosphorylation
was unregulated. To evaluate the downstream target of
STAT3 upregulation, expression of the intestinal trefoil
factor TFF3 which is a NF-jB regulator and STAT3
downstream target was studied. LC-EAS was observed to
elevate TFF3 mRNA expression. Overall the study shows
that the anti-inflammatory potential of LC-EAS is through
inhibition of NF-kB in different cell types.
1) A study investigated the vasodilatory and toxic effects of a crude extract of Ruta graveolens (Ruta) on rat aortas and CRL1730 endothelial cells.
2) The Ruta extract generated vasodilation in rat aortas at subtoxic concentrations, partially dependent on the endothelium. It caused a loss of cell viability in CRL1730 cells at high concentrations but did not induce oxidative stress or DNA fragmentation.
3) The results suggest Ruta extract regulates vascular tone through a complex, partially endothelium-dependent mechanism and has vasodilatory activity at subtoxic levels without damaging cell membranes or viability.
This document summarizes a study that investigated the effects of melatonin on inflammatory responses induced by Prevotella intermedia lipopolysaccharide (LPS) in murine macrophages. The study found that melatonin suppressed the LPS-induced production of nitric oxide and interleukin-6 by inhibiting the NF-κB and STAT1 signaling pathways. Melatonin blocked NF-κB signaling by inhibiting the nuclear translocation and DNA-binding activity of the NF-κB p50 subunit. This suggests that melatonin may help reduce host inflammatory responses associated with periodontal disease by modulating these signaling pathways and inflammatory mediators.
Mice lacking the integrin β3 gene (Itgb3-/-) were subjected to unpredictable chronic mild stress (UCMS) to examine their behavioral and neurochemical responses to stress compared to wild-type mice (Itgb3+/+). Itgb3-/- mice displayed increased anxiety-like behaviors and decreased activity in an open field test after UCMS compared to Itgb3+/+ mice. UCMS led to reductions in dopamine turnover in the midbrains of Itgb3+/+ mice but not Itgb3-/- mice, suggesting disrupted stress regulation of dopamine homeostasis. Chronic stress also altered synaptic expression of proteins in the midbrains of Itgb3-/- mice compared to Itgb3+/+
The successful of pregnancy in humans and rodents occur between the interaction maternal and fetal
interface, specially involving the participation of uNK cells. This interaction involved neo angiogenesis,
placentation and presence of mediators like nitric oxide. During the pregnancy the administration of LPS
in the dams can results in necrosis, preterm birth, IUGR, miscarriage or neurological problem. Once the
uNK cells are activated, they can produce vasodilators, like NO. So, the main purpose of this study was
to evaluate if LPS cause alteration in the uNK cells in pregnant mice and if the same behaviour can be
detected by NO in the blood. Also we evaluated the effect of LPS to cause neurological injuries. To do that
we used pregnant mice on gd 10th and those was treated with LPS for different times. Uterine samples
were collected at 0.5,1,2 and 6hr after LPS treated and processed for paraffin embedding and tissue
homogenate. The samples designated for paraffin embedding was performed the Dolichos biflorus (DBA)
lectin cytochemistry and anti-iNOS immunocytochemistry. The samples designated to tissue homogenates
were processed for SDS-PAGE and Western-blot using anti-iNOS and evaluate of NO concentration. We
found after 2h LPS exposure the mice showed fever and low capacity to explore different environment.
At the same time, we found increase in the nitrate/nitrito ratio in a dose dependent manner in the uterus
after 2h LPS exposure.
This study investigated the effects of neuropeptide Y (NPY) on signaling proteins in C. elegans. NPY is a neurotransmitter that regulates anxiety in humans. The researchers extracted proteins from C. elegans exposed to NPY and analyzed them using ELISA. They found a statistically significant upregulation of Akt1 protein levels in the presence of NPY, but no significant effects on other proteins like p38 MAP kinase and Stat3. This suggests NPY may activate Akt1 signaling in C. elegans. The results help validate the proposed pathway of NPY's anxiolytic effects.
This study investigated the effects of neuropeptide Y (NPY) on signaling proteins in C. elegans. NPY is a neurotransmitter that regulates anxiety in humans. The researchers extracted proteins from C. elegans exposed to NPY and analyzed them using ELISA. They found a statistically significant upregulation of Akt1 protein levels in the presence of NPY, but no significant effects on other proteins like p38 MAP kinase and Stat3. This suggests NPY specifically increases Akt1, a regulator of cell growth and survival, in C. elegans. The results help validate the proposed pathway of NPY's anxiolytic effects.
This study investigated the effects of neuropeptide Y (NPY) on signaling proteins in C. elegans. NPY is a neurotransmitter that regulates anxiety in humans. The researchers extracted proteins from C. elegans exposed to NPY and analyzed them using ELISA. They found a statistically significant upregulation of Akt1 protein levels in the presence of NPY, but no significant effects on other proteins like p38 MAP kinase and Stat3. This suggests NPY may upregulate Akt1 through a CRE element in its gene. Further experiments are needed to validate the proposed signaling pathway and determine if NPY increases phospho-CREB levels.
Topical Delivery of Fenoprofen Proliposomes: Preparation, Evaluation and In V...inventionjournals
International Journal of Pharmaceutical Science Invention (IJPSI) is an international journal intended for professionals and researchers in all fields of Pahrmaceutical Science. IJPSI publishes research articles and reviews within the whole field Pharmacy and Pharmaceutical Science, new teaching methods, assessment, validation and the impact of new technologies and it will continue to provide information on the latest trends and developments in this ever-expanding subject. The publications of papers are selected through double peer reviewed to ensure originality, relevance, and readability. The articles published in our journal can be accessed online.
This study examined neurons in the medulla oblongata related to gastric mucosal lesions in rats subjected to restraint water-immersion stress (RWIS). The study found that compared to controls, RWIS rats had: 1) increased cholinergic neurons in the dorsal motor nucleus of the vagus and nucleus ambiguous, and increased catecholaminergic neurons in the nucleus of the solitary tract; 2) increased oxytocin receptor- and vasopressin 1b receptor-expressing neurons in the dorsal motor nucleus and nucleus of the solitary tract; but 3) no difference in methionine-enkephalin-expressing neurons in the nucleus of the solitary tract. This suggests hyperactivity of cholinergic and cate
International Journal of Pharmaceutical Science Invention (IJPSI)inventionjournals
International Journal of Pharmaceutical Science Invention (IJPSI) is an international journal intended for professionals and researchers in all fields of Pahrmaceutical Science. IJPSI publishes research articles and reviews within the whole field Pharmacy and Pharmaceutical Science, new teaching methods, assessment, validation and the impact of new technologies and it will continue to provide information on the latest trends and developments in this ever-expanding subject. The publications of papers are selected through double peer reviewed to ensure originality, relevance, and readability. The articles published in our journal can be accessed online
Background: It is often difficult to predict which newborn with HIE will develop neurological sequlae so there is an urgent need for predictors for adverse neurological outcomes in these infants. Aim of Study: To evaluate the serum levels of serum amyloid A (SAA) protein in newborns with HIE during the first week of life and after 3 and 6 months of follow up to assess its correlation with degree of HIE neurological sequlee. Patients and Methods; This case-control study was conducted on 72 infants; group (1) included 36 full term neonates diagnosed as HIE and group (2)included 36 age and sex matched, infants as a control group, Serum amyloid A by ELIZA technique was measured at post natal age of 1 and 7 days, CT scan was done in justified cases .with follow up at age of 3 and 6 months for neurological sequlee. Results: SAA protein level was elevated in the asphyxiated group in comparison to the control group at day 1 and day 7, SAA level was significantly correlated to the Sarnat scoring system of HIE. SAA level significantly differ on follow up of developmental milestone at age of 3 and 6 months. ROC curve for validity of SAA for severity of HIE at cut off point > 25μg/ml at day 1 and at cut off point > 20 μg/ml at day 7 of HIE diagnosis reported sensitivity 100% and specificity 100% .Conclusion: SAA correlates with the severity of HIE and higher SAA expression is a prognostic marker for morbidity in these infants.
Similar to 2011 repeated restraint stress reduces the ig a producing cells in peyers patches (20)
This article examines the effects of repeated restraint stress on the structure and function of the nasal-associated lymphoid tissue (NALT) in mice. The study found that 4 days of restraint stress decreased the percentage of CD3+ and CD4+ T cells in the NALT as well as IgA+ cells and nasal IgA levels, while 8 days of stress increased the numbers of these cells and IgA levels compared to the control group. Repeated restraint stress was found to selectively affect immune cells in the nasal mucosa and basal IgA production.
Este documento presenta la información de contacto y las líneas de investigación de la Dra. Luvia Enid Sánchez Torres en el Posgrado de Inmunología de la Escuela Nacional de Ciencias Biológicas. Sus líneas de investigación incluyen la respuesta inmunológica a la malaria, la participación de la apoptosis en diferentes patologías y la evaluación de la actividad antineoplásica de fitoquímicos y compuestos sintéticos. También incluye cinco de sus publicaciones más recientes sobre estos temas.
Cas IIgly induces apoptosis in glioma C6 cells through both caspase-dependent and caspase-independent pathways. When exposed to Cas IIgly, glioma C6 cells showed inhibited proliferation, increased reactive oxygen species formation, and induced apoptosis in a dose-dependent manner. Both the mitochondria-nuclear translocation of apoptosis induction factor and endonuclease G, as well as the fragmentation of nucleosomal DNA and caspase-3 activation were involved in the apoptotic effects of Cas IIgly. Reactive oxygen species formation induced by Cas IIgly may also contribute to its apoptotic effects. Administration of Cas IIgly to glioma-bearing rats reduced tumor size and increased apoptosis.
The study examines apoptosis in mouse splenic T cell and B cell populations during infection with Plasmodium chabaudi chabaudi AS malaria. High levels of apoptosis were found to correlate with high parasitemia and splenomegaly, particularly in CD4+ T cells. Apoptosis levels decreased as parasitemia was cleared but remained elevated compared to normal mice, with CD8+ T cells and B cells returning to basal apoptosis levels while CD4+ T cells remained higher.
This document summarizes research testing the ability of eight diindolylmethane derivatives to induce apoptosis in murine L5178Y lymphoma cells. The derivatives were synthesized with different substituted phenyl groups attached to the methane carbon. Testing found compound 3a, with a meta-hydroxyl group, was the most active, inhibiting 93% of cell growth and inducing 71.04% apoptosis. In general, substituents able to form hydrogen bonds and in the meta position were most effective at arresting the cell cycle. The preliminary results provide insight into how the substituent and position impact potency against lymphoma cells.
This article examines the cytotoxic effects of perezone and its isomer isoperezone on human leukemia cells. The key findings are:
1. Both perezone and isoperezone induce cytotoxicity in a concentration-dependent manner through both caspase-dependent and caspase-independent mechanisms.
2. The compounds cause changes in cell size, granularity, phosphatidylserine translocation, mitochondrial membrane potential disruption, and do not induce apoptosis inducing factor.
3. Perezone showed greater cytotoxic effects than isoperezone, but the mechanisms of cell death induction differed between the two compounds depending on concentration.
This article examines the effects of repeated restraint stress on the structure and function of the nasal-associated lymphoid tissue (NALT) in mice. The study found that 4 days of restraint stress decreased the percentage of CD3+ and CD4+ T cells in the NALT as well as IgA+ cells and nasal IgA levels, while 8 days of stress increased the numbers of these cells and IgA levels compared to the control group. Repeated restraint stress was found to selectively affect immune cells in the nasal mucosa and basal IgA production.
La Dra. Luvia Enid Sánchez Torres es una investigadora de nivel SNI I en el Laboratorio de Inmunología Celular y de Inmunología de los Microorganismos. Sus líneas de generación y aplicación del conocimiento incluyen la inmunología de las enfermedades infecciosas, vacunas e inmunoterapia. Algunas de sus líneas de investigación son la muerte celular, la evaluación de compuestos naturales y sintéticos, y la respuesta inmune contra parásitos.
Strategies for Effective Upskilling is a presentation by Chinwendu Peace in a Your Skill Boost Masterclass organisation by the Excellence Foundation for South Sudan on 08th and 09th June 2024 from 1 PM to 3 PM on each day.
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𝐃𝐢𝐬𝐜𝐮𝐬𝐬 𝐭𝐡𝐞 𝐄𝐏𝐏 𝐂𝐮𝐫𝐫𝐢𝐜𝐮𝐥𝐮𝐦 𝐢𝐧 𝐭𝐡𝐞 𝐏𝐡𝐢𝐥𝐢𝐩𝐩𝐢𝐧𝐞𝐬:
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𝐄𝐱𝐩𝐥𝐚𝐢𝐧 𝐭𝐡𝐞 𝐍𝐚𝐭𝐮𝐫𝐞 𝐚𝐧𝐝 𝐒𝐜𝐨𝐩𝐞 𝐨𝐟 𝐚𝐧 𝐄𝐧𝐭𝐫𝐞𝐩𝐫𝐞𝐧𝐞𝐮𝐫:
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বিসিএস ও ব্যাংক এর লিখিত পরীক্ষা ...+এছাড়া মাধ্যমিক ও উচ্চমাধ্যমিকের স্টুডেন্টদের জন্য অনেক কাজে আসবে ...
This document provides an overview of wound healing, its functions, stages, mechanisms, factors affecting it, and complications.
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There are 4 phases of wound healing: hemostasis, inflammation, proliferation, and remodeling. This document also describes the mechanism of wound healing. Factors that affect healing include infection, uncontrolled diabetes, poor nutrition, age, anemia, the presence of foreign bodies, etc.
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Walmart Business+ and Spark Good for Nonprofits.pdfTechSoup
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Special TechSoup offer for a free 180 days membership, and up to $150 in discounts on eligible orders.
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Answers about how you can do more with Walmart!"
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How to Setup Warehouse & Location in Odoo 17 Inventory
2011 repeated restraint stress reduces the ig a producing cells in peyers patches
1. 1
REPEATED RESTRAINT STRESS REDUCES THE IgA PRODUCING CELLS IN
PEYER’S PATCHES
Beatriz Elina Martínez-Carrillo1, Marycarmen Godinez-Victoria1,2, Adriana Jarillo-Luna3,
Rigoberto Oros-Pantoja1, Edgar Abarca-Rojano4, Víctor Rivera-Aguilar5, Judith Pacheco-
Yepez6, Luvia Enid Sánchez-Torres2 and Rafael Campos-Rodríguez1*.
1
Departamento de Bioquímica y Sección de Estudios de Posgrado e Investigación, Escuela
Superior de Medicina-IPN, Salvador Díaz Mirón y Plan de San Luis s/n, Colonia Santo
Tomás, México, D.F., C.P. 11340, México.
2
Departamento de Inmunología, Escuela Nacional de Ciencias Biológicas-IPN, Prol.
Carpio y Plan de Ayala s/n, Colonia Santo Tomás, México, D.F., C.P. 11340, México
3
Departamento de Morfología, Escuela Superior de Medicina-IPN, Salvador Díaz Mirón y
Plan de San Luis s/n, Colonia Santo Tomás, México, D.F., C.P. 11340, México.
4
Laboratorio de Inmunobiología del Endotelio, Escuela Superior de Medicina-IPN,
Salvador Díaz Mirón y Plan de San Luis s/n, Colonia Santo Tomás, México, D.F., C.P.
11340, México.
5
Departamento de Microbiología, Unidad de Biología Tecnología y Prototipos (UBIPRO),
Facultad de Estudios Superiores (FES)-Iztacala, Universidad Nacional Autónoma de
México, Avenida de los Barrios s/n, Tlalnepantla Estado. de México. CP. 54090, México,
D. F.
2. 2
6
Laboratorio de Microscopía Electrónica, Facultad de Medicina, Universidad La Salle
Fuentes 17, Tlalpan, CP. 14000, México, D. F.
*Corresponding author: Rafael Campos-Rodríguez PhD. Escuela Superior de Medicina-
IPN, Salvador Díaz Mirón y Plan de San Luis s/n, Colonia Santo Tomás, México, D.F.,
C.P. 11340, México. Tel. +52(55) 57482004; Fax. +52(55) 57145455. E-mail:
citli@prodigy.net.mx
3. 3
Abstract
The few reports that analyze the effects of stress on the immune cells of the intestinal
mucosa or the functions of these cells tend to focus on S-IgA levels in saliva, and these
studies have shown contradictory results. The principal objective of this study was to
analyze the effects of repeated restraint stress on the number and distribution of immune
cells of Peyer’s patches (PPs), and also the effects of glucocorticoid and catecholamine
administration on the same stress-related parameters. Upon analyzing the effect of repeated
restraint stress on PPs, it was found that there was no modification in the morphological
structure of the PPs, but that restraint stress reduced the total number of lymphocytes, the
number of CD8+ T cells, B cells and plasma cells in PPs. Only in the site of PPs where
IgA-producing plasma cells are most numerous (the dome) was a decrease found in this
type of cell. These effects were due at least in part to the effect of glucocorticoids and
catecholamines. Since IgA produced in the Peyer’s patches is a natural antibody that
impedes bacterial infections, repeated stress may favor the entrance of pathogens through
the intestine.
Keywords: Repeated restraint stress, Peyer’s patches, IgA, glucocorticoids,
catecholamines.
4. 4
Introduction
The neuroendocrine system regulates the immune responses, including those of the
intestinal mucosa [1, 2]. Studies done on the neuroendocrine regulation of the mucosal
immune system have focused mainly on the quantity of IgA secreted in saliva [3, 4], the
synthesis and secretion of the secretory component of the tear glands [5, 6], and the
regulation of the synthesis of the secretory component in the female reproductive apparatus
[7, 8]. On the other hand, studies done on the effects of stress on the mucosal immune
system have focused almost exclusively in relation to inflammatory diseases of the intestine
and the secretion of IgA in saliva.
The abundant information available confirms that psychological stress plays a fundamental
role in the physiopathology and clinical symptoms of intestinal inflammatory diseases in
humans [9, 10]. However, in relation to IgA levels in human saliva, both reductions and
increases have been reported, depending on the type of psychological and/or physical stress
protocol employed [4, 11, 12]. These contradictions make it difficult to reach any
conclusion about the effects of stress on the humoral immune response of the mucosas,
which is represented by secretory IgA (sIgA) levels. Our recent studies show that restraint
stress reduces IgA levels in mouse intestine as well as the intraepithelial lymphocyte
population in mouse duodenal mucosa [13, 14].
The immune system of the mucosa can be divided into inductor and effector sites. In the
former, consisting of principally of lymphoid tissue in PPs, the appendix and solitary nodes
[15, 16], the antigens captured from the mucosa surface stimulate a response from T and B
lymphocytes. In the latter, the effector cells perform their action, such as the production of
5. 5
S-IgA [15, 17]. PPs, the most studied inductor site, have fundamental importance in the
capture of antigens in the intestinal lumen and in the induction of the humoral immune
response, mainly through the production of S-IgA, in the intestine [18, 19]. It is known that
PPs are innervated by nerve fibers containing norepinephrine, vasoactive intestinal
polypeptide (VIP), substance P (SP) and somatostatin (SOM) [20, 21], and that the
lymphocytes of PPs express receptors for neuropeptides (e.g., SOM and SP). In vitro
studies have demonstrated that the production of IgA, IgG and IgM by PP lymphocytes is
regulated by β-endorphin, ACTH and various peptides, which also regulate the migration
and proliferative response of lymphocytes [21].
Stress-related effects on the structure and functions of PPs have been done in the context of
GALT, with a focus on changes in T lymphocyte populations [22, 23]. Since the
neuroendocrine regulation of the function of inductor sites has been little studied, the aim
of the current study was to analyze the effects of repeated restraint stress on the
morphological structure as well as the number and percentage of lymphocytes, including
the IgA-producing plasma cells, in the different regions of Peyer´s patches, and to compare
these stress-related effects with those produced by treatment with glucocorticoids and
catecholamines.
6. 6
Materials and Methods
2.1 Animals
Two-month old male Balb/c mice (from the animal house of the Escuela Superior de
Medicina, ESM), weighing 25 to 30 g, were treated with the appropriate dose of
mebendazole and metronidazole to eliminate parasites. They were housed 6 per cage in a
room with little noise and kept on a 12:12 h light/dark cycle (lights on at 6 am). All
handling and assays were carried out between 8 am and 12 am to avoid the influence of the
circadian cycles of ACTH and cortisol. The mice were handled in accordance with the
norms of the Institutional Commission for the Care and Use of Lab Animals of the ESM.
2.2 Restraint stress protocol
Mice were placed in cylindrical plexiglass containers 9 cm long and 3.5 cm in diameter,
with many ventilation holes to prevent hyperthermia (n = 6). Animals could move to back
and forward freely in the container, but could not turn around. The duration of the restraint
cycle was 1 or 4 h for four successive days. Unrestrained mice were left undisturbed in
their home cages without access to food or water during the same period.
2.3 Hormonal treatment
Groups of 6 mice were treated subcutaneously with epinephrine at 0.1 or 0.5 mg/kg/day for
4 consecutive days. Other groups of 6 mice were treated subcutaneously with
dexamethasone at 5 or 50 mg/kg/day, also for 4 consecutive days. Control mice (n = 6)
received only the vehicle (NaCl 0.89%).
2.4 Isolation of Peyer’s patches and purification of lymphocytes
7. 7
On the fourth day, all animals were sacrificed and the small intestine was extracted and
washed with PBS. The segments that contained Peyer’s patches were separated with fine
dissection scissors, obtaining 3 to 5 patches, depending on the mouse. One PP from the
proximal segment was fixed in 4% formaldehyde and processed for cuts on a wax block,
while another was frozen in isopentane and stored at -70°C until use. The rest of the PPs
were used to quantify the number of lymphocytes by flow cytometry.
Briefly, PPs that were not fixed or frozen were immediately disaggregated and resuspended
in Hank´s balanced salt solution (HBSS). The cell suspension was filled with gauze twice,
with the aim of eliminating epithelial cells and remaining tissue. After that, suspension cells
were washed three times with the same solution, then the button cells were resuspended in
1 mL of HBSS to conduct a count. The total number of cells in each patch was counted by
employing a violet crystal solution, while the viability was evaluated by Trypan blue
exclusion analysis. Finally, the suspension was adjusted to a concentration of 10x106
cells/mL of HBSS.
2.5 Reverse Hemolytic Plaque Assay
The reverse hemolytic assay for the detection of IgA was adapted from a previously
described method [24, 25]. Briefly 0.5 mL of washed and packed sheep erythrocytes
(SRBC) was incubated with 0.1 mL solution of 10 mg rabbit anti-mouse IgA antibody
dissolved in 1 mL of PBS, used as a coupler, and 0.5 mL of CrCl3 solution (1 mg/mL in
0.15 M NaCl) at 37oC for 1 hr with continuous shaking. One-tenth milliliter of cell
suspension of PPs in HBSS was mixed with 20 μL of 20% v/v suspension in 0.1 5 M NaCl
of freshly anti-IgA-coupled SRBC, 20 μL of the serum of rabbit anti-mouse IgA
8. 8
(“developer”, diluted 1:150), and 20 μL of guinea pig serum (complement source). The
mixture was incubated in Cunningham´s chambers at 37oC for 90 min, then the hemolytic
plates were counted and the mean of IgA producing cells per million viable cells was
calculated.
2.6 Topographic staining
The PPs fixed in formaldehyde were cut in slices of 7 µm and stained with Hematoxiline-
Eosine (H-E) and Gomori's trichomic technique for morphological analysis.
2.7 In situ detection of lymphocytes by immunohistochemical techniques
The PPs frozen in isopentane were cut in slices of 7 µm. The cells were stained using an
immunohistochemical technique with monoclonal biotynilated antibodies specific for
identifying T CD4+ and T CD8+ cells, followed by the application of streptavidin
conjugated with HPR. To detect the IgA-producing plasma cells in situ, a monoclonal
antibody to the heavy α-chain of mice and an antibody conjugated to HPR were used.
Finally, the reactions were revealed for 10 min with diaminobenzidine and counterstained
with Harris’ hematoxylin. After dehydration, cells were covered with synthetic resin and
counted by tissue area, each area being measured by an ocular micrometer calibrated with a
hemocytometer.
2.8 Immunophenotyping by flow cytometry
For cell immunophenotyping, directly labeled monoclonal antibodies were used: anti-
CD19-APC or -PE, CD45-PercCP, CD138-APC, IgA-FITC, CD3-FITC, CD8-PE and
CD4-PercCP (all from BD Biosciences, San Jose, CA, USA).
9. 9
Cells were harvested, washed twice with PBS and 0.5% BSA, and then stained for T cell
phenotype with a cocktail of anti-CD3, -CD4 and -CD8 mAb, or for B cell phenotype using
anti-B220 and anti-CD19 mAb, for 30 min at room temperature in darkness. The cells were
then washed with PBS and fixed in 1 % formaldehyde in PBS. IgA-producing plasma cells
(CD138+ cells) and B cells (CD19+/B220+ cells) were fixed, permeabilized and stained
according to BD Bioscience’s protocol for intracellular staining. The fluorescent signal
intensity was recorded and analyzed in a FACSCalibur flow cytometer (Becton Dickinson).
For each sample 15,000 events were collected. Data were analyzed using the Summit
software v4.3 (Dako, Colorado Inc.). The total number of lymphocytes was calculated from
the percentage of cells located in the lymphocyte region in the dot-plot of FSC vs SSC, and
the total number of cell/patch according to the following formula: (# total cells/Peyer´s
patch) x (% lymphocytes)/100. The absolute number of positive cells (subsets of
lymphocytes) was calculated from the total number of lymphocytes, according to the
following formula: (# total cells/Peyer´s patch) x (% positive cells)/100. The percentage
and number of CD4+ cells and CD8+ cells were calculated from the CD3+ cells.
2.9 Statistical Analysis
The differences between two groups were determined by the Student’s t test. The analysis
of data from 3 or more groups was done with one-way ANOVA. All values were presented
as the mean ± SD of at least three independent assays. Statistical analyses were performed
by using the statistical program Sigma Stat for Windows Version 2.03 software (SPSS Inc).
A P-value equal or less than 0.05 was considered statistically significant.
10. 10
Results
3.1 Restraint stress did not modify the morphology of PPs
It is well established that repeated stress in the short or long run modifies the number and
function of immune and inflammatory cells [26, 27]. However, there have been no studies
on the effects of repeated stress on PPs of the mouse intestinal mucosa. Since PPs have
fundamental importance in the capture of antigens in the intestinal lumen and in the
induction of the humoral immune response, we evaluated the structure as well as the
number and percentage of lymphocytes in this tissue. We observed that the normal structure
of the germinal center, internodal regions and dome remained intact in the PPs of stressed
mice (Figure 1). Also, there was no significant difference in this parameter between the
mice stressed for 1 or 4 h.
3.2. Restraint stress modified the cellular composition of PPs
The percentage of B cells was significantly lower in mice stressed for 4 h than in the other
two groups: animals stressed for 1 h or those non-stressed (Figure 2A, P < 0.001), as
detected by flow cytometry. There were no differences in the percentages of the other
subsets of lymphoid cells, including plasma cells, among the three groups. However, when
the absolute number of each cellular subset was determined, the total number of
lymphocytes in the PPs was found to be lower in both groups of stressed mice than the
control animals (Figure 2B, P < 0.001, Bonferroni t-test). The absolute number of CD8+ T
cells, B cells and plasma cells was significantly lower in mice stressed for 4 h than the other
two groups: animals stressed for 1 h or those non-stressed (Figure 2B, P < 0.001). Among
these parameters, only plasma cells were found to be lower in mice stressed for 1 h than in
11. 11
control animals (P < 0.05). The total number of T cells and CD4+ T cells was not affected
by the restraint stress protocol employed.
3.3. Restraint stress reduced the number of IgA-producing plasma cells
Considering that Peyer’s patches contain antibody-producing effector cells [28-30], we
evaluated the effect of repeated restraint stress on the total number of IgA-producing
plasma cells in PPs, as well as the number of these cells in each region. The
immunohistochemical assay demonstrated that repeated restraint stress diminished the
number of IgA-producing plasma cells in the dome, but not in the corona, germinal center
or intermodal region of PPs (Figure 3A and 3B; P < 0.001, Bonferroni t-test). The flow
cytometric analysis confirmed that the total number of IgA-producing plasma cells
(CD138+/IgA+) in PPs was lower in mice stressed for 1 and 4 h than in the control animals
(Figure 3C, P < 0.001). However, when we determined the percentage of IgA-producing
plasma cells, the only significant difference between the three groups was the lower
percentage of these cells found in mice stressed for 1h compared to the control animals
(Figure 3D).
3.4. Effects of dexamethasone and epinephrine
In several studies it has been demonstrated that changes observed in the immune response
induced by stress are mediated principally through the release of glucocorticoids and
catecholamines in different kinds of tissues. Therefore in the present study the effect of
dexamethasone (a glucocorticoid) and epinephrine (a catecholamine) on the number of
lymphocytes and subsets of T and B cells in PPs was evaluated.
12. 12
The dose of 5 mg dexamethasone reduced the percentage of CD4+ T cells (Figure 4A, P <
0.001), while increasing that of CD8+ T cells (P < 0.001) and plasma cells (P < 0.05). The
dose of 50 mg increased the percentage of CD8+ T cells (P < 0.001) and plasma cells (P <
0.05). Not only the percentage but also the number of cells was evaluated. Dexamethasone
at doses of 5 and 50 mg/kg/day significantly reduced: (i) the size of the patches, in which
no germinal centers were found (Figure 1C), (ii) the total number of lymphocytes (Figure
4A, P < 0.001), (iii) the number of all subsets of T cells (Figure 4A, CD3+/CD4+, *P <
0.001; and CD3+/CD8+ cells, **P < 0.05), and (iv) the number of B cells (*P < 0.001;
Bonferroni t-test). However, only the higher dose of dexamethasone reduced the number of
plasma cells (Figure 4A P < 0.001).
In relation to epinephrine, the structure of the PPs remained normal (Figure 1D) and there
were no differences in the percentage of T cells, CD4+ T cells and plasma cells among the
three groups of mice (Figure 5A). The dose of 0.5 mg increased the percentage of CD8+ T
cells (**P < 0.05) and both doses reduced the percentage of B cells (*P < 0.001). There
were no significant changes in the percentages of the other cell subpopulations.
Regarding the absolute number of cells, both doses of epinephrine (0.1 and 0.5 mg/kg/day)
significantly reduced the total number of lymphocytes and the different subpopulations of
lymphocytes, including T cells and subsets of T cells, B cells, and plasma cells (Figure 5B,
*P < 0.001, **P < 0.05, Bonferroni t-test).
Dexamethasone at doses of 5 mg and 50 mg significantly reduced the absolute number of
IgA-producing plasma cells in PPs, as determined by immunohistochemistry (Figure 6A;
*P < 0.01). In agreement with these results, flow cytometry also showed that both doses of
13. 13
dexamethasone decreased the number of IgA-producing plasma cells (Figure 6B, **P <
0.05) as well as their percentage, compared to the control animals (Figure 6C, *P < 0.001).
Epinephrine at the higher dose (0.5 mg/Kg/day) significantly reduced the number of IgA-
producing plasma cells in PPs, as detected by immunohistochemistry (Figure 6D; *P <
0.01). However, no significant change was found with the lower dose (0.1 mg/Kg/day).
When using flow cytometry, both doses of epinephrine were found to reduce the number of
IgA-producing plasma cells (Figure 6E, **P < 0.05), whereas only the 0.1 mg/Kg/day dose
reduced the percentage of IgA-producing plasma cells (Figure 6F, P < 0.001).
3.5. The reverse hemolytic assay
To confirm the findings of immunohistochemistry and flow cytometry, we performed a
functional assay, the reverse hemolytic assay, to quantify IgA antibody secreting cells (IgA-
SCs). This test confirmed that the number of IgA-SCs was significantly lower in stressed
animals (1 or 4 hours) compared to the control group. Also, it confirmed that both doses of
dexamethasone (*P < 0.001) and the higher dose of epinephrine (0.5 mg/Kg/day)
significantly reduced the number of IgA-SCs in PPs (Figure 7; *P < 0.001, **P < 0.05).
14. 14
Discussion
In some lymphoid organs, such as the thymus, spleen, and nodes, diverse types of stress
cause atrophy due to a notable reduction in the number of lymphocytes [26, 27, 31].
Contrarily, in relation to the protocol of the present study, whether applied for 1 or 4 h
restraint stress did not cause atrophy of the PPs (Figure 1B), although it did indeed result in
a decrease in the total number of lymphocytes (Figure 2B).Similarly, both doses of
epinephrine administered in the current contribution caused a decrease of the total number
of lymphocytes in the PPs (Figure 5B) without producing atrophy in this tissue (Figure 1D).
However, both doses of dexamethasone caused atrophy of PPs (Figure 1C) due to a more
significant decrease in the total number of lymphocytes (Figure 4A) than that found with
epinephrine.
In the present study, the number and percentage of T cells (CD3+ cells) and their subsets
(CD4+ T cells and CD8+ T cells), B cells (CD19+/B220+ cells) and plasma cells (CD138+
cells) were evaluated in the PPs of mice. Compared to the control group, in the mice
restrained for 1 h, a reduction was found in the total lymphocytes and the number of plasma
cells and IgA-producing plasma cells. In the mice restrained for 4 h, these same reductions
were observed along with decreases in CD8+ T cells and B cells (Figure 2). Similar results
were reported from another study, where 12 h of restraint stress caused a decrease in the
number of B cells (B220+ cells), CD8+ T cells and total T cells in PPs of mice [22]. In that
study, plasma cells were not evaluated. The fact that a decrease in total T cells was
observed was likely due to the greater time of restraint stress.
15. 15
Even though PPs are an inductor site, they also have antibody-producing effector cells [28-
30]. Consequently, the number and percentage of IgA-producing plasma cells were
evaluated in the present study. The flow cytometric analysis showed that compared to the
non-stressed animals, the total number of IgA-producing plasma cells in PPs was lower in
both groups of restraint-stressed mice. The immunohistochemical study confirmed these
results, and also revealed that of the regions of the PPs, only in the dome was there a
decrease in the number of these cells in stressed mice. In others sites of the PPs, such as the
corona, germinal center and internodal region, there were no significant differences in the
number of IgA-producing plasma cells between stressed and non-stressed groups (Figure
3). The fact that the only change in the number of IgA-producing plasma cells was found in
the dome is to be expected, as it is known that IgA-producing plasma cells are
predominantly located in this region of PPs.
The reduction in the response of IgA-producing plasma cells was not significantly greater
in the mice stressed for 4 hours than those stressed for 1 hour, which could be due to the
effect of habituation. In previous studies, rats were exposed to stress by electric shock [32,
33], finding that the initial exposure significantly reduced the T-cell proliferative response
to mitogens in the spleen (but not in the blood), whereas such effect caused by subsequent
exposures was only minor.
Whereas some IgA antibodies are directed against endogenous antigens (e.g., DNA),
others, such as natural polyreactive IgA, also react with exogenous antigens [29, 30]. The
function of the latter antibodies is not clear, but they could be important for reducing the
frequency of allergies, as well as inflammatory and autoimmune diseases in the intestine
16. 16
[34]. Furthermore, these innate secretory antibodies may protect against some infections,
such as Salmonella typhimurium [35]. Since repeated restraint stress reduced the number of
IgA-producing plasma cells, it may facilitate an invasion by pathogenic micro-organisms.
It is known that stress-induced changes in the immune response are mediated principally
through the release of glucocorticoids and catecholamines in different kinds of tissues, that
administration of high doses of glucocorticoids notably suppresses the humoral immune
response [36, 37], and that dexamethasone and epinephrine at least partially mediate some
of the effects of stress on the systemic immune response [38-40]. Therefore, an evaluation
of the effects of these hormones was included in the present study. The doses of
dexamethasone (5 and 50 mg/kg) were much higher than those normally used [41, 42]
Compared to control animals, both doses of dexamethasone and epinephrine reduced the
number of IgA-producing plasma cells in PPs, as determined by flow cytometry (Figure 6).
The immunohistochemical analysis confirmed these results, with the exception of finding
no significant difference in this parameter with the lower dose of epinephrine.
Compared to control animals, dexamethasone and epinephrine at both doses induced a
decrease in the number of all other evaluated sub-populations of lymphoid cells (Figure 4
and Figure 5). The only exception was with dexamethasone at the lower dose, which
produced no significant reduction in total plasma cells. Interestingly, the effects on the
populations of lymphocytes were progressively greater, considering the 1 hr restraint stress
group, the 4 h restraint stress group, and the application of dexamethasone or epinephrine.
Hence, the effects of restraint stress in the present study were due at least in part to the
effect of glucocorticoids and catecholamines, and it is possible that the doses of these
17. 17
hormones administered represent the effect of a more intense stress than that provoked by
the restraint stress protocol of the current contribution.
In previous studies on rats, dexamethasone significantly reduced the S-IgA levels in the bile
and the number of IgA-producing plasma cells in the ileum [43], and favored the adherence
of bacteria to the epithelium as well as the invasion of the mucosa [44]. There have not
been any previous reports, to the best of our knowledge, regarding the effect of
dexamethasone on the IgA-producing plasma cells in PPs of mice. However, it has been
reported that a single injection of dexamethasone reduces the number of Ig-producing
plasma cells (IgM, IgG and IgA) in the spleen and mesenteric lymph nodes of mice [45],
and that a treatment with a dose between 30 µg/kg and 2.5 mg/kg of BW reduces the
number of T and B lymphocytes in PPs in animals other than mice (pigs, neonatal calves,
sheep and rabbits) [41, 42, 46, 47].
The reduction in the number of IgA-producing plasma cells caused by both doses of this
glucocorticoid could owe itself to three mechanisms: a) a decrease in the number of
lymphoid cells, principally by apoptosis [42, 48], b) an inhibitory effect on macrophages
and helper T cells, as these are responsible for the induction of the immune response [49],
and c) changes in the distribution and migration of lymphocytes, since these are able to
alter the capacity of an organ or tissue to mount a specific immune response [22, 50].
No report was found in the literature about the effect of a catecholamine on the production
of antibodies in PPs. Contradictory results have been reported regarding the effect of
catecholamines on the systemic immune response. Whereas some studies report an
inhibition of the proliferation of B cells and the production of antibodies, others show the
18. 18
opposite effect [51-53]. In one study the effect varied according to the moment of the
administration of the immunogen [54]. Although the precise mechanism by which the doses
of epinephrine employed in the present study inhibited the production of antibodies in the
PPs is unclear, it may be through direct action on mature cells [55]. It seems relevant that
PPs are innervated by fibers that contain epinephrine [20, 56, 57], which in turn modulate
the internalization of pathogenic bacteria [57].
Conclusion
In summary, whereas repeated restraint stress of different intensities did not have any
notable effect on the morphological structure of the PPs, it did change the number of
lymphocytes in this lymphoid tissue. Usually when a stress response causes such effects, it
is at least partially through an increased production of glucocorticoids and catecholamines.
There was a progressive scale of stress-related effects for the 1-hour restraint stress group
(expressing a decrease in total lymphocytes and the number of plasma cells and IgA-
producing plasma cells), the 4-hour restraint-stressed mice (expressing the same decreases
as the 1-hour group, plus a decrease in B cells and CD8+ T cells), and the mice treated with
dexamethasone and epinephrine (expressing the same decreases as the 4-hour group, plus a
decrease in CD4+ T cells and total T cells). Therefore, these effects were due at least in part
to the effect of glucocorticoids and catecholamines, and it seems likely that the effects of
epinephrine and dexamethasone represent the equivalent of an even greater stress than that
produced by the 4 h restraint stress protocol.
19. 19
Acknowledgements
We thank Bruce Allan Larsen for reviewing the use of English in this manuscript. This
research was financially supported by SIP-IPN and CONACYT, Mexico.
Conflict of Interest Statement
The authors declare that they have no conflicts of interest regarding any of the products or
techniques employed in this study.
20. 20
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24. 24
Figure Legends
Figure 1: Effect of restraint stress and treatment with dexamethasone and
epinephrine on the morphology of Peyer’s patches. Mice were unrestrained (A),
restrained for 4 h (B), or were treated with epinephrine (0.5 mg/kg, C) o dexamethasone
(50 mg/kg, D) daily for 4 d. The Peyer’s patches were removed and fixed in formaldehyde
and processed for paraffin embedding and stained with H&E and Gomori's trichomic
techniques. Germinative center (GC), mantle zone (MZ), and dome (Do). Restraint stress or
epinephrine treatment did not cause atrophy of the Peyer’s patches (B and D). However, the
higher dose of dexamethasone caused atrophy and morphological alterations of the patch
(C). (H-E) 100x.
Figure 2. Effects of repeated restraint stress on the on the percentage (A) and number (B)
of total lymphocytes, total and sub-sets of T cells (CD3+ cells), B cells and plasma cells in
Peyer’s patches determined by flow cytometry. The percentage and the number of CD4+ T
cells and CD8+ T cells were calculated from T cells. Results are the media± SD at least 3
independent experiments. *P < 0.001, **P < 0.05.
25. 25
Figure 3. Effects of repeated restraint stress on the number of IgA-producing plasma
cells in Peyer’s patches. Mice were restrained for 1h or 4 h, or unrestrained. (A)
morphological structure of PPs of stressed mice for 4 h by immunohistochemical
technique; observe that the majority of the IgA+ cells are located in the dome (a: 100x, b:
200x). (B) Number of IgA+ cells according to region of PPs of stressed and unstressed mice
detected by immunohistochemistry. (C) Absolute number and (D) percentage of IgA-
producing plasma cells in PPs in stressed and unstressed mice determined by flow
cytometry. Data represent the media ± SD of three experiments. *P < 0.001, **P < 0.05.
Figure 4 .Effect of dexamethasone on the percentage (A) and number (B) of total
lymphocytes and subsets of T cells (CD3+ cells), B cells and plasma cells in Peyer´s
Patches analyzed by flow cytometry. The percentage and the number of CD4+ T cells and
CD8+ T cells were calculated from T cells. Data represent the media ± SD of three
experiments. *P < 0.001, **P < 0.05.
26. 26
Figure 5. Effect of epinephrine on the percentage (A) and number (B) of total lymphocytes,
subsets of T (CD3+ cells) and B cells in Peyer´s Patches analyzed by flow cytometry. The
percentage and the number of CD4+ T cells and CD8+ T cells were calculated from T cells.
Data represent the media ± DS of three experiments. *P < 0.001, **P < 0.05.
Figure 6. Effect of dexamethasone and epinephrine on IgA-producing plasma cells in
PPs. Mice were treated with dexamethasone (5 or 50 mg/kg) or epinephrine (0.1 or 0.5
mg/kg) daily for 4 days. Control animals were treated with the vehicle. Data were obtained
from 6 to 12 mice/group and are presented as the mean ± SD. (A) Number of IgA+ cells in
PPs of treated and untreated mice with dexamethasone, determined by
immunohistochemistry. (B) Number and (C) percentage of IgA-producing plasma cells in
PPs of treated and untreated mice with dexamethasone, determined by flow cytometry. (D)
Number of IgA+ cells in PPs of treated and untreated mice with epinephrine, determined by
immunohistochemistry. (E) Number and (F) percentage of IgA-producing plasma cells in
PPs of treated and untreated mice with ephinephrine, determined by flow cytometry. *P <
0.001, ** P < 0.01, Bonferroni’s t-test.
27. 27
Figure 7.- Effects of chronic restraint stress on the number of IgA-producing cells in
Peyer’s patches detected by an plaque hemolytic assay. Restraint stress, both doses of
dexamethasone (5 and 50 mg) and epinephrine (0.5 mg) significantly reduced (* P <
0.001) the number of IgA-APC detected by a plaque hemolytic assay in a suspension of
Peyer's patch lymphocytes. Similar results were obtained in four independent experiments.