Stem sel pada hewan dan aplikasinya
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Stem sel pada hewan dan aplikasinya

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Stem Cell on animal & it's application

Stem Cell on animal & it's application

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Stem sel pada hewan dan aplikasinya Stem sel pada hewan dan aplikasinya Presentation Transcript

  • Drh. Yuda Heru Fibrianto, MP., PhD.Bagian Fisiologi Fakultas Kedokteran Hewan Universitas Gadjah Mada Yogyakarta 19012013
  • The Stem CellConceptA stem cell is an undifferentiated,dividing cell that gives rise to adaughter cell like itself and adaughter cell that becomes aspecialized cell type.
  • Cell Therapy A treatment intended to regenerate or rejuvenate the body by injecting it with healthy live or freeze-dried cells derived from organs or embryos. Sometimes called fresh or live cell therapy. Performed to treat specific diseases and disorders: arthritis, lupus, cancer, HIV infection, cardiovascular and neurological disorders, and Parkinsons disease. Also used to stimulate the immune system, revitalize bodily organs, and slow the effects of aging, including memory loss and sexual dysfunction.
  • Human and Animal Stem Cell Embryonic Infant Fetal Adult Umbilical Wharton’sBlastocyst Gonadal ridge Abortus Jelly Germ line cord blood Somatic(5-7 days) (6 weeks) (Fetal tissues) Spermatogonia OogoniaEmbrionic Embrionic Fetal stem Umbilical cord Umbilical cordstem cells germ cells cells blood stem cells matrix stem cellsHemopoietic Mesenchymal Pancreas? Liver Epidermal Neuronal Eye Gut Bone Peripheral (skin, hair) marrow blood Bone marrow stroma
  • Therap sel dan produk sel padahewan Lebih berkembang Sebagai hewan coba Hewan model Etika dapat diterima
  • Effect of advanced kidney cell-derived proteinextract (AKCPE) on treatment of cronic renalfailure in dog and cat (fibrianto dkk., 2012)
  • Effect of advanced kidney cell-derived protein extract (AKCPE) on treatment of cronic renal failure in dog and cat (fibrianto dkk., 2012) SDM Hb PCV MCH MCV MCHC BUN Creat (106/μL) (g/dL) (%) (pg) (fl) (%) (mg/dL) (mg/dL)Ke-0 5,19 9,74 31,46 18,92 63,59 29,21 102,32 3,87Ke-11 5,09 10,75 36,26 22,07 77,93 29,33 57,85 2,59Ke-18 4,09 6,1 27,5 14,66 66,13 22,4 34,5 2,5
  • Dog I Dog II Dog III Dog IV Dog V CatBUN 0 69 46 62.3 174 160.3 50 11 59.4 11.5 48.7 - 111.8 46 18 - 46 23 - - 23Creatinin 0 2 2 4.7 4.97 5.7 1 11 2.07 1 4.2 - 3.1 1 18 - 1 4 - - 1RBC 0 2.13 6.65 3.43 9.19 4.53 1.18 11 9.11 3.13 3.76 - 4.36 1.79 18 - 4.83 3.34 - - 2.71Hb 0 3.5 6.1 8.6 21.1 9.4 1.7 11 17.6 7.5 9.2 8.7 2.8 18 - 7.8 4.4 - 3.5
  • Tissue engineering and therapeutic cloning in an effort toproduce genetically identical renal tissue in an animal model(Bos taurus)  Bovine skin fibroblasts from adult Holstein steers were obtained by ear notch and single donor cells were isolated and microinjected into the perivitelline space of donor enucleated oocytes (nuclear transfer).  Blastocysts were transferred to the uterus of progestin- synchronized recipients permit further in vivo growth.  After 12 weeks cloned renal cells were harvested, expanded in vitro, then seeded onto biodegradable scaffolds.  The constructs (consisting of cells + scaffolds) were then implanted into the subcutaneous space of the same steer from which the cells were cloned to allow for tissue growth (Hipp and Atala 2004)
  • a cb d a. Combining therapeutic cloning and tissue engineering to produce kidney tissue, an illustration of the tissue- engineered renal unit b. Renal unit seeded with cloned cells, three months after implantation, showing the accumulation of urinelike fluid c. Clear unidirectional continuity between the mature glomeruli, their tubules, and the polycarbonate membrane. d. Elispot analyses of the frequencies of T-cells that secrete IFN-gamma after primary and secondary stimulation with allogeneic renal cells, cloned renal cells, or nuclear donor fibroblasts. (Hipp and Atala 2004)
  • Succesful transplantation of bovine testicularcells to heterologous recipientsHistology of donor testes. In calves with SC between 18-20 cm, 45% of tubulescontained a single layer of epithelium composed of Spermatogonia (arrows) andSertoli cells (arrowhead) (A), while the remaining tubuleswerecomposedof2–4 layerswith spermatocytes (arrows) (B). In calves with SC between 21 and 22 cm, nearly53% of tubules contained 4–6 layers epithelium and spermatogenesis hadprogressed to Production of spermatids (arrow) (C). Bar Z 50 mm. Herrid et al.Reproduction 2006; 132:617-624
  • Stem cell based therapeutical approach of male infertility by teratocarcinoma derived germ cells Nayernia et al., 2004. Human Molecular Genetics 13, (14):1451–1460Analysis of differentiation of SSC1 cells in vivo. (A) A fluorescent microscopic picture of a section of a 3 week transplanted testisshowed proliferation of GFP positive cells (2.1% of tubuli). (B) Eight weeks after transplantation, GFP positive cells migrated to thebasement membrane and colonized the tubules (3.8 colonies per 107 cells). These cells were able to initiate spermatogenesis and todifferentiate into spermatids after 3 months (D) and into mature sperm after 7 months (F) of transplantation (3.0 colonies per 107cells). A higher magnification of a spermatid and a sperm cell are shown at the right corner of the pictures. The other non-transplantedtestis served as an internal control (C and E), no regeneration of spermatogenesis was observed. Hemalaun-eosin staining (G) and thecorresponding fluorescence picture (H) showed the GFP positive cells in the periphery of seminiferous tubules (arrow) and theirdifferentiation into sperm (arrow). A higher magnification of a sperm cell is shown at the right corner of the picture. (I) RT–PCRanalysis of transplanted testis (TR) 3 months after transplantation, compared with a germ cell depleted but non-transplanted testis(NTR). Expression of genes specific for premeiotic (EGFP, Stra8), meiotic (SCP3, Pgk2) and postmeiotic (Tp2) stages shows that SSC1cells were differentiated into postmeiotic cells. For control, RNA from SSC1 cells and testicular RNA (T) were used. GAPDH served as aninternal standard. (J) DNA image cytometry analysis. The DNA contents were quantified by assigning an optical density to each pixel inthe image and summing the optical density values for each nucleus. One hundred cells in luminal layer of the same sections in (G) and(H) were evaluated for DNA ploidy analysis. The presence of a haploid cell population (arrow) was confirmed. (K) As reference cells,mouse epidermis and lymphocytes were measured. (L and M) Characterization of differentiated cells. Testis sections were subjected toindirect immunofluorescence with antibodies to outer acrosomal membrane protein (L) and transition protein 2 (M) as primaryantibodies and Cy3-conjugated (red) secondary IgG antibodies. Positive cells are shown (arrows).
  •  Injeksi jantung tikus dengan sel jantung dari hESC + PSC teridentifikasi sel jantung manusia disokong oleh pembuluh darah tikus dan memperbaiki kemampuan untuk memompa darah (Murry. Heart Cells Derived from Human Embryonic Stem Cells Help Restore Rat Heart Function. Nature Biotechnology 2007; 25 (9):1015- 1024. ).
  • Ability of hESDCMs to survive, function andintegrate in the in vivo heart as “biologicalpacemaker” Generation of a reproducible spontaneous cardiomyocyte differentiating system (Kehat et al., 2001)  EBs (7-10 days in suspension) plated on top of gelatin-coated culture dishes and observe microscopically for the appearance of spontaneous contraction  4-22days after plating, in 8.1% EBs rhythmically contracting appear Cell transplated to the posterolateral region of the left ventricle in swine model of slow heart rate   after grafting, a new ectopic ventricular rhythm was detected in 11 of 13 animal studies, in 6 was characterized by sustained and long term activity  Electrophysiological mapping: ectopic ventricular rhythm originated from the area of cell transplantation and pathologic studies validated the presence and integration of the grafted (Caspi and Gepstein, 2006)
  • BM-derived cells used in various animal models
  •  The generation of hepatocytes from mesenchymal stem cells and engraftment into murine liver. Stock et al. Nat Protoc. 2010 Apr;5(4):617-27 iPS cells can be efficiently differentiated into neural precursor cells, giving rise to neuronal and glial cell types in culture. Upon transplantation into the fetal mouse brain, the cells migrate into various brain regions and differentiate into glia and neurons, including glutamatergic, GABAergic, and catecholaminergic subtypes (Wernig et al., 2008) Telomere elongation in induced pluripotent stem cells from dyskeratosis congenita patients Suneet agarwal et al. Nature 464, 292-296 (11 March 2010) Functional mesenchymal stem cells derived from human induced pluripotent stem cells attenuate limb ischemia in mice. Lian et al. Circulation. 2010 Mar 9;121(9):1113-23
  • hES cell-derived oligodendrocytes and theirability to remyelinate and restore function of the spinal cord in mice after injury by Keirstead et al., 2005 approved by FDA for phase I clinical trial Neurons derived from reprogrammed fibroblast functionally integrate into the fetal brain and improve sympton of rats with Parkinson’s disease. iPS cells can be efficiently differentiated into neural precursor cells, giving rise to neuronal and glial cell types in culture. Upon transplantation into the fetal mouse brain, the cells migrate into various brain regions and differentiate into glia and neurons, including glutamatergic, GABAergic, and catecholaminergic By Wernig et al., 2008. (PNAS ) PLoS One. 2011 Mar 4;6(3):e17560 Functional integration of grafted neural stem cell- derived dopaminergic neurons monitored by optogenetics in an in vitro Parkinson model. Tønnesen J, Parish CL, Sørensen AT, Andersson A, Lundberg C, Deisseroth K, Arenas E, Lindvall O, Kokaia M.
  • Neurosurgery. 2011 Jan;68(1):213-22; discussion 222.Predifferentiated brain-derived adult human progenitor cells migrate towardischemia after transplantation to the adult rat brain. Olstorn H, Varghese M, Murrell W,Moe MC, Langmoen IA. The adult human brain contains neural stem/progenitor cells (AHNPCs) that can survivetransplantation into the adult rat brain, migrate toward a lesion, and display limitedneuronal differentiation in vivo Epilepsia. 2010 Jul;51 Suppl 3:71-5.  Effect of neuronal precursor cells derived from medial ganglionic eminence in an acute epileptic seizure model. Calcagnotto ME, Ruiz LP, Blanco MM, Santos-Junior JG, Valente MF, Patti C, Frussa-Filho R, Santiago MF, Zipancic I, Alvarez-Dolado M, Mello LE, Longo BM . Stem Cells. 2010 Jul;28(7):1153-64. Medial ganglionic eminence-derived neural stem cell grafts ease spontaneous seizures and restore GDNF expression in a rat model of chronic temporal lobe epilepsy. Waldau B, Hattiangady B, Kuruba R, Shetty AK.
  • Neurol Med Chir (Tokyo). 2010;50(2):98-105 Seizure suppression in amygdala-kindled mice by transplantation of neural stem/progenitor cells derived from mouse embryonic stem cells. Shindo A, Nakamura T, Matsumoto Y, Kawai N, Okano H, Nagao S, Itano T, Tamiya T.  ScienceDaily (Aug. 30, 2008) — Oregon Health & Science University scientists have successfully produced functional auditory hair cells in the cochlea of the mouse inner ear. The breakthrough suggests that a new therapy may be developed in the future to successfully treat hearing loss. The results of this research was recently published by the journal Nature.ScienceDaily (Aug. 3, 2009) — University of Florida researchers were able toprogram bone marrow stem cells to repair damaged retinas in mice, suggesting apotential treatment for one of the most common causes of vision loss in olderpeople.
  • Functional mesenchymal stem cellsderived from human induced pluripotentstem cells attenuate limb ischemia in mice  Human iPSCs were induced to MSC differentiation with a clinically compliant protocol. Three monoclonal, karyotypically stable, and functional MSC-like cultures were successfully isolated using a combination of CD24(-) and CD105(+) sorting. They did not express pluripotent-associated markers but displayed MSC surface antigens and differentiated into adipocytes, osteocytes, and chondrocytes.  Transplanting iPSC-MSCs into mice significantly attenuated severe hind-limb ischemia and promoted vascular and muscle regeneration. The benefits of iPSC-MSCs on limb ischemia were superior to those of adult bone marrow MSCs. The greater potential of iPSC-MSCs may be attributable to their superior survival and engraftment after transplantation to induce vascular and muscle regeneration via direct de novo differentiation and paracrine mechanisms.  Functional MSCs can be clonally generated, beginning at a single-cell level, from human iPSCs. Patient-specific iPSC-MSCs can be prepared as an "off-the-shelf" format for the treatment of tissue ischemia.  By Lian Q, Zhang Y, Zhang J, Zhang HK, Wu X, Zhang Y, Lam FF, Kang S, Xia JC, Lai WH, Au KW, Chow YY, Siu CW, Lee CN, Tse HF. Circulation. 2010 Mar 9;121(9):1113-23
  • Umbilical Cord Blood-Derived Multipotent StemCells for Buergers Disease and Ischemic LimbDisease Animal Model. Sung-Whan Kim , Hoon Han , Gue-Tae Chae , 1 2 1Sung-Hoon Lee3, Sun Bo3, Jung-Hee Yoon4, Yong-Soon Lee3, Kwang-Soo Lee5, Hwon-Kyum Park M.D., Ph.D.5,*, Kyung-Sun Kang Ph.D.3,*STEM CELLS Volume 24, Issue 6, pages1620–1626, June 2006 Human umbilical cord blood-derived multipotent stem cells can salvage limbs in ischemic hind limb mouse model. Representative photographs of control medium (left) showed autoamputation within a week after ligation. Umbilical cord blood-derived mesenchymal stem cell treated ischemic limb showed limb necrosis (middle) and limb salvage (right) on day 28 after ligation. 1. Necrotic lesion on right thumb of Patient 2. Patient showed necrotic lesion of right thumb before treatment (left). Umbilical cord blood-derived mesenchymal stem cell treatment can cure the necrotic lesion on day 120 after injecting (right).
  • Angiographic analysis of patient with Buergers disease after transplantation with umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs). Collateral branches and vascularities increased strikingly at the ankle and foot before (upper left), 30 days after (right), and 120 days after (lower left) UCB-MSC implantationAngiography of hind limb mouse model onday 28 after femoral artery ligation.Umbilical cord blood-derived MSC-treatedhind limb shows the artery (red arrow).
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