Proliferation and differentiation of fetal liver epithelial progenitor cells after transplantation into adult rat liver.
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To identify cells that have the ability to proliferate and differentiate into all epithelial components of the liver lobule, we isolated fetal liver epithelial cells (FLEC) from ED 14 Fischer (F) 344 rats and transplanted these cells in conjunction with two-thirds partial hepatectomy into the liver of normal and retrorsine (Rs) treated syngeneic dipeptidyl peptidase IV mutant (DPPIV(-)) F344 rats. Using dual label immunohistochemistry/in situ hybridization, three subpopulations of FLEC were identified: cells expressing both alpha-fetoprotein (AFP) and albumin, but not CK-19; cells expressing CK-19, but not AFP or albumin, and cells expressing AFP, albumin, and cytokeratins-19 (CK-19). Proliferation, differentiation, and expansion of transplanted FLEC differed significantly in the two models. In normal liver, 1 to 2 weeks after transplantation, mainly cells with a single phenotype, hepatocytic (expressing AFP and albumin) or bile ductular (expressing only CK-19), had proliferated. In Rs-treated rats, in which the proliferative capacity of endogenous hepatocytes is impaired, transplanted cells showed mainly a dual phenotype (expressing both AFP/albumin and CK-19). One month after transplantation, DPPIV(+) FLEC engrafted into the parenchyma exhibited an hepatocytic phenotype and generated new hepatic cord structures. FLEC, localized in the vicinity of bile ducts, exhibited a biliary epithelial phenotype and formed new bile duct structures or were incorporated into pre-existing bile ducts. In the absence of a proliferative stimulus, ED 14 FLEC did not proliferate or differentiate. Our results demonstrate that 14-day fetal liver contains lineage committed (unipotential) and uncommitted (bipotential) progenitor cells exerting different repopulating capacities, which are affected by the proliferative status of the recipient liver and the host site within the liver where the transplanted cells become engrafted. These findings have important implications in future studies directed toward liver repopulation and ex vivo gene therapy. (+info)
Therapeutic liver repopulation in a mouse model of hypercholesterolemia.
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Liver repopulation constitutes an attractive approach for the treatment of liver disorders or of diseases requiring abundant secretion of an active protein. We have described previously a model of selective repopulation of a normal liver by Fas/CD95-resistant hepatocytes, in which we achieved up to 16% hepatocyte repopulation. In the present study, we investigated the therapeutic efficacy of this strategy. With this aim, apolipoprotein E (ApoE) knockout mice were transplanted with Fas/CD95-resistant hepatocytes which constitutively express ApoE. Transplanted mice were submitted to weekly injections of non-lethal doses of the Fas agonist antibody Jo2. After 8 weeks of treatment, we obtained up to 30% of the normal level of plasma ApoE. ApoE secretion was accompanied by a drastic and significant decrease in total plasma cholesterol, which even fell to normal levels. Moreover, this secretion was sufficient to markedly reduce the progression of atherosclerosis. These results demonstrate the efficacy of this repopulation approach for correcting a deficiency in a protein secreted by the liver. (+info)
Lim1 activity is required for intermediate mesoderm differentiation in the mouse embryo.
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During gastrulation and early organogenesis, Lim1 is expressed in the visceral endoderm, the anterior mesendoderm, and the lateral mesoderm that comprises the lateral plate and intermediate mesoderm. A previous study has reported that kidneys and gonads are missing in the Lim1 null mutants (W. Shawlot and R. R. Behringer, 1995, Nature 374, 425-430). Results of the present study show that in the early organogenesis stage mutant embryo, the intermediate mesoderm that contains the urogenital precursor tissues is disorganized and displays diminished expression of PAX2 and the Hoxb6-lacZ transgene. When posterior epiblast cells of the Lim1 null mutant embryo were transplanted to the primitive streak of wild-type host embryos, they were able to colonize the lateral plate and intermediate mesoderm of the host, suggesting that Lim1 activity is not essential for the allocation of epiblast cells to these mesodermal lineages. However, most of the mutant cells that colonized the lateral and intermediate mesoderm of the host embryo did not express the Hoxb6-lacZ transgene, except for some cells that were derived from the distal part of the posterior epiblast. Lim1 activity may therefore be required for the full expression of this transgene that normally marks the differentiation of the lateral plate and intermediate mesoderm. (+info)
Distribution of dorsal-forming activity in precleavage embryos of the Japanese newt, Cynops pyrrhogaster: effects of deletion of vegetal cytoplasm, UV irradiation, and lithium treatment.
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Two types of axis-deficient embryos developed after deletion of the vegetal cytoplasm: wasp-shaped embryos and permanent-blastula-type embryos. In situ hybridization revealed that neither type of axis-deficient embryo expressed goosecoid or pax-6. brachyury was expressed in the constricted waist region of the wasp-shaped embryos but was not expressed in the permanent-blastula-type embryos. Further, we examined the effect of UV irradiation on Japanese newt embryos. Surprisingly, UV-irradiated Japanese newt eggs formed hyperdorsalized embryos. These embryos gastrulated in an irregular circular fashion with goosecoid expression in the circular equatorial region. At tailbud stage, these embryos formed a proboscis which is very reminiscent of that formed in hyperdorsalized Xenopus embryos. Transplantation of the marginal region of the UV-irradiated embryos revealed that the entire marginal zone had organizer activity. Thus we conclude that UV hyperdorsalizes Japanese newt embryos. Finally, lithium treatment of normal embryos at the 32-cell stage also resulted in hyperdorsalization. Lithium treatment of vegetally deleted embryos had two distinct results. Lithium treatment of permanent-blastula-type embryos did not result in the formation of dorsal axial structures, while the same treatment reinduced gastrulation and dorsal axis formation in the wasp-shaped embryos. Based on these results, we propose a model for early axis specification in Japanese newt embryos. The model presented here is fundamentally identical to the Xenopus model, with some important modifications. The vegetally located determinants required for dorsal development (dorsal determinants, DDs) are distributed over a wider region at fertilization in Japanese newt embryos than in Xenopus embryos. The marginal region of the Japanese newt embryo at the beginning of development overlaps with the field of the DDs. Gastrulation is very likely to be a dorsal marginal-specific property, while self-constriction is most probably a ventral marginal-specific property in Japanese newt embryos. (+info)
Reconstruction of damaged corneas by transplantation of autologous limbal epithelial cells.
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BACKGROUND: Stevens-Johnson syndrome, ocular pemphigoid, and thermal or chemical burns can cause scarring and opacification of the cornea and loss of vision. Transplantation of epithelial cells from the limbus of the contralateral cornea can restore useful vision. However, this procedure requires a large limbal graft from the healthy eye and is not possible in patients who have bilateral lesions. METHODS: We took specimens of limbal epithelial cells from the healthy contralateral eyes of six patients with severe unilateral corneal disease. The epithelial cells were cultured and expanded on amniotic membrane. The amniotic membrane, together with the sheet of limbal epithelial cells, was transplanted to the denuded corneal surface of the damaged eye after superficial keratectomy to remove fibrovascular ingrowth. The mean (+/-SD) follow-up period was 15+/-2 months. RESULTS: Complete reepithelialization of the corneal surface occurred within two to four days of transplantation in all six eyes receiving transplants. By one month, the ocular surface was covered with corneal epithelium, and the clarity of the cornea was improved. In five of the six eyes receiving transplants (83 percent), the mean visual acuity improved from 20/112 to 20/45. In one patient with a chemical burn who had total opacification of the cornea, the acuity improved from the ability to count fingers at 40 cm to 20/200. No patient had recurrent neovascularization or inflammation in the transplanted area during the follow-up period. CONCLUSIONS: Transplantation of autologous limbal epithelial cells cultured on amniotic membrane is a simple and effective method of reconstructing the corneal surface and restoring useful vision in patients with unilateral deficiency of limbal epithelial cells. (+info)
Auto iris pigment epithelial cell transplantation in patients with age-related macular degeneration: short-term results.
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Autologous iris pigment epithelial cell transplantation was performed on patients with exudative age-related macular degeneration (AMD). Autologous IPE cell culture was performed using autologous serum after iridectomy in 7 patients with AMD. The cell suspensions (2 approximately 20 x 10(4) cells) were transplanted into the submacular lesion of individuals after removal of neovascular membranes. Subsequent ophthalmological examinations, including best corrected visual acuity and fluorescein or indocyanine green angiography, were performed. In addition, 15 patients with AMD, who underwent removal of neovascular membrane without transplantation, were evaluated as non randomized controls. Varying degrees of atrophy or defects of choriocapillaris and retinal pigment epithelium were observed in all of the patients. No cystoid macular edema or fluorescein leakage was observed after treatment, but window defects were present. No patient had decreased visual acuity. One treated patient developed mild subretinal fibrosis and an other patient developed mild preretinal fibrosis, however no difference was significant when compared with the control. In conclusion, the treatment resulted in no significant improvement in macular function, as compared with the control; however, no rejection or deterioration in visual acuity occurred up to the 13 month follow up. (+info)
Experimental inflammatory bowel disease--role of T cells.
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BACKGROUND: Our experiments were aimed to test: 1. which lymphocyte subpopulations participate in mouse colitis, produced by intrarectal (i.r.) deposition of trinitrobenzene sulphonic acid (TNBSA, TNP hapten); 2. the expression of cell adhesion molecules on lymphocytes draining the site of reaction; 3. the influence of mouse haplotype on the development of colitis. METHODS: CBA/J, BALB/c and C57BI/6 inbred and outbred Swiss Webster strains were used. Mesentheric lymph node (MLN) cells of immunized animals, unseparated or separated into CD4+, CD8+ or gammadelta+ and alphabeta+ T cell subpopulations or depleted of B lymphocytes, were transferred into recipients which were challenged i.r. with TNBSA. Inflammatory reaction in the colon was confirmed macro- and microscopically and by myeloperoxidase (MPO) level. MLN lymphocyte surface markers were tested cytofluorimetrically using appropriate antibodies. RESULTS: Sensitization with TNP results in chronic colitis (hapten dose-dependent colon weight gain and cellular infiltrate, significant increase of MPO level) only in CBA/J and BALB/c strains and can be adoptively transferred in a cell-dose dependent manner into syngeneic recipients by T alphabeta+ cells of both CD4+ and CD8+ subpopulations. T gammadelta+ cells were ineffective and B lymphocytes do not participate in the passive transfer reaction. In MLN the number of T lymphocytes positive for cell adhesion molecules particularly LPAM-1 (V-CAM1) and LPAM-2 increases significantly. CONCLUSIONS: Both CD4+ and CD8+ lymphocytes participate in the development of TNP-induced colitis. High MPO level may suggests that both Th1 and Th2 cells are involved. Colitis is accompanied by a significant accumulation in MLN of T lymphocytes positive for several cell surface adhesion molecules characteristic for memory T cells. Significant differences in susceptibility to develop colitis were found between different strains of mice. (+info)
Transplacental injection of somite-derived cells in mdx mouse embryos for the correction of dystrophin deficiency.
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Duchenne muscular dystrophy (DMD) is a lethal recessive disease caused by the absence of dystrophin in skeletal muscle, heart and other tissues. No cure is available at present for DMD. Here we describe a new strategy for the correction of dystrophin deficiency based on the transplantation of normal somite-derived cells into mdx mouse embryos. Somite-derived cells were isolated from E11.5 transgenic mouse embryos expressing the LacZ gene under the control of the muscle-specific desmin promoter and injected into the uterine circulation of pregnant mdx mice at gestational days E11.5-E17. Approximately 30% of the injected mdx embryos survived the procedure. Donor somite-derived cells were able to cross the placenta and migrate into host embryonic tissues. The pattern of donor cell distribution in host tissues depended on the gestational age of the transplanted embryos. Cells were found in hindlimb muscles, diaphragm, heart and ribs in E11.5 treated embryos and in the skull, ribs, vertebrae and lung of E15-E17 treated embryos. Normal dystrophin transcripts were detected in muscle and bone by RT-PCR. Histochemical analysis showed co-localization of LacZ and dystrophin expression in 5% of soleus and quadriceps muscle fibres and in 4% of heart myocytes of two of seven 8-week-old treated mdx mice. (+info)