AIM： To enrich hepatic progenitors using epithelial cell adhesion molecule （EpCAM） as a marker from human fetal liver and investigate the expression of human leukocyte antigen （HLA） and their markers associated w...AIM： To enrich hepatic progenitors using epithelial cell adhesion molecule （EpCAM） as a marker from human fetal liver and investigate the expression of human leukocyte antigen （HLA） and their markers associated with hepatic progenitor cells. METHODS： EpCAM ＋ve cells were isolated using magnetic cell sorting （MACS） from human fetuses （n = 10） at 15-25 wk gestation. Expression of markers for hepatic progenitors such as albumin, alpha-fetoprotein （AFP）, CD29 （integrin ~1）, CD49f （integrin c^6） and CD90 （Thy 1） was studied by using flow cytometry, immunocytochemistry and RT-PCR; HLA class Ⅰ （A, B, C） and class Ⅱ （DR） expression was studied by flow cytometry only. RESULTS： FACS analysis indicated that EpCAM ＋ve cells were positive for CD29, CD49f, CD90, CD34, HLA class I, albumin and AFP but negative for HLA class Ⅱ （DR） and CD45. RT PCR showed that EpCAM ＋ve cells expressed liver epithelial markers （CK18）, biliary specific marker （CK19） and hepatic markers （albumin, AFP）. On immunocytochemical staining, EpCAM ＋ve cells were shown positive signals for CK18 and albumin. CONCLUSION： Our study suggests that these EpCAM ＋ve cells can be used as hepatic progenitors for cell transplantation with a minimum risk of alloreactivity and these cells may serve as a potential source for enrichment of hepatic progenitor.展开更多
Cell-based regenerative medicine is of growing interest in biomedical research. The role of stem cells in this context is under intense scrutiny and may help to define principles of organ regeneration and develop inno...Cell-based regenerative medicine is of growing interest in biomedical research. The role of stem cells in this context is under intense scrutiny and may help to define principles of organ regeneration and develop innovative therapeutics for organ failure. Utilizing stem and progenitor cells for organ replacement has been conducted for many years when performing hematopoietic stem cell transplantation. Since the first successful transplantation of umbilical cord blood to treat hematological malignancies, non-hematopoietic stem and progenitor cell populations have recently been identified within umbilical cord blood and other perinatal and fetal tissues. A cell population entitled mesenchymal stromal cells (MSCs) emerged as one of the most intensely studied as it subsumes a variety of capacities: MSCs can differentiate into various subtypes of the mesodermal lineage, they secrete a large array of trophic factors suitable of recruiting endogenous repair processes and they are immunomodulatory.Focusing on perinatal tissues to isolate MSCs, we will discuss some of the challenges associated with these cell types concentrating on concepts of isolation and expansion, the comparison with cells derived from other tissue sources, regarding phenotype and differentiation capacity and finally their therapeutic potential.展开更多
Stem cell therapy is a promising approach to clinical healing in several diseases. A great variety of tissues(bone marrow, adipose tissue, and placenta) arepotentially sources of stem cells. Placenta-derived stem cell...Stem cell therapy is a promising approach to clinical healing in several diseases. A great variety of tissues(bone marrow, adipose tissue, and placenta) arepotentially sources of stem cells. Placenta-derived stem cells(p-SCs) are in between embryonic and mesenchymal stem cells, sharing characteristics with both, such as non-carcinogenic status and property to differentiate in all embryonic germ layers. Moreover, their use is not ethically restricted as fetal membranes are considered medical waste after birth. In this context, the present review will be focused on the biological properties, culture and potential cell therapy uses of placental-derived stem cells. Immunophenotype characterization, mainly for surface marker expression, and basic principles of p-SC isolation and culture(mechanical separation or enzymatic digestion of the tissues, the most used culture media, cell plating conditions) will be presented. In addition, some preclinical studies that were performed in different medical areas will be cited, focusing on neurological, liver, pancreatic, heart, muscle, pulmonary, and bone diseases and also in tissue engineering field. Finally, some challenges for stem cell therapy applications will be highlighted. The understanding of the mechanisms involved in the p-SCs differentiation and the achievement of pure cell populations(after differentiation) are key points that must be clarified before bringing the preclinical studies, performed at the bench, to the medical practice.展开更多
AIM: To enrich putative hepatic progenitors from the developing human fetal liver using CD34 as a marker. METHODS: Aborted fetuses of 13-20 wk were used for the isolation of liver cells. The cells were labeled with an...AIM: To enrich putative hepatic progenitors from the developing human fetal liver using CD34 as a marker. METHODS: Aborted fetuses of 13-20 wk were used for the isolation of liver cells. The cells were labeled with anti CD34; a marker used for isolating progenitor population and the cells were sorted using magnetic cell sorting. The positive fractions of cells were assessed for specific hepatic markers. Further, these cells were cultured in vitro for long term investigation. RESULTS: Flow cytometric and immunocytochemical analysis for alphafetoprotein (AFP) showed that the majority of the enriched CD34 positive cells were positive for AFP. Furthermore, these enriched cells proliferated in the long term and maintained hepatic characteristics in in vitro culture. CONCLUSION: The study shows that aborted human fetal liver is a potential source for isolation of hepatic progenitors for clinical applications. The study also demonstrates that CD34 can be a good marker for the enrichment of progenitor populations.展开更多
Fetal liver （FL） is an intricate and highly vascularized hematopoietic organ, which can support the extensive expansion of hematopoietic stem cells （HSCs） without loss of stenmess, as well as of the downstream lin...Fetal liver （FL） is an intricate and highly vascularized hematopoietic organ, which can support the extensive expansion of hematopoietic stem cells （HSCs） without loss of stenmess, as well as of the downstream lineages of HSCs. This powerful function of FL largely benefits from the niche （or microenvironment）, which provides a residence for HSC expansion. Numerous studies have demonstrated that the FL niche consists of heterogeneous cell populations that associate with HSCs spatially and regulate HSCs functionally. At the molecular level, a complex of cell extrinsic and intrinsic signaling network within the FL niche cells maintains HSC expansion. Here, we summarize recent studies on the analysis of the FL HSCs and their niche, and specifically on the molecular regulatory network for HSC expansion. Based on these studies, we hypothesize a strategy to obtain a large number of functional HSCs via 3D reconstruction of FL organoid ex vivo for clinical treatment in the future.展开更多
AIM To understand the molecular mechanism of esophageal cancer development and provide molecular markers for screening high-risk populations and early diagnosis. METHODS Two-dimensional electrophoresis combined with m...AIM To understand the molecular mechanism of esophageal cancer development and provide molecular markers for screening high-risk populations and early diagnosis. METHODS Two-dimensional electrophoresis combined with mass spectrometry were adopted to screen differentially expressed proteins in nine cases of fetal esophageal epithelium, eight cases of esophageal cancer, and eight cases of tumor-adjacent normal esophageal epithelium collected from fetuses of different gestational age, or esophageal cancer patients from a high-risk area of esophageal cancer in China. Immunohistochemistry(avidin-biotin-horseradish peroxidase complex method) was used to detect the expression of peroxiredoxin(PRX)6 in 91 cases of esophageal cancer, tumoradjacent normal esophageal tissue, basal cell hyperplasia, dysplasia, and carcinoma in situ, as well as 65 cases of esophageal epithelium from fetuses at a gestational age of 3-9 mo.RESULTS After peptide mass fingerprint analysis and search of protein databases, 21 differential proteins were identified; some of which represent a protein isoform. Varying degrees of expression of PRX6 protein, which was localized mainly in the cytoplasm, were detected in adult and fetal normal esophageal tissues, precancerous lesions, and esophageal cancer. With the progression of esophageal lesions, PRX6 protein expression showed a declining trend(P < 0.05). In fetal epithelium from fetuses at gestational age 3-6 mo, PRX6 protein expression showed a declining trend with age(P < 0.05). PRX6 protein expression was significantly higher in well-differentiated esophageal cancer tissues than in poorly differentiated esophageal cancer tissues(P < 0.05).CONCLUSION Development and progression of esophageal cancer result from interactions of genetic changes(accumulation or superposition). PRX6 protein is associated with fetal esophageal development and cancer differentiation.展开更多
Background： Endothelial cell damage is an important pathophysiological step of restenosis after angioplasty and stenting. Cell transplantation has great therapeutic potential for endothelial recovery. We investigated...Background： Endothelial cell damage is an important pathophysiological step of restenosis after angioplasty and stenting. Cell transplantation has great therapeutic potential for endothelial recovery. We investigated the effect of transplanting endothelial progenitor cells （EPCs） derived from human early fetal aortas in rat injured arteries. Methods： The carotid arterial endothelium of Sprague-Dawley rats was damaged by dilatation with a 1.5 F balloon catheter, and then EPCs derived from human early fetal aortas （〈14 weeks） were injected into the lumen of the injured artery in transplanted rats, with an equal volume of normal saline injected into control rats. Rats were sacrificed at 2 and 4 weeks after treatment and transplanted cells were identified by immunohistochemical staining with anti-human CD31 and anti-human mitochondria antibodies. Arterial cross-sections were analyzed by pathology, immunohistochemistry, and morphometry. Results： Green fluorescence-labeled EPCs could be seen in the endovascular surface of balloon-injured vessels after transplantation. The intimal area and intimal/medial area ratio were significantly smaller in the transplanted group than in the control （P 〈 0.05） and the residual lumen area was larger （P 〈 0.05）. After EPC transplantation, a complete vascular endothelial layer was formed, which was positive for human yon Willebrand factor after immunohistochemical staining, and immunohistochemical staining revealed many CD31- and mitochondria-positive cells in the re-endothelialized endothelium with EPC transplantation but not control treatment. Conclusion： EPCs derived from human early fetal aorta were successfully transplanted into injured vessels and might inhibit neointimal hyperplasia after vascular injury.展开更多
基金Council of Scientific and Industrial Research Network Grant CMM002ICMR Grant (GAP 0215)
文摘AIM： To enrich hepatic progenitors using epithelial cell adhesion molecule （EpCAM） as a marker from human fetal liver and investigate the expression of human leukocyte antigen （HLA） and their markers associated with hepatic progenitor cells. METHODS： EpCAM ＋ve cells were isolated using magnetic cell sorting （MACS） from human fetuses （n = 10） at 15-25 wk gestation. Expression of markers for hepatic progenitors such as albumin, alpha-fetoprotein （AFP）, CD29 （integrin ~1）, CD49f （integrin c^6） and CD90 （Thy 1） was studied by using flow cytometry, immunocytochemistry and RT-PCR; HLA class Ⅰ （A, B, C） and class Ⅱ （DR） expression was studied by flow cytometry only. RESULTS： FACS analysis indicated that EpCAM ＋ve cells were positive for CD29, CD49f, CD90, CD34, HLA class I, albumin and AFP but negative for HLA class Ⅱ （DR） and CD45. RT PCR showed that EpCAM ＋ve cells expressed liver epithelial markers （CK18）, biliary specific marker （CK19） and hepatic markers （albumin, AFP）. On immunocytochemical staining, EpCAM ＋ve cells were shown positive signals for CK18 and albumin. CONCLUSION： Our study suggests that these EpCAM ＋ve cells can be used as hepatic progenitors for cell transplantation with a minimum risk of alloreactivity and these cells may serve as a potential source for enrichment of hepatic progenitor.
基金Supported by Research Funds of the German Federal Ministry of Education and Research (01GN0531 and 01GN0939)Proj-ect Commissioned by the European Community ("CASCADE"HEALTH-F5-2009-223236).
文摘Cell-based regenerative medicine is of growing interest in biomedical research. The role of stem cells in this context is under intense scrutiny and may help to define principles of organ regeneration and develop innovative therapeutics for organ failure. Utilizing stem and progenitor cells for organ replacement has been conducted for many years when performing hematopoietic stem cell transplantation. Since the first successful transplantation of umbilical cord blood to treat hematological malignancies, non-hematopoietic stem and progenitor cell populations have recently been identified within umbilical cord blood and other perinatal and fetal tissues. A cell population entitled mesenchymal stromal cells (MSCs) emerged as one of the most intensely studied as it subsumes a variety of capacities: MSCs can differentiate into various subtypes of the mesodermal lineage, they secrete a large array of trophic factors suitable of recruiting endogenous repair processes and they are immunomodulatory.Focusing on perinatal tissues to isolate MSCs, we will discuss some of the challenges associated with these cell types concentrating on concepts of isolation and expansion, the comparison with cells derived from other tissue sources, regarding phenotype and differentiation capacity and finally their therapeutic potential.
基金Supported by Coordenacao de Aperfeicoamento de Pessoal de Nível Superior(CAPES)Conselho Nacional de Desenvolvmento Científico e Tecnológico(CNPq)Fundacao de AmparoàPesquisa do estado de Minas Gerais(FAPEMIG)
文摘Stem cell therapy is a promising approach to clinical healing in several diseases. A great variety of tissues(bone marrow, adipose tissue, and placenta) arepotentially sources of stem cells. Placenta-derived stem cells(p-SCs) are in between embryonic and mesenchymal stem cells, sharing characteristics with both, such as non-carcinogenic status and property to differentiate in all embryonic germ layers. Moreover, their use is not ethically restricted as fetal membranes are considered medical waste after birth. In this context, the present review will be focused on the biological properties, culture and potential cell therapy uses of placental-derived stem cells. Immunophenotype characterization, mainly for surface marker expression, and basic principles of p-SC isolation and culture(mechanical separation or enzymatic digestion of the tissues, the most used culture media, cell plating conditions) will be presented. In addition, some preclinical studies that were performed in different medical areas will be cited, focusing on neurological, liver, pancreatic, heart, muscle, pulmonary, and bone diseases and also in tissue engineering field. Finally, some challenges for stem cell therapy applications will be highlighted. The understanding of the mechanisms involved in the p-SCs differentiation and the achievement of pure cell populations(after differentiation) are key points that must be clarified before bringing the preclinical studies, performed at the bench, to the medical practice.
基金Supported by Department of Biotechnology, Government of India
文摘AIM: To enrich putative hepatic progenitors from the developing human fetal liver using CD34 as a marker. METHODS: Aborted fetuses of 13-20 wk were used for the isolation of liver cells. The cells were labeled with anti CD34; a marker used for isolating progenitor population and the cells were sorted using magnetic cell sorting. The positive fractions of cells were assessed for specific hepatic markers. Further, these cells were cultured in vitro for long term investigation. RESULTS: Flow cytometric and immunocytochemical analysis for alphafetoprotein (AFP) showed that the majority of the enriched CD34 positive cells were positive for AFP. Furthermore, these enriched cells proliferated in the long term and maintained hepatic characteristics in in vitro culture. CONCLUSION: The study shows that aborted human fetal liver is a potential source for isolation of hepatic progenitors for clinical applications. The study also demonstrates that CD34 can be a good marker for the enrichment of progenitor populations.
基金supported by the National Natural Science Foundation of China (81530004, 31425016)the Ministry of Science and Technology of China (2016YFA0100500)the Strategic Priority Research Program of the Chinese Academy of Sciences, China (XDA16010104)
文摘Fetal liver （FL） is an intricate and highly vascularized hematopoietic organ, which can support the extensive expansion of hematopoietic stem cells （HSCs） without loss of stenmess, as well as of the downstream lineages of HSCs. This powerful function of FL largely benefits from the niche （or microenvironment）, which provides a residence for HSC expansion. Numerous studies have demonstrated that the FL niche consists of heterogeneous cell populations that associate with HSCs spatially and regulate HSCs functionally. At the molecular level, a complex of cell extrinsic and intrinsic signaling network within the FL niche cells maintains HSC expansion. Here, we summarize recent studies on the analysis of the FL HSCs and their niche, and specifically on the molecular regulatory network for HSC expansion. Based on these studies, we hypothesize a strategy to obtain a large number of functional HSCs via 3D reconstruction of FL organoid ex vivo for clinical treatment in the future.
基金Supported by National Natural Science Foundation of Chinathe Guangdong Provincial People’s Government of the Joint Natural Science Fund,U1301227Major Project of Science and Technology of Henan Province,161100311300
文摘AIM To understand the molecular mechanism of esophageal cancer development and provide molecular markers for screening high-risk populations and early diagnosis. METHODS Two-dimensional electrophoresis combined with mass spectrometry were adopted to screen differentially expressed proteins in nine cases of fetal esophageal epithelium, eight cases of esophageal cancer, and eight cases of tumor-adjacent normal esophageal epithelium collected from fetuses of different gestational age, or esophageal cancer patients from a high-risk area of esophageal cancer in China. Immunohistochemistry(avidin-biotin-horseradish peroxidase complex method) was used to detect the expression of peroxiredoxin(PRX)6 in 91 cases of esophageal cancer, tumoradjacent normal esophageal tissue, basal cell hyperplasia, dysplasia, and carcinoma in situ, as well as 65 cases of esophageal epithelium from fetuses at a gestational age of 3-9 mo.RESULTS After peptide mass fingerprint analysis and search of protein databases, 21 differential proteins were identified; some of which represent a protein isoform. Varying degrees of expression of PRX6 protein, which was localized mainly in the cytoplasm, were detected in adult and fetal normal esophageal tissues, precancerous lesions, and esophageal cancer. With the progression of esophageal lesions, PRX6 protein expression showed a declining trend(P < 0.05). In fetal epithelium from fetuses at gestational age 3-6 mo, PRX6 protein expression showed a declining trend with age(P < 0.05). PRX6 protein expression was significantly higher in well-differentiated esophageal cancer tissues than in poorly differentiated esophageal cancer tissues(P < 0.05).CONCLUSION Development and progression of esophageal cancer result from interactions of genetic changes(accumulation or superposition). PRX6 protein is associated with fetal esophageal development and cancer differentiation.
文摘Background： Endothelial cell damage is an important pathophysiological step of restenosis after angioplasty and stenting. Cell transplantation has great therapeutic potential for endothelial recovery. We investigated the effect of transplanting endothelial progenitor cells （EPCs） derived from human early fetal aortas in rat injured arteries. Methods： The carotid arterial endothelium of Sprague-Dawley rats was damaged by dilatation with a 1.5 F balloon catheter, and then EPCs derived from human early fetal aortas （〈14 weeks） were injected into the lumen of the injured artery in transplanted rats, with an equal volume of normal saline injected into control rats. Rats were sacrificed at 2 and 4 weeks after treatment and transplanted cells were identified by immunohistochemical staining with anti-human CD31 and anti-human mitochondria antibodies. Arterial cross-sections were analyzed by pathology, immunohistochemistry, and morphometry. Results： Green fluorescence-labeled EPCs could be seen in the endovascular surface of balloon-injured vessels after transplantation. The intimal area and intimal/medial area ratio were significantly smaller in the transplanted group than in the control （P 〈 0.05） and the residual lumen area was larger （P 〈 0.05）. After EPC transplantation, a complete vascular endothelial layer was formed, which was positive for human yon Willebrand factor after immunohistochemical staining, and immunohistochemical staining revealed many CD31- and mitochondria-positive cells in the re-endothelialized endothelium with EPC transplantation but not control treatment. Conclusion： EPCs derived from human early fetal aorta were successfully transplanted into injured vessels and might inhibit neointimal hyperplasia after vascular injury.
