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Transglutaminase 3 expression in C57BL/6J mouse embryo epidermis and the correlation with its differentiation

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Cell Research 2005年第2期15卷 105-110页

作者： Jian ZHANG1,2, Hui Ying ZHI1, Fang DING1, Ai Ping LUO1, Zhi Hua LIU1*1National Laboratory of Molecular Oncology, Cancer Institute, Chinese Academy of Medical Sciences & Peking UnionMedical College, Beijing 100021, China.2Life Science Institute, Hebei Normal University, Shijiazhuang 050016, China National Laboratory of Molecular Oncology Cancer Institute Chinese Academy of Medical Sciences & Peking Union Medical College Beijing China Life Science Institute Hebei Normal University Shijiazhuang China

Epidermal-type transglutaminase 3 (TGM3) is involved in the cross-linking of structural proteins to form the cornifiedenvelope in the epidermis. In the present study, we detected the expression of TGM3 in the mouse embryo using RT-PCR.TGM3 mRNA is weakly presented from E11.5 to E14.5 and increases significantly from E15.5 to birth. Then wedetermined the spatial and temporal expression pattern of TGM3 in the skin and other organs by in situ hybridization. Wefound a deprivation of TGM3 in skin at E11.5, while a rich supply in periderm cells and a weak expression in basal cellsfrom E12.5 to E14.5. From the period of E15.5 to E16.5, after keratinization in the epidermis, TGM3 was expressed inthe granular and cornified layers. The electron microscopic observation of the C57BL/6J mouse limb bud skin develop-ment provided several morphological evidences for the epidermal differentiation. The above findings suggest that theexpression of TGM3 plays a important role in the epidermis differentiation in embryogenesis.

关键词： transglutaminase 3 epidermis differentiation C57BL/6J mouse embryo.

Improved osteoblast proliferation, differentiation and mineralization on nanophase Ti6Al4V

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Chinese Medical Journal 2011年第2期124卷 273-279页

作者： HAN Pei JI Wei-ping ZHAO Chang-li ZHANG Xiao-nong JIANG Yao Department of Orthopaedics Affiliated No. 6 People's Hospital of Shanghai Jiao Tong University Shanghai 200233 China

Background Previous studies have demonstrated increased functions of osteoblasts on nanophase materials compared to conventional ceramics or composites.Nanophase materials are unique materials that simulate dimensions of constituent components of bone as they possess particle or grain sizes less than 100 nm.However,to date,interactions of osteoblasts on nanophase materials compared to conventional metals remain to be elucidated.The objective of the present in vitro study was to synthesize nanophase metals （Ti6Al4V）,characterize,and evaluate osteoblast functions on Ti6Al4V.Such metals in conventional form are widely used in orthopedic applications.Methods In this work,nanophase Ti6Al4V surfaces were processed by the severe plastic deformation （SPD） principle and used to investigate osteoblast long-term functions.Primary cultured osteoblasts from neonatal rat calvaria were cultured on both nanophase and conventional Ti6Al4V substrates.Cell proliferation,total protein content,and alkaline phosphatase （ALP） activity were evaluated after 1,3,7,10 and 14 days.Calcium deposition,gene expression of type Ⅰ collagen （Col-Ⅰ）,osteocalcin （OC）,osteopontin （OP） and the production of insulin-like growth factor-Ⅰ （IGF-Ⅰ） and transforming growth factor-beta 1 （TGF-β1） were also investigated after 14 days of culture.Results Functions of osteoblasts including proliferation,synthesis of protein,and ALP activity were improved on the nanophase compared to the conventional Ti6Al4V.The expression of Col-Ⅰ,OC and OP mRNA was also increased on nanophase Ti6Al4V after 14 days of culture.Calcium deposition was the same;the average number of the calcified nodules on the two Ti6Al4V surfaces was similar after 14 days of culture;however,highly significant size calcified nodules on the nanophase Ti6Al4V was observed,Of the growth factors examined,only TGF-β1 showed a difference in production on the nanophase surface.Conclusion Nanophase Ti6Al4V surfaces improve proliferation,differentiation and mineralization of osteoblast cells and should be f

关键词： titanium alloy nanophase osteoblast proliferation differentiation mineralization

Autophagy activator promotes neuronal differentiation of adult adipose-derived stromal cells

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Neural Regeneration Research 2013年第10期8卷 882-889页

作者： Yanhui Lu Xiaodong Yuan Qiaoyu Sun Ya Ou Department of Neurology Kailuan General HospitalHebei Union University

Preliminary research from our group found altered autophagy intensity during adipose-derived stromal cell differentiation into neuronal-like cells, and that this change was associated with morphological changes in differentiated cells. This study aimed to verify the role of rapamycin, an autophagy activator, in the process of adipose-derived stromal cell differentiation into neuronal-like cells. Immunohistochemical staining showed that expression of neuron-specific enolase and neurofilament-200 were gradually upregulated in adipose-derived stromal cells after 5 mM 13-mercaptoethanol induction, and the differentiation rate gradually increased with induction time. Using transmission electron microscopy, induced cells were shown to exhibit cytoplasmic autophagosomes, with bilayer membranes, and autolysosomes. After rapamycin （200 IJg/L） induction for 1 hour, adipose-derived stromal cells began to extend long processes, similar to the morphology of neuronal-like cells, while untreated cells did not exhibit similar morphologies until 3 hours after induction. Moreover, the differentiation rate was significantly increased after rapamycin treatment. Compared with untreated cells, expression of LC3, an autophagy protein, was also significantly upregulated. Positive LC3 expression tended to concentrate at cell nuclei with increasing induction times. Our experimental findings indicate that autophagy can significantly increase the speed of adipose-derived stromal cell differentiation into neuronal-like cells.

关键词： neural regeneration stem cells adult adipose-derived sllomal cells neuronal-like cells autophagy autophagy activator autophagy protein rapamycin differentiation neuroprotection neuroregeneration

AATYK is a Novel Regulator of Oligodendrocyte differentiation and Myelination

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Neuroscience Bulletin 2018年第3期34卷 527-533页

作者： Chunxia Jiang Wanqing Yang Zhihong Fan Peng Teng Ruyi Mei Junlin Yang Aifen Yang Mengsheng Qiu Xiaofeng Zhao Institute of Developmental and Regenerative Biology Zhejiang Key Laboratory of Organ Development and Regeneration College of Life and Environmental SciencesHangzhou Normal University Hangzhou 310036 China Department of Anatomical Sciences and Neurobiology School of Medicine University of Louisville LouisvilleKY 40202 USA College of Life Sciences Zhejiang UniversityHangzhou 310036 China

Oligodendrocytes（OLs） are myelinating glial cells that form myelin sheaths around axons to ensure rapid and focal conduction of action potentials. Here, we found that an axonal outgrowth regulatory molecule, AATYK（apoptosis-associated tyrosine kinase）, was up-regulated with OL differentiation and remyelination. We therefore studied its role in OL differentiation. The results showed that AATYK knockdown inhibited OL differentiation and the expression of myelin genes in vitro. Moreover, AATYKdeficiency maintained the proliferation status of OLs but did not affect their survival. Thus, AATYK is essential for the differentiation of OLs.

关键词： AATYK differentiation Oligodendrocytes

Neuronal-like cell differentiation of non-adherent bone marrow cell-derived mesenchymal stem cells

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Neural Regeneration Research 2013年第22期8卷 2078-2085页

作者： Yuxin Wu Jinghan Zhang Xiaoming Ben Department of Neurosurgery Nanjing Children's Hospital of Nanjing Medical University Department of Newborn Medicine Nanjing Children's Hospital of Nanjing Medical University Department of Newborn Medicine Shanghai First Maternity and Infant HospitalTongji University School of Medicine

Non-adherent bone marrow cell-derived mesenchymal stem cells from C57BL/6J mice were sepa- rated and cultured using the ＂pour-off＂ method. Non-adherent bone marrow cell-derived mesen- chymal stem ceils developed colony-forming unit-fibroblasts, and could be expanded by supple- mentation with epidermal growth factor. Immunocytochemistry showed that the non-adherent bone marrow cell-derived mesenchymal stem cells exposed to basic fibroblast growth factor/epidermal growth factor/nerve growth factor expressed the neuron specific markers, neurofilament-200 and NeuN, in vitro. Non-adherent bone marrow cell-derived mesenchymal stem cells from 13-galactosidase transgenic mice were also transplanted into focal ischemic brain （right corpus striatum） of C57BL/6J mice. At 8 weeks, cells positive for LacZ and 13-galactosidase staining were observed in the ischemic tissues, and cells co-labeled with both 13-galactosidase and NeuN were seen by double immunohistochemical staining. These findings suggest that the non-adherent bone marrow cell-derived mesenchymal stem cells could differentiate into neuronal-like cells in vitro and in vivo.

关键词： neural regeneration stem cells non-adherent bone marrow cell-derived mesenchymal stem cells neuronal-like cells colony-forming unit-fibroblasts proliferation differentiation beta-galactosidasetransgenic mouse cell transplantation cerebral ischemia bone marrow cells-derived mesenchymalstem cells grants-supported paper neuroregeneration

Erythropoietin modulates bone marrow stromal cell differentiation

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Bone Research 2019年第3期7卷 272-285页

作者： Sukanya Suresh Luis Fernandez de Castro Soumyadeep Dey Pamela G. Robey Constance Tom Noguchi Molecular Medicine Branch National Institute of Diabetes and Digestive and Kidney Diseases National Institutes of Health Bethesda MD 20892 USA Skeletal Biology Section National Institute of Dental and Craniofacial Research National Institutes of Health Bethesda MD 20892 USA

Erythropoietin is essential for bone marrow erythropoiesis and erythropoietin receptor on non-erythroid cells including bone marrow stromal cells suggests systemic effects of erythropoietin. Tg6 mice with chronic erythropoietin overexpression have a high hematocrit, reduced trabecular and cortical bone and bone marrow adipocytes, and decreased bone morphogenic protein 2 driven ectopic bone and adipocyte formation. Erythropoietin treatment(1 200 IU·kg–1) for 10 days similarly exhibit increased hematocrit,reduced bone and bone marrow adipocytes without increased osteoclasts, and reduced bone morphogenic protein signaling in the bone marrow. Interestingly, endogenous erythropoietin is required for normal differentiation of bone marrow stromal cells to osteoblasts and bone marrow adipocytes.ΔEpoR_E mice with erythroid restricted erythropoietin receptor exhibit reduced trabecular bone, increased bone marrow adipocytes, and decreased bone morphogenic protein 2 ectopic bone formation. Erythropoietin treated ΔEpoR_E mice achieved hematocrit similar to wild-type mice without reduced bone, suggesting that bone reduction with erythropoietin treatment is associated with non-erythropoietic erythropoietin response. Bone marrow stromal cells from wild-type,Tg6, and ΔEpoR_E-mice were transplanted into immunodeficient mice to assess development into a bone/marrow organ. Like endogenous bone formation, Tg6 bone marrow cells exhibited reduced differentiation to bone and adipocytes indicating that high erythropoietin inhibits osteogenesis and adipogenesis, while ΔEpoR_E bone marrow cells formed ectopic bones with reduced trabecular regions and increased adipocytes, indicating that loss of erythropoietin signaling favors adipogenesis at the expense of osteogenesis. In summary, endogenous erythropoietin signaling regulates bone marrow stromal cell fate and aberrant erythropoietin levels result in their impaired differentiation.

关键词： bone marrow adipocyte normal differentiation

Globoside accelerates the differentiation of dental epithelial cells into ameloblasts

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International Journal of Oral Science 2016年第4期8卷 205-212页

作者： Takashi Nakamura Yuta Chiba Masahiro Naruse Kan Saito Hidemitsu Harada Satoshi Fukumoto Division of Molecular Pharmacology and Cell Biophysics Department of Oral Biology Tohoku University Graduate School of Dentistry Division of Pediatric Dentistry Department of Oral Health and Development Sciences Tohoku University Graduate School of Dentistry Division of Developmental Biology and Regenerative Medicine Department of Anatomy Iwate Medical University

Tooth crown morphogenesis is tightly regulated by the proliferation and differentiation of dental epithelial cells. Globoside （Gb4）, a globo-series glycosphingolipid, is highly expressed during embryogenesis as well as organogenesis, including tooth development. We previously reported that Gb4 is dominantly expressed in the neutral lipid fraction of dental epithelial cells. However, because its functional role in tooth development remains unknown, we investigated the involvement of Gb4 in dental epithelial cell differentiation. The expression of Gb4 was detected in ameloblasts of postnatal mouse molars and incisors. A cell culture analysis using HAT-7 cells, a rat-derived dental epithelial cell line, revealed that Gb4 did not promote dental epithelial cell proliferation. Interestingly, exogenous administration of Gb4 enhanced the gene expression of enamel extracellular matrix proteins such as ameloblastin, amelogenin, and enamelin in dental epithelial cells as well as in developing tooth germs. Gb4 also induced the expression of TrkB, one of the key receptors required for ameloblast induction in dental epithelial cells. In contrast, Gb4 downregulated the expression of p75, a receptor for neurotrophins （including neurotrophin-4） and a marker of undifferentiated dental epithelial cells. In addition, we found that exogenous administration of Gb4 to dental epithelial cells stimulated the extracellular signal-regulated kinase and p38 mitogen-activated protein kinase signalling pathways. Furthermore, Gb4 induced the expression of epiprofin and Runx2, the positive regulators for ameloblastin gene transcription. Thus, our results suggest that Gb4 contributes to promoting the differentiation of dental epithelial cells into ameloblasts.

关键词： ameloblast differentiation enamel matrix epiprofin glycosphingolipids tooth development

Supportive angiogenic and osteogenic differentiation of mesenchymal stromal cells and endothelial cells in monolayer and co-cultures

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International Journal of Oral Science 2016年第4期8卷 223-230页

作者： Florian Bohrnsen Henning Schliephake Clinic of Oral and Maxillofacial Surgery Georg August University Gottingen Germany

Sites of implantation with compromised biology may be unable to achieve the required level of angiogenic and osteogenic regeneration. The specific function and contribution of different cell types to the formation of prevascularized, osteogenic networks in co-culture remains unclear. To determine how bone marrow-derived mesenchymal stromal cells （BMSCs） and endothelial cells （ECs） contribute to cellular proangiogenic differentiation, we analysed the differentiation of BMSCs and ECs in standardized monolayer, Transwell and co-cultures. BMSCs were derived from the iliac bone marrow of five patients, characterized and differentiated in standardized monolayers, permeable Transwells and co-cultures with human umbilical vein ECs （HUVECs）. The expression levels of CD31, von Willebrand factor, osteonectin （ON） and Runx2 were assessed by quantitative reverse transcriptase polymerase chain reaction. The protein expression of alkaline phosphatase, ON and CD31 was demonstrated via histochemical and immunofluorescence analysis. The results showed that BMSCs and HUVECs were able to retain their lineage-specific osteogenic and angiogenic differentiation in direct and indirect co-cultures. In addition, BMSCs demonstrated a supportive expression of angiogenic function in co-culture, while HUVEC was able to improve the expression of osteogenic marker molecules in BMSCs.

关键词： angiogenic co-culture differentiation endothelial cell mesenchymal stromal cell osteogenic

ERK1 and ERK2 are involved in recruitment and maturation of human mesenchymal stem cells induced to adipogenic differentiation

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分子细胞生物学报:英文版 2011年第2期3卷 123-131页

作者： Elisabetta Donzelli Caterina Lucchini Elisa Ballarini Arianna Scuteri Fabrizio Carini Giovanni Tredici Mariarosaria Miloso Department of Neuroscience and Biomedical Technologies University of Milano-BicoccaVia Cadore 48-20052Monza(MB)Italy Reproductive Medicine Nuova Villa Claudia ClinicVia Flaminia Nuova 280RomeItaly

Adipocytes’biology and the mechanisms that control adipogenesis have gained importance because of the need to develop therapeutic strategies to control obesity and the related pathologies.Human mesenchymal stem cells(hMSCs),undifferentiated stem cells present in the bone marrow that are physiological precursors of adipocytes,were induced to adipogenic differentiation.The molecular mechanisms on the basis of the adipogenesis were evaluated,focusing on the MAPKinases ERK1 and ERK2,which are involved in many biological and cellular processes.ERK1 and ERK2 phosphorylation was reduced with different timing and intensity for the two isoforms in treated hMSCs in comparison with control cells until day 10 and then at 14–28 days,it reached the level of untreated cultures.The total amount of ERK1 was also decreased up to day 10 and then was induced to the level of untreated cultures,whereas the expression of ERK2 was not changed following adipogenic induction.Treatment with the specific ERK1/2 inhibitor U0126 during the whole differentiation period hampered hMSCs’adipogenic differentiation,as lipid droplets appeared in very few cells and were reduced in number and size.When U0126 was administered only during the initial phase of differentiation,the number of hMSCs recruited to adipogenesis was reduced while,when it was administered later,hMSCs did not acquire a mature adipocytic phenotype.ERK1 and ERK2 are important for hMSC adipogenic differentiation since any alteration to the correct timing of their phosphorylation affects either the recruitment into the differentiation program and the extent of their maturation.

关键词： mesenchymal stem cells ERK adipogenesis MAPK obesity differentiation