scholarly journals The secretome of induced pluripotent stem cells reduces lung fibrosis in part by hepatocyte growth factor

2014 ◽  
Vol 5 (6) ◽  
pp. 123 ◽  
Author(s):  
Amiq Gazdhar ◽  
Iwona Grad ◽  
Luca Tamò ◽  
Mathias Gugger ◽  
Anis Feki ◽  
...  
Keyword(s):  
Stem Cells ◽  
Growth Factor ◽  
Hepatocyte Growth Factor ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Lung Fibrosis ◽  
Induced Pluripotent ◽  
Hepatocyte Growth

scholarly journals Growth Factor-free Chondrogenic Differentiation from Induced Pluripotent Stem Cells using Minicircle Vectors

2018 ◽  
Author(s):  
Yeri Alice Rim ◽  
Yoojun Nam ◽  
Ji Hyeon Ju
Keyword(s):  
Stem Cells ◽  
Growth Factors ◽  
Growth Factor ◽  
Gene Delivery ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Bone Morphogenetic Protein 2 ◽  
Viral Gene ◽  
Induced Pluripotent ◽  
Viral Gene Delivery

The human degenerative cartilage has low regenerative potential. Chondrocyte transplantation offers a promising strategy for cartilage treatment and regeneration. Currently chondrogenesis using human pluripotent stem cells are accomplished using human recombinant growth factors. Here, we differentiated human induced pluripotent stem cells (hiPSCs) into chondrocytes and cartilage pellet using minicircle vectors. Minicircles are used as a non-viral gene delivery system for gene therapy in various diseases. Non-viral gene delivery can produce growth factors without integrating into the host genome. Minicircle vectors containing bone morphogenetic protein 2 (BMP2) and transforming growth factor, beta 3 (TGFβ3) were successfully generated and delivered to hiPSC-derived outgrowth (OG) cells. Cell pellets generated using minicircle-transfected OG cells successfully differentiated into chondrogenic lineage. Chondrogenic pellets transfected with growth factor-encoding minicircles effectively recovered osteochondral defect in rat models. Taken together, this work shows the potential application of minicircles in cartilage regeneration using hiPSCs.

188 PLURIPOTENCY OF PORCINE EMBRYONIC AND INDUCED PLURIPOTENT STEM CELLS RELATED WITH THE EXPRESSIONS OF EPITHELIAL–MESENCHYMAL TRANSITION AND APOPTOTIC MARKERS

2014 ◽  
Vol 26 (1) ◽  
pp. 208
Author(s):  
Y.-S. Kim ◽  
B.-R. Yi ◽  
S.-H. Hyun ◽  
C.-K. Lee ◽  
K.-C. Choi
Keyword(s):  
Stem Cells ◽  
Growth Factor ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Epithelial Mesenchymal Transition ◽  
Transgenic Pig ◽  
Rt Pcr ◽  
Mesenchymal Transition ◽  
Apoptotic Markers ◽  
Induced Pluripotent

In transgenic pig production for generating animal models of human diseases, apoptosis of an early implantation embryo disturbs the transgenic pig production. Porcine embryonic stem cells (pESC) and porcine induced pluripotent stem cells (piPSC) have an advantage for the generation of transgenic pigs; however, porcine stem cells have not yet been established. In addition, epithelial–mesenchymal transition (EMT) may play a critical role in embryo development and apoptosis. Thus, in this study we generated pESC and pIPSC and further examined the changes in EMT and apoptotic markers. We cultured pESC and piPSC in pESC media containing basic fibroblast growth factor (bFGF), doxicyclin, and leukemia inhibitory factor (LIF), and performed RT-PCR and alkaline phosphatase (AP) test to measure pluripotency markers. The RT-PCR results show that OCT-4, NANOG, and SOX2 were expressed in these stem cells, characteristic of stem cells. AP-positive cells were observed in pESCs and piPSC. In addition, we performed immunocytochemistry (ICC) to examine the expression of surface markers, such as SSEA-1 and SSEA-4. We found that pESC and piPSC expressed these markers, indicating that they have a stem cell property similar to rodent and human stem cells. Second, we treated pESC and piPSC with transforming growth factor beta (TGF-β) to examine the relationship between EMT and apoptotic markers, and confirmed a significant variation of EMT and apoptotic markers, i.e. bcl-2, bax, E-cadherin, and vimentin, by Western blot analysis. In a future study, we will examine the effect(s) of various endocrine hormones secreted by the ovary, such as E2 or P4, on the expressions of EMT and apoptotic markers in pESC and piPSC. Consequently, this study will contribute to elucidate underlying mechanism(s) of EMT and apoptosis by endocrine factors to prevent early apoptosis of pig embryos in these porcine stem cells. This work was supported by a grant from the Next-Generation BioGreen 21 Program (No. PJ009599), Rural Development Administration, Republic of Korea.

219 Invitro culture environment influences the ability to generate porcine primordial germ cell-like from induced pluripotent stem cells

2020 ◽  
Vol 32 (2) ◽  
pp. 237
Author(s):  
N. Pieri ◽  
R. Botigelli ◽  
A. de Souza ◽  
K. Recchia ◽  
R. de Castro ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Growth Factor ◽  
Germ Cell ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Culture Condition ◽  
Primordial Germ Cell ◽  
Serum Replacement ◽  
Induced Pluripotent

The ability to generate primordial germ cell-like (PGCLs) from induced pluripotent stem cells (iPSCs) in swine could greatly contribute to regenerative medicine. Herein, we aimed to generate porcine PGCLs (ipPGCLs) from iPSCs derived from different culture systems. Porcine (p)iPSCs from fibroblasts of stillborn animals (n=3) were transduced with lentiviral vectors containing murine OCT4, SOX2, c-MYC, and KLF4 cDNAs and maintained in iPSC medium on mouse embryonic fibroblasts (MEFs). The cells were divided into three groups: (1) supplemented with 10ngmL−1 basic fibroblast growth factor (bFGF) and murine leukemia inhibitory factor (LIF), (2) only bFGF, or (3) only LIF. The piPSC colonies were generated and characterised for pluripotency. To induce piPSCs into ipPGCLs, three or more cell lines from each culture condition (after passage 20) were differentiated into epiblast stem cell-like cells (EpiLCs) by culture with 20ngmL−1 Activin A, 12ngmL−1 bFGF, and 1% knockout serum replacement (KSR) for 2 days. Then, cells were further induced to differentiate by nonadherent culture and supplementation with 500ngmL−1 bone morphogenetic protein (BMP)4, 500ngmL−1 BMP8a, LIF, 100ngmL−1 stem cell factor (SCF), and 50ngmL−1 epidermal growth factor for 4 days. The ipPGCLs were characterised by cell morphology and detection of germ cell markers by immunofluorescence and gene expression. Statistical analysis was determined by one-way ANOVA (Prism Software). Co-location quantification was determined using the plugin Colocalization Threshold in Image J software (National Institutes of Health). On average, the efficiency rate of iPSC generation was 71% for the iPSCs-bFGF group, 17% for the LIF group, and 85% for the bFGF+LIF group. All iPSCs colonies were positive for alkaline phosphatase and OCT4, SOX2, NANOG, TRA1-60, TRA1-81, SSEA1, and SSEA4 by immunofluorescence. Embryoid body assay revealed that the piPSCs were able to differentiate into three germ layers. The culture condition did not influence the expression of OCT4, NANOG, and KLF4 based on qRT-PCR, however; SOX2 was upregulated in the LIF group (P<0.05). The ipPGCLs generated showed a round morphology. Analysis of endogenous pluripotent genes OCT4, SOX2, and NANOG throughout differentiation (fibroblasts, iPSCs, EpiLCs, and PGCLs) revealed a mild upregulation in ipPGCLs, while OCT4 was slightly downregulated in ipPGCLs from iPSCs-LIF group. PRDM14 and STELLA were not observed in ipPGCLs, although BLIMP1 was present; DAZL and VASA were mildly upregulated. The STELLA, VASA, OCT4, and SOX2 proteins were detected in ipPGCLs, and DAZL was detected only in ipPGCLs from the iPSCs-FGF group. Protein co-localization analysis showed that ipPGCLs from the iPSCs-FGF group were 100% OCT4+STELLA-positive, 55% positive for DAZL+SOX2, and 66% positive for VASA+NANOG; for the LIF group: 99.3% were OCT4+STELLA positive, DAZL was not detected, 95.2% were positive for SOX2 and 85.6% for VASA+NANOG. In the bFGF+LIF group, 95.8% were positive for OCT4+STELLA, DAZL and SOX2 were not observed, and 70% were positive for VASA+NANOG. Exogenous reprogramming factors were still expressed and did not differ between groups. These results indicate that, under our conditions, the iPSCs-FGF group may represent the best culture condition for induction into ipPGCLs. Financial support for this study was provided by FAPESP (2015/25564-0 and 2015/26818-5).

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