• Journal of Animal Reproduction and Biotechnology
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• v.38 no.2
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• pp.54-61
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• 2023

Background: Germ cells undergo towards male or female pathways to produce spermatozoa or oocyte respectively which is essential for sexual reproduction. Mesenchymal stem cells (MSCs) have the potential of trans-differentiation to the multiple cell lineages. Methods: Herein, rat MSCs were isolated from bone marrow and characterized by their morphological features, expression of MSC surface markers, and in vitro differentiation capability. Results: Thereafter, we induced these cells only by retinoic acid supplementation in MSC medium and, could able to show that bone marrow derived MSCs are capable to trans-differentiate into male germ cell-like cells in vitro. We characterized these cells by morphological changes, the expressions of germ cell specific markers by immunophenotyping and molecular biology tools. Further, we quantified these differentiated cells. Conclusions: This study suggests that only Retinoic acid in culture medium could induce bone marrow MSCs to differentiate germ cell-like cells in vitro. This basic method of germ cell generation might be helpful in the prospective applications of this technology.

• Archives of Pharmacal Research
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• v.18 no.6
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• pp.376-380
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• 1995

Monocyte adhesion involves specific cell surface receptors, integrins and results in cell differentiation. We have studied expression and regulation of integrin .${\alpha}_5{\beta}_1$ during differntiation of U937 as in vitro model. To determine expression of integrin ${\alpha}_5{\beta}_1$ during differentiation of U937 as in vitro model. To determine expression of integrin ${\alpha}_5{\beta}_1$ genes by RT-PCR (reverse transcription and polymerase chain reaction) method. We determined expression of integrin ${\alpha}_5{\beta}_1$ genes by RT-PCE (reverse transcription and polymerase chain reaction) method. We found that expression of integrin .alpha.5.betha.1 was greatly increased during PMA-induced differentiation of U937 cells and also found that PMA-induced expression of integrin ${\alpha}_5{\beta}_1$ was inhibited by colchicine, microtubule depoly merizing agent. These results indicate that microtubular integrity is associated with expression of integrin. ${\alpha}_5{\beta}_1$ during PMA-induced differentiation of U937 cells.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1996.04a
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• pp.80-104
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• 1996

The development of routine techniques for the isolation and in vitro maintenance of conducting airway epithelial cells in a differentiated state provides an ideal model to study the factors involved in the regulation of the expression of mucocilicary differentiation. Several key factors and conditions have been identified. These factors and conditions include the use of biphasic culture technique to achieve mucociliary differentiation and the use of such stimulators, the thickness of collagen gel substratum, the calcium level, and vitamin A, and such inhibitors, the growth factors EGF and insulin, and steroid hormones, for mucous cell differentiation. Using the defined culture medium, the life cycle of the mucous cell population in vitro was investigated. It was demonstrated that the majority of the mucous cell population in primary cultures is not involved in DNA replication. However, the mucous cell type is capable of self-renewal in culture and this reproduction is vitamin A dependent. furthermore, differentiation from non-mucous cell type to mucous cell type can be demonstrated by adding back a positive regulator such as vitamin A to the “starved” culture. Cell kinetics data suggest that vitamin A-dependent mucous cell differentiation in culture is a DNA replication-independent process and the process is inhibited by TGF-${\beta}$1.

• Asian-Australasian Journal of Animal Sciences
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• v.18 no.12
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• pp.1708-1714
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• 2005

Keratinocyte growth factor (KGF) functions in epithelial growth and differentiation in many tissues and organs. KGF is expressed in the uterine endometrial epithelial cells during the estrous cycle and pregnancy in pigs, and receptors for KGF (KGFR) are expressed by conceptus trophectoderm and endometrial epithelia. KGF has been shown to stimulate the proliferation and differentiation of conceptus trophectoderm. However, the role of KGF on the endometrial epithelial cells has not been determined. Therefore, this study determined the effect of KGF on proliferation and differentiation of endometrial epithelial cells in vitro and in vivo using an immortalized porcine luminal epithelial (pLE) cell line and KGF infusion into the uterine lumen of pigs between Days 9 and 12 of estrous cycle. Results showed that KGF did not stimulate proliferation of uterine endometrial epithelial cells in vitro and in vivo determined by the $^3$H]thymidine incorporation assay and the proliferating cell nuclear antigen staining, respectively. Effects of KGF on expression of several markers for epithelial cell differentiation, including integrin receptor subunits $\alpha$4, $\alpha$5 and $\beta$1, plasmin/trypsin inhibitor, uteroferrin and retinol-binding protein were determined by RT-PCR, Northern and slot blot analyses, and immunohistochemisty, and KGF did not affect epithelial cell differentiation in vitro and in vivo. These results show that KGF does not induce epithelial cell proliferation and differentiation, suggesting that KGF produced by endometrial epithelial cells acts on conceptus trophectoderm in a paracrine manner rather than on endometrial epithelial cells in an autocrine manner.

• The Journal of Internal Korean Medicine
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• v.38 no.6
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• pp.1035-1048
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• 2017

Objectives: This study was performed to investigate the effects of Gardenia jasminoides extract (GJ) on osteoclast differentiation and bone resorption in vitro. Methods: To investigate the effect of GJ on osteoclast differentiation, the mouse leukemic myeloid cell line RAW 264.7 was stimulated by RANKL (receptor activator of nuclear factor kB ligand). Osteoclast differentiation was measured by counting TRAP (+) MNC in the presence of RANKL. To elucidate the mechanism of the inhibitory effect of GJ on osteoclast differentiation, gene expression of TRAP, Cathepsin K, MMP-9, NFATc1, c-Fos, MITF, DC-STAMP, CTR, OC-STAMP and Atp6v0d2 was measured using reverse transcription-PCR (RT-PCR). Bone resorption was measured using the bone pit formation assay. Results: GJ decreased the number of TRAP (+) MNCs in the presence of RANKL. GJ inhibited the expression of cathepsin K, MMP-9, TRAP, MITF, NFATc1, c-Fos, iNON, OC-STAMP, Atp6v0d2, and DC-STAMP in the osteoclast, and inhibited bone pit formation in vitro. Conclusions: The results suggest that GJ has inhibitory effects on bone resorption resulting from inhibition of osteoclast differentiation and gene expression.

• Journal of the Korean Society of Food Science and Nutrition
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• v.24 no.1
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• pp.1-9
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• 1995

1, 25(OH)2-23ene-D3 is a novel vitamine D3 analog which has a double bond between C-23 and C-24. We describe the effects of this analog on cell differentiation and cell proliferation in vitro using the human histiocytic lymphoma cell line U937, and on calcium metabolism in rats in vivo. In the present investigation 1, 25(OH)2-23ene-D3 was compared to the natural metabolite of vitamin D3, 1$\alpha$, 25-dihydroxycholecalciferol[1, 25(OH)2-23ene-D3 was more potent than 1, 25(OH)2-23ene-D3 for inhibition of proliferation and induction of differentiation of U937 cells. Especially, its effect on induction of differentiation, as measured by superoxide production and nonspecific esterase(NSE) activity, was about 20-fold more potent that 1, 25(OH)2-23ene-D3. This analog morphologically and functionally differentiated U937 cells to monocyte-macrophage phenotype showing a decrease of N/C ratio in Giemsa staining and the increase of adherence ability to surface. Intraperitoneal administration of 1, 25(OH)2-23ene-D3 to rats showed that the compound had at least 50 times less activity than 1, 25(OH)2-23ene-D3 in causing hypercalcemia and hypercalciuria. The strong direct effects of 1, 25(OH)2-23ene-D3 on cell proliferation and cell differentiation, coupled with its decreased activity of calcium metabolism make this compound an interesting candidate for clinical studies including patients with leukemia, as well as several skin disorders, such as psoriasis.

• International Journal of Oral Biology
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• v.30 no.3
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• pp.99-103
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• 2005

Bone morphogenetic proteins (BMPs), members of a large group of TGF-beta family, are important molecular regulators of morphogenesis of numerous tissues and organs, including bones and teeth. Most BMPs are capable of inducing bone formation in vivo and therefore are of considerable clinical interest for regenerating mineralized tissues. Recently, we have developed a method to culture cells from human cementum (human cementum-derived cells, HCDCs). HCDCs, when attached to synthetic hydroxyapatite/tricalcium phosphate (HA/TCP) ceramic and transplanted into immunodeficient mice, formed histologically identifiable cementum-like tissue. Since it is unclear to what extent BMPs are involved in cementogenesis, the aim of this study was to establish which BMPs are expressed by cementogenic HCDCs and whether the expression of BMPs is related to the degree of cellular differentiation in vitro. HCDCs were maintained in growth medium (DMEM/F12 supplemented with 10% FBS) until confluent (proliferation stage). Upon reaching confluence, cells were incubated in the differentiation medium (DMEM/F12 medium containing 10% FBS and 50 mg/ml ascorbic acid) for 14 days (differentiation stage). Next, HCDCs were incubated in mineralization medium (DMEM/F12, 50 mg/ml ascorbic acid, 2.5 mg/ml of ITS (insulin-transferrinselenium), 5 mM beta-glycerophosphate and $10^{-8}M$ dexamethasone) for another 14 days (mineralization stage). At the end of each differentiation stage, total RNA was isolated and evaluated for BMPs (2 through 8) expression by employing real time RT-PCR. HCDCs expressed most of BMPs examined except BMP-7 and BMP-8. Furthermore, on average, the highest levels of BMPs were expressed at the earlier differentiation stage, prior to the initiation of mineralization in vitro. These results indicate that several BMPs are expressed during cementoblastic differentiation and suggest that BMPs may be involved in the homeostasis of human cementum.

• Journal of Embryo Transfer
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• v.33 no.2
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• pp.85-97
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• 2018

The trans-differentiation potential of mesenchymal stem cells (MSCs) is employed, but there is little understanding of the cell source-dependent trans-differentiation potential of MSCs into corneal epithelial cells. In the present study, we induced trans-differentiation of MSCs derived from umbilical cord matrix (UCM-MSCs) and from dental tissue (D-MSCs), and we comparatively evaluated the in vitro trans-differentiation properties of both MSCs into corneal epithelial-like cells. Specific cell surface markers of MSC (CD44, CD73, CD90, and CD105) were detected in both UCM-MSCs and D-MSCs, but MHCII and CD119 were significantly lower (P < 0.05) in UCM-MSCs than in D-MSCs. In UCM-MSCs, not only expression levels of Oct3/4 and Nanog but also proliferation ability were significantly higher (P < 0.05) than in D-MSCs. In vitro differentiation abilities into adipocytes and osteocytes were confirmed for both MSCs. UCM-MSCs and D-MSCs were successfully trans-differentiated into corneal epithelial cells, and expression of lineage-specific markers (Cytokeratin-3, -8, and -12) were confirmed in both MSCs using immunofluorescence staining and qRT-PCR analysis. In particular, the differentiation capacity of UCM-MSCs into corneal epithelial cells was significantly higher (P < 0.05) than that of D-MSCs. In conclusion, UCM-MSCs have higher differentiation potential into corneal epithelial-like cells and have lower expression of CD119 and MHC class II than D-MSCs, which makes them a better source for the treatment of corneal opacity.

• The Journal of Internal Korean Medicine
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• v.40 no.1
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• pp.58-69
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• 2019

Objective: This study investigated the effects of Hansu-Daebowon (HDW) on bone resorption in vitro and bone loss in vivo. Methods: Osteoclast differentiation was measured by counting TRAP (+) MNC formed from RAW 264.7 in the presence of RANKL. Bone pit formation was determined in an artificial bone slice loaded with RANKL-stimulated osteoclasts. To elucidate the mechanisms of the inhibitory effects of HDW on bone resorption and osteoclast differentiation, osteoclastogenic genes (i.e. TRAP, MMP-9, NFATc1, c-Fos, and Cathepsin K) were measured using real time PCR. Furthermore, bone loss was observed using micro-CT in an LPS-treated mammal model. Results: HDW inhibited the bone pit formation in vitro and inhibited bone loss in vivo. Moreover, HDW decreased the number of TRAP (+) MNCs in the presence of RANKL, and HDW inhibited the expressions of cathepsin K, MMP-9, TRAP, NFATc1, and c-Fos in the osteoclasts. Conclusion: HDW exerts inhibitory effects on bone loss and bone resorption resulting from the inhibitions of osteoclast differentiation and osteoclastogenic gene expression.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.35 no.5
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• pp.304-309
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• 2009

Purpose: Fibrous dysplasia (FD) is a fibro-osseous disease associated with activating missense mutations of the gene encoding the $\alpha$-subunit of stimulatory G protein. FD may affect a single bone (called monostotic form) or multiple bones (called polyostotic form). The extent of lesions reflects the onset time of mutation. In this study, cells from monostotic FD in maxilla of a patient were isolated and cultured in vitro for characterization. Materials and Methods: The single cells were released from FD lesion which was surgical specimen from 15 years-old boy. These isolated cells were cultured in vitro and tested their proliferation activity with MTT assay. In osteogenic media, these cells underwent differentiation process comparing with its normal counterpart i.e. bone marrow stromal cells. The proliferated FD cells were detached and transplanted into the dordsal pocket of nude mouse and harvested in 6 weeks and 12 weeks. Results and Summary: FD cells have an increased proliferation rate and poor differentiation. As a result, cells isolated from FD lesion decreased differentiation into osteoblast and increased proliferation capacity. MTT assay presented that proliferation rate of FD cells were higher than control. However, the mineral induction capacity of FD was lesser than that of control. Monostotic FD cells make fewer amounts of bone ossicles and most of them are woven bone rather than lamellar bone in vivo transplantation. In transplanted FD cells, hematopoietic marrow were not seen in the marrow space and filled with the organized fibrous tissue. Therefore, they were recapitulated to the original histological features of FD lesion. Collectively, these results indicated that the FD cells were shown that the increased proliferation and decreased differentiation potential. These in vitro and in vivo system can be useful to test FD cell's fate and possible.