• Clinical and Experimental Reproductive Medicine
• /
• v.45 no.2
• /
• pp.75-81
• /
• 2018

Objective: Recapitulation of the spermatogenesis process in vitro is a tool for studying the biology of germ cells, and may lead to promising therapeutic strategies in the future. In this study, we attempted to transdifferentiate Wharton's jelly-derived mesenchymal stem cells (WJ-MSCs) into male germ cells using all-trans retinoic acid and Sertoli cell-conditioned medium. Methods: Human WJ-MSCs were propagated by the explant culture method, and cells at the second passage were induced with differentiation medium containing all-trans retinoic acid for 2 weeks. Putative germ cells were cultured with Sertoli cell-conditioned medium at $36^{\circ}C$ for 3 more weeks. Results: The gene expression profile was consistent with the stage-specific development of germ cells. The expression of Oct4 and Plzf (early germ cell markers) was diminished, while Stra8 (a premeiotic marker), Scp3 (a meiotic marker), and Acr and Prm1 (postmeiotic markers) were upregulated during the induction period. In morphological studies, approximately 5% of the cells were secondary spermatocytes that had completed two stages of acrosome formation (the Golgi phase and the cap phase). A few spermatid-like cells that had undergone the initial stage of tail formation were also noted. Conclusion: Human WJ-MSCs can be transdifferentiated into more advanced stages of germ cells by a simple two-step induction protocol using retinoic acid and Sertoli cell-conditioned medium.

• Biomolecules & Therapeutics
• /
• v.6 no.3
• /
• pp.276-282
• /
• 1998

Retinoids regulate a wide variety of biological processes such as cellular proliferation and differentiation in many cell types. They have also shown to stimulate replication of several viruses including human cytomegalovirus (CMV). Retinoid signalling pathway involves two distinct subfamilies of nuclear receptors, retinoic acid receptors (RARs) and retinoid X receptors (RXRs) that bind to specific retinoic acid response elements (RAREs) in the promoter regions of retinoid-target genes. Here, we characterized RAREs in the regulatory regions of the CMV and of the hepatitis B vi.us (HBV). The viral RAREs, i.e., CMV-RARE and HBV-RARE, are composed of two consensus RARE half-sites (A/GGGTCA) arranged as a direct repeat separated by 5-bp and 1-bp, respectively. The RAREs were activated by both RAR/RXR heterodimers and RXR homodimers in transient transfection experiments. We also found that COUP-TF$\alpha$ (chicken ovalbumin upstream promoter-transcription factor u) and COUP-TF$\beta$ repressed the retinoid response of the viral elements. Further we demonstrated that previously known retinoid antagonist, SRI 1330, repressed retinoid-induced transactivation of the CMV-RARE. These results implicate Vitamin A, it's nuclear receptors and COUP-TFs as important regulators of the CMV and HBV pathogenesis and the SRl1330 as potential negative modulator of such retinoid-dependent processes.

• YAKHAK HOEJI
• /
• v.43 no.3
• /
• pp.378-384
• /
• 1999

Retinoic acid (RA) and dibutyryl cyclic AMP (dbcAMP) induce the differentiation of the multipotent embryonic carcinoma cell line, F9 cells, into parietal endoderm like cell. The F9 cells are highly proliferative doubling approximately 12 hourse. S Phase is predominant, lasting 10 hours and G2/M phase occupies most of the remaining cycle (2 hours) and G1 phase is nearly non-existent. In this study, we showed the effect of RA and dbcAMPon the cell cycle associated molecules (especially around G1 phase) during F9 cell differentiation. Differentiation of F9 cells was induced by the combined addition of RA ($10^{-7}M$) and dbcAMP (0.5mM), and cells were harvested daily up to 4 days. Flow cytometric analysis showed the prolongation of G1 phase around 30 hours after induction. Western blot analysis revealed that the amount of cyclin D1 and cdk2 were increased at day 4. However, histone H1 kinase activity of cdk2 was decreased. These data strongly suggest that RA and dbcAMP induce the growth arrest of F9 cells at G1 phase by decreasing the activity of cdk2, although they have increased the protein contents of cyclin D1 and cdk2. The reason for the discrepancy between the H1 kinase activity and protein contents are not clear yet.

• The Korean Journal of Zoology
• /
• v.37 no.4
• /
• pp.545-553
• /
• 1994

척추동물의 초기 발생에 관여하는 물질로 알려져 있는 retinoic acid(RA)를 Xenopus 배(embryo)에 처리한 결과, 체절의 형성에 이상 발생이 나타났다 즉 낭배 초기에 RA에 노출된 개체의 체절은 대조군의 화살머리 모양((((() 대신 기등 모양(1111)을 형성하고 있었으며, 고농도일수록 직선에 가까운 모습을 하고 있었다. 신경배 시기에 노출된 개체의 체절은 정상적인 분절이 이루어지지 않았으며, 이들 이상 체질의 위치는 처리 시기에 따라 다르게 나타났으나 농도에는 영향을 받지 않았다. 그러나 분절 과정중 일어나는 체절 세포들의 회전은 RA에 의해 억제되지 않았다. 이와 같은 결과는 체절의 형성에 분절 이전의 세포의 행동을 주도하는 프로그램과 분절을 주도하는 프로그램이 분리되어 있음을 암시한다. 한 가지 특이한 사항은 열처리 등으로 유발된 혼란이 수 개의 체절에 국한되어 나타나는 점과는 달리 RA의 처리는 어느 지점 이 후부터 모든 체절에 혼란을 야기시킨다는 점이다. 이와 같은 연속적인 분절의 흔란 현상은 RA가 다른 저해제에 의해 영향을 받는 유전자의 활동을 조절하는 전 단계의 유전자에 영향을 주기 때문인 것으로 사료된다. 또한 신경배 시기에 RA에 노출된 배의 a-cardiac actin은 정상적으로 발현되는데 비해 낭배 초기에 RA에 노출된 배는 감소된 양의 a-cardiac actin myNA의 축적을 보였다. 그러나 EFI-u은 RA의 처리 시기와는 무관하게 정상적으로 발현되었다.

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

• The Korean Journal of Zoology
• /
• v.33 no.3
• /
• pp.322-329
• /
• 1990

The effects of retinoic acid (RA) on the regenerating limbs of Korean newt (Hynobius leechii) larvae have been studied. Intraperitoneal injection of RA at 4 days post-amputation caused the proximalization of regenerate structures in the proximodistal (PD) axis of the limbs amputated through either the distal zeugopodium or the distal stylopodium. The mean level of proximalization (MLP) increased as a dose-dependent manner, and the MLP was also dependent on the level of amputation at a given dose of RA. At the dose of 150 $\mu$ g/g body wt., MLP's in either amputation level reached close to a peak value, and an increasing number of inhibition of regeneration or duplication in the transverse plane occurred at the dose of 200 $\mu$ g/ g body wt. and above. The changes induced by RA treatment suggest that positional values in the cells of the regenerating limbs have been modified in the three cardinal axes including PD and transverse axes in a graded mode. Furthermore, from the comparison of data in two different amputation levels, it can be speculated that the sensitivity of cells to RA effect along the PD axis might not be linear.

• Journal of Life Science
• /
• v.21 no.4
• /
• pp.534-541
• /
• 2011

Retinoic acid (RA), a metabolite of vitamin A, is known to regulate dedifferentiation of rabbit articular chondrocytes. The regulatory mechanism of dedifferentiation by RA is not yet understood. Thus, the effect of RA on the regulation of nitric oxide (NO)-induced dedifferentiation was investigated in rabbit articular chondrocytes. RA caused loss of the differentiated chondrocyte phenotype as demonstrated by inhibition of type II collagen expression and proteoglycan synthesis. RA also accelerated NO-induced dedifferentiation in rabbit articular chondrocytes as detected by expression of type II collagen and Sox-9 using Western blot analysis and production of sulfated proteoglycan using Alcain blue staining. Further, RA potentiated NO-induced activation of ERK. Inhibition of ERK with PD98059 (PD) recovered the expression of type II collagen and Sox-9 and production of sulfate proteoglycan in NO-induced dedifferentiated chondrocytes by RA treatment. Our findings suggest that RA accelerates NO-induced dedifferentiation of rabbit articular chondrocytes via the ERK pathway.

• BMB Reports
• /
• v.36 no.3
• /
• pp.265-268
• /
• 2003

Retinoic acid (RA) can transform the Golgi apparatus (GA) into a diffuse vacuolar aggregate and increase the toxicity of some immunotoxins that enter into cells by receptor-mediated endocytosis. An ultramorphological study of the RA-induced GA disruption was performed on F2000 fibroblasts. Cultures were treated with 0.11 to $30\;{\mu}M$ RA for 7 - 180 min. The endocytosis of Limax flavus agglutinin-peroxidase conjugate (LFA), and the interactions between a phorbol ester (PMA) and RA concerning GA disruption, were examined. Exposure to $0.33\;{\mu}M$ RA for 20 min transformed the GA into vacuolar aggregate. These vacuoles were not involved in endocytosis since they remained unstained after endocytosis of LFA. However, the lysosomes were involved in endocytosis, as they were strongly stained. Therefore, a RA-induced shift towards lysosomal routing of the entered LFA was presumed. Exposure to PMA made cells resistant to the Golgi-disturbing effects of RA, indicating that protein kinase C plays an important role in this process.

• Macromolecular Research
• /
• v.16 no.8
• /
• pp.682-685
• /
• 2008

The aim of this work was to develop and evaluate solid lipid nanoparticles (SLN) containing all-trans-retinoic acid (ATRA) for topical delivery. SLN composed of coconut oil and curdlan improved the suspension instability of ATRA in aqueous solution. The photodegradation of ATRA by light was reduced by incorporation in SLN. The loading efficiency of ATRA in SLN was higher than 95% (w/w). The amounts of ATRA released from SLN at $4^{\circ}C$ and at $37^{\circ}C$ were less than 15% and more than 60% (w/w) for 96 h, respectively. The ATRA-loaded SLN can be used as a potential carrier for topical delivery.

• Animal cells and systems
• /
• v.6 no.4
• /
• pp.301-304
• /
• 2002

In our previous study, we have shown that Fgf-8 is expressed in the basal layer of the apical epithelial cap (AEC) and in the underlying thin layer of mesenchymal tissue of the regenerating limbs of Mexican axolotl, Amby-stoma mexicanum. Our present RT-PCR data also demonstrate that Fgf-8 transcript is localized both in the mesenchymal and epidermal tissues. To understand the effect of retinoic acid (RA) on the expression of Fgf-8 in the regenerating axolotl limbs, RA was injected intraperitoneally at the dediffer-entiation stage of limb regeneration. The RA treatment caused 8 change in the Fgf-8 expression profile of the regenerating limbs. In RA-treated limbs, duration of Fgf-8 expression was prolonged and a high level of expression was maintained during dedifferentiation and blastema formation stages. These results suggest that Fgf-8 is an important molecule in the process of pattern duplication of regenerating salamander limbs evoked by RA treatment.