• Journal of Life Science
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• v.29 no.9
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• pp.1002-1009
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• 2019

Early life stress (ELS) increases the risk of depression. ELS may be involved in the susceptibility to subsequent stress exposure during adulthood. We investigated whether epigenetic mechanisms of p11 promoter affect the vulnerability to chronic unpredictable stress (CUS) induced by the maternal separation (MS). Mice pups were separated from their dams (3 hr/day from P1-P21). When the pups reached adulthood, we applied CUS (daily for 3 weeks). The levels of hippocampal p11 expression were analyzed by quantitative real-time PCR. The levels of acetylated and methylated histone H3 at p11 promoter were measured by chromatin immunoprecipitation. Depression-like behavior was measured by the forced swimming test (FST). The MS and CUS group exhibited significant decreases in p11 mRNA level and the MS plus CUS group had a greater reduction in this level than the CUS group. The MS plus CUS group also resulted in greater reduction in H3 acetylation than the CUS group. This reduction was associated with an upregulation of histone deacetylase 5. Additionally, the MS plus CUS group showed a greater decrease in H3K4met3 level and a greater increase in H3K27 met3 level than the CUS group. Consistent with the reduction of p11 expression, the MS plus CUS group displayed longer immobility times in the FST compared to the control group. Mice exposed to MS followed by CUS had much greater epigenetic alterations in the hippocampus compared to adult mice that only experienced CUS. ELS can exacerbate the effect of stress exposure during adulthood through histone modification of p11 gene.

• Asian-Australasian Journal of Animal Sciences
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• v.32 no.7
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• pp.922-929
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• 2019

Objective: Mutations in low-density lipoprotein receptor (LDLR), which encodes a critical protein for cholesterol homeostasis and lipid metabolism in mammals, are involved in cardiometabolic diseases, such as familial hypercholesterolemia in pigs. Whereas microRNAs (miRNAs) can control LDLR regulation, their involvement in circulating cholesterol and lipid levels with respect to cardiometabolic diseases in pigs is unclear. We aimed to identify and analyze LDLR as a potential target gene of SSC-miR-20a. Methods: Bioinformatic analysis predicted that porcine LDLR is a target of SSC-miR-20a. Wild-type and mutant LDLR 3'-untranslated region (UTR) fragments were generated by polymerase chain reaction (PCR) and cloned into the pGL3-Control vector to construct pGL3 Control LDLR wild-3'-UTR and pGL3 Control LDLR mutant-3'-UTR recombinant plasmids, respectively. An miR-20a expression plasmid was constructed by inserting the porcine premiR-20a-coding sequence between the HindIII and BamHI sites in pMR-mCherry, and constructs were confirmed by sequencing. HEK293T cells were co-transfected with the miR-20a expression or pMR-mCherry control plasmids and constructs harboring the corresponding 3'-UTR, and relative luciferase activity was determined. The relative expression levels of miR-20a and LDLR mRNA and their correlation in terms of expression levels in porcine liver tissue were analyzed using reverse-transcription quantitative PCR. Results: Gel electrophoresis and sequencing showed that target gene fragments were successfully cloned, and the three recombinant vectors were successfully constructed. Compared to pMR-mCherry, the miR-20a expression vector significantly inhibited wild-type LDLR3'-UTR-driven (p<0.01), but not mutant LDLR-3'-UTR-driven (p>0.05), luciferase reporter activity. Further, miR-20a and LDLR were expressed at relatively high levels in porcine liver tissues. Pearson correlation analysis revealed that porcine liver miR-20a and LDLR levels were significantly negatively correlated (r = -0.656, p<0.05). Conclusion: LDLR is a potential target of miR-20a, which might directly bind the LDLR 3'-UTR to post-transcriptionally inhibit expression. These results have implications in understanding the pathogenesis and progression of porcine cardiovascular diseases.

• Journal of Life Science
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• v.31 no.8
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• pp.710-718
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• 2021

Placenta-derived mesenchymal stem cells (PD-MSCs) are promising candidates for cell-based therapy in regenerative medicine. The migration and homing potential of PD-MSCs to injured sites is a critical property of MSC engraftment. MicroRNAs (miRNAs) have recently been shown to regulate the critical functions of MSCs, such as proliferation, survival, and migration. The objective of the present study was to identify the miRNA and target genes involved in PD-MSCs homing in a bile duct ligation (BDL) rat model. We selected candidate miRNAs targeting genes for PD-MSCs homing based on microarray analysis. PD-MSC engraftment in BDL-injured rat liver was identified by immunofluorescence assay and human-specific Alu gene expression by quantitative real-time polymerase chain reaction (qRT-PCR) one week after transplantation. Compared with migrated naïve PD-MSCs under hypoxic and normoxic conditions (Hyp/Nor), the transplanted group with PD-MSCs (Tx) showed distinct differences in miRNA expressions in BDL-injured rat liver. We also validated the miRNAs and their target genes for PD-MSCs homing. The expressions of integrin α4 (ITGA4) and integrin α5 (ITGA5) target genes for miR-199a-5p and miR-148a-3p were significantly upregulated in the Tx group (p<0.05). In addition, integrin β1 (ITGB1) and integrin β8 (ITGB8) were upregulated by suppressing miR-183-5p and miR-145-5p, respectively. These results demonstrated that PD-MSCs regulate miRNA expression related to the integrin family for their homing effects on the BDL-injured rat liver. The findings further suggest that miRNA-mediated regulation of the integrin family contributes to the therapeutic efficacy of PD-MSCs in the rat hepatic fibrosis model by BDL.

• Journal of Life Science
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• v.33 no.11
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• pp.905-914
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• 2023

To evaluate the effectiveness of the skin barrier improvement of lactic acid (LA) and gluconolactone (GL), the expression of filaggrin, loricrin, hyaluronic acid (HA), hyaluronan syhthase-2 (HAS2), and aquaporine-3 (AQP3) in keratinocytes, and the moisture content and transepidermal water loss (TEWL) by clinical trials were evaluated. The expression levels of filaggrin and locricrin, which are the main factors affecting the proper functioning of skin barrier function, and HA, HAS2, and AQP3, which are skin moisturizing-related proteins measured by quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting. The results showed that the expression levels of the factors that decreased by H2O2 treatment were significantly increased by LA, GL, and a mixture of LA and GL at the mRNA and protein levels (p<0.05). The nanoemulsion containing a mixture of LA and GL was prepared using the emulsion inversion method, and the average particle size was 299.9 ± 0.287 nm. After measuring the TEWL of nanoemulsion using Vapometer, it was found that TEWL significantly decreased by 15.53% and 26.73% after two weeks and four weeks of product use, respectively, compared to TEWL before product use (p<0.001). Similarly, the skin moisture content of the nanoemulsion significantly increased by 15.40% and 26.59% after two weeks and four weeks of product use, respectively, compared to skin moisture content before product use (p<0.001). Therefore, the skin barrier function and moisturizing effect of a mixture of LA and GL are shown by increasing the moisture content and decreasing the TEWL by increasing the expression of filaggrin, loricrin, HA, HAS2, and AQP3. This suggests the possibility for the development of functional cosmetic ingredients in the future.

• Journal of Life Science
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• v.25 no.8
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• pp.861-869
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• 2015

One of the domesticated species; the dog has been selectively bred for various aims by human. The dog has many breeds, which are artificially selected for specific behaviors and morphologies. Dogs contribute their life to human as working dogs for guide, rescue, detection or etc. Working dogs requires good personality, such as gentleness, robustness and patience for performing their special duty. Many studies have concentrated on finding genetic marker for selecting the high-quality working dog. In this study, we confirmed quantitative expression patterns of eight genes (ABAT; 4-Aminobutyrate Aminotransferase, PLCB1; Phospholipase C, Beta 1, SLC10A4; Solute Carrier Family 10, Member 4, WNT1; Wingless-Type MMTV Integration Site Family, Member 1, BARX2; BarH-Like Homeobox 2, NEUROD6; Neuronal Differentiation 6, SEPT9; Septin 9 and TBR1; T-Box, Brain, 1) among brains tissues from four dog breeds (Beagle, Sapsaree, Shepherd and Jindo), because these genes were expressed and have functions in brain mostly. Specially, BARX2, SEPT9, SLC10A4, TBR1 and WNT1 genes were highly expressed in Beagle and Jindo, and Sapsaree and German Shepherd were vice versa. The biological significance of total genes was estimated by database for annotation, visualization and integrated discovery (DAVID) to determine a different gene ontology (GO) class. In these analyses, we suppose to these eight genes could provide influential information for brain development, and intelligence of organisms. Taken together, these results could provide clues to discover biomarker related to functional traits in brain, and beneficial for selecting superior working dogs.

• Development and Reproduction
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• v.11 no.1
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• pp.49-54
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• 2007

Ethane 1,2-dimethane sulfonate(EDS), a toxin which specifically kills Leydig cells(LC), has been widely used to prepare the reversible testosterone(T) depletion rat model. In the present study, we monitored the gene expression profiles of pituitary gonadotropins, LH and FSH, up to 7 weeks after EDS injection. Adult male Sprague-Dawley rats($300{\sim}350\;g$ B.W.) were injected with a single dose of EDS(75 mg/kg i.p.) and sacrificed on weeks 0, 1, 2, 3, 4, 5, 6 and 7. Total RNAs were purified from each pituitary, and the message levels of common alpha subunit($C{\alpha}$) of pituitary glycoprotein hormones, LH beta subunit($LH{\beta}$), FSH beta subunit($FSH{\beta}$) and GnRH receptor(GnRH-R) were evaluated by semi-quantitative RT-PCRs. The message levels of $C{\alpha}$ increased sharply during weeks 1-4, then return to the control level on week 5. The mRNA levels of $LH{\beta}$ were elevated after week 2, reached the peak at week 4, then declined to the control level after week 5. The message levels of $FSH{\beta}$ were elevated after week 2, reached the peak at week 3, then declined to the nadir at week 5. Similarly, the mRNA levels of GnRH-R were elevated after week 2, reached the peak at week 3, then gradually declined to the control level after week 5. The present study indicated that EDS treatment could induce reversible alterations in the transcriptional activities of gonadotropin subunits and GnRH-R in the anterior pituitary from male rats. EDS injection model might be useful to understand the mechanism of hormonal regulation of hypothalamus- pituitary neuroendocrine axis in male rats.

• Korean Journal of Poultry Science
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• v.41 no.2
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• pp.115-125
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• 2014

With Single Comb White Leghorn (WL) and Korean Native Chicken (KNC) breeds, we compared the stress response with chicken breeds that were subjected to a high stocking density. Stress response was analyzed by the quantity of telomeric DNA, the rate of DNA damage and the expression levels of heat shock proteins (HSPs) and hydroxyl-3-methyl-glutaryl coenzyme A reductase (HMGCR) genes on tissues and blood. The telomere length and telomere shortening rates were analyzed by quantitative fluorescence in situ hybridization on the nuclei of lymphocytes and tissues. The DNA damage rate of lymphocytes was quantified by the comet assay. The expression levels of HSP70, HSP90-${\alpha}$, HSP90-${\beta}$ and HMGCR genes were measured by quantitative real-time polymerase chain reaction in lymphocytes. There was no significant difference between KNC and WL in body weight, weight gain, telomere shortening rate and DNA damage rate. However, the growth rate significantly decreased in chickens raised under high stocking density conditions, as compared to the control group. The telomere-shortening rate, DNA damage and HSPs expression of the lymphocytes were significantly higher in the high stocking density group than the control. The stress condition and breeds had a significant effect on the expressions of HSP70, HSP90-${\alpha}$ and HSP90-${\beta}$ in lymphocytes, except HMGCR. The stress response of WL was higher than that of KNC, as analyzed to the expression of HSP70 and HSP90-${\alpha}$. Therefore, we concluded that the chickens which were exposed to a high stocking density had increased the individual physiological stress response regardless of breeds, and White Leghorns are more susceptible to stress condition than Korean Native Chickens.

• Journal of Life Science
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• v.19 no.5
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• pp.593-597
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• 2009

The uterus plays a critical role in pregnancy and steroid hormones, and both estrogen (E2) and progesterone (P4) especially play important roles in the cross-talk between embryos and uterus to support the pregnancy. E2 stimulates uterine growth during early pregnancy to prepare for implantation of embryos. This cross-talk during the implantation period involves hormones (E2 and P4) and growth factors, including insulin-like growth factor-I (IGF-I). In the uterus of a pregnant pig, the action of E2 is mediated by estrogen receptor-${\beta}$ (ER-${\beta}$). The expression of ER-a was much higher in early pregnancy than in mid- and late- pregnancy, suggesting E2 secretion from embryos enhances transcription of ER-a during early pregnancy. In order to prove whether IGF-I is an E2 target gene, quantitative real-time PCR was performed on ovariectomized murine uterus with E2 and/or P4 treatment(s). Increased IGF-I mRNA expression was observed with E2 treatment, however, it was not significantly induced by P4 treatment, which clearly demonstrates that, in mice, E2 depends on the activation of uterine IGF-I gene expression. The expression of IGF-I in the uterus of pigs was much higher in early pregnancy than in mid- and late- pregnancy and these data exhibited the same expression pattern with the ER-${\beta}$ gene expression in the uterus. It suggests that a positive co-relationship between IGF-I and ER-${\beta}$ expression exists in the uterus, and that both gene expressions of IGF-I and ER-${\beta}$ are regulated by E2. It further suggests that uterine the IGF-I gene expression might be initiated by E2 secreted from embryos to increase ER-${\beta}$ gene expression, and that this increased ER-${\beta}$ further stimulates the expression of IGF-I in the uterus during early pregnancy.

• Restorative Dentistry and Endodontics
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• v.29 no.5
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• pp.479-484
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• 2004

Immunoinhibitory protein extracted from sonicated Treponema denticola have been shown to suppress cell cycle progression of human lymphocytes. To study in detail about the effect of this microorganism on the function of lymphocytes. we investigated the levels of Interleukin-2 (IL-2) and Interleukin-4 (IL-4) production by T lymphocytes before and after the addition of $12.5{\;}\mu\textrm{g}/ml$ T. denticola sonicated extracts. In this study. levels of IL-2 and IL-4 produced from T cells pretreated with sonicated extracts were evaluated by using the quantitative sandwich enzyme immunoassay technique. In response to phytohemagglutinin (PHA) stimulation. T cell produced increased levels of IL-2 and IL-4. However. the expressions of both cytokines were significantly inhibited when PHA activated-T cells were pre-exposed to sonicated T. denticola extracts (p < 0.05). These findings suggest that the T. denticola sonicated extracts induced-immunosuppression in Th1 and Th2 cell functions could be a part of the pathogenic mechanism of the endodontic failure associated with this microorganism.

• Journal of Life Science
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• v.16 no.2 s.75
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• pp.231-239
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• 2006

Human mesenchymal stem cells (hMSCs) in bone marrow (BM) can be induced to differentiate into a variety of mesenchymal tissues, including adipocytes, osteoblasts and chondroblasts, under the influence of certain growth or environmental factors. In this study, we analyzed the differentiation process and the associated gene expression profiles inherent to the process by which hMSCs differentiate into osteoblasts. We conducted a comparison of gene expression profiles of the normal human BM MSCs, using human 8K cDNA microarray, incubated in media containing either a combination of $\beta$-glycerol phosphate, L-ascorbic acid, and dexamethasone, or in medium lacking these osteogenic supplements. During the osteoblastic differentiation process, 36 genes were determined to be up-regulated, and 59 genes were shown to be down-regulated. Osteoprotegerin, LRP5, and metallothionein 2A, all of which are associated with the osteogenetic process, were up-regulated, and genes associated with the differentiation of MSCs into other lineages, including muscle, adipose tissue and vascular structure were down-regulated. The set of differentially expressed genes reported in this work should contribute to our current understanding of the processes inherent to the differentiation of MSCs into osteoblasts.