scholarly journals GSK-3β at the Crossroads in Regulating Protein Synthesis and Lipid Deposition in Zebrafish

Cells ◽  
2019 ◽  
Vol 8 (3) ◽  
pp. 205 ◽  
Author(s):  
Yaqi Gu ◽  
Lili Gao ◽  
Qiang Han ◽  
Ao Li ◽  
Hairui Yu ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Protein Expression ◽  
Danio Rerio ◽  
Fatty Acid Synthase ◽  
Binding Protein ◽  
Lipid Deposition ◽  
Nonesterified Fatty Acids ◽  
Mrna And Protein Expression ◽  
Ribosomal S6 Kinase ◽  
Gsk 3Β

In this study, the mechanism by which GSK-3β regulates protein synthesis and lipid deposition was investigated in zebrafish (Danio rerio). The vector of pEGFP-N1-GSK-3β was constructed and injected into the muscle of zebrafish. It was found that the mRNA and protein expression of tuberous sclerosis complex 2 (TSC2) was significantly increased. However, the mRNA and protein expression of mammalian target of rapamycin (mTOR), p70 ribosomal S6 kinase 1 (S6K1), and 4E-binding protein 1 (4EBP1) was significantly decreased by the pEGFP-N1-GSK-3β vector in the muscle of zebrafish. In addition, the mRNA and protein expression of β-catenin, CCAAT/enhancer binding protein α (C/EBPα), and peroxisome proliferators-activated receptor γ (PPARγ) was significantly decreased, but the mRNA expression of fatty acid synthase (FAS), acetyl-CoA carboxylase (ACC), ATP-citrate lyase (ACL), and HMG-CoA reductase (HMGCR) was significantly increased by the pEGFP-N1-GSK-3β vector. The activity of FAS, ACC, ACL, and HMGCR as well as the content of triglyceride (TG), total cholesterol (TC), and nonesterified fatty acids (NEFA) were significantly increased by the pEGFP-N1-GSK-3β vector in the muscle of zebrafish. The content of free amino acids Arg, Lys, His, Phe, Leu, Ile, Val, and Thr was significantly decreased by the pEGFP-N1-GSK-3β vector. The results indicate that GSK-3β may participate in regulating protein synthesis via TSC2/mTOR signaling and regulating lipid deposition via β-catenin in the muscle of zebrafish (Danio rerio).

scholarly journals S-nitrosoglutathione inhibits adipogenesis in 3T3-L1 preadipocytes by S-nitrosation of CCAAT/enhancer-binding protein β

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Marion Mussbacher ◽  
Heike Stessel ◽  
Teresa Pirker ◽  
Antonius C. F. Gorren ◽  
Bernd Mayer ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Protein Expression ◽  
Binding Protein ◽  
Total Protein Content ◽  
Fat Cell ◽  
No Donor ◽  
Enhancer Binding Protein ◽  
Ccaat Enhancer Binding Protein ◽  
Mrna And Protein Expression ◽  
Adipogenic Effect

Abstract Murine 3T3-L1 adipocytes share many similarities with primary fat cells and represent a reliable in vitro model of adipogenesis. The aim of this study was to probe the effect of S-nitrosoglutathione (GSNO) on adipocyte differentiation. Adipogenesis was induced with a mixture of insulin, dexamethasone, and 3-isobutyl-1-methylxanthine in the absence and presence of increasing GSNO concentrations. Biochemical analysis after 7 days of differentiation showed a prominent anti-adipogenic effect of GSNO which was evident as reduced cellular triglycerides and total protein content as well as decreased mRNA and protein expression of late transcription factors (e.g. peroxisome proliferator activated receptor γ) and markers of terminal differentiation (e.g. leptin). By contrast, the nitrosothiol did not affect mRNA and protein expression of CCAAT/enhancer-binding protein β (C/EBPβ), which represents a pivotal early transcription factor of the adipogenic cascade. Differentiation was also inhibited by the NO donor (Z)-1-[2-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-ium-1,2-diolate. Biotin switch experiments showed significantly increased S-nitrosation of C/EBPβ variants indicating that posttranslational S-nitrosative modification of this transcription factor accounts for the observed anti-adipogenic effect of NO. Our results suggest that S-nitrosation might represent an important physiological regulatory mechanism of fat cell maturation.

scholarly journals Hypothermal Effects on Expression of Regucalcin, A Calcium Binding Protein, in Livers of Seawater- and Fresh Water-Acclimated Milkfish, Chanos Chanos

2021 ◽  
Author(s):  
Chia-Hao Chang ◽  
Tsung-Han Lee
Keyword(s):  
Protein Expression ◽  
Fresh Water ◽  
Calcium Binding ◽  
Binding Protein ◽  
Calcium Binding Protein ◽  
Calcium Dependent ◽  
Protein Levels ◽  
Binding Function ◽  
Mrna And Protein Expression ◽  
Apoptotic Pathways

Abstract Regucalcin (RGN) is a calcium-binding protein mainly expressed in the liver. It functions in regulating activities of several calcium-dependent enzymes related to energy metabolism, antioxidant mechanisms, and apoptotic pathways. Previous proteomics analyses revealed downregulation of regucalcin in milkfish livers when acclimated to low temperature (18°C) from normal temperature (28°C). This study first identified the full-length sequence of milkfish regucalcin from livers with high similarity in the protein structure and calcium-binding function compared to the regucalcin of other animals. The mRNA and protein expression of regucalcin in livers of fresh water (FW)- and seawater (SW)-acclimated milkfish under hypothermal acclimation were further analyzed. In FW milkfish, upregulation of regucalcin was found in mRNA and protein levels from two and four days, respectively, to one week after transfer to 18°C for the two. However, in SW milkfish, upregulation of regucalcin occurred quickly and returned to the basal levels in one (mRNA expression) or two days (protein expression) up until one week after transfer. These results suggested potential roles of regucalcin in maintaining calcium homeostasis and its correlation to differential physiological responses in livers of milkfish when they were acclimated to FW and SW.

scholarly journals MicroRNA 33 Potentially Participates in the Development of Goose Fatty Liver via Its Target Gene CROT

2021 ◽  
Author(s):  
Xiao Lin ◽  
Ya Xing ◽  
Yihui Zhang ◽  
Biao Dong ◽  
Minmeng zhao ◽  
...  
Keyword(s):  
Protein Expression ◽  
Fatty Liver ◽  
Target Gene ◽  
Liver Cells ◽  
Acid Oxidation ◽  
Lipid Deposition ◽  
Primary Hepatocytes ◽  
Regulatory Relationship ◽  
Mrna And Protein Expression ◽  
High Concentrations

Abstract Background Previous studies indicate that microRNA33 (miR-33) and its target gene, CROT, are implicated in hepatic lipid metabolism, but it is unclear whether miR-33 participates in the development of goose fatty liver via CROT. Methods The expression of miR-33 in goose fatty liver, muscle and fat tissues, as well as the mRNA and protein expression of CROT in goose fatty liver was determined by q-PCR or Western-blot. The targeting regulatory relationship between miR-33 and CROT in goose liver cells was validated by miR-33 overexpression and interference assays. The effects of miR-33 mimic and CROT overexpression on lipid deposition and the expression of downstream genes were determined in goose primary hepatocytes. The treatment of high concentrations of glucose and insulin was performed to determine their regulation on the expression of miR-33 and CROT in goose primary hepatocytes. Results Here, data showed that miR-33 expression was significantly increased in the liver, muscle and fat tissues of overfed geese. Consistently, miR-33 mimic promoted lipid deposition in goose primary hepatocytes. Moreover, the regulatory targeting relationship between miR-33 and CROT was validated in goose primary hepatocytes. Consistently, the mRNA and protein expression of CROT were significantly reduced in goose fatty liver. Interestingly, CROT overexpression could induce the expression of fatty acid oxidation associated genes including CRAT, PEX5, EHHADH, CAT and ACOT8 in goose primary hepatocytes, but only the expression of PEX5 was significantly inhibited in goose fatty liver. However, it seemed conflicting that CROT overexpression increased lipid deposition and reduced lipid peroxidation in goose primary hepatocytes. Additionally, high glucose inhibited miR-33 expression and induced CROT expression in goose primary hepatocytes. Conclusions These findings suggest that miR-33 potentially participates in the development of goose fatty liver via CROT, and that miR-33/CROT may partially mediate the effect of glucose in goose liver cells.

scholarly journals A D-vitamin metabolizmusa humán hepatocellularis carcinomában és az azt körülvevő tumormentes májszövetben

2016 ◽  
Vol 157 (48) ◽  
pp. 1910-1918 ◽  
Author(s):  
Evelin Horváth ◽  
Bernadett Balla ◽  
János Kósa ◽  
Péter András Lakatos ◽  
Áron Lazáry ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Vitamin D ◽  
Protein Synthesis ◽  
Protein Expression ◽  
Mrna Expression ◽  
Gene Activation ◽  
Human Hepatocellular Carcinoma ◽  
Rt Pcr ◽  
Antitumor Effects ◽  
Mrna And Protein Expression

Introduction: 1,25-Dihydroxy vitamin D3 mediates antitumor effects in hepatocellular carcinoma. Aim: We examined mRNA and protein expression differences in 1,25-Dihydroxy vitamin D3-inactivating CYP24A1, mRNA of activating CYP27B1 enzymes, and that of VDR between human hepatocellular carcinoma and surrounding non-tumorous liver. Methods: Snap-frozen tissues from 13 patients were studied for mRNA and protein expression of CYP24A1. Paraffin-embedded tissues from 36 patients were used to study mRNA of VDR and CYP27B1. mRNA expression was measured by RT-PCR, CYP24A1 protein was detected by immunohistochemistry. Results: Expression of VDR and CYP27B1 was significantly lower in hepatocellular carcinoma compared with non-tumorous liver (p<0.05). The majority of the HCC samples expressed CYP24A1 mRNA, but neither of the non-tumorous liver. The gene activation was followed by CYP24A1 protein synthesis. Conclusions: The presence of CYP24A1 mRNA and the reduced expression of VDR and CYP27B1 mRNA in human hepatocellular carcinoma samples indicate decreased bioavailability of 1,25-Dihydroxy vitamin D3, providing an escape mechanism from the anti-tumor effect. Orv. Hetil., 2016, 157(48), 1910–1918.

scholarly journals Bioactive Fraction of Aronia melanocarpa Fruit Inhibits Adipogenic Differentiation of Cultured 3T3-L1 Cells

2021 ◽  
Vol 11 (19) ◽  
pp. 9224
Author(s):  
Hwa-Young Lee ◽  
Kwang Sik Suh ◽  
Young Il Kim ◽  
Bong-Keun Jang ◽  
Bo-Hyung Kim ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Protein Expression ◽  
Lipid Accumulation ◽  
Binding Protein ◽  
Fatty Acid Binding ◽  
Aronia Melanocarpa ◽  
Mrna And Protein Expression ◽  
Bioactive Fraction ◽  
Oil Red O Staining ◽  
Oil Red O

Obesity is caused by excessive fat cells and the overgrowth of adipocytes and is a major risk factor for several chronic illnesses. Aronia melanocarpa fruit is rich in anthocyanins and polyphenols and has protective effects against various diseases. In this study, we examined the effect of Aronia extract (Aronia bioactive fraction, ABF®) on the biomarkers of the adipogenic pathway during adipocyte differentiation of 3T3-L1 cells. Lipid accumulation was verified by Oil Red O staining. mRNA and protein expression of lipoprotein lipase (LPL), CCAAT/enhancer-binding protein α (C/EBPα), peroxisome proliferator-activated receptor γ (PPARγ), fatty acid-binding protein 2 (FABP2), and fatty acid synthase (FAS) were assayed by RT-qPCR and Western blot analyses. Adiponectin and leptin secretion were measured using enzyme-linked immunosorbent assays. ABF® treatment downregulated lipid accumulation based on Oil Red O staining. ABF®-treated cells exhibited decreased mRNA and protein expression of LPL, C/EBPα, PPARγ, FABP2, and FAS. Moreover, ABF® treatment significantly increased adiponectin secretion and decreased leptin secretion. In conclusion, ABF® has anti-adipogenic effects on the differentiation of 3T3-L1 cells and may be used as an anti-obesity nutraceutical.

scholarly journals Regulation of (pro)renin receptor expression in mIMCD via the GSK-3β-NFAT5-SIRT-1 signaling pathway

2014 ◽  
Vol 307 (5) ◽  
pp. F593-F600 ◽  
Author(s):  
Syed Quadri ◽  
Helmy M. Siragy
Keyword(s):  
Protein Expression ◽  
Signaling Pathway ◽  
Collecting Duct ◽  
Receptor Expression ◽  
Low Salt ◽  
Mrna And Protein Expression ◽  
Collecting Duct Cells ◽  
Medullary Collecting Duct ◽  
Gsk 3Β ◽  
Si Rna

The localization and regulation of (pro)renin receptor (PRR) expression in kidney collecting duct cells are not well established. We hypothesized that low salt (LS) contributes to the regulation of PRR expression in these cells via the GSK-3β-NFAT5-sirtuin1 (SIRT-1) signaling pathway. Mouse inner medullary collecting duct (mIMCD) cells were treated with NaCl at 130 (normal salt; NS), 63 (LS), or 209 mM (high salt; HS) alone or in combination with NFAT5 scrambled small interfering (si) RNA, NFAT5 siRNA, or the SIRT-1 inhibitor EX-527. Compared with NS, LS increased the mRNA and protein expression of PRR by 71% and 69% ( P < 0.05), and reduced phosphorylation of GSK-3β by 62% ( P < 0.01), mRNA and protein expressions of NFAT5 by 65% and 45% ( P < 0.05), and SIRT-1 by 44% and 50% ( P < 0.01), respectively. LS also enhanced p65 NF-κB by 102% ( P < 0.01). Treatment with HS significantly reduced the mRNA and protein expression of PRR by 32% and 23% ( P < 0.05), and increased the mRNA and protein expression of NFAT5 by 39% and 45% ( P < 0.05) and SIRT-1 by 51% and 56% ( P < 0.05), respectively. HS+NFAT5 siRNA reduced the mRNA and protein expression of NFAT5 by 51% and 35% ( P < 0.01) and increased the mRNA and protein expression of PRR by 148% and 70% ( P < 0.01), respectively. HS+EX-527 significantly increased the mRNA and protein expression of PRR by 96% and 58% ( P < 0.05), respectively. We conclude that expression of PRR in mIMCD cells is regulated by the GSK-3β-NFAT5- SIRT-1 signaling pathway.

scholarly journals Pulmonary artery smooth muscle hypertrophy: roles of glycogen synthase kinase-3β and p70 ribosomal S6 kinase

2010 ◽  
Vol 298 (6) ◽  
pp. L793-L803 ◽  
Author(s):  
Huan Deng ◽  
Marc B. Hershenson ◽  
Jing Lei ◽  
Anuli C. Anyanwu ◽  
David J. Pinsky ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Protein Expression ◽  
Cell Size ◽  
Translational Control ◽  
Muscle Hypertrophy ◽  
Glycogen Synthase ◽  
Contractile Protein ◽  
Ribosomal S6 Kinase ◽  
Gsk 3Β ◽  
Tgf Β1

Increased medial arterial thickness is a structural change in pulmonary arterial hypertension (PAH). The role of smooth muscle hypertrophy in this process has not been well studied. Bone morphogenetic proteins (BMPs), transforming growth factor (TGF)-β1, serotonin (or 5-hydroxytryptamine; 5-HT), and endothelin (ET)-1 have been implicated in PAH pathogenesis. We examined the effect of these mediators on human pulmonary artery smooth muscle cell size, contractile protein expression, and contractile function, as well on the roles of glycogen synthase kinase (GSK)-3β and p70 ribosomal S6 kinase (p70S6K), two proteins involved in translational control, in this process. Unlike epidermal growth factor, BMP-4, TGF-β1, 5-HT, and ET-1 each increased smooth muscle cell size, contractile protein expression, fractional cell shortening, and GSK-3β phosphorylation. GSK-3β inhibition by lithium or SB-216763 increased cell size, protein synthesis, and contractile protein expression. Expression of a non-phosphorylatable GSK-3β mutant blocked BMP-4-, TGF-β1-, 5-HT-, and ET-1-induced cell size enlargement, suggesting that GSK-3β phosphorylation is required and sufficient for cellular hypertrophy. However, BMP-4, TGF-β1, 5-HT, and ET-1 stimulation was accompanied by an increase in serum response factor transcriptional activation but not eIF2 phosphorylation, suggesting that GSK-3β-mediated hypertrophy occurs via transcriptional, not translational, control. Finally, BMP-4, TGF-β1, 5-HT, and ET-1 treatment induced phosphorylation of p70S6K and ribosomal protein S6, and siRNAs against p70S6K and S6 blocked the hypertrophic response. We conclude that mediators implicated in the pathogenesis of PAH induce pulmonary arterial smooth muscle hypertrophy. Identification of the signaling pathways regulating vascular smooth muscle hypertrophy may define new therapeutic targets for PAH.

scholarly journals Targeting Glycogen Synthase Kinase 3 Beta Regulates CD47 Expression After Myocardial Infarction in Rats via the NF-κB Signaling Pathway

2021 ◽  
Vol 12 ◽  
Author(s):  
Li-Na Xu ◽  
Shu-Hui Wang ◽  
Xue-Ling Su ◽  
Sumra Komal ◽  
Hong-Kun Fan ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Protein Expression ◽  
Glycogen Synthase ◽  
Cell Damage ◽  
H9c2 Cell ◽  
H9c2 Cells ◽  
Rat Cardiomyocytes ◽  
Hypoxic Conditions ◽  
Mrna And Protein Expression ◽  
Gsk 3Β

The aim of this study was to investigate the effects of the GSK-3β/NF-κB pathway on integrin-associated protein (CD47) expression after myocardial infarction (MI) in rats. An MI Sprague Dawley rat model was established by ligating the left anterior descending coronary artery. The rats were divided into three groups: Sham, MI, and SB + MI (SB216763) groups. Immunohistochemistry was used to observe the changes in cardiac morphology. A significant reduction in the sizes of fibrotic scars was observed in the SB + MI group compared to that in the MI group. SB216763 decreased the mRNA and protein expression of CD47 and NF-κB during MI. Primary rat cardiomyocytes (RCMs) and the H9c2 cell line were used to establish in vitro hypoxia models. Quantitative real-time PCR and western blotting analyses were conducted to detect mRNA and protein expression levels of CD47 and NF-κB and apoptosis-related proteins, respectively. Apoptosis of hypoxic cells was assessed using flow cytometry. SB216763 reduced the protein expression of CD47 and NF-κB in RCMs and H9c2 cells under hypoxic conditions for 12 h, and alleviated hypoxia-induced apoptosis. SN50 (an NF-κB inhibitor) also decreased CD47 protein expression in RCMs and H9c2 cells under hypoxic conditions for 12 h and protected cells from apoptosis. GSK-3β upregulates CD47 expression in cardiac tissues after MI by activating NF-κB, which in turn leads to myocardial cell damage and apoptosis.

scholarly journals Effects of WNT1 c.110 T>C and c.505G>T mutations on osteoblast differentiation via the WNT1/β-catenin signaling pathway

2021 ◽  
Vol 16 (1) ◽  
Author(s):  
Bashan Zhang ◽  
Rong Li ◽  
Wenfeng Wang ◽  
Xueming Zhou ◽  
Beijing Luo ◽  
...  
Keyword(s):  
Protein Expression ◽  
Signaling Pathway ◽  
Osteoblast Differentiation ◽  
Missense Mutations ◽  
Wild Type ◽  
Expression Levels ◽  
Type Group ◽  
In The Wild ◽  
Mrna And Protein Expression ◽  
Gsk 3Β

Abstract Background WNT1 c.110 T>C and c.505G>T missense mutations have been identified in patients with osteogenesis imperfecta (OI). Whether these mutations affect osteoblast differentiation remains to be determined. This study aimed to investigate the effects of WNT1 c.110 T>C and c.505G>T mutations on osteoblast function, gene expression, and pathways involved in OI. Methods Empty vector (negative control), wild-type WNT1, WNT1 c.110 T>C, WNT1 c.505G>T, and WNT1 c.884C>A (positive control) mutant plasmids were constructed and transfected into preosteoblast (MC3T3-E1) cells to investigate their effect on osteoblast differentiation. The expressions of osteoblast markers, including BMP2, RANKL, osteocalcin, and alkaline phosphatase (ALP), were determined using quantitative real-time polymerase chain reaction (RT-qPCR), western blotting (WB), enzyme-linked immunosorbent assay, and ALP staining assay, respectively. The mRNA and protein expression levels of WNT1 or the expression levels of the relevant proteins involved in the WNT1/β-catenin signaling pathway were also determined using RT-qPCR, WB, and immunofluorescence (IF) assays after the different plasmids were transfected into MC3T3-E1 cells. Results Compared with those in the wild-type group, in the mutation groups, the mRNA and protein expression levels of BMP2 were suppressed, the expressions of osteocalcin and ALP were inhibited, and the mRNA and protein expression levels of RANKL were enhanced in MC3T3-E1 cells. WB and IF assays revealed that the protein expression levels of WNT1 in MC3T3-E1 cells were downregulated in the mutation groups compared with those in the wild-type WNT1 group. Furthermore, the expression levels of nonphosphorylated β-catenin (non-p-β-catenin) and phosphorylated GSK-3β (p-GSK-3β) were downregulated in the mutation groups compared with those in the wild-type group. However, no significant changes in the expression level of non-p-β-catenin or p-GSK-3β were observed in the mutation groups. Conclusions WNT1 c.110 T>C and c.505G>T mutations may alter the proliferation and osteogenic phenotype of MC3T3-E1 linked to the progression of OI via the inhibition of the WNT1/β-catenin signaling pathway. This is the first study to confirm the effect of WNT1 c.110 T>C and c.505G>T missense mutations on osteoblast differentiation and propose a new molecular mechanism for OI development.

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