Effect of Neudesin Neurotrophic Factor on Differentiation of Bovine Preadipocytes and Myoblasts in a Co-Culture System

In this study, we successfully established a co-culture system of bovine preadipocytes and myoblasts to explore the effect of exogenous addition of Neudesin neurotrophic factor (NENF) recombinant protein on the differentiation of adipocytes and myoblasts in co-culture. The optimal concentration of NENF recombinant protein was 100 pg/mL. NENF promoted the differentiation of bovine preadipocytes and inhibited the differentiation of bovine myoblasts when the cells were cultured separately. After adding NENF recombinant protein to the co-culture system, the accumulation of lipid droplets in bovine preadipocytes decreased, but the differentiation of bovine myoblasts did not change significantly. The results of real-time quantitative PCR (RT-qPCR) and Western blot showed that the expression levels of adipogenesis-related factors such as PPARγ, FABP4 and FASN were significantly down-regulated at the mRNA and protein levels in adipocytes, while myogenic marker genes such as MYOD1, MYOG and MYHC had no significant changes at the mRNA or protein levels in myoblasts. This differs from, and potentially conflicts with, the monoculture system, where NENF expression in each cell type changed with the cell microenvironment. Consequently, the molecular mechanism of marbling beef formation cannot be fully revealed using monocultures of adipocytes or myocytes.

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4-Hydroxyisoleucine relieves inflammation through iRhom2-dependent pathway in co-cultured macrophages and adipocytes with LPS stimulation

BMC Complementary Medicine and Therapies ◽

10.1186/s12906-020-03166-1 ◽

2020 ◽

Vol 20 (1) ◽

Author(s):

Cong Zhou ◽

Rui Chen ◽

Feng Gao ◽

Jiaoyue Zhang ◽

Furong Lu

Keyword(s):

Culture System ◽

Real Time Quantitative Pcr ◽

Protein Levels ◽

Raw264.7 Macrophages ◽

Chemotactic Protein ◽

Tnf Α ◽

Mrna And Protein Expression ◽

Anti Inflammatory ◽

Factor Α ◽

Dependent Pathway

Abstract Background 4-Hydroxyisoleucine (4-HIL) is an active ingredient extracted from Trigonella foenum-graecum L., a Chinese traditional herbal medicine, which exerts the efficacy of anti-obesity and anti-diabetes. We previously reported that 4-HIL potentiates anti-inflammatory and anti-insulin resistance effects through down-regulation of TNF-α and TNF-α converting enzyme (TACE) in 3 T3-L1 adipocytes and HepG2 cells. In the present study, we further investigate the effects and mechanisms of 4-HIL on obesity-induced inflammation in RAW264.7 macrophages and 3 T3-L1 adipocytes co-culture system. Methods RAW264.7 macrophages and 3 T3-L1 adipocytes were co-cultured to mimic the microenvironment of adipose tissue. siRNA-iRhom2 transfection was performed to knockdown iRhom2 expression in RAW264.2 macrophages. The mRNA and protein expression of iRhom2 and TACE were measured by real-time quantitative PCR (RT-qPCR) and western blotting. The production of tumor necrosis factor-α (TNF-α), monocyte chemotactic protein-1 (MCP-1), IL-6 and IL-10 were evaluated by ELISA. The ratio of M2/M1 was detected by flow cytometry. Results 4-HIL significantly repressed the mRNA and protein levels of iRhom2 and TACE in RAW264.7 macrophages after LPS stimulated. Meanwhile, the levels of pro-inflammatory cytokines, including TNF-α, MCP-1, and IL-6, were substantially suppressed by 4-HIL in the co-culture system. Moreover, the level of anti-inflammatory cytokine IL-10 was increased significantly by 4-HIL in the co-culture system after LPS stimulation. Additionally, the ratio of M2/M1 was also increased by 4-HIL in the co-culture system after LPS stimulation. Finally, these effects of 4-HIL were largely enhanced by siRNA-iRhom2 transfection. Conclusion Taken together, our results indicated that obesity-induced inflammation was potently relieved by 4-HIL, most likely through the iRhom2-dependent pathway.

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Brain-derived neurotrophic factor is associated with coronary macrophage infiltrates in patients with acute coronary syndrome

European Heart Journal ◽

10.1093/ehjci/ehaa946.1300 ◽

2020 ◽

Vol 41 (Supplement_2) ◽

Author(s):

R.A Montone ◽

M Camilli ◽

M Russo ◽

M Del Buono ◽

F Gurguglione ◽

...

Keyword(s):

Acute Coronary Syndrome ◽

Neurotrophic Factor ◽

Serum Levels ◽

Brain Derived Neurotrophic Factor ◽

C Reactive Protein ◽

St Segment Elevation ◽

Protein Levels ◽

Reactive Protein ◽

Coronary Syndrome ◽

St Segment

Abstract Background Brain-derived neurotrophic factor (BDNF) is a neurotrophine that plays a key role in the regulation of both central and peripheral nervous system. Moreover, BDNF is secreted in multiple tissues and exerts systemic, autocrine, and paracrine effects in the cardiovascular system. Of importance, BDNF expression was enhanced in macrophages and smooth muscle cells in atherosclerotic coronary arteries and may be involved in thrombus formation. Thus, BDNF has been suggested as an important link between inflammation and thrombosis, potentially involved in the pathogenesis of acute coronary syndrome (ACS). Purpose In our study we aimed at assessing serum levels of BDNF in patients with ACS, evaluating differences according to clinical presentation [ST-segment elevation myocardial infarction (STEMI) vs. Non-ST-segment elevation ACS (NSTE-ACS)]. Moreover, we assessed the presence of optical coherence (OCT)-defined macrophage infiltrates (MØI) in the culprit vessel of ACS patients and evaluated their relationship with BDNF levels. Methods ACS patients were prospectively selected. Blood samples were collected at admission and serum levels of BDNF were subsequently assessed. Presence of OCT-defined MØI along the culprit vessel was assessed. Results 166 ACS patients were enrolled [mean age 65.3±11.9 years, 125 (75.3%) male, 109 STEMI, 57 NSTE-ACS]. Serum levels of BDNF were higher among STEMI patients compared with NSTE-ACS [median (IQR) 2.48 pg/mL (1.54–3.34) vs. 2.12 pg/mL (1.34–2.47), p=0.007], while C-reactive protein levels did not differ between the two groups. OCT assessment was performed in 53 patients and MØI were detected in 27 patients. Of importance, patients with MØI in the culprit vessel had higher levels of BDNF compared with patients without MØI [median (IQR) 2.23 pg/mL (1.38–2.53) vs. 1.41 pg/mL (0.93–2.07), p=0.023], while C-reactive protein levels did not differ between the two groups. Of note, at multivariate regression analysis BDNF levels were independent predictor of MØI [OR: 2.20; 95% CI (1.02–4.74), p=0.043]. Conclusions Serum levels of BDNF may reliable identify the presence of local macrophage inflammatory infiltrates in patients with ACS. Moreover, BDNF levels are higher in patients with STEMI compared with NSTE-ACS. Taken together, these data suggest that BDNF may represent an interesting link between local inflammatory activation and enhanced thrombosis in ACS. BDNF serum levels Funding Acknowledgement Type of funding source: None

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Alterations in BDNF Protein Concentrations in the Hippocampus do not Explain the Pro-Neurogenic Effect of Citalopram on Adult Neurogenesis

Pharmacopsychiatry ◽

10.1055/a-1291-8079 ◽

2020 ◽

Author(s):

Markus Petermann ◽

Golo Kronenberg ◽

Valentina Mosienko ◽

Michael Bader ◽

Natalia Alenina ◽

...

Keyword(s):

Cell Survival ◽

Mechanism Of Action ◽

Adult Neurogenesis ◽

Neurotrophic Factor ◽

Selective Serotonin Reuptake Inhibitors ◽

Daily Injection ◽

Wild Type ◽

Protein Levels ◽

Serotonin System ◽

Antidepressant Pharmacotherapy

Abstract Introduction Brain-derived neurotrophic factor (BDNF) has been implicated in the pro-neurogenic effect of selective serotonin reuptake inhibitors. In this study, we used Tph2 −/− mice lacking brain serotonin to dissect the interplay between BDNF and the serotonin system in mediating the effects of antidepressant pharmacotherapy on adult neurogenesis in the hippocampus. Methods Besides citalopram (CIT), we tested tianeptine (TIA), an antidepressant whose mechanism of action is not well understood. Specifically, we examined cell survival and endogenous concentrations of BDNF following daily injection of the drugs. Results Twenty-one days of CIT, but not of TIA, led to a significant increase in the survival of newly generated cells in the dentate gyrus of wild-type mice, without a significant effect on BDNF protein levels by either treatment. In Tph2 −/− mice, adult neurogenesis was consistently increased. Furthermore, Tph2 −/− mice showed increased BDNF protein levels, which were not affected by TIA but were significantly reduced by CIT. Discussion We conclude that the effects of CIT on adult neurogenesis are not explained by changes in BDNF protein concentrations in the hippocampus.

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Traditional Herbal Formula Oyaksungi-San Inhibits Adipogenesis in 3T3-L1 Adipocytes

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2015/949461 ◽

2015 ◽

Vol 2015 ◽

pp. 1-10 ◽

Cited By ~ 4

Author(s):

Sae-Rom Yoo ◽

Chang-Seob Seo ◽

Hyeun-Kyoo Shin ◽

Soo-Jin Jeong

Keyword(s):

Binding Protein ◽

Peroxisome Proliferator ◽

Herbal Formula ◽

Related Factors ◽

Peroxisome Proliferator Activated Receptor ◽

Fatty Acid Binding ◽

Gpdh Activity ◽

Protein Levels ◽

Intracellular Lipid Accumulation ◽

Glycerol 3 Phosphate Dehydrogenase

Background. Oyaksungi-san (OYSGS) is a herbal formula that has been used for treating cardiovascular diseases in traditional Asian medicine. Here, we investigated the antiadipogenic effect of OYSGS extract in 3T3-L1 adipose cells.Methods. 3T3-L1 preadipocytes were differentiated into adipocytes with or without OYSGS. After differentiation, we measured Oil Red O staining, glycerol-3-phosphate dehydrogenase (GPDH) activity, leptin production, mRNA, and protein levels of adipogenesis-related factors.Results. OYSGS extract dramatically inhibited intracellular lipid accumulation in the differentiated adipocytes. It also significantly suppressed the (GPDH) activity, triglyceride (TG) content, and leptin production by reducing the expression of adipogenesis-related genes including lipoprotein lipase, fatty acid binding protein 4, CCAAT/enhancer-binding protein-alpha (C/EBP-α), and peroxisome proliferator-activated receptor gamma (PPAR-γ). Furthermore, OYSGS clearly enhanced phosphorylation of AMP-activated protein kinase (AMPK) as well as its substrate acetyl CoA (ACC) carboxylase.Conclusions. Our results demonstrate that OYSGS negatively controls TG accumulation in 3T3-L1 adipocytes. We suggest antiadipogenic activity of OYSGS and its potential benefit in preventing obesity.

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Exosomal KLF3-AS1 from hMSCs promoted cartilage repair and chondrocyte proliferation in osteoarthritis

Biochemical Journal ◽

10.1042/bcj20180675 ◽

2018 ◽

Vol 475 (22) ◽

pp. 3629-3638 ◽

Cited By ~ 43

Author(s):

Yubao Liu ◽

Rui Zou ◽

Zhen Wang ◽

Chuanyang Wen ◽

Fan Zhang ◽

...

Keyword(s):

Flow Cytometry ◽

Cartilage Repair ◽

Rat Model ◽

Human Mesenchymal Stem Cells ◽

Type Ii Collagen ◽

Morphological Observation ◽

Chondrocyte Proliferation ◽

Real Time Quantitative Pcr ◽

Protein Levels

The present study was designed to explore whether exosomal lncRNA-KLF3-AS1 derived from human mesenchymal stem cells (hMSCs) can serve as a positive treatment for osteoarthritis (OA). hMSCs and MSC-derived exosomes (MSC-exo) were prepared for morphological observation and identification by transmission electron microscopy and flow cytometry. IL-1β-induced OA chondrocytes and collagenase-induced rat model of OA were established for the further experiments. Lentivirus-mediated siRNA targeting KLF3-AS1 was transfected into MSCs for silencing KLF3-AS1. The real-time quantitative PCR and western blotting analysis were performed to examine the mRNA and protein levels of type II collagen alpha 1 (Col2a1), aggrecan, matrix metalloproteinase 13 and runt-related transcription factor 2. Cell proliferation, apoptosis and migration were evaluated by CCK-8 assay, flow cytometry and transwell assay. HE (hematoxylin and eosin) staining and immunohistochemistry were used for histopathological studies. MSC-exo ameliorated IL-1β-induced cartilage injury. Furthermore, lncRNA KLF3-AS1 was markedly enriched in MSC-exo, and exosomal KLF3-AS1 suppressed IL-1β-induced apoptosis of chondrocytes. Further in vivo investigation indicated that exosomal KLF3-AS1 promoted cartilage repair in a rat model of OA. Exosomal KLF3-AS1 promoted cartilage repair and chondrocyte proliferation in a rat model of OA, which might be an underlying therapeutic target for OA.

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Role of Luteolin-Induced Apoptosis and Autophagy in Human Glioblastoma Cell Lines

Medicina ◽

10.3390/medicina57090879 ◽

2021 ◽

Vol 57 (9) ◽

pp. 879 ◽

Cited By ~ 1

Author(s):

Hye-Sung Lee ◽

Bong-Soo Park ◽

Hae-Mi Kang ◽

Jung-Han Kim ◽

Sang-Hun Shin ◽

...

Keyword(s):

Cell Lines ◽

Therapeutic Effects ◽

Dependent Manner ◽

Glioblastoma Cell ◽

Related Factors ◽

Green Pepper ◽

Real Time Quantitative Pcr ◽

Anti Inflammatory ◽

Induced Apoptosis ◽

Glioblastoma Cell Lines

Background and Objectives: Malignant glioblastoma (GBM) is caused by abnormal proliferation of glial cells, which are found in the brain. The therapeutic effects of surgical treatment, radiation therapy, and chemo-therapy against GBM are relatively poor compared with their effects against other tumors. Luteolin is abundant in peanut shells and is also found in herbs and other plants, such as thyme, green pepper, and celery. Luteolin is known to be effective against obesity and metabolic syndrome. The anti-inflammatory, and anti-cancer activities of luteolin have been investigated. Most studies have focused on the antioxidant and anti-inflammatory effects of luteolin, which is a natural flavonoid. However, the association between the induction of apoptosis by luteolin in GBM and autophagy has not yet been investigated. This study thus aimed to confirm the occurrence of luteolin-induced apoptosis and autophagy in GBM cells and to assess their relationship. Materials and Methods: A172 and U-373MG glioblastoma cell lines were used for this experiment. We confirmed the apoptosis effect of Luteolin on GBM cells using methods such as 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, immunofluorescence, Flow cytometry (FACS) western blot, and real-time quantitative PCR (qPCR). Results: In the luteolin-treated A172 and U-373MG cells, cell viability decreased in a concentration- and time-dependent manner. In addition, in A172 and U-373MG cells treated with luteolin at concentrations greater than 100 μM, nuclear fragmentation, which is a typical morphological change characterizing apoptosis, as well as fragmentation of caspase-3 and Poly (ADP-ribose) polymerase (PARP), which are apoptosis-related factors, were observed. Autophagy was induced after treatment with at least 50 μM luteolin. Inhibition of autophagy using 3MA allowed for a low concentration of luteolin to more effectively induce apoptosis in A172 and U-373MG cells. Conclusions: Results showed that luteolin induces apoptosis and autophagy and that the luteolin-induced autophagy promotes cell survival. Therefore, an appropriate combination therapy involving luteolin and an autophagy inhibitor is expected to improve the prognosis of GBM treatment.

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Early Alcohol Withdrawal Reverses the Abnormal Levels of proBDNF/mBDNF and their Receptors

10.21203/rs.3.rs-337394/v1 ◽

2021 ◽

Author(s):

Li Zhou ◽

Jing Xiong ◽

Chang-qing Gao ◽

Jian-jun Bao ◽

Xin-Fu Zhou

Keyword(s):

Serum Level ◽

Alcohol Dependence ◽

Alcohol Withdrawal ◽

Neurotrophic Factor ◽

Ethanol Exposure ◽

Enzyme Linked Immunosorbent Assay ◽

Western Blots ◽

Time Points ◽

Protein Levels ◽

Chronic Ethanol Exposure

Abstract ObjectiveProlonged excessive ethanol intake impairs learning, memory and also causes brain atrophy. Brain-derived neurotrophic factor (BDNF) plays pivotal roles in the pathology of alcohol dependence. Our previous work found that chronic ethanol exposure altered the metabolism of BDNF, leading to the imbalance of proBDNF and mature BDNF (mBDNF). In this study, we hypothesized that early alcohol withdrawal would reverse the abnormal levels of proBDNF, mBDNF and their receptors.Method30 male alcohol dependence patients were recruited. Peripheral blood was sampled from all the subjects before and one week after alcohol withdrawal. The lymphocyte protein levels of proBDNF, p75NTR, sortilin and TrkB were analyzed by western blots and the serum level of mBDNF and TrkB was assayed by sandwich enzyme-linked immunosorbent assay (ELISA) at two different time points. ResultsThe levels of mBDNF and its receptor (TrkB) increased, oppositely the levels of proBDNF and its receptors (p75NTR and sortilin) decreased one week after alcohol withdrawal. ConclusionsEarly alcohol withdrawal reversed the abnormal levels of proBDNF, mBDNF and their receptors. The shift levels of proBDNF and mBDNF were both taken in the pathology of alcohol withdrawal.

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Bone Marrow Stromal Cells (BMSCs) Inhibit Retinal Ganglion Cell Apoptosis and Alleviate Inflammation Through Up-Regulation of MicroRNA-43 and Brain-Derived Neurotrophic Factor in Glaucoma

Journal of Biomaterials and Tissue Engineering ◽

10.1166/jbt.2021.2850 ◽

2021 ◽

Vol 11 (12) ◽

pp. 2478-2483

Author(s):

Xiang Ji ◽

Kai-Wen Zhou

Keyword(s):

Neurotrophic Factor ◽

Vision Loss ◽

Luciferase Reporter ◽

Retinal Ganglion ◽

Luciferase Reporter Gene ◽

Down Regulation ◽

Related Factors ◽

Gene Assay

Glaucoma is a leading cause of vision loss mainly due to retinal ganglion cells (RGC) loss. MicroRNAs (miRNAs) are highlighted as potential biomarkers in diseases. This study aims to investigate the role of miR-43 and BMSCs in the RGC apoptosis and glaucoma.RGCs were transfected with miR-43 inhibitors and mimics, and then co-cultured with BMSCs. RT-qPCR analysis was conducted to determine miR-43 expression, whilst Western blot, and flow cytometry were carried out to assess the role of miR-43 in apoptosis and inflammation. The interaction between miR-43 and BDNF, a neurotrophic factor, was detected by dual-luciferase reporter gene assay. Overexpression of miR-43 promoted RGC proliferation and decreased apoptosis. Furthermore, miR-43 overexpression diminished the contents of apoptosis- and inflammatory-related factors, and elevated the expression of BDNF. Down-regulation of BDNF exerted similar effect as down-regulation of miR-43, enhancing apoptosis and aggravating inflammation. Importantly, BMSC treatment reversed the in vitro inhibitory effect of si-BDNF on RGC with enhancement of miR-43 expression. Mechanically, miR-43 was indicated to target BDNF in glaucoma. Collectively, miR-43 delivered by BMSCs plays an important role in the inflammatory injury and abnormal apoptosis of RGC by regulating the expression of BDNF. These findings might help development of new treatment for glaucoma and provide a promising biomarker for diagnosis and treatment.

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Browning Effects of a Chronic Pterostilbene Supplementation in Mice Fed a High-Fat Diet

International Journal of Molecular Sciences ◽

10.3390/ijms20215377 ◽

2019 ◽

Vol 20 (21) ◽

pp. 5377 ◽

Cited By ~ 3

Author(s):

Martina La Spina ◽

Eva Galletta ◽

Michele Azzolini ◽

Saioa Gomez Zorita ◽

Sofia Parrasia ◽

...

Keyword(s):

Adipose Tissue ◽

White Adipose Tissue ◽

High Fat Diet ◽

Uncoupling Protein ◽

Protein A ◽

Health Concern ◽

Marker Genes ◽

High Fat ◽

Protein Levels

Obesity and related comorbidities are a major health concern. The drugs used to treat these conditions are largely inadequate or dangerous, and a well-researched approach based on nutraceuticals would be highly useful. Pterostilbene (Pt), i.e., 3,5-dimethylresveratrol, has been reported to be effective in animal models of obesity, acting on different metabolic pathways. We investigate here its ability to induce browning of white adipose tissue. Pt (5 µM) was first tested on 3T3-L1 mature adipocytes, and then it was administered (352 µmol/kg/day) to mice fed an obesogenic high-fat diet (HFD) for 30 weeks, starting at weaning. In the cultured adipocytes, the treatment elicited a significant increase of the levels of Uncoupling Protein 1 (UCP1) protein—a key component of thermogenic, energy-dissipating beige/brown adipocytes. In vivo administration antagonized weight increase, more so in males than in females. Analysis of inguinal White Adipose Tissue (WAT) revealed a trend towards browning, with significantly increased transcription of several marker genes (Cidea, Ebf2, Pgc1α, PPARγ, Sirt1, and Tbx1) and an increase in UCP1 protein levels, which, however, did not achieve significance. Given the lack of known side effects of Pt, this study strengthens the candidacy of this natural phenol as an anti-obesity nutraceutical.

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LINC01128 expedites cervical cancer progression by regulating miR-383-5p/SFN axis

BMC Cancer ◽

10.1186/s12885-019-6326-5 ◽

2019 ◽

Vol 19 (1) ◽

Cited By ~ 5

Author(s):

Yi Hu ◽

Yan Ma ◽

Jie Liu ◽

Yanlin Cai ◽

Mengmeng Zhang ◽

...

Keyword(s):

Cervical Cancer ◽

Cell Proliferation ◽

Cancer Progression ◽

Cell Apoptosis ◽

High Expression ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Real Time Quantitative Pcr ◽

Protein Levels ◽

Normal Tissues

Abstract Background Cervical cancer (CC), causing significant morbidity and mortality worldwide, is one of the most common gynecological malignancies in women. SFN has been reported as a potential prognostic marker with apparent high expression in tumors. Nevertheless, the function mechanism of SFN is not clear yet in CC. Methods The relative expressions of RNAs were detected by real-time quantitative PCR (RT-qPCR). Colony formation assay, EdU stained assay and CCK-8 assay were to check cell proliferation ability in CC. Flow cytometry and apoptosis related proteins analysis were used to measure cells apoptosis capacity. Luciferase reporter assay and RNA pull down assay were to verify the molecular mechanism. Results SFN was highly expressed in CC tissues and CC cell lines compared with normal tissues and normal cell line. After interfering SFN, cell proliferation, migration and invasion ability was inhibited as well as cell apoptosis ability was promoted. In subsequence, miR-383-5p exhibited conspicuous low expression in CC tissues. And miR-383-5p was found to bind to SFN and have anti-cancerous effects in CC. Moreover, LINC01128 displayed remarkable high expression in CC tissues. Besides, LINC01128 shortage could reduce the expression of SFN at mRNA and protein levels. And the affinity between LINC01128 and miR-383-5p was verified. In the end, it was proved that LINC01128 could enhance cell proliferation, migration and invasion as well as inhibit cell apoptosis by binding with miR-383-5p and upregulating SFN. Conclusion LINC01128 expedited cells cellular process in CC by binding with miR-383-5p to release SFN. Graphical Abstract

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