scholarly journals EPA and DHA Inhibit Myogenesis and Downregulate the Expression of Muscle-related Genes in C2C12 Myoblasts

Genes ◽  
2019 ◽  
Vol 10 (1) ◽  
pp. 64 ◽  
Author(s):  
Jing Zhang ◽  
Xin Xu ◽  
Yan Liu ◽  
Lin Zhang ◽  
Jack Odle ◽  
...  
Keyword(s):  
Gene Expression ◽  
Skeletal Muscle ◽  
Cell Proliferation ◽  
Cell Viability ◽  
Rna Sequencing ◽  
Cell Counting ◽  
Sequencing Analysis ◽  
C2c12 Myoblasts ◽  
Epa And Dha ◽  
Proliferation And Differentiation

This study was conducted to elucidate the biological effects of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) on cell proliferation, differentiation and gene expression in C2C12 myoblasts. C2C12 were treated with various concentrations of EPA or DHA under proliferation and differentiation conditions. Cell viability was analyzed using cell counting kit-8 assays (CCK-8). The Edu assays were performed to analyze cell proliferation. To analyze cell differentiation, the expressions of myogenic marker genes were determined at the transcriptional and translational levels by qRT-PCR, immunoblotting and immunofluorescence. Global gene expression patterns were characterized using RNA-sequencing. Phosphorylation levels of ERK and Akt were examined by immunoblotting. Cell viability and proliferation was significantly inhibited after incubation with EPA (50 and 100 μM) or DHA (100 μM). Both EPA and DHA suppressed C2C12 myoblasts differentiation. RNA-sequencing analysis revealed that some muscle-related genes were significantly downregulated following EPA or DHA (50 μM) treatment, including insulin-like growth factor 2 (IGF-2), troponin T3 (Tnnt3), myoglobin (Mb), myosin light chain phosphorylatable fast skeletal muscle (Mylpf) and myosin heavy polypeptide 3 (Myh3). IGF-2 was crucial for the growth and differentiation of skeletal muscle and could activate the PI3K/Akt and the MAPK/ERK cascade. We found that EPA and DHA (50 μM) decreased the phosphorylation levels of ERK1/2 and Akt in C2C12 myoblasts. Thus, this study suggested that EPA and DHA exerted an inhibitory effect on myoblast proliferation and differentiation and downregulated muscle-related genes expression.

scholarly journals The influence of direct and indirect fibroblast cell contact on human myogenic cell behavior and gene expression in vitro

2019 ◽  
Vol 127 (2) ◽  
pp. 342-355 ◽  
Author(s):  
Cecilie J. L. Bechshøft ◽  
Peter Schjerling ◽  
Michael Kjaer ◽  
Abigail L. Mackey
Keyword(s):  
Gene Expression ◽  
Skeletal Muscle ◽  
Cell Proliferation ◽  
Myogenic Cell ◽  
Mrna Levels ◽  
Indirect Contact ◽  
Permeable Membrane ◽  
Primary Myoblasts ◽  
Cell Proliferation And Differentiation ◽  
Proliferation And Differentiation

Underpinning skeletal muscle plasticity is the interplay between many cell types, of which fibroblasts are emerging as potent players, both negatively in the development of fibrosis but also positively in stimulating muscle repair through enhancing myogenesis. The mechanisms behind this interaction however remain unknown. To investigate this, waste hamstring muscle tissue was obtained from eight healthy young men undergoing reconstructive anterior cruciate ligament surgery and primary myoblasts and fibroblasts were isolated. Myoblasts were cultured alone or with fibroblasts, either in direct or indirect contact (separated by an insert with a permeable membrane). The myogenesis parameters proliferation, differentiation, and fusion were determined from immunostained cells, while, in replicate samples, gene expression levels of GAPDH, Ki67, Pax7, MyoD, myogenin, myomaker, MHC-Iβ, TCF7L2, COL1A1, and p16 were determined by RT-PCR. We found only trends for an influence of skeletal muscle fibroblasts on myogenic cell proliferation and differentiation. While greater mRNA levels of GAPDH, Pax7, MyoD, myogenin, and MHC-Iβ were observed in myogenic cells in indirect contact with fibroblasts (insert) when compared with cells cultured alone, a similar effect of an empty insert was also observed. In conclusion we find very little influence of skeletal muscle fibroblasts on myoblasts derived from the same tissue, although it cannot be excluded that a different outcome would be seen under less optimal myogenic growth conditions. NEW & NOTEWORTHY Using passage one primary myoblasts and fibroblasts isolated from human skeletal muscle, we found only a trend for an effect of skeletal muscle fibroblasts on myogenic cell proliferation and differentiation. This is contrary to previous reports and raises the possibility that fibroblasts of different tissue origins exert distinct roles.

scholarly journals Olfactomedin 4 mediation of prostate stem/progenitor-like cell proliferation and differentiation via MYC

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Hongzhen Li ◽  
Vijender Chaitankar ◽  
Jianqiong Zhu ◽  
Kyung Chin ◽  
Wenli Liu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Epithelial Cells ◽  
Signaling Pathway ◽  
Rna Sequencing ◽  
Progenitor Cell ◽  
Sequencing Analysis ◽  
Normal Prostate ◽  
Prostate Epithelial Cells ◽  
Cell Proliferation And Differentiation ◽  
Proliferation And Differentiation

AbstractOlfactomedin 4 (OLFM4) is expressed in normal prostate epithelial cells and immortalized normal human prostate epithelial cells (RWPE1), but the identity of OLFM4-expressing cells within these populations and OLFM4’s physiological functions in these cells have not been elucidated. Using single-cell RNA sequencing analysis, we found here that OLFM4 was expressed in multiple stem/progenitor-like cell populations in both the normal prostate epithelium and RWPE1 cells and was frequently co-expressed with KRT13 and LY6D in RWPE1 cells. Functionally, OLFM4-knockout RWPE1 cells exhibited enhanced proliferation of the stem/progenitor-like cell population, shifts stem/progenitor-like cell division to favor symmetric division and differentiated into higher levels PSA expression cells in organoid assays compared with OLFM4-wild RWPE1 cells. Bulk-cell RNA sequencing analysis pinpointed that cMYC expression were enhanced in the OLFM4-knockout RWPE1 cells compared with OLFM4-wild cells. Molecular and signaling pathway studies revealed an increase in the WNT/APC/MYC signaling pathway gene signature, as well as that of MYC target genes that regulate multiple biological processes, in OLFM4-knockout RWPE1 cells. These findings indicated that OLFM4 is co-expressed with multiple stem/progenitor cell marker genes in prostate epithelial cells and acts as a novel mediator in prostate stem/progenitor cell proliferation and differentiation.

scholarly journals MiR-146a-5p targeting SMAD4 and TRAF6 inhibits adipogenensis through TGF-β and AKT/mTORC1 signal pathways in porcine intramuscular preadipocytes

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Que Zhang ◽  
Rui Cai ◽  
Guorong Tang ◽  
Wanrong Zhang ◽  
Weijun Pang
Keyword(s):  
Cell Proliferation ◽  
Adipogenic Differentiation ◽  
Signal Pathways ◽  
Pork Quality ◽  
Sequencing Analysis ◽  
Novel Mirna ◽  
Mtorc1 Signaling ◽  
Proliferation And Differentiation ◽  
Mirna Sequencing ◽  
Imf Content

Abstract Background Intramuscular fat (IMF) content is a vital parameter for assessing pork quality. Increasing evidence has shown that microRNAs (miRNAs) play an important role in regulating porcine IMF deposition. Here, a novel miRNA implicated in porcine IMF adipogenesis was found, and its effect and regulatory mechanism were further explored with respect to intramuscular preadipocyte proliferation and differentiation. Results By porcine adipose tissue miRNA sequencing analysis, we found that miR-146a-5p is a potential regulator of porcine IMF adipogenesis. Further studies showed that miR-146a-5p mimics inhibited porcine intramuscular preadipocyte proliferation and differentiation, while the miR-146a-5p inhibitor promoted cell proliferation and adipogenic differentiation. Mechanistically, miR-146a-5p suppressed cell proliferation by directly targeting SMAD family member 4 (SMAD4) to attenuate TGF-β signaling. Moreover, miR-146a-5p inhibited the differentiation of intramuscular preadipocytes by targeting TNF receptor-associated factor 6 (TRAF6) to weaken the AKT/mTORC1 signaling downstream of the TRAF6 pathway. Conclusions MiR-146a-5p targets SMAD4 and TRAF6 to inhibit porcine intramuscular adipogenesis by attenuating TGF-β and AKT/mTORC1 signaling, respectively. These findings provide a novel miRNA biomarker for regulating intramuscular adipogenesis to promote pork quality.

scholarly journals Receptor-driven invasion profiles in diffuse intrinsic pontine glioma (DIPG)

2021 ◽  
Author(s):  
Anju Karki ◽  
Noah E Berlow ◽  
Jin-Ah Kim ◽  
Esther Hulleman ◽  
Qianqian Liu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Growth Factor ◽  
Protein Expression ◽  
Cell Viability ◽  
Rna Sequencing ◽  
Cell Invasion ◽  
Growth Factor Receptor ◽  
Diffuse Intrinsic Pontine Glioma ◽  
Sequencing Data ◽  
Pontine Glioma

Abstract Background Diffuse intrinsic pontine glioma (DIPG) is a devastating pediatric cancer with unmet clinical need. DIPG is invasive in nature, where tumor cells interweave into the fiber nerve tracts of the pons making the tumor unresectable. Accordingly, novel approaches in combating the disease is of utmost importance and receptor-driven cell invasion in the context of DIPG is under-researched area. Here we investigated the impact on cell invasion mediated by PLEXINB1, PLEXINB2, platelet growth factor receptor (PDGFR)α, PDGFRβ, epithelial growth factor receptor (EGFR), activin receptor 1 (ACVR1), chemokine receptor 4 (CXCR4) and NOTCH1. Methods We used previously published RNA-sequencing data to measure gene expression of selected receptors in DIPG tumor tissue versus matched normal tissue controls (n=18). We assessed protein expression of the corresponding genes using DIPG cell culture models. Then, we performed cell viability and cell invasion assays of DIPG cells stimulated with chemoattractants/ligands. Results RNA-sequencing data showed increased gene expression of receptor genes such as PLEXINB2, PDGFRα, EGFR, ACVR1, CXCR4 and NOTCH1 in DIPG tumors compared to the control tissues. Representative DIPG cell lines demonstrated correspondingly increased protein expression levels of these genes. Cell viability assays showed minimal effects of growth factors/chemokines on tumor cell growth in most instances. Recombinant SEMA4C, SEM4D, PDGF-AA, PDGF-BB, ACVA, CXCL12 and DLL4 ligand stimulation altered invasion in DIPG cells. Conclusions We show that no single growth factor-ligand pair universally induces DIPG cell invasion. However, our results reveal a potential to create a composite of cytokines or anti-cytokines to modulate DIPG cell invasion.

EPA and DHA confer protection against DON-induced endoplasmic reticulum stress and iron imbalance in IPEC-1 cells

2021 ◽  
pp. 1-29
Author(s):  
Jia Lin ◽  
Feifei Huang ◽  
Tianzeng Liang ◽  
Qin Qin ◽  
Qiao Xu ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Rna Sequencing ◽  
Cell Injury ◽  
Cell Damage ◽  
Binding Protein ◽  
Sequencing Analysis ◽  
Epa And Dha ◽  
Expression Of Genes ◽  
Transferrin Receptor 1

Abstract This study assessed the molecular mechanism of eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA) protection against IPEC-1 cell damage induced by deoxynivalenol (DON). The cells were divided into six groups, including the CON group, the EPA group, the DHA group, the DON group, the EPA+DON group, and the DHA+DON group. RNA sequencing was used to investigate the potential mechanism, and qRT-PCR was employed to verify the expression of selected genes. Changes in ultrastructure were used to estimate pathological changes and endoplasmic reticulum (ER) injury in IPEC-1 cells. Transferrin receptor 1 (TFR1) was tested by ELISA. Fe2+ and malondialdehyde (MDA) contents were estimated by spectrophotometry, and reactive oxygen species (ROS) was assayed by fluorospectrophotometry. RNA sequencing analysis showed that EPA and DHA had a significant effect on the expression of genes involved in ER stress and iron balance during DON-induced cell injury. The results showed that DON increased ER damage, the content of MDA and ROS, the ratio of X-box binding protein 1s (XBP-1s)/X-box binding protein 1u (XBP-1u), the concentration of Fe2+, and the activity of TFR1. However, the results also showed that EPA and DHA decreased the ratio of XBP-1s/XBP-1u to relieve DON-induced ER damage of IPEC-1 cells. Moreover, EPA and DHA (especially DHA) reversed the factors related to iron balance. It can be concluded that EPA and DHA reversed IPEC-1 cell damage induced by DON. DHA has the potential to protect IPEC-1 cells from DON-induced iron imbalance by inhibiting ER stress.

AKIP1 promotes cell proliferation, invasion, stemness and activates PI3K-Akt, MEK/ERK, mTOR signaling pathways in prostate cancer

2021 ◽  
Author(s):  
Jiabin Zhao ◽  
Binjiahui Zhao ◽  
Limin Hou
Keyword(s):  
Prostate Cancer ◽  
Cell Proliferation ◽  
Signaling Pathways ◽  
Cancer Patients ◽  
Rna Sequencing ◽  
Mtor Signaling ◽  
Sequencing Analysis ◽  
Lncap Cells ◽  
Oncogenic Signaling ◽  
Interacting Protein

Abstract Background: The study aimed to examine the molecular mechanism and clinical significance of A-kinase interacting protein 1 (AKIP1) in prostate cancer. Methods: The effect of AKIP1 on cell proliferation, migration, invasion, apoptosis and stemness was determined by overexpressing and knocking down AKIP1 in LNCaP and 22Rv1 cells via lentivirus infection. Furthermore, differentially expressed genes (DEGs) by AKIP1 modification were determined using RNA sequencing. Besides, the correlation of AKIP1 with clinicopathological features and prognosis in 130 prostate cancer patients was assessed. Results: AKIP1 expression was increased in VCaP, LNCaP, DU145 cells while similar in 22Rv1 cells compared with RWPE-1 cells. Furthermore, AKIP1 overexpression promoted 22Rv1 and LNCaP cell proliferation, invasion, but inhibited apoptosis; meanwhile, AKIP1 overexpression increased CD133+ cell rate and enhanced spheres formation efficiency in 22Rv1 and LNCaP cells. Reversely, AKIP1 knockdown exhibited the opposite effect in 22Rv1 and LNCaP cells. Further RNA sequencing analysis exhibited that AKIP1-modified DEGs were enriched in the oncogenic signaling pathways related to prostate cancer, such as PI3K-Akt, MEK/ERK, mTOR signaling pathways. The following western blot indicated that AKIP1 overexpression activated while its knockdown blocked PI3K-Akt, MEK/ERK, mTOR signaling pathways in prostate cancer cells. Clinically, AKIP1 was upregulated in the prostate tumor tissues compared with paired adjacent tissues, and its tumor high expression correlated with increased pathological T, pathological N stage and poor prognosis in prostate cancer patients. Conclusion: AKIP1 promotes cell proliferation, invasion, stemness, activates PI3K-Akt, MEK/ERK, mTOR signaling pathways and correlates with worse tumor features and prognosis in prostate cancer.

Abstract W P73: Feasibility and Preliminary Results of Whole Blood RNA-Sequencing Analysis in Patients With Intracranial Aneurysms

Stroke ◽  
2014 ◽  
Vol 45 (suppl_1) ◽  
Author(s):  
Blake Haas ◽  
Nestor R Gonzalez ◽  
Elina Nikkola ◽  
Mark Connolly ◽  
William Hsu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Network Analysis ◽  
Rna Sequencing ◽  
Whole Blood ◽  
Intracranial Aneurysms ◽  
Small Sample Size ◽  
Expression Patterns ◽  
Small Sample ◽  
Cellular Respiration ◽  
Sequencing Analysis

Introduction: Intracranial aneurysms (IA) growth and rupture have been associated with chronic remodeling of the arterial wall. However, the pathobiology of this process remains poorly understood. The objective of the present study was to evaluate the feasibility of analyzing gene expression patterns in peripheral blood of patients with ruptured and unruptured saccular IAs. Materials and Methods: We analyzed human whole blood transcriptomes by performing paired-end, 100 bp RNA-sequencing (RNAseq) using the Illumina platform. We used STAR to align reads to the genome, HTSeq to count reads, and DESeq to normalize counts across samples. Self-reported patient information was used to correct expression values for ancestry, age, and sex. We utilized weighted gene co-expression network analysis (WGCNA) to identify gene expression network modules associated with IA size and rupture. The DAVID tool was employed to search for Gene Ontology enrichment in relevant modules. Results: Samples from 12 patients (9 females, age 57.6 +/-12) with IAs were analyzed. Four had ruptured aneurysms. RNA isolation and application of the methodology described above was successful in all samples. Although the small sample size prevents us from drawing definite conclusions, we observed promising novel co-expression networks for IAs: WCGNA analysis showed down-regulation of two transcript modules associated with ruptured IA status (r=-0.78, p=0.008 and r=-0.77, p=0.009), and up-regulation of two modules associated with aneurysm size (r=0.86, p=0.002 and r=0.9, p=4e-04), respectively. DAVID analyses showed that genes upregulated in an IA size-associated module were enriched with genes involved in cellular respiration and translation, while genes involved in transcription were down-regulated in a module associated with ruptured IAs. Conclusions: Whole blood RNAseq analysis is a feasible tool to capture transcriptome dynamics and achieve a better understanding of the pathophysiology of IAs. Further longitudinal studies of patients with IAs using network analysis are justified.

Photobiomodulation of inflamed dental pulp stem cells under different nutritional conditions

2021 ◽  
Author(s):  
Seyedeh Sareh Hendi ◽  
Leila Gholami ◽  
Massoud Saidijam ◽  
Roghayeh Mahmoudi ◽  
Ali Asghar Arkian ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Cell Proliferation ◽  
Laser Irradiation ◽  
Dental Pulp ◽  
Culture Conditions ◽  
Dental Pulp Stem Cells ◽  
Reverse Transcription Pcr ◽  
Nutritional Conditions ◽  
Proliferation And Differentiation

Aim: The present study aimed to investigate photobiomodulation's (PBM) effect on inflamed dental pulp stem cells (IDPSCs) under different nutritional conditions. Methods: Cell proliferation and odontogenic differentiation were evaluated using the MTT assay and real-time quantitative reverse transcription PCR, respectively after laser PBM of cells in 5 or 10% fetal bovine serum (FBS) culture conditions. Results: A significant positive effect of laser irradiation on cell proliferation under both nutritional conditions after 24 and 48 h was observed. DMP-1 gene expression increased in the groups with laser irradiation and 5% FBS. Comparison of gene expression levels in the four groups revealed no statistically significant stimulatory effect. The highest gene expression was observed in the non-laser group with 5% FBS. Conclusion: Further studies are required to obtain an irradiation setup to ideally improve inflamed dental pulp stem cells' proliferation and differentiation.

On the Origins of AML Relapse

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 223-223
Author(s):  
Liran I. Shlush ◽  
Amanda Mitchell ◽  
Lawrence Heisler ◽  
Sagi Abelson ◽  
Monica Doedens ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Rna Sequencing ◽  
Genetic Variants ◽  
Gene Expression Pattern ◽  
The Other ◽  
Specific Cell ◽  
Sequencing Analysis ◽  
Major Clone ◽  
A Minor

Abstract While induction into remission is effective in the majority of acute myeloid leukemia (AML) patients, disease recurrence is common, especially among the elderly. Understanding the origins of AML relapse would permit better treatments targeting the specific cells that survive chemotherapy. While some evidence suggests that AML relapse can originate either from a minor or a major clone that is already present at diagnosis, the exact origins of AML relapse are still obscure. In the current study we aimed at identifying the origins of AML by identifying genetic variants that appear at relapse, and to then track these variants back into specific cell populations present at diagnosis. We hypothesized that relapse might have multiple origins: from the major blast population, from rare leukemia imitating cells (LIC) as detected using xenografting, or from preleukemic stem cells (preL-HSCs). Methods: The bulk diagnosis and relapse samples of peripheral blood from eleven AML patients were analyzed, first by whole genome sequencing (50X coverage) to identify somatic mutations and genetic variants which were specifically present at relapse (relapse variants-RVs). The presence RVs was then reassessed in phenotypically defined sub-fractions sorted from the diagnosis samples, at a sensitivity of 1 in 1000 by digital PCR. The following sub-populations were genotyped: 1) isolated CD33+ blasts (the major population) 2) phenotypically defined leukemic and preleukemic stem cells 3) functionally defined leukemia initiating cells (LICs) harvested from xenografts (an average of 30 xenografts were generated from each diagnosis and relapse sample). Results: LICs, but not the dominant blast population from diagnosis carried the RVs in 3 of 11 cases. In these patients CD33-CD34+CD45RA+ immature cells from diagnosis also carried the RVs. In a second subset of 3 of 11 AML samples, relapse originated from a minor clone present within the CD33+ leukemic blasts; these samples did not produce exnografts. Other samples (2/11) exhibited relapse samples that arose from a combined origin (both LICs, and CD33+ blasts, or from the major clone (1/11). In two cases we could not identify the origins of relapse. As our initial results suggested that the cells responsible for AML relapse can come from distinct origins within the diagnosis sample, we next asked whether other functional and phenotypic differences might be present between the patients that have different relapse origins. RNA sequencing analysis of bulk cells from diagnosis demonstrated a remarkable clustering of the global gene expression that correlated with the origin of relapse. Unsupervised hierarchical clustering grouped together the AML samples who relapsed from the LICs, while all other samples were in a very distinct second cluster. The gene expression signature of the samples that relapsed from LICs was consistent with a monocytic phenotypic signature, while the other samples were more progenitor-like. To further expand and validate our findings we used the same unsupervised clustering on the RNA sequencing data of AML samples who relapsed in the TCGA dataset (n=86). Remarkably, the similar two main clusters were generated; comparison by GSEA provided evidence that the gene expression clusters in our study were generated by the same genes as in the TCGA clusters. Conclusion: Our results provide for the first time evidence that AML can relapse from distinct, predictable and pre-existing origins: AMLs with a monocytic phenotype relapse from chemo-resistant LICs; and AMLs with a progenitor gene expression pattern (yet lacking xenografting capacity) that relapse from CD33+ cells. These results pose a series of predictions as to the success of different therapies. For example, in the former group the major monocytic clone is sensitive to chemotherapy, yet relapse originates from CD33-CD34+CD45RA+ cells and would therefore be predicted to be resistant to Anti-CD33 therapeutics. On the other hand, relapse in the latter group originates from CD33+ cells and these are predicted to be sensitive to Anti-CD33 therapeutics. The results of this study document the complexity in origins of AML relapse and have important implications for the design of future more effective and personalized strategies for preventing AML relapse. Disclosures No relevant conflicts of interest to declare.

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