Exosome-mediated transfer of microRNAs promotes mouse iPSCs differentiation into therapeutic insulin-producing cells

Abstract Purpose Exosome-based therapeutic approaches have been applied in diabetes. In the present study, we explored the effect of exosomes on iPSCs differentiation into insulin-producing cells and its underlying mechanisms. Methods Exosomes were isolated by ultracentrifugation from MIN6 cells and identified by Transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA) and Western blot. PKH67 tracer and transwell assay were used to confirm exosome delivery into iPSCs. QRT-PCR was applied to detect key pancreatic gene expression and miRNAs expression in differentiated iPSCs. Insulin expression was assessed by flow cytometry (FCM) and immunofluorescence. The mechanism underlying exosome induction capacity for iPSCs was determined via RNA-interference of Argonaute-2 (Ago2). Streptozotozin(STZ) was used to establish diabetic mouse model to verify the function of differentiated β-like cells. Results MIN6-derived exosomes promoted the key pancreatic gene expression and immunofluorescence for Nkx6.1 and insulin remarkably, confirming the capability of exosomes for iPSCs differentiation. Moreover, transplantation of differentiated iPSCs efficiently enhanced IPGTT and partially control hyperglycemia in T1D mice. Knockdown of Ago2 in MIN6 cells affect exosomal miRNAs expression and pancreatic gene expression and insulin secretion in iPSCs.The therapeutic effect in vivo was weakened, further indicating decreased exosomal miRNA affect iPSCs differentiation.7 specific exosomal miRNAs were selected for single-assay validation. MiR-706, miR-709, miR-466c-5p and miR-423-5p were found dynamic changed during differentiation stages. Conclusion Exosomes is an effective and convenient induction approach for iPSCs differentiation into functional insulin secreting cells.The effect was downregulated via Ago2 knockdown illustrates the mechanisms are highly relevant to specific miRNAs enriched in exosomes.

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CircFAM13B promotes the proliferation of hepatocellular carcinoma by sponging miR-212, upregulating E2F5 expression and activating the P53 pathway

Cancer Cell International ◽

10.1186/s12935-021-02120-6 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Ying Xie ◽

Xiaofeng Hang ◽

Wensheng Xu ◽

Jing Gu ◽

Yuanjing Zhang ◽

...

Keyword(s):

Gene Expression ◽

Hepatocellular Carcinoma ◽

Gene Expression Omnibus ◽

Differentially Expressed ◽

In Vivo Studies ◽

Circular Rnas ◽

Novel Target ◽

Underlying Mechanisms

Abstract Background Most of the biological functions of circular RNAs (circRNAs) and the potential underlying mechanisms in hepatocellular carcinoma (HCC) have not yet been discovered. Methods In this study, using circRNA expression data from HCC tumor tissues and adjacent tissues from the Gene Expression Omnibus database, we identified out differentially expressed circRNAs and verified them by qRT-PCT. Functional experiments were performed to evaluate the effects of circFAM13B in HCC in vitro and in vivo. Results We found that circFAM13B was the most significantly differentially expressed circRNA in HCC tissue. Subsequently, in vitro and in vivo studies also demonstrated that circFAM13B promoted the proliferation of HCC. Further studies revealed that circFAM13B, a sponge of miR-212, is involved in the regulation of E2F5 gene expression by competitively binding to miR-212, inhibits the activation of the P53 signalling pathway, and promotes the proliferation of HCC cells. Conclusions Our findings revealed the mechanism underlying the regulatory role played by circFAM13B, miR-212 and E2F5 in HCC. This study provides a new theoretical basis and novel target for the clinical prevention and treatment of HCC.

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Argonaute2 promotes myeloma angiogenesis by deregulation of microRNAs

Blood ◽

10.1182/blood.v122.21.5334.5334 ◽

2013 ◽

Vol 122 (21) ◽

pp. 5334-5334

Author(s):

Chen Lijuan ◽

Xu Jiaren ◽

Shuang Wu ◽

Rong Wang ◽

Qu Xiaoyan ◽

...

Keyword(s):

Conflicts Of Interest ◽

Tube Formation ◽

Cancer Cell Proliferation ◽

Cam Assay ◽

Core Component ◽

Over Expression ◽

Argonaute 2 ◽

The Core ◽

Mirnas Expression

Abstract Dysregulation of miRNAs expression contributes to cancer cell proliferation, apoptosis and angiogenesis. Angiogenesis is a hallmark of multiple myeloma development and progression. Argonaute 2 (AGO2) protein, the core component of RNA-induced silencing complex (RISC), can directly bind miRNAs and mediate target mRNAs degradation. Previous study showed that AGO2 knockdown suppressed the growth and tube formation of HUVECs. In current study, the supernatant of AGO2 over-expression MM lines could induce HUVECs migration and accelerated tube formation. Conversely, the supernatant of AGO2 knockdown MM lines could suppress cell migration and tube formation of HUVECs. Moreover, CAM assay also demonstrated AGO2 can drive neovessel formation of MM in vivo. Using miRNAs microarray, we observed that 25 miRNAs were up-regulated and 14 miRNAs were down-regulated by AGO2 protein. Among these AGO2-associated miRNAs, most Let-7 family and two miR-17-92 cluster members (miR-17a and miR-92-1), known as pro-angiogenetic miRNAs, were the dominant positively regulated miRNAs by AGO2, and the anti-angiogenetic miRNAs, such as miR-145 and miR-361 were inversely regulated by AGO2 protein, which play crucial role in AGO2 mediating angiogenesis. Disclosures: No relevant conflicts of interest to declare.

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Effect of Ambient pH on Growth, Pathogenicity, and Patulin Production of Penicillium expansum

Toxins ◽

10.3390/toxins13080550 ◽

2021 ◽

Vol 13 (8) ◽

pp. 550

Author(s):

Carelle Kouasseu Jimdjio ◽

Huali Xue ◽

Yang Bi ◽

Mina Nan ◽

Lan Li ◽

...

Keyword(s):

Gene Expression ◽

Growth Factors ◽

Soft Rot ◽

Penicillium Expansum ◽

Expression Of Genes ◽

Transmission Electron ◽

Expression Studies ◽

Gene Expression Studies

Penicillium expansum is an important postharvest pathogen of pomaceous fruit and a causal agent of blue mold or soft rot. In this study, we investigated the effect of ambient pH on growth, ultrastructure alteration, and pathogenicity of P. expansum, as well as accumulation of patulin and expression of genes involved in patulin biosynthesis. Under different pH, the fungus was routinely cultured and collected for growth, pathogenicity, patulin production, and gene expression studies using transmission electron microscopy, apple inoculation, HPLC, and RT-qPCR methods. Different ambient pH had significant impact on expression of genes and growth factors involved in patulin biosynthesis. Under same range of pH, gene expression profile, growth factors, and patulin accumulation (in vivo and in vitro) all showed similar changing trends. A well-developed cell was observed in addition to upregulation of genes at pH between pH 5.0 and 7.0, while the opposite was observed when pH was too basic (8.5) or too acid (2.5). Additionally, ambient pH had direct or indirect influence on expression of PecreaA, PelaeA, and PepacC. These findings will help in understanding the effect of ambient pH on growth, pathogenicity, and patulin production and support the development of successful methods for combating P. expansum infection on apple fruits.

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Ago2-seq identifies new microRNA targets for seizure control

10.1101/777144 ◽

2019 ◽

Author(s):

Morten T. Venø ◽

Cristina R. Reschke ◽

Gareth Morris ◽

Niamh M. C. Connolly ◽

Junyi Su ◽

...

Keyword(s):

Gene Expression ◽

Animal Models ◽

Seizure Control ◽

Noncoding Rnas ◽

Tgfβ Signaling ◽

Argonaute 2 ◽

Proteomic Data ◽

Microrna Targets ◽

Spontaneous Recurrent Seizures

AbstractMicroRNAs (miRNAs) are short noncoding RNAs that shape the gene expression landscape, including during the pathogenesis of temporal lobe epilepsy (TLE). In order to provide a full catalog of the miRNA changes that happen during experimental TLE, we sequenced Argonaute 2-loaded miRNAs in the hippocampus of three different animal models at regular intervals between the time of the initial precipitating insult to the establishment of spontaneous recurrent seizures. The commonly upregulated miRNAs were selected for a functional in vivo screen using oligonucleotide inhibitors. This revealed anti-seizure phenotypes upon inhibition of miR-10a-5p, miR-21a-5p and miR-142a-5p as well as neuroprotection-only effects for inhibition of miR-27a-3p and miR-431-5p. Proteomic data and pathway analysis on predicted and validated targets of these miRNAs indicated a role for TGFβ signaling in a shared seizure-modifying mechanism. Together, these results identify functional miRNAs in the hippocampus and a pipeline of new targets for seizure control in epilepsy.

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HSP90α Mediates Sorafenib Resistance in Human Hepatocellular Carcinoma by Necroptosis Inhibition under Hypoxia

Cancers ◽

10.3390/cancers13020243 ◽

2021 ◽

Vol 13 (2) ◽

pp. 243

Author(s):

Yan Liao ◽

Yue Yang ◽

Di Pan ◽

Youxiang Ding ◽

Heng Zhang ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Imaging System ◽

Morphological Changes ◽

Critical Role ◽

Tumor Xenograft ◽

Target Drug ◽

Transmission Electron ◽

Underlying Mechanisms

As one of the most common malignancies worldwide, Hepatocellular carcinoma (HCC) has been treated by Sorafenib, which is the first approved target drug by FDA for advanced HCC. However, drug resistance is one of the obstacles to its application. As a typical characteristic of most solid tumors, hypoxia has become a key cause of resistance to chemotherapy and radiotherapy. It is important to elucidate the underlying mechanisms of Sorafenib resistance under hypoxia. In this study, the morphological changes of hepatocellular carcinoma cells were observed by Live Cell Imaging System and Transmission Electron Microscope; Sorafenib was found to induce necroptosis in liver cancer. Under hypoxia, the distribution of necroptosis related proteins was changed, which contributed to Sorafenib resistance. HSP90α binds with the necrosome complex and promotes chaperone-mediated autophagy (CMA) degradation, which leads necroptosis blocking and results in Sorafenib resistance. The patient-derived tumor xenograft (PDX) model has been established to investigate the potential therapeutic strategies to overcome Sorafenib resistance. 17-AAG inhibited HSP90α and presented obvious reversal effects of Sorafenib resistance in vivo and in vitro. All the results emphasized that HSP90α plays a critical role in Sorafenib resistance under hypoxia and 17-AAG combined with Sorafenib is a promising therapy for hepatocellular carcinoma.

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The importance of cellular and exosomal miRNAs in mesenchymal stem cell osteoblastic differentiation

Scientific Reports ◽

10.1038/s41598-021-85306-2 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Sajjad Shirazi ◽

Chun-Chieh Huang ◽

Miya Kang ◽

Yu Lu ◽

Sriram Ravindran ◽

...

Keyword(s):

Gene Expression ◽

Osteoblastic Differentiation ◽

Hippo Signaling ◽

Paracrine Signaling ◽

Wild Type ◽

Argonaute 2 ◽

Kegg Analysis ◽

Exosomal Mirnas ◽

Complex Regulation ◽

Global Reduction

AbstractThe differentiation of osteoblasts is under complex regulation that includes autocrine and paracrine signaling from MSCs. Exosomes are important components of the MSC secretome and their cargo contains numerous miRNAs. In this study, the importance of MSC miRNAs in modulation of osteoblastic differentiation was examined by global reduction of miRNA biosynthesis in Dicer knock down hMSCs. We additionally impaired hMSC responses to miRNAs by knockdown of Argonaute 2 expression. Knockdown of Dicer and Argonaute 2 both reduced osteoblastic differentiation of hMSCs. This was observed at the levels of hMSC culture mineralization and osteoblastic gene expression. The treatment of Dicer deficient hMSCs with wild type hMSC exosomes effectively recovered the impaired osteoblastic differentiation. Dicer knockdown reduced the quantity and diversity of miRNAs present in hMSC exosomes. miRSeq data and KEGG analysis implicated the miRNA-dependent effects on multiple osteoinductive pathways in Dicer deficient cells, including the Hippo signaling and TGF-beta signaling pathways. Treatment of hMSCs with mimics of miRNAs significantly downregulated in Dicer knockdown cells recovered functions of exosome-mediated signaling in hMSCs. These results indicate that hMSC exosomes exert miRNA-dependent control that contributes to osteoblastic differentiation.

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Mechanism of Hypoxia-Activated MiR-194 /FoxO3a Inducing Glycolytic Metabolism Reprogramming in Gastric Precancerous Lesions

10.21203/rs.3.rs-788905/v1 ◽

2021 ◽

Author(s):

Sheng-xiong Zhang ◽

Wei Liu ◽

Bo Ai ◽

Hua-feng Pan ◽

Zi-ming Zhao ◽

...

Keyword(s):

Gastric Mucosa ◽

Precancerous Lesions ◽

Metabolic Reprogramming ◽

Key Factors ◽

Ki 67 ◽

Expression Levels ◽

Transmission Electron ◽

Hematoxylin And Eosin ◽

Underlying Mechanisms

Abstract Background: Early diagnosis and treatment of gastric precancerous lesions (GPL) are key factors for reducing the incidence and morbidity of gastric cancer. The study aimed to examine GPL in mice induced by MNU and to illustrate the underlying mechanisms of tumorigenesis.Methods: In this study, we utilized an in vivo MNU (N-methyl-N-nitroso-urea)-induced GPL mouse model, Histopathological changes of the gastric mucosa were observed by Hematoxylin and Eosin (H&E-stain) and alcian blue (AB-PAS-stain); The level of miR-194-5p in the gastric mucosa were determined by real‐time polymerase chain reaction; We used transmission electron microscopy to observe the effects of MNU on gastric chief cells and parietal cells; We performed immunohistochemical detection of HIF-1α, vWF, Ki-67 and P53, while the changes in protein expression of key genes in LKB1-AMPK and AKT-FoxO3 signaling pathways were detected by western blot analysis.Results: We demonstrated that miR-194-5p expression was upregulated under hypoxia in GPL gastric tissues, and that a high miR-194-5p expression level closely related with tumorigenesis. Mechanistically, miR-194-5p exerted the acceleration of activities related to metabolic reprogramming through LKB1-AMPK and AKT-FoxO3 pathways. Furthermore, similar to miR-194-5p, high expression levels of AMPK and AKT were also related to the metabolic reprogramming of GPL. Moreover, we revealed the correlation between the expression levels of miR-194-5p, p-AMPKα, p-AKT, and FoxO3a. Conclusions: These findings suggest that miR-194-5p/FoxO3 pathway is important for the reversal of metabolic reprogramming in GPL. Thus, exploring strategies to regulate the miR-194-5p/FoxO3a pathway may provide an efficient strategy for the prevention and treatment of GPL.

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Abstract 457: Transcription Coactivator MED1 Protects Against Atherosclerosis by Modulation of Macrophage Polarization

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvb.36.suppl_1.457 ◽

2016 ◽

Vol 36 (suppl_1) ◽

Author(s):

Liang Bai ◽

Enqi Liu

Keyword(s):

Gene Expression ◽

Macrophage Polarization ◽

Low Density Lipoprotein ◽

Ldl Receptor ◽

Density Lipoprotein ◽

Peritoneal Macrophages ◽

Gene Markers ◽

Underlying Mechanisms ◽

High Level

Mediator 1 (MED1), a key subunit of the Mediator complex, interacts with several nuclear receptors and transcription factors to direct gene-specific transcription. It is well-known that MED1 play important roles in lipid metabolism. However, the role and underlying mechanisms of MED1 in atherosclerosis remains unclear. Here, macrophage-specific MED1 knockout (MED1 ΔMac ) mice were generated to investigate the contribution of MED1 on atherogenesis in vivo. We demonstrate that among mice deficient in apolipoprotein E (Apoe), the additional loss of macrophage MED1 (MED1 ΔMac /ApoE -/- ) exhibited significantly larger atherosclerotic lesions in the whole aortic tree and aortic root compared with MED1 fl/fl /ApoE -/- littermates on either the chow or the western diet, and these effects were also found in low-density lipoprotein (LDL) receptor-deficient (LDLR -/- ) mice reconstituted with bone marrow from MED1 ΔMac mice. Peritoneal macrophages from MED1 ΔMac /ApoE -/- mice had significantly increased expression of gene markers for M1-like macrophages, including IL-1β, IL6,COX2, iNOS, Gro1, MCP1 and TNFα, etc, whereas decreased levels of anti-inflammatory genes for M2-like macrophages, such as Arg1, Mrc1, Retnla, Chi3l3 and PPARγ. Over-expression of MED1 using adenovirally-driven MED1 (Ad/MED1) in MED1 fl/fl /ApoE -/- macophages repressed the proinflammatory gene expression. Re-expression of MED1 using Ad/MED1 counteracted the high level of inflammatory gene in MED1 ΔMac /ApoE -/- macophages. Furthermore, gene expression profiling and PCR array showed that MED1-deficient macrophages exhibited the increased M1 targets and decreased M2 targets. These data demonstrate that MED1 expression by macrophages has anti-atherogenic effects via regulation of macrophage polarization. MED1 may be considered as a potential therapeutic target to treat atherosclerosis.

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Raddeanin a Suppresses Glioblastoma Growth by Inducing ROS Generation and Subsequent JNK Activation to Promote Cell Apoptosis

Cellular Physiology and Biochemistry ◽

10.1159/000490187 ◽

2018 ◽

Vol 47 (3) ◽

pp. 1108-1121 ◽

Cited By ~ 11

Author(s):

Fei Peng ◽

Xiaoxiong Wang ◽

Mengting Shu ◽

Mingfei Yang ◽

Ligang Wang ◽

...

Keyword(s):

Xenograft Model ◽

Ros Generation ◽

Transmission Electron ◽

Glioma Xenograft ◽

Potential Strategy ◽

Underlying Mechanisms ◽

Induced Apoptosis ◽

Hoechst Staining ◽

Jnk Activation

Background/Aims: Raddeanin A (RA), an active pharmacological ingredient from Anemone raddeana Regel, plays an important role in tumor suppression. In this study, we assessed the potentially therapeutic effect of RA on glioblastoma and its underlying mechanisms. Methods: Cell viability was examined using the MTT assay. Invasive and migratory capacities were examined using Transwell and wound healing assays. Apoptosis was determined by Hoechst staining, flow cytometry, DCFH-fluorescent probe and immunohistochemical staining. Autophagy was detected by transmission electron microscopy and western blotting. A U251 glioma xenograft model was established to evaluate the effect of RA in vivo. Results: The data demonstrated that RA inhibited viability, and abrogated the invasive/migratory abilities of glioblastoma cells. In addition, RA induced apoptosis by reactive oxygen species (ROS)/ Jun N-terminal kinase (JNK) signaling in glioblastoma. Conversely, the antioxidant N-Acetyl-L-cysteine (NAC) and pan-caspase inhibitor z-VAD-fmk attenuated RA-induced apoptosis by scavenging ROS and inactivating caspase-3. Furthermore, the inhibition of autophagy by 3-MA exacerbated apoptosis through ROS generation and JNK phosphorylation. In vivo, RA exhibited a curative effect on U251-derived xenografts in nude mice. Conclusions: The results of this study suggest that RA suppressed the growth of glioblastoma, thus serving as a promising and potential strategy for glioblastoma chemotherapy.

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Effects of BDNF, T3, and corticosterone on expression of the hypothalamic obesity gene network in vivo and in vitro

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.90813.2008 ◽

2009 ◽

Vol 296 (4) ◽

pp. R1180-R1189 ◽

Cited By ~ 30

Author(s):

Mardi S. Byerly ◽

Jean Simon ◽

Elisabeth Lebihan-Duval ◽

Michel J. Duclos ◽

Larry A. Cogburn ◽

...

Keyword(s):

Gene Expression ◽

Body Composition ◽

Leptin Receptor ◽

Receptor Gene ◽

Energy Regulation ◽

Underlying Mechanisms ◽

Agouti Related Protein ◽

Receptor Gene Expression

Hypothalamic neuropeptides, neurotrophins, and systemic hormones modulate food intake and body composition. Although advances toward elucidating these interactions have been made, many aspects of the underlying mechanisms remain vague. Hypothalami from fat and lean chicken lines were assessed for differential expression of anabolic/orexigenic and catabolic/anorexigenic genes. Effects of triiodothyronine (T3), corticosterone (Cort), and brain-derived neurotrophic factor (BDNF) on expression of anabolic/orexigenic and catabolic/anorexigenic genes were tested in cultures of hypothalamic neurons. From this, we found that BDNF increased and T3 decreased gene expression for BDNF, leptin receptor (LEPR), pro-opiomelanocortin (POMC), thyrotropin releasing hormone (TRH), and agouti-related protein (AGRP). Thyroid hormone levels were manipulated during development to show that T3 inhibited BDNF, TRH, and BDNF receptor gene expression. Delivery of T3, Cort, T3 plus Cort, or vehicle in vivo continuously for 72 h indicated that Cort and T3 have overlapping roles in regulating TRH, LEPR, and POMC gene expression and that Cort and T3 regulate BDNF, neuropeptide Y, and AGRP in opposite directions. Collectively, these findings suggest that interactions between the neuropeptide BDNF and the hormones T3 and/or Cort may constitute a homeostatic mechanism that links hypothalamic energy regulation controlling body composition.

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