Exploring the role of circ-0091579/miR-1225-5p and circ-HIPK3/miR-338-3p axes in the pathogenesis of postmenopausal osteoporosis.

CircRNAs have been increasingly appreciated as modulators of osteoporosis. This study investigated the expression of circ-0091579 and circ-HIPK3 in PBMCs of postmenopausal women with osteopenia and osteoporosis, aiming to underline their molecular mechanisms involved in pathogenesis of the disease. Seventy patients were stratified into two groups: 35 with osteopenia and 35 with osteoporosis, along with 30 healthy controls. Expressions of circ-0091579 and circ-HIPK3, miR-1225-5p and miR-338-3p, together with NF-κB, were assessed using RT-PCR. Keap1, Nrf2, and MAFB were determined using Western blot, while RANKL, OPG, IL-1ß, and IL-6 were measured by ELISA. GSH and MDA were estimated colorimetrically. Data revealed that circ-0091579 was markedly upregulated, whereas miR-1225-5p was downregulated in patients relative to controls. Additionally, circ-HIPK3 was significantly decreased, while miR-338-3p was increased in the diseased groups. Circ-0091579 was directly correlated with RANKL/OPG, NF-κB, IL-1ß, IL-6 and MDA, while inversely correlated with miR-1225-5p, T-score, BMD and GSH. Meanwhile, circ-HIPK3 and miR-338-3p were interrelated in an opposite manner. Eventually, the interplay among these downstream players induced an imbalance in bone homeostasis, triggering osteoporosis. Notably, these circRNAs differentiated patients from controls and those with osteopenia from osteoporotic ones. Thus, they could serve as biomarkers for early detection and tracking of osteoporosis.

Profile of plasma microRNAs as a potential biomarker of Wilson's disease.

BACKGROUND: Wilson's disease (WD) is a rare condition resulting from autosomal recessive mutations in ATP7B, a copper transporter, manifesting with hepatic, neurological, and psychiatric symptoms. Timely diagnosis and appropriate treatment yield a positive prognosis, while delayed identification and/or insufficient therapy lead to a poor outcome. Our aim was to establish a prognostic method for WD by characterising biomarkers based on circulating microRNAs. METHODS: We conducted investigations across three cohorts: discovery, validation (comprising unrelated patients), and follow-up (revisiting the discovery cohort 3 years later). All groups were compared to age- and gender-matched controls. Plasma microRNAs were analysed via RNA sequencing in the discovery cohort and subsequently validated using quantitative PCR in all three cohorts. To assess disease progression, we examined the microRNA profile in Atp7b-/- mice, analysing serum samples from 6 to 44 weeks of age and liver samples at three time points: 20, 30, and 40 weeks of age. RESULTS: In patients, elevated levels of the signature microRNAs (miR-122-5p, miR-192-5p, and miR-885-5p) correlated with serum activities of aspartate transaminase, alanine aminotransferase and gamma-glutamyl transferase. In Atp7b-/- mice, levels of miR-122-5p and miR-192-5p (miR-885-5p lacking a murine orthologue) increased from 12 weeks of age in serum, while exhibiting fluctuations in the liver, possibly attributable to hepatocyte regenerative capacity post-injury and the release of hepatic microRNAs into the bloodstream. CONCLUSIONS: The upregulation of the signature miR-122-5p, miR-192-5p, and miR-885-5p in patients and their correlation with liver disease progression in WD mice support their potential as biomarkers of WD.

TGF-ß1-Induced LINC01094 promotes epithelial-mesenchymal transition in hepatocellular carcinoma through the miR-122-5p/TGFBR2-SAMD2-SMAD3 Axis.

Hepatocellular carcinoma (HCC) is a common malignancy with a poor prognosis. It has been proven that long non-coding RNAs (lncRNAs) play an essential role in regulating HCC progression. However, the involvement of LINC01094 in regulating epithelial-mesenchymal transition (EMT) in HCC remains unclear. LINC01094 expression in HCC patients was retrieved from the Cancer Genome Atlas database. Overexpressing and downregulating LINC01094 were conducted to investigate its biological functions using Hep3B, SNU-387, and HuH-7 cells. Western blotting and morphological observation were performed to study the EMT in HCC cells. Transwell assay was adopted to determine the migration and invasion of HCC cells. The underlying mechanism of competitive endogenous RNAs (ceRNAs) was investigated using bioinformatics analysis, quantitative reverse-transcription polymerase chain reaction, and rescue experiments. Elevated LINC01094 expression was observed in HCC and associated with a poor prognosis. Knockdown of LINC01094 expression in SNU-387 and HuH-7 cells could inhibit migration, invasion, and EMT markers. Overexpression of LINC01094 indicated that LINC01094 promoted EMT via the TGF-ß/SMAD signaling pathway. The bioinformatics analysis revealed that miR-122-5p was a target of LINC01094. The miRWalk database analysis showed that TGFBR2, SMAD2, and SMAD3 were downstream targets of miR-122-5p. Mechanically, LINC01094 acted as a ceRNA that facilitated HCC metastasis by sponging miR-122-5p to regulate the expression of TGFBR2, SMAD2, and SMAD3. Further, TGF-ß1 could enhance the expression of LINC01094, forming a positive feedback loop. TGF-ß1-induced LINC01094 expression promotes HCC cell migration and invasion by targeting the miR-122-5p/TGFBR2-SMAD2-SMAD3 axis. LINC01094 may be a potential prognostic biomarker and therapeutic target for HCC metastasis.

MiR-122-5p regulates erastin-induced ferroptosis via CS in nasopharyngeal carcinoma.

Nasopharyngeal carcinoma (NPC) is a tumor that occurs in the nasopharynx. Although advances in detection and treatment have improved the prognosis of NPC the treatment of advanced NPC remains challenging. Here, we explored the effect of microRNA (miR)-122-5p on erastin-induced ferroptosis in NPC cells and the role of ferroptosis in the development of NPC. The effect of miR-122-5p silencing and overexpression and the effect of citrate synthase on erastin-induced lipid peroxidation in NPC cells was analyzed by measuring the amounts of malondialdehyde, Fe2+, glutathione, and reactive oxygen species and the morphological alterations of mitochondria. The malignant biological behavior of NPC cells was examined by cell counting kit-8, EDU, colony formation, Transwell, and wound healing assays. The effects of miR-122-5p on cell proliferation and migration associated with ferroptosis were examined in vivo in a mouse model of NPC generated by subcutaneous injection of NPC cells. We found that erastin induced ferroptosis in NPC cells. miR-122-5p overexpression inhibited CS, thereby promoting erastin-induced ferroptosis in NPC cells and decreasing NPC cell proliferation, migration, and invasion.

Would Combination Be Better: Swimming Exercise and Intermittent Fasting Improve High-Fat Diet-Induced Nonalcoholic Fatty Liver Disease in Obese Rats via the miR-122-5p/SREBP-1c/CPT1A Pathway.

Background: Swimming and intermittent fasting can both improve obesity-induced NAFLD, but which of the two is more effective and whether the combination of the two has a superimposed effect is inconclusive. Methods: The model of NAFLD in obese rats was established by a high-fat diet and performed swimming, intermittent fasting, and a combination of both interventions for 8 weeks. Serum lipids and enzyme activity were measured by an automatic biochemical analyzer. Liver morphostructural analysis was observed by transmission electron microscopy, and morphology was observed by HE staining. RT‒PCR was used to detect the mRNA level. Results: Morphology and microstructure of the liver of model rats were impaired, with the upregulation of miR-122-5p, SREBP-1c, FASN and ACC1. Eight weeks of swimming exercise, intermittent fasting and the combination of both attenuate these effects, manifested by the downregulation of miR-122-5p and upregulation of CPT1A mRNA levels. There was no significant stacking effect of the combination of the swimming and intermittent fasting interventions. Conclusion: NAFLD leads to pathology in model rats. Eight weeks of swimming exercise, intermittent fasting and the combination of both can inhibit miR-122-5p and improve hepatic lipid metabolism, while no significant additive effects of combining the interventions were found.

CircPHGDH downregulation decreases papillary thyroid cancer progression through miR-122-5p/PKM2 axis.

BACKGROUND: Although papillary thyroid carcinoma (PTC) has a favorable prognosis, it could affect patient life quality and become a serious threat because of invasion and metastasis. Many investigations have suggested that circular RNAs (circRNAs) are involved in different cancer regulations. Nevertheless, circRNAs role in invasive PTC remains unclear. METHODS: In the present investigation, next-generation sequencing was applied to explore abnormal circRNA expression. The expression of circRNA phosphoglycerate dehydrogenase (circPHGDH) in PTC cell lines and tissues were examined. Then, we investigated regulatory mechanism and circPHGDH downstream targets using bioinformatics analysis and luciferase reporting analysis. Then transwell migration, Cell Counting Kit-8 (CCK8) and 5-ethynyl-2'-deoxyuridine (EdU) assays were used for cells migration and proliferation analysis. In vivo metastasis and tumorigenesis assays were also employed to evaluate the circPHGDH role in PTC. RESULTS: The data showcased that circPHGDH expression increased in both PTC cell lines and tissues, which suggested that circPHGDH functions in PTC progression. circPHGDH downregulation suppressed PTC invasion and proliferation in both in vivo and in vitro experiments. Bioinformatics and luciferase reporter results confirmed that both microRNA (miR)-122-5p and pyruvate kinase M2 subtype (PKM2) were downstream targets of circPHGDH. PKM2 overexpression or miR-122-5p suppression reversed PTC cell invasion and proliferation post silencing circPHGDH by restoring aerobic glycolysis. CONCLUSION: Taken together, our research found that circPHGDH downregulation reduced PTC progression via miR-122-5p/PKM2 axis regulation mediated by aerobic glycolysis.

Serum Levels of miR-122-5p and miR-125a-5p Predict Hepatotoxicity Occurrence in Patients Undergoing Autologous Hematopoietic Stem Cell Transplantation.

Hepatic complications are an acknowledged cause of mortality and morbidity among patients undergoing hematopoietic stem cell transplantation. In this study, we aimed to evaluate the potential role in the prediction of liver injury of five selected microRNAs (miRNAs)-miR-122-5p, miR-122-3p, miR-15b-5p, miR-99b-5p, and miR-125a-5p-in the setting of autologous hematopoietic stem cell transplantation (ASCT). A total of 66 patients were included in the study: 50 patients (75.8%) with multiple myeloma (MM) and 16 (24.2%) with lymphoma. Blood samples were collected after the administration of the conditioning regimen, on the day of transplant (day 0). The expression levels of selected miRNAs were quantified by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) using the miRCURY LNA miRNA Custom PCR Panels (QIAGEN). In a multivariate logistic regression analysis adjusted for age, sex, and the administered conditioning regimen, two miRNAs, hsa-miR-122-5p (odds ratio, OR 2.10, 95% confidence interval, CI: 1.29-3.42, p = 0.0029) and hsa-miR-125a-5p (OR 0.27, 95% CI: 0.11-0.71, p = 0.0079), were independent for hepatic toxicity occurrence during the 14 days after transplant. Our model in 10-fold cross-validation preserved its diagnostic potential with a receiver operating characteristics area under the curve (ROC AUC) of 0.75, 95% CI: 0.63-0.88 and at optimal cut-off reached 72.0% sensitivity and 74.4% specificity. An elevated serum level of miR-122-5p and decreased level of miR-125a-5p on day 0 are independent risk factors for hepatotoxicity in ASCT recipients, showing promise in accurately predicting post-ASCT complications. Identifying patients susceptible to complications has the potential to reduce procedure costs and optimize the selection of inpatient or outpatient procedures.

Tumor-Derived Exosomal Circular RNA Pinin Induces FGF13 Expression to Promote Colorectal Cancer Progression through miR-1225-5p.

Background/Aims: : Colorectal cancer (CRC) is a common malignant tumor, and circular RNAs (circRNAs) are abnormally expressed in CRC. However, the function and underlying mechanism of circRNA pinin (circ-PNN; hsa_circ_0101802) in CRC remain unclear. Methods: : Exosomes were isolated from the plasma of CRC patients and identified by transmission electron microscopy and Western blotting. The RNA expression levels of circ-PNN, miR-1225-5p, and fibroblast growth factor 13 (FGF13) were measured by quantitative real-time polymerase chain reaction. Cell proliferation was detected by Cell Counting K-8, colony formation, and 5-ethynyl-2'-deoxyuridine assays. Cell apoptosis was assessed by flow cytometry. The expression of apoptosis and metastasis-related proteins was evaluated by Western blotting. The associations among circ-PNN, miR-1225-5p, and FGF13 were confirmed by dual-luciferase report assay and RNA immunoprecipitation assay. A xenograft model was used to verify the function of circ-PNN in tumor formation in vivo. Results: : circ-PNN expression was upregulated in plasmic exosomes derived from CRC patients. The expression of circ-PNN and FGF13 was upregulated, while miR-1225-5p expression was downregulated in CRC cells incubated with plasmic exosomes derived from CRC patients. Tumor-derived exosomes promoted the proliferation, migration, and invasion but inhibited apoptosis of CRC cells. Moreover, the addition of tumor-derived exosomes partly reversed the inhibitory effect of circ-PNN knockdown on CRC tumor progression in vitro and in vivo. Thus, circ-PNN acts as a sponge for miR-1225-5p to regulate FGF13 expression. Conclusions: : Tumor-derived exosomal circ-PNN promoted CRC progression through the regulation of the miR-1225-5p/FGF13 pathway, providing a potential therapeutic target for CRC.

HSCs-derived exosomes regulate the levels of inflammatory cytokines in HIBECs through miR-122-5p mediated p38 MAPK signaling pathway.

PBC is an autoimmune-mediated liver disease, and intrahepatic biliary epithelial cells (IBECs) are the target cells of early damage. Previous studies found that miRNAs and inflammation is closely related to PBC. In this study, we extracted exosomes from serum and human IBECs supernatant, and RNA-sequence analyzed the expression profiles of miRNAs. Elisa measured the levels of inflammatory cytokines. RT- qPCR and western blot detected the levels of miR-122-5p, p38 and p-p38. The results showed that 263 differentially expressed (DE) miRNAs were identified in serum exosomes of PBC patients. The levels of IL-1ß, IL-6, IL-12, IL-17 A, IFN-γ, TNF-α and TGF-ß1 in peripheral blood of PBC patients were higher than those of normal controls. According to the validation results and previous literature, exosomal miR-122-5p was finally selected as the study object, and correlated with inflammatory factors. In vitro experiments further found that exosomal miR-122-5p may derive from hepatic stellate cells (HSCs), and can be HIBECs intake, and influence HIBECs inflammatory factor levels though p38 MAPK signaling pathways. This may provide a new strategy for the treatment of PBC.

Activation of IGFBP4 via unconventional mechanism of miRNA attenuates metastasis of intrahepatic cholangiocarcinoma.

BACKGROUND: Intrahepatic cholangiocarcinoma (ICC) is the second most common primary liver malignancy. Although its incidence is lower than that of hepatocellular carcinoma (HCC), ICC has a worse prognosis, and it is more prone to recur and metastasize, resulting in a far greater level of malignancy. METHODS: Bioinformatics analysis and qRT-PCR were applied to assess the level of miR-122-5p and IGFBP4. Western blot, transwell assays, wound-healing assays, real-time cellular invasion monitoring, in vivo study were applied to explore the function of miR-122-5p and IGFBP4. Dual luciferase reporter assays and chromatin isolation by RNA purification (ChiRP) were applied to explore the regulation of IGFBP4 by miR-122-5p. RESULTS: Using The Cancer Genome Atlas (TCGA) data set, Sir Run Run Shaw hospital data set and bioinformatics analyses, we identified miR-122-5p as a potential tumor suppressor in ICC and validated its suppressive effect in metastasis and invasion of ICC. Transcriptome sequencing, rescue and complement experiments were used to identify insulin-like growth factor binding protein 4 (IGFBP4) as a target of miR-122-5p. The mechanism by which miR-122-5p regulates IGFBP4 was clarified by chromatin separation RNA purification technology, and dual-luciferase reporter assays. We discovered a rare novel mechanism by which miR-122-5p promotes IGFBP4 mRNA transcription by binding to its promoter region. Furthermore, in mouse orthotopic metastasis model, miR-122-5p inhibited the invasion of ICC. CONCLUSION: In summary, our study revealed a novel mechanism of miR-122-5p and function of the miR-122-5p/IGFBP4 axis in the metastasis of ICC. We also highlighted the clinical value of miR-122-5p and IGFBP4 in inhibiting ICC invasion and metastasis.

Serum exosomes from hepatitis B virus-infected patients inhibit glycolysis in Sertoli cells via miR-122-5p/ALDOA axis.

Hepatitis B virus (HBV) infection is associated with male infertility. The mechanism includes an increase in chromosomal instability in sperm, which has an adverse effect on sperm viability and function. Sertoli cells (SCs) are vital in spermatogenesis because they use glycolysis to provide energy to germ cells and themselves. HBV infection impairs sperm function. However, whether HBV infection disrupts energy metabolism in SCs remains unclear. This study aimed to determine the role of serum exosomes of HBV-infected patients in SC viability and glycolysis. Serum exosomes were obtained from 30 patients with (HBV+_exo) or without (HBV-_exo) HBV infection using high-speed centrifugation and identified by transmission electron microscopy and western blot analysis. Cell viability is determined by CCK-8 assay. Glycolysis is determined by detecting extracellular acidification rate and ATP levels. miR-122-5p expression levels are detected by quantitative RT-PCR, and a dual-luciferase gene reporter assay confirms the downstream target gene of miR-122-5p. Protein expression is determined by western blot analysis. The results show that HBV+ _exo inhibited cell viability, extracellular acidification rate, and ATP production of SCs. miR-122-5p is highly expressed in HBV+ _exo compared with that in HBV-_exo. Furthermore, HBV+ _exo is efficiently taken up by SCs, whereas miR-122-5p is efficiently transported to SCs. miR-122-5p overexpression downregulates ALDOA expression and inhibits SC viability and glycolysis. However, ALDOA overexpression reverses the effects of miR-122-5p and HBV+ _exo on SC viability and glycolysis. HBV+ _exo may deliver miR-122-5p to target ALDOA and inhibit SC viability and glycolysis, thus providing new therapeutic ideas for treating HBV-associated male infertility.

MiR-122-5p as a potential regulator of pulmonary vascular wall cell in idiopathic pulmonary arterial hypertension.

MicroRNAs (miRNAs) are versatile regulators of pulmonary arterial remodeling in idiopathic pulmonary arterial hypertension (IPAH). We herein aimed to characterize miRNAs in peripheral blood mononuclear cell (PBMC) and plasma exosomes, and investigate specific miRNA expression in pulmonary artery cells and lung tissues in IPAH. A co-dysregulated miRNA was identified from the miRNA expression profiles of PBMC and plasma exosomes in IPAH. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed the potential function of differentially expressed miRNAs. Real-time quantitative reverse transcription polymerase chain reaction was used to validate the expression of specific miRNAs in hypoxia-induced pulmonary microvascular endothelial cells (PMECs), pulmonary artery smooth muscle cells (PASMCs), pericyte cells (PCs), and lung tissues of patients with IPAH and rats. Finally, the miRNA-mRNA mechanisms of miR-122-5p were predicted. MiR-122-5p was the only co-upregulated miRNA in PBMC and plasma exosomes in patients with IPAH. Functional analysis of differentially expressed miRNAs revealed associations with the GO terms "transcription, DNA-templated," "cytoplasm," and "metal ion binding" in both PBMC and plasma exosomes, KEGG pathway MAPK signaling in PBMC, and KEGG-pathway human papillomavirus infection in plasma exosomes. Hypoxic PMECs and PCs, lung tissue of patients with IPAH, and rats showed increased expression of miR-122-5p, but hypoxic PASMCs showed decreased expression. And miR-122-5p mimics and inhibitor affected cell proliferation. Finally, miR-122-5p was found to potentially target DLAT (in lung tissue) and RIMS1 (in PMECs) in IPAH. According to the dual-luciferase assay, miR-122-5p bound to DLAT or RIMS1. In studies, DLAT imbalance was associated with cell proliferation and migration, RIMS1 is differentially expressed in cancer and correlated with cancer prognosis. Our findings suggest that the miR-122-5p is involved in various biological functions in the adjacent vascular wall cells in IPAH.

LncRNA xist regulates sepsis associated neuroinflammation in the periventricular white matter of CLP rats by miR-122-5p/PKCη Axis.

Background: Neuroinflammation is a common feature of many neurological diseases, and remains crucial for disease progression and prognosis. Activation of microglia and astrocytes can lead to neuroinflammation. However, little is known about the role of lncRNA xist and miR-122-5p in the pathogenesis of sepsis-associated neuroinflammation (SAN). This study aims to investigate the role of lncRNA xist and miR-122-5p in the pathogenesis of SAN. Methods: Levels of miR-122-5p and proinflammatory mediators were detected in the cerebrospinal fluid (CSF) of patients with intracranial infection (ICI) by ELISA and qRT-PCR. miRNA expression in the periventricular white matter (PWM) in rats was analyzed by high-throughput sequencing. Levels of lncRNA xist, miR-122-5p and proinflammatory mediators in the PWM were measured using qRT-PCR and western blot. Bioinformatics analysis was used to predict the upstream and downstream of miR-122-5p. The interaction between miR-122-5p and its target protein was validated using luciferase reporter assay. BV2 and astrocytes were used to detect the expression of lncRNA xist, miR-122-5p. Results: The level of miR-122-5p was significantly decreased in the CSF of ICI patients, while the expression of IL-1ß and TNF-α were significantly upregulated. Furthermore, it was found that the expression of IL-1ß and TNF-α were negatively correlated with the level of miR-122-5p. A high-throughput sequencing analysis showed that miR-122-5p expression was downregulated with 1.5-fold changes in the PWM of CLP rats compared with sham group. Bioinformatics analysis found that lncRNA xist and PKCη were the upstream and downstream target genes of miR-122-5p, respectively. The identified lncRNA xist and PKCη were significantly increased in the PWM of CLP rats. Overexpression of miR-122-5p or knockdown of lncRNA xist could significantly downregulate the level of PKCη and proinflammatory mediators from activated microglia and astrocytes. Meanwhile, in vitro investigation showed that silencing lncRNA xist or PKCη or enhancing the expression of miR-122-5p could obviously inhibit the release of proinflammatory mediators in activated BV2 cells and astrocytes. Conclusion: LncRNA xist could regulate microglia and astrocytes activation in the PWM of CLP rats via miR-122-5p/PKCη axis, further mediating sepsis associated neuroinflammation.

Hypoxic tumour-derived exosomal miR-1225-5p regulates M2 macrophage polarisation via toll-like receptor 2 to promote ovarian cancer progress.

Tumor-secreted exosomes are critical for the functional regulation of tumor-associated macrophages (TAMs). This study aimed to explore how exosomes secreted by ovarian carcinoma cells regulate the phenotype and function of macrophages. Hypoxic treatment of A2780 cells was postulated to mimic the tumor microenvironment, and exosomes were co-cultured with TAMs. miR-1225-5p was enriched in hypoxic exosomes and contributed to M2 macrophage polarizationby modulating Toll-like receptor 2 expression (TLR2). Furthermore, hypoxia-treated macrophages promote ovarian cancer cell viability, migration, and invasion via the wnt/ß-catenin pathway. This study clarified that exosomal miR-1225-5p promotes macrophage M2-like polarization by targeting TLR2 to promote ovarian cancer, which may via the wnt/ß-catenin pathway.

Extracellular Vesicles from Human Adipose Tissue-Derived Mesenchymal Stem Cells Suppress RANKL-Induced Osteoclast Differentiation via miR122-5p.

Researchers are increasingly interested in cell therapy using mesenchymal stem cells (MSCs) as an alternative remedy for osteoporosis, with fewer side effects. Thus, we isolated and characterized extracellular vesicles (EVs) from human adipose tissue-derived MSCs (hMSCs) and investigated their inhibitory effects on RANKL-induced osteoclast differentiation. Purified EVs were collected from the supernatant of hMSCs by tangential flow filtration. Characterization of EVs included typical evaluation of the size and concentration of EVs by nanoparticle tracking analysis and morphology analysis using transmission electron microscopy. hMSC-EVs inhibited RANKL-induced differentiation of bone marrow-derived macrophages (BMDMs) into osteoclasts in a dose-dependent manner. F-actin ring formation and bone resorption were also reduced by EV treatment of osteoclasts. In addition, EVs decreased RANKL-induced phosphorylation of p38 and JNK and expression of osteoclastogenesis-related genes in BMDMs treated with RANKL. To elucidate which part of the hMSC-EVs plays a role in the inhibition of osteoclast differentiation, we analyzed miRNA profiles in hMSC-EVs. The results showed that has-miR122-5p was present at significantly high read counts. Overexpression of miR122-5p in BMDMs significantly inhibited RANKL-induced osteoclast differentiation and induced defects in F-actin ring formation and bone resorption. Our results also revealed that RANKL-induced phosphorylation of p38 and JNK and osteoclast-specific gene expression was decreased by miR122-5p transfection, which was consistent with the results of hMSC-EVs. These findings suggest that hMSC-EVs containing miR122-5p inhibit RANKL-induced osteoclast differentiation via the downregulation of molecular mechanisms and could be a preventive candidate for destructive bone diseases.

Peptide OA-VI12 restrains melanogenesis in B16 cells and C57B/6 mouse ear skin via the miR-122-5p/Mitf/Tyr axis.

Excessive melanogenesis leads to hyperpigmentation, which is one of the common skin conditions in humans. Existing whitening cosmetics cannot meet market needs due to their inherent limitations. Thus, the development of novel skin-whitening agents continues to be a challenge. The peptide OA-VI12 from the skin of amphibians at high altitude has attracted attention due to its remarkable anti light damage activity. However, whether OA-VI12 has the skin-whitening effect of inhibiting melanogenesis is still. Mouse melanoma cells (B16) were used to study the effect of OA-VI12 on cell viability and melanin content. The pigmentation model of C57B/6 mouse ear skin was induced by UVB and treated with OA-VI12. Melanin staining was used to observe the degree of pigmentation. MicroRNA sequencing, quantitative real-time PCR (qRT-PCR), immunofluorescence analysis and Western blot were used to detect the change of factor expression. Double luciferase gene report experiment was used to prove the regulatory relationship between miRNA and target genes. OA-VI12 has no effect on the viability of B16 cells in the concentration range of 1-100 µM and significantly inhibits the melanin content of B16 cells. Topical application of OA-VI12, which exerted transdermal potency, prevented UVB-induced pigmentation of ear skin. MicroRNA sequencing and double luciferase reporter analysis results showed that miR-122-5p, which directly regulated microphthalmia-associated transcription factor (Mitf), had significantly different expression before and after treatment with OA-VI12. Mitf is a simple helix loop and leucine zipper transcription factor that regulates tyrosinase (Tyr) expression by binding to the M-box promoter element of Tyr. qRT-PCR, immunofluorescence analysis and Western blot showed that OA-VI12 up-regulated the expression of miR-122-5p and inhibited the expression of Mitf and Tyr. The effects of OA-VI12 on melanogenesis inhibition in vitro and in vivo may involve the miR-122-5p/Mitf/tyr axis. OA-VI12 represents the first report on a natural amphibian-derived peptide with skin-whitening capacity and the first report of miR-122-5p as a target for regulating melanogenesis, thereby demonstrating its potential as a novel skin-whitening agent and highlighting amphibian-derived peptides as an underdeveloped resource.

miR-122-5p Restrains Pancreatic Cancer Cell Growth and Causes Apoptosis by Negatively Regulating ASCT2.

BACKGROUND/AIM: System ASC amino acid transporter-2 (ASCT2) is abnormally highly expressed in tumor cells and closely associated with a poor prognosis, but the regulatory mechanism of abnormally high ASCT2 expression is scarcely investigated. MicroRNAs (miRNAs) that are abnormally expressed regulate gene expression to have either oncogenic or tumor-suppressive effects in pancreatic cancer (PC). MicroRNA-122-5p (miR-122-5p) dysregulation has been seen in various cancer entities, but the biological function of miR-122-5p in PC and its regulation mechanisms remain unknown. MATERIALS AND METHODS: Western blot and quantitative RT-PCR were used to measure the expression of miR-122-5p, ASCT2, and apoptosis-related proteins. CCK-8 assays were used to elucidate the effect on cell proliferation. Flow cytometry (FCM) assays were utilized to evaluate cell apoptosis. A dual-luciferase reporter assay was utilized to determine if miR-122a-5p directly targeted ASCT2. Glutamine consumption and the α-ketoglutarate (α-KG) and adenosine triphosphate (ATP) contents were determined using respective assays. RESULTS: MiR-122-5p expression was low whereas ASCT2 expression was high in PC tissues and cells. Overexpressing miR-122-5p restrained pancreatic cancer cell proliferation, accelerated apoptosis, and decreased glutamine consumption, α-ketoglutarate (α-KG) production and ATP generation, whereas suppressing miR-122-5p had the opposite effect. Moreover, the reporter gene test established ASCT2 as a miR-122-5p target. Overexpression of miR-122-5p decreased ASCT2 expression, whereas miR-122-5p repression increased ASCT2 expression. In addition, miR-122-5p also regulated apoptosis-related pathways. CONCLUSION: MiR-122-5p may function as a tumor suppressor by inhibiting the proliferation, glutamine metabolism, and inducing apoptosis via altering the expression of ASCT2 in pancreatic cancer cells.

miR-122-5p Promotes Peripheral and Central Nervous System Inflammation in a Mouse Model of Intracerebral Hemorrhage via Disruption of the MLLT1/PI3K/AKT Signaling.

Intracerebral hemorrhage (ICH) is a recognized central nervous system inflammation complication. Several microRNAs (miRNAs or miRs) have been documented to be vital modulators in peripheral and central nervous system inflammation. Based on whole transcriptome sequencing and bioinformatics analysis, this study aims to reveal the possible molecular mechanisms by which miR-122-5p affects the inflammatory response in the peripheral and central nervous system in a mouse model of ICH. Differentially expressed ICH-related miRNAs were screened. Adeno-associated viral vectors were used to knock down miR-122-5p in mice to evaluate the effect of miR-122-5p on peripheral and central nervous system inflammation. The downstream target gene of miR-122-5p was analyzed. Neurons were isolated from mice and treated with hemin to construct an in vitro model of ICH, followed by transduction with miR-122-5p mimic or combined with oe-MLLT1. The neurons were then co-cultured with microglia BV2 to assess their activation. It was found that miR-122-5p was highly expressed in ICH, and MLLT1 was lowly expressed. In vivo experiments showed that miR-122-5p knockdown decreased neurological deficits, BBB permeability, and inflammation in the peripheral and central nervous system in ICH mice. It involved its binding to MLLT1 and downregulation of the activity of the PI3K/AKT pathway. In vitro data exhibited that miR-122-5p stimulated the generation of inflammatory factors and microglia activation by targeting MLLT1 and inhibiting the PI3K/AKT pathway. Collectively, our work reveals a novel miR-122-5p/MLLT1-mediated regulatory network in ICH that may be a viable target for neuroinflammation alleviation.

Circ_LRP6 facilitates osteosarcoma progression via the miR-122-5p/miR-204-5p/HMGB1 axis.

Circ_LRP6 is participated in the occurrence and development of numerous tumors. Nevertheless, its roles and mechanism in osteosarcoma (OS) is unknown. This study aims to illustrate this point. With the use of qRT-PCR, the level of circ_LRP6, miR-122-5p, miR-204-5p and HMGB1 was identified. To observe cell proliferation, migration and invasion, we adopted CCK-8 and Transwell assays in the present study. Besides, to prove the existing interaction, bioinformatics analysis and dual luciferase reporting assays were employed. The influence of circ_LRP6 on osteosarcoma in vivo was evaluated by subcutaneous tumor formation model in nude mice. In osteosarcoma tissues, circ_LRP6 and HMGB1 are strongly denoted, whereas miR-122-5p and miR-204-5p are under-expressed. Circ_LRP6 knockdown could significantly hinder the proliferation, migration and invasion of osteosarcoma cells. Circ_LRP6 hindered the proliferation of osteosarcoma in vivo. Bioinformatics predicted that miR-122-5p and miR-204-5p functioned as direct targets of circ_LRP6, and HMGB1 were possible target genes of miR-122-5p and miR-204-5p. The findings indicated that the low level of miR-122-5p and miR-204-5p and the overexpression of HMGB1 could partially restore and reduce the inhibitory impact of circ_LRP6 on the proliferation, migration and invasion of osteosarcoma cells. Circ_LRP6 affects osteosarcoma progression via the miR-122-5p/miR-204-5p/HMGB1 axis, and is shown to be a molecular biomarker.

Transcriptome analysis reveals FABP5 as a key player in the development of chicken abdominal fat, regulated by miR-122-5p targeting.

BACKGROUND: The development of abdominal fat and meat quality are closely related and can impact economic efficiency. In this study, we conducted transcriptome sequencing of the abdominal fat tissue of Gushi chickens at 6, 14, 22, and 30 weeks, and selected key miRNA-mRNA regulatory networks related to abdominal fat development through correlation analysis. RESULTS: A total of 1893 differentially expressed genes were identified. Time series analysis indicated that at around 6 weeks, the development of chicken abdominal fat was extensively regulated by the TGF-ß signaling pathway, Wnt signaling pathway, and PPAR signaling pathway. However, at 30 weeks of age, the apoptosis signaling pathway was the most significant, and correlation analysis revealed several genes highly correlated with abdominal fat development, including Fatty Acid Binding Protein 5 (FABP5). Based on miRNA transcriptome data, it was discovered that miR-122-5p is a potential target miRNA for FABP5. Cell experiments showed that miR-122-5p can directly target FABP5 to promote the differentiation of preadipocytes. CONCLUSION: The present study confirms that the key gene FABP5 and its target gene miR-122-5p are critical regulatory factors in the development of chicken abdominal fat. These results provide new insights into the molecular regulatory mechanisms associated with the development of abdomen-al fat in chickens.