MicroRNA-218-5p relieves postmenopausal osteoporosis through promoting the osteoblast differentiation of bone marrow mesenchymal stem cells

MicroRNAs (miRs) are short noncoding RNAs that play key regulatory roles in osteoblast differentiation. In this study, the specific regulatory roles of miR-218-5p on postmenopausal osteoporosis (PMOP) were investigated. The mouse model of PMOP was established by bilateral ovariectomy, and the injection of miR-218-5p mimics significantly relieved PMOP degree. Then, bone marrow mesenchymal stem cells (BMMSCs) isolated from PMOP mice were induced into osteoblasts. When compared with normal BMMSCs , PMOP BMMSCs exhibited significantly lower alkaline phosphatase (ALP) activity and less mineralized nodules, as well as downregulated miR-218-5p, Runx2, Osterix, COL1A1, and OCN after induction (P < .05). The transfection of miR-218-5p mimics, and inhibitor significantly promoted, inhibited the osteoblast differentiation of PMOP BMMSCs, respectively. In addition, COL1A1 was a target of miR-218-5p. The transfection of miR-218-5p mimics into PMOP BMMSCs significantly upregulated COL1A1 at 14th and 21st day post-induction, but not at 7th day. Our findings suggest miR-218-5p may relieve PMOP through promoting the osteoblast differentiation of BMMSCs.     

慢性牙周炎患者血清miR-205-5p、miR-28-5p的表达与诊断价值研究

摘要 目的：探讨慢性牙周炎（CP）患者血清微小核糖核酸（miRNA）-205-5p、miR-28-5p表达与牙周指标、炎症反应的关系及诊断价值。方法：选取2021年1月～2022年12月我院口腔科收治的102例CP患者为CP组,根据病情严重程度分为轻度组31例、中度组37例、重度组34例,另选取同期我院65名体检健康者为对照组。采用实时荧光定量聚合酶链式反应（PCR）检测血清miR-205-5p、miR-28-5p表达,酶联免疫吸附法检测白细胞介素（IL）-1β、IL-6、肿瘤坏死因子-α（TNF-α）水平,检查各组牙周指标[菌斑指数（PLI）、牙龈出血指数（BI）、附着丧失（AL）、探诊深度（PD）]。CP患者血清miR-205-5p、miR-28-5p表达与牙周指标和炎症指标的相关性采用Pearson相关性分析。受试者工作特征（ROC）曲线分析血清miR-205-5p、miR-28-5p表达诊断CP的价值。结果：与对照组比较,CP组血清miR-205-5p、miR-28-5p表达降低,PLI、BI、AL、PD和IL-1β、IL-6、TNF-α水平升高（P＜0.05）。轻度组、中度组、重度组血清miR-205-5p、miR-28-5p表达依次降低,PLI、BI、AL、PD和IL-1β、IL-6、TNF-α水平依次升高（P＜0.05）。Pearson相关性分析显示,CP患者血清miR-205-5p、miR-28-5p表达与PLI、BI、AL、PD、IL-1β、IL-6、TNF-α呈负相关（P＜0.05）。ROC曲线分析显示,血清miR-205-5p联合miR-28-5p诊断CP的曲线下面积分别为0.885大于miR-205-5p、miR-28-5p单独检测的0.792、0.790。结论：CP患者血清miR-205-5p、miR-28-5p低表达,与牙周指标、炎症反应密切相关,血清miR-205-5p、miR-28-5p的表达情况可能成为CP辅助诊断指标,且miR-205-5p联合miR-28-5p诊断CP的价值较高。     

miR-205-5p in exosomes divided from chondrogenic mesenchymal stem cells alleviated rheumatoid arthritis via regulating MDM2 in fibroblast-like synoviocytes

Objective:To explore the role and mechanism of chondrogenic bone marrow mesenchymal stem cells (BMSCs)-derived exosomes on Rheumatoid arthritis (RA).Methods:The chondrogenesis of BMSCs was induced by chondrogenic medium. Exosomes from BMSCs and chondrogenic BMSCs were isolated and characterized by transmission electron microscope (TEM), laser particle size analyzer and western blot. ELISA was used to analyze the expression levels of pro-inflammatory cytokines and matrix metalloproteinases (MMPs). Western bolt was performed to assess MAPK and NF-κB pathways expression. The inflammation score and the pathological damage of RA mice were evaluated. Luciferase reporter assay and RIP were carried out to examine the relationship between microRNA-205-5p (miR-205-5p) and mouse double minute 2 (MDM2).Results:Chondrogenic BMSCs-derived exosomes suppressed pro-inflammatory cytokines, MMPs and MAPK and NF-κB pathways in RA-FLSs. miR-205-5p had a high expression in chondrogenic BMSCs-derived exosomes. Functionally, exosomal miR-205-5p also played the anti-inflammation effects. Besides, MDM2 was a direct target of miR-205-5p. Additionally, chondrogenic BMSCs-secreted exosomal miR-205-5p suppressed the inflammation score, joint destruction, and inflammatory response in collagen-induced arthritis (CIA) mice through MDM2.Conclusion:Chondrogenic BMSCs-derived exosomal miR-205-5p suppressed inflammatory response, MAPK and NF-κB pathways through MDM2 in RA, indicating exosomal miR-205-5p might be a potential target for RA treatment.     

miR-15a-5p和miR-16-5p在骨肉瘤组织中的表达分析

目的:使用microRNAs基因芯片及实时定量PCR法测定骨肉瘤组织中miR-15a-5p和miR-16-5p的相对表达含量,并与瘤旁组织对比,分析骨肉瘤细胞内miR-15a-5p和miR-16-5p的表达变化。方法:选取34例骨肉瘤组织蜡块样本,使用microRNAs基因芯片观察miR-15a-5p和miR-16-5p在骨肉瘤和瘤旁组织内的表达差异;实时定量PCR法测定骨肉瘤组织和瘤旁组织中miR-15a-5p和miR-16-5p的相对表达含量,并将两种结果对比分析。结果:microRNAs基因芯片结果显示,在骨肉瘤组织中,miR-15a-5p在肿瘤中的表达较瘤旁组织低1.79倍,miR-16-5p较瘤旁组织低1.62倍。实时定量PCR实验结果表明,miR-15a-5p和miR-16-5p表达较瘤旁组织降低,差异有统计学意义(P0.05)。经过统计学计算,miR-15a-5p在肿瘤中的表达较瘤旁组织低3.14倍,miR-16-5p较瘤旁组织低5.65倍。结论:在骨肉瘤中,miR-15a-5p和miR-16-5p表达含量降低,提示这两种microRNAs在骨肉瘤中可能做为抑癌因子存在。     

miR-305-3p和miR-71-5p通过调控谷胱甘肽代谢途径参与斜纹夜蛾应对植物次生物质

昆虫和植物在长期进化过程中形成相互作用的关系。其中植物次生物质是植物防御昆虫的主要机制,而昆虫则以解毒酶来应对。为了发现可用于害虫防治的miRNA,通过对农业害虫斜纹夜蛾Spodoptera litura进食芥菜Brassica juncea后中肠的测序分析,获得了一系列差异表达的miRNA。通过生物信息学分析,发现斜纹夜蛾miR-305-3p靶向了谷胱甘肽代谢中的谷氨酸半胱氨酸连接酶的催化亚基(Slgclc),这是一个谷胱甘肽从头合成的限速酶;而miR-71-5p靶向调控gclc和解毒酶表达的转录因子SlNrf2。用芥菜的次生物质吲哚-3-甲醇处理斜纹夜蛾Spli-221细胞株后,实时荧光定量PCR证明Slgclc和SlNrf2表达上调,miR-305-3p和miR-71-5p表达下调。分别在斜纹夜蛾Spli-221细胞中超表达miR-305-3p及miR-71-5p mimics,Slgclc或SlNrf2转录水平下调,并且miR-71-5p mimic处理抑制了SlNrf2下游基因Slgclc和解毒酶谷胱甘肽S-转移酶(GSTs)的表达。结果表明:miR-305-3p和miR-71-5p响应植物次生物质而分别促进Slgclc和SlNrf2表达。然而双荧光素酶实验显示miR-305-3p并不与Slgclc直接结合,miR-71-5p也不与SlNrf2直接结合,推测miR-305-3p和miR-71-5p可能间接调控Slgclc和SlNrf2的表达。研究结果表明,斜纹夜蛾miR-305-3p和miR-71-5p通过调控解毒酶GSTs表达及其底物谷胱甘肽的生成,而参与昆虫抵抗植物次生物质。     

The long noncoding RNA ZFAS1 promotes the progression of glioma by regulating the miR-150-5p/PLP2 axis

Numerous studies have reported that long noncoding RNA (lncRNA) dysregulation is involved in the progression of many malignant tumors, including glioma. The lncRNA ZNFX1 antisense RNA 1 (ZFAS1) plays an oncogenic role in various malignant tumors, such as gastric cancer and hepatocellular carcinoma. However, the underlying molecular mechanism of ZFAS1 in glioma has not been fully clarified. In this study, we found that the expression of ZFAS1 was upregulated in both glioma tissues and cell lines. Functional experiments revealed that ZFAS1 promoted glioma proliferation, migration and invasion, and increased resistance to temozolomide in vitro. By using online databases, RNA pull-down assays and luciferase reporter assays, ZFAS1 was demonstrated to act as a sponge of miR-150-5p. Furthermore, proteolipid protein 2 (PLP2) was shown to be the functional target of miR-150-5p. Rescue experiments revealed that ZFAS1 regulated the expression of PLP2 by sponging miR-150-5p. Finally, a xenograft tumor assay demonstrated that ZFAS1 promoted glioma growth in vivo. Our results showed that ZFAS1 promoted glioma malignant progression by regulating the miR-150-5p/PLP2 axis, which may provide a potential therapeutic target for the treatment of glioma.     

Circular RNA circ_0011269 sponges miR-122 to regulate RUNX2 expression and promotes osteoporosis progression

Circular RNAs (circRNAs) are a novel class of noncoding RNAs that are widely expressed in human disease. However, circRNAs expression profile and potential mechanism in osteoporosis pathogenesis remain to be further studied. In the present study, a total of 69 circRNAs were identified to be abnormally expressed in osteoporosis patient samples by microarray and bioinformatics analyses. We found that circ_0011269 was notably downregulated in osteoporosis (fold change, 3.94). By means of miRanda algorithm, we constructed the interaction network of circ_0011269-miRNAs in osteoporosis based on target binding and miR-122 was enrolled in the network. Dual-luciferase reporter assay verified the target relationship of miR-122 and circ_0011269/RUNX2. The expression of circ_0011269 and RUNX2 were gradually increased during osteogenic differentiation while miR-122 exhibited a decreased expression. Moreover, overexpression of circ_0011269 could promote RUNX2 expression and inhibit osteoporosis. In summary, this study found that circ_0011269 sponges miR-122 to regulate RUNX2 expression and promotes osteoporosis progression.     

miR-145-5p affects the differentiation of gastric cancer by targeting KLF5 directly

Krüppel-like factor 5 (KLF5) takes part in the pathologic processes of many types of cancer; however, its expression and roles in the biological behavior of gastric cancer remain unknown. TargetScan suggested that miR-145-5p is the predicted effective and conserved microRNA (miRNA) that binds to KLF5 through its 3′-untranslated region (UTR). We investigated the expression of KLF5 and miR-145-5p messenger RNA (mRNA) in gastric cancer and then analyzed its role in the biological behavior of gastric cancer cells. Our results indicated that KLF5 expression was detected by immunohistochemistry in 39.7% of the gastric cancer cases and was increased compared with that of the corresponding noncancerous normal mucosa (0.01 < p < 0.05). The poorly differentiated subtype showed positive KLF5 expression, whereas the differentiated subtype showed negative KLF5 expression (p < 0.05). Dual-luciferase reporter assay suggested KLF5 3′-UTR was the direct target of miR-145-5p. Compared with the differentiated gastric cancer, miR-145-5p was downregulated in undifferentiated gastric cancer (p < 0.05). The downregulation of KLF5 expression and differentiation of MGC-803 and BGC-823 caused by siKLF5 or miR-145-5p mimic transfection. Our results indicated that miR-145-5p/KLF5 3′-UTR affected the differentiation of gastric cancer. miR-145-5p was able to promote gastric cancer differentiation by targeting KLF5 3′-UTR directly. Our data suggest a novel mechanism for cancer differentiation and a new facet to the role of miR-145-5p/KLF5 in gastric cancer.     

MiR-138-5p Targets MACF1 to Aggravate Aging-related Bone Loss

Senile osteoporosis is one of the major health problems in an aging society. Decreased bone formation due to osteoblast dysfunction may be one of the causes of aging-related bone loss. With increasing evidence suggesting that multiple microRNAs (miRNAs) play important roles in osteoblast function, the relationship between miRNAs and senile osteoporosis has become a popular research topic. Previously, we confirmed that mechanoresponsive miR-138-5p negatively regulated bone anabolic action. In this study, the miR-138-5p level was found to be negatively correlated with BMD and osteogenic markers in bone specimens of senile osteoporotic patients by bioinformatic analysis and experimental verification. Furthermore, high miR-138-5p levels aggravated the decrease of aged osteoblast differentiation in vitro and led to worse bone loss in aged osteoblastic miR-138-5p transgenic mice in vivo. We also previously identified that the target of miR-138-5p, microtubule actin cross-linking factor 1 (MACF1), could attenuate senile osteoporosis. Here, miR-138-5p was demonstrated to regulate aged osteoblast differentiation by targeting MACF1. Finally, the therapeutic inhibition of miR-138-5p counteracted the decrease in bone formation and aging-related bone loss in aged mice. Overall, our results highlight the crucial roles and the molecular mechanism of miR-138-5p in aging-related bone loss and may provide a powerful therapeutic target for ameliorating senile osteoporosis.     

Circulating miR-3613-5p but not miR-125b-5p,miR-199a-3p,and miR-451a are biomarkers of endometriosis

ObjectiveThis study aimed to assess the utility of circulating miR-125b-5p, miR-199a-3p, miR-451a, and miR-3613-5p as biomarkers of endometriosis.Study designPatients with stage III or IV of endometriosis according to the revised American Society of Reproductive Medicine (rASRM) staging classification, as well as control women, were recruited. We created a prospective study conducted on a group of 48 patients (n = 25 controls, n = 24 endometriosis) who had laparoscopic surgery. Blood samples were taken and plasma miRNA levels were measured by quantitative real-time polymerase chain reaction (RT-qPCR) and assessed with AUC and ROC curves.ResultsMiR-451a and miR-3613-5p were significantly decreased in the plasma of endometriosis patients. miR-451a had a receiver-operating characteristic (ROC) area under the curve 0.8283 and miR-3613-5p had a ROC area under the curve 0.7617. The concentration of circulating miR-125b-5p and miR-199-3p did not differ between endometriosis patients and controls. Plasma miRNA levels did not change with BMI, smoking status, fertility problems, or menstrual pain according to the VAS scale (p > 0.05).ConclusionCirculating miR-451a and miR-3613-5p levels significantly differed between endometriosis and controls. However, the levels of miR-451a were discordant with previous studies. Therefore, miR-3613-5p may have better potential as the endometriosis biomarker. Circulating miR-125b-5p and miR-199a-3p cannot be used as reliable markers of endometriosis.     

miR-625-5p/PKM2 negatively regulates melanoma glycolysis state

PKM2 plays an important role in cancer glycolysis, however, the link of PKM2 and microRNAs (miRNAs) in melanoma is still unclear. The study will investigate the role of miRNAs in regulating PKM2 mediated melanoma cell glycolysis. We found that high PKM2 expression in melanoma tissues and cell lines was positively associated with glycolysis. Further study indicated that miR-625-5p regulated PKM2 expression on mRNA and protein levels in melanoma cells. There was a negative relationship between miR-625-5p and PKM2 expression in the clinical melanoma samples. These findings provide an evidence that miR-625-5p/PKM2 plays a role in melanoma cell glucose metabolism.     

Circular RNA CircSLC8A1 contributes to osteogenic differentiation in hBMSCs via CircSLC8A1/miR-144-3p/RUNX1 in periprosthetic osteolysis

Circular RNAs (circRNAs) are often found in eukaryocyte and have a role in the pathogenesis of a variety of human disorders. Our related research has shown the differential expression of circRNAs in periprosthetic osteolysis (PPOL). However, the involvement of circRNAs in the exact process is yet unknown. CircSLC8A1 expression was evaluated in clinical samples and human bone marrow mesenchymal stem cells (hBMSCs) in this investigation using quantitative real-time PCR. In vitro and in vivo studies were conducted to explicate its functional role and pathway. We demonstrated CircSLC8A1 is involved in PPOL using gain- and loss-of-function methods. The association of CircSLC8A1 and miR-144-3p, along with miR-144-3p and RUNX1, was predicted using bioinformatics. RNA pull-down and luciferase assays confirmed it. The impact of CircSLC8A1 in the PPOL-mouse model was also investigated using adeno-associated virus. CircSLC8A1 was found to be downregulated in PPOL patients' periprosthetic tissues. Overexpression of CircSLC8A1 promoted osteogenic differentiation (OD) and inhibited apoptosis of hBMSCs in vitro. The osteogenic markers of RUNX1, osteopontin (OPN) and osteocalcin (OCN) were significantly upregulated in hBMSCs after miR-144-3p inhibitor was transferred. Mechanistic analysis demonstrated that CircSLC8A1 directly bound to miR-144-3p and participated in PPOL through the miR-144-3p/RUNX1 pathway in hBMSCs. Micro-CT and quantitative analysis showed that CircSLC8A1 markedly inhibited PPOL, and osteogenic markers (RUNX1, OPN and OCN) were significantly increased (P<0.05) in the mice model. Our findings prove that Circ SLC8A1 exerted a regulatory role in promoting osteogenic differentiation in hBMSCs, and CircSLC8A1/miR-144-3p/RUNX1 pathway may provide a potential target for prevention of PPOL.     

miR-3065-5p regulates mouse odontoblastic differentiation partially through bone morphogenetic protein receptor type II

Illumination of the molecular mechanisms regulating odontoblastic differentiation of dental papilla cells is of great significance for proper dentinogenesis and dental pulp regeneration. In this study, we discovered that microRNA (miR)-3065-5p is up-regulated during odontoblastic differentiation. Overexpression of miR-3065-5p promoted odontoblastic differentiation in vitro. Dual luciferase report assay verified that miR-3065-5p could bind to the 3′UTR of bone morphogenetic protein receptor type II (BMPR2), which dramatically increased in the beginning of odontoblastic differentiation but decreased in the terminal differentiation stage. Inhibition of Bmpr2 in the early stage retarded odontoblastic differentiation while knockdown of Bmpr2 in the terminal stage enhanced odontoblastic differentiation, resembling the effect of miR-3065-5p. Taken together, our present study suggests that miR-3065-5p positively regulates odontoblastic differentiation by directly binding to Bmpr2 in the terminal differentiation stage.     

Effect of miR-143-3p on C2C12 myoblast differentiation

MicroRNAs are a class of 18–22 nucleotide non-coding RNAs that modulate gene expression by associating with the 3′ untranslated regions of mRNAs. A large number of microRNAs are involved in the regulation of myoblast differentiation, many of which remain undiscovered. In this study, we found that miR-143-3p was upregulated during C2C12 myoblast differentiation and over-expression of miR-143-3p significantly inhibited the relative expression levels of MyoD, MyoG, myf5, and MyHC genes, especially in the later stages of differentiation. In addition, miR-143-3p inhibited expression of genes involved in the endogenous Wnt signaling pathway during C2C12 myoblast differentiation, including Wnt5a, LRP5, Axin2, and β-catenin. These results indicate that miR-143-3p represents a new myogenic differentiation-associated microRNA that can inhibit C2C12 myoblast differentiation, especially in the later stages of differentiation.