bims-mirbon Biomed News
on MicroRNAs in bone
Issue of 2022–01–30
nine papers selected by
Japneet Kaur, Mayo Clinic



  1. Arch Biochem Biophys. 2022 Jan 24. pii: S0003-9861(22)00019-4. [Epub ahead of print] 109134
      Osteoporosis is a common complication accompanied by spinal cord injury (SCI) occurrence. MicroRNAs (miRNAs) have been proved to play a crucial role in the progression of osteoporosis, but their regulating mechanism is unclear. The present study investigated miRNA-19b-3p level in SCI rats induced by modified Allen method, as well as the role of miRNA-19b-3p in osteogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs). MiRNA-19b-3p expression and bone mineral density (BMD) of femurs were measured at day 21 and day 60 after SCI in rats. Obvious miRNA-19b-3p up-regulation and aggravated bone loss were observed. MiRNA-19b-3p overexpression suppressed BMSC-derived osteoblast differentiation, which was confirmed by the decrease in alkaline phosphatase (ALP) activity, EBF2 expression, osteoprotegrin (OPG) to receptor activator of nuclear factor kappa-B ligand (RANKL) ratio and cell mineralization degree. Besides, MiRNA-19b-3p knockdown could reverse this phenomenon. Dual-luciferase reporter assays verified the targeting relationship between miRNA-19b-3p and EBF2. The in vivo experiments confirmed miRNA-19b-3p down-regulation could significantly attenuate osteoporosis after SCI, which was verified by increased BMD, collagen content, femur mineralization degree, EBF2 protein, and OPG-to-RANKL ratio. The results show that miRNA-19b-3p plays an important role in the osteoporosis process after SCI through regulating EBF2 expression.
    Keywords:  EBF2; Osteoblast differentiation; Osteoporosis; SCI; miRNA-19b-3p
    DOI:  https://doi.org/10.1016/j.abb.2022.109134
  2. Front Genet. 2021 ;12 783352
      Objective: Osteoporosis is caused by the dysregulation of bone homeostasis which is synergistically mediated by osteoclasts and osteoblasts. MiR-27a-3p is a key inhibitor of bone formation. Hence, unearthing the downstream target gene of miR-27a-3p is of great significance to understand the molecular mechanism of osteoporosis. Methods: Bioinformatics analysis was utilized to find the downstream target gene of miR-27a-3p, and dual-luciferase reporter assay was conducted to validate the interplay of miR-27a-3p and GLP1R. Besides, qRT-PCR, Western blot, and enzyme-linked immunosorbent assay (ELISA) were employed to verify the impact of miR-27a-3p on GLP1R expression and the differentiation, autophagy, and inflammatory response of MC3T3-E1 pre-osteoblasts. Results: Dual-luciferase assay validated that miR-27a-3p directly targeted GLP1R. Additionally, posttreatment of MC3T3-E1 cells with miR-27a-3p mimics resulted in a remarkable decrease in expression levels of GLP1R, cell differentiation marker gene, autophagy marker gene, and AMPK. These results indicated that miR-27a-3p targeted GLP1R to inhibit AMPK signal activation and pre-osteoblast differentiation and autophagy, while promoting the release of inflammatory factors. Conclusion: The miR-27a-3p/GLP1R regulatory axis in pre-osteoblasts contributes to understanding the molecular mechanism of osteoporosis.
    Keywords:  GLP1R; MiR-27a-3p; autophagy; differentiation; inflammation; osteoblast
    DOI:  https://doi.org/10.3389/fgene.2021.783352
  3. Biochem Biophys Res Commun. 2022 Jan 13. pii: S0006-291X(21)01716-2. [Epub ahead of print]594 117-123
       BACKGROUND: Research on the role of lncRNAs in the process of bone metastasis in breast cancer (BM-BCa) has just begun at an early stage, and an increasing number of lncRNAs have been proved to play a regulatory role in the process of BM-BCa. Our study focused on the balance of osteogenic-osteoclast regulated by lncRNA-SNHG3 in bone metastasis microenvironment.
    METHODS: SNHG3 level of clinical tissues and breast cancer cell lines was determined by RT-qPCR. ALP staining, ALP activity identification and western blotting of OPG, OSX, RUNX2, BMP2 together with BMP3 was performed to verify the osteogenesis of bone marrow mesenchymal stem cells (BMSCs) both in vitro and in vivo. Exosomes derived from MDA-MB-231 were characterized and sequenced, followed by RT-qPCR. Dual luciferase reporter gene assay was utilized to analyze the binding sites of miR-1273g-3p on SNHG3 and BMP3.
    RESULTS: Expression of BMP3 was positively regulated by SNHG3 via exosomal miR-1273g-3p.
    CONCLUSION: The overexpression of SNHG3 in breast cancer cells may be responsible for osteolytic metastasis Thus, knockdown of SNHG3 might be a potential target for improvement of BM-BCa Treatment.
    DOI:  https://doi.org/10.1016/j.bbrc.2021.12.075
  4. J Proteome Res. 2022 Jan 25.
      Recent studies have shown the promotive effect of resveratrol on wound healing. This study aims to explore the underlying molecular mechanism of resveratrol in type 1 diabetes mellitus (T1DM) through microRNA (miR)-129-containing extracellular vesicles (EVs) derived from mesenchymal stem cells (MSCs) based on in silico analysis. The rat model of T1DM was established by intraperitoneal injection of sodium citrate containing streptozotocin, and the wound was made around the deep fascia. Rat MSCs were isolated and treated with resveratrol (SRT501), and the corresponding EVs (SRT501-EVs) were isolated, where the expression of miR-129 was determined. By performing function experiments, the effect of SRT501-EVs and miR-129 on the biological functions of human umbilical vein endothelial cells (HUVECs) was determined. Finally, the binding relationship between miR-129 and tumor necrosis factor receptor-associated factor 6 (TRAF6) was also determined by the dual-luciferase reporter gene assay. miR-129 was shown as a candidate related to both resveratrol and wound healing in T1DM. SRT501-EVs promoted the skin wound healing of T1DM rats and also further improved the proliferative, migratory, and tube formation potentials of HUVECs. Resveratrol inhibited the expression of TRAF6 in HUVECs stimulated by MSC-conditioned medium and promoted the transfer of miR-129 via EVs, while TRAF6 was confirmed as a target gene of miR-129. Furthermore, inhibition of miR-129 attenuated the proangiogenic effect of resveratrol on HUVECs. Resveratrol exerts promotive role in wound healing in T1DM through downregulation of TRAF6 via MSC-EV-carried miR-129, suggesting a regulatory network involved in the wound healing process in T1DM.
    Keywords:  extracellular vesicles; mesenchymal stem cells; microRNA-129; resveratrol; tumor necrosis factor receptor-associated factor 6; type 1 diabetes mellitus; wound healing
    DOI:  https://doi.org/10.1021/acs.jproteome.1c00248
  5. Bioengineered. 2022 Feb;13(2): 3194-3206
      Mesenchymal stem cells (MSCs)-derived exosomes (Exo) are beneficial in the use of brain damages. Restrictively, the mechanism of Exo expressing miR-124-3p in hypoxic-ischemic brain damage (HIBD) is not completely comprehended. Thereupon, this work was put forward to reveal the action of bone marrow MSCs-derived Exo (BMSCs-Exo) expressing miR-124-3p in the illness. BMSCs were isolated and transfected with miR-124-3p agomir. Then, BMSCs-Exo were extracted and identified. The newborn HIBD rats were injected with miR-124-3p-modified BMSCs-Exo or tumor necrosis factor receptor associated factor 6 (TRAF6)-related vectors. Next, neurological functions, neuron pathological and structural damages, oxidative stress and neuronal apoptosis were observed. miR-124-3p and TRAF6 expression was tested, along with their targeting relationship. miR-124-3p was down-regulated, and TRAF6 was up-regulated in newborn HIBD rats. miR-124-3p targeted TRAF6. BMSCs-Exo improved neurological functions, alleviated neuron pathological and structural damages, suppressed oxidative stress and reduced neuronal apoptosis in newborn HIBD rats, whereas BMSCs-Exo-mediated effects were enhanced by restoring miR-124-3p. Silencing TRAF6 attenuated HIBD in newborn rats, but overexpression of TRAF6 reversed the protective role of miR-124-3p-overexpressing BMSCs-Exo. This work makes it comprehensive that up-regulated exosomal miR-124-3p ameliorates HIBD in newborn rats by targeting TRAF6, which replenishes the potential agents for curing HIBD.
    Keywords:  Hypoxic-ischemic brain damage; MicroRNA-124-3p; bone marrow mesenchymal stem cells; exosomes; tumor necrosis factor receptor associated factor 6
    DOI:  https://doi.org/10.1080/21655979.2021.2016094
  6. Invest Ophthalmol Vis Sci. 2022 Jan 03. 63(1): 41
       Purpose: Increasing evidence suggested that microRNAs (miRs) are implicated in the regulation of the inflammatory response and autophagy in multiple diseases. The present study aimed to explore the effect of miR-223-3p on inflammation and autophagy in fungal keratitis (FK).
    Methods: An FK mouse model was established, and primary corneal stromal cells were isolated by inoculation with Fusarium solani. The expression of miR-223-3p was determined by quantitative RT-PCR. Subsequently, the target gene of miR-223-3p was identified by a dual-luciferase reporter assay. The levels of miR-223-3p were altered by transfecting miR agomir/antagomir to evaluate its effects. Slit-lamp biomicroscopy and hematoxylin and eosin staining were employed to detect corneal damage. The levels of autophagy were assessed by immunofluorescence, Western blotting, mRFP-GFP-LC3 fluorescence microscopy, and electron microscopy. In addition, inflammation was demonstrated by determining the proinflammatory mediators IL-1β and TNF-ɑ.
    Results: Our data suggested that miR-223-3p was increased and that autophagic flux was impaired in mouse FK. Then, we confirmed that autophagy-related gene 16L1 (ATG16L1) was a potential target of miR-223-3p and that this miR negatively regulated the expression of ATG16L1. The inhibition of miR-223-3p attenuated inflammation in FK, reduced P62 expression, and increased the ratio of LC3-II/LC3-I, whereas the overexpression of miR-223-3p displayed the opposite results.
    Conclusions: Taken together, miR-223-3p might regulate autophagy via targeting ATG16L1 in experimental F. solani keratitis and is associated with the inflammatory response. MiR-223-3p might be a potential therapeutic target for FK.
    DOI:  https://doi.org/10.1167/iovs.63.1.41
  7. Cell Prolif. 2022 Jan 27. e13191
       OBJECTIVES: Evidences have suggested that the metabolic function is the key regulator to the fate of MSCs, but its function in senescence of MSC and the underlying mechanism is unclear. Therefore, the purpose of this study was to investigate the metabolic activity of MSCs and its possible mechanism during aging.
    MATERIALS AND METHODS: We used the Seahorse XF24 Analyzer to understand OCR and ECAR in BMSCs and used RT-PCR to analyze the gene expression of mitochondrial biogenesis and key enzymes in glycolysis. We analyzed BMSC mitochondrial activity by MitoTracker Deep Red and JC-1 staining, and detected NAD+/NADH ratio and ATP levels in BMSCs. Microarray and proteomic analyses were performed to detect differentially expressed genes and proteins in BMSCs. The impact of aging on BMSCs through mitochondrial electron transport chain (ETC) was evaluated by Rotenone and Coenzyme Q10.
    RESULTS: Our results demonstrated that the oxidative phosphorylation and glycolytic activity of BMSCs in aged mice were significantly decreased when compared with young mice. BMSCs in aged mice had lower mitochondrial membrane potential, NAD+/NADH ratio, and ATP production than young mice. FABP4 may play a key role in BMSC senescence caused by fatty acid metabolism disorders.
    CONCLUSIONS: Taken together, our results indicated the dysfunction of the metabolic activity of BMSCs in aged mice, which would play the important role in the impaired biological properties. Therefore, the regulation of metabolic activity may be a potential therapeutic target for enhancing the regenerative functions of BMSCs.
    Keywords:  aging; bone marrow mesenchymal stem cell; metabolic activity; metabolism; mitochondrial; osteogenesis
    DOI:  https://doi.org/10.1111/cpr.13191
  8. Neurosci Lett. 2022 Jan 24. pii: S0304-3940(22)00032-5. [Epub ahead of print] 136475
      Hypoxia-ischemia brain damage (HIBD) is a leading cause of neonatal death worldwide, which significantly influences the development of newborns; however, effective treatment strategies remain limited. Recent studies have discovered that microRNAs (miRNAs) play essential roles in the progression of HIBD. Our study was designed to explore whether miR-17-5p was involved in the pathological development of HIBD. In our study, HIBD mouse experimental model was established by carotid artery ligation combined with a hypoxic environment. RT-qPCR and western blot analyses found that Casp2 was high expressed while miR-17-5p was poorly expressed in the cerebral cortical tissue of HIBD mice. Knockdown of Casp2 significantly alleviated brain injury and cell apoptosis. Additionally, the luciferase reporter assay confirmed that miR-17-5p targeted the 3' UTR of Casp2 and negatively regulated Casp2 expression. The rescue experiment demonstrated that miR-17-5p mimic significantly relieved brain tissue damage and improved memory ability in the HIBD mouse model, while these functions of miR-17-5p were blocked by overexpression of Casp2. In summary, our results indicated that miR-17-5p exerted protective effects on HIBD by targeting Casp2.
    Keywords:  Casp2; Hypoxia-ischemia brain damage; microRNA-17-5p
    DOI:  https://doi.org/10.1016/j.neulet.2022.136475
  9. Bone. 2022 Jan 20. pii: S8756-3282(21)00474-9. [Epub ahead of print] 116308
      Fragility fractures are an important hallmark of aging and an increasingly recognized complication of Type 2 diabetes (T2D). T2D individuals have been found to exhibit an increased fracture risk despite elevated bone mineral density (BMD) by dual X-ray absorptiometry (DXA). However, BMD and FRAX-scores tend to underestimate fracture risk in T2D. New, reliable biomarkers are therefore needed. MicroRNAs (miRNAs) are secreted into the circulation from cells of various tissues proportional to local disease severity. Serum miRNA-classifiers were recently found to discriminate T2D women with and without prevalent fragility fractures with high specificity and sensitivity (AUC > 0.90). However, the association of circulating miRNAs with incident fractures in T2D has not been examined yet. In 168 T2D postmenopausal women in the AGES-Reykjavik cohort, miRNAs were extracted from baseline serum and a panel of 10 circulating miRNAs known to be involved in diabetic bone disease and aging was quantified by qPCR and Ct-values extracted. Adjusted Cox proportional hazard models assessed the associations between serum miRNAs and incident fragility fracture. Covariates included age, BMI, aBMD, clinical FRAX, and FRAX with aBMD (FRAXaBMD). Additionally, Receiver operating curve (ROC) analyses were performed. Of the included 168 T2D postmenopausal women who were on average 77.2 ± 5.6 years old, 70 experienced at least one incident fragility fracture during the mean follow-up of 5.8 ± 2.7 years. We found that 3 miRNAs were significantly associated with incident diabetic fragility fracture: while miR-19b-1-5p was associated with significantly lower risk of incident fragility fracture (HR 0.84 (95% CI: 0.71-0.99, p = 0.0323)), miR-203a and miR-31-5p were each significantly associated with a higher risk of incident fragility fracture per unit increase in Ct-value (miR-203a: HR 1.29 (95% CI: 1.12-1.49), p = 0.0004, miR-31-5p HR 1.27 (95% CI: 1.06-1.52), p = 0.009). Hazard ratios of the latter two miRNAs remained significant after adjustments for age, BMI, areal bone mineral density (aBMD), clinical FRAX or FRAXaBMD. When stratified into Ct-quartiles, postmenopausal women in the highest miR-203a and miR-31-5p Ct-quartiles exhibited a 2.4-3.4-fold larger fracture risk than women in the lowest Ct-quartile (0.002 ≤ p ≤ 0.039). Women with both miR-203a and miR-31-5p Ct levels above the median had an increased fracture risk (Unadjusted HR 3.26 (95% CI: 1.57-6.78, p = 0.001)) compared to those with both levels below the median, stable to adjustments. We next built a diabetic fragility signature consisting of the 3 miRNAs that showed the largest associations with incident fracture (miR-203a, miR-31-5p, miR-19b-1-5p). This 3-miRNA signature showed with an AUC of 0.722 comparable diagnostic accuracy in identifying incident fractures to any of the clinical parameters such as aBMD, Clinical FRAX or FRAXaBMD alone. When the 3 miRNAs were combined with aBMD, this combined 4-feature signature performed with an AUC of 0.756 (95% CI 0.680, 0.823) significantly better than aBMD alone (AUC 0.666, 95% CI: (0.585, 0.741)) (p = 0.009). Our data indicate that specific serum microRNAs including senescent miR-31-5p are associated with incident fragility fracture in older diabetic women and can significantly improve fracture risk prediction in diabetics when combined with aBMD measurements of the femoral neck.
    Keywords:  Diabetic bone disease; FRAX; Fracture risk; Incident fragility fracture; Secondary osteoporosis; Serum microRNA; Type 2 diabetes mellitus
    DOI:  https://doi.org/10.1016/j.bone.2021.116308