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Cardiac fibrosis, characterized by the excessive deposition of extracellular matrix proteins, significantly contributes to the morbidity and mortality associated with cardiovascular diseases. This article explores the complex interplay between Runt-related transcription factor 2 (RUNX2), bone morphogenetic protein 2 (BMP2), and Notch signaling pathways in the pathogenesis of cardiac fibrosis. Each of these pathways plays a crucial role in the regulation of cellular functions and interactions that underpin fibrotic processes in the heart. Through a detailed review of current research, we highlight how the crosstalk among RUNX2, BMP2, and Notch not only facilitates our understanding of the fibrotic mechanisms but also points to potential biomolecular targets for intervention. This article delves into the regulatory networks, identifies key molecular mediators, and discusses the implications of these signaling pathways in cardiac structural remodeling. By synthesizing findings from recent studies, we provide insights into the cellular and molecular mechanisms that could guide future research directions, aiming to uncover new therapeutic strategies to manage and treat cardiac fibrosis effectively. [ABSTRACT FROM AUTHOR]

BACKGROUND: There is an internal relationship between hyperhomocysteinemia and vascular calcification. However, the pathogenesis of hyperhomocysteinemia promoting vascular calcification is still unclear. OBJECTIVE: To investigate the role of bone morphogenetic protein-2 in hyperhomocysteinemia-induced vascular calcification. METHODS: Human carotid wax samples were divided into a calcified group (n=29) and a non-calcified group (n=13) according to the presence or absence of calcified plaque. Sixteen ApoE-/- mice were randomly divided into a control group and a hyperhomocysteinemia group, with 8 mice in each group. Bone morphogenetic protein-2 vector was used to transfect rat thoracic artery smooth muscle A7r5 cells, and gradient concentration of homocysteine (50, 100, 200, and 400 μmol/L) was utilized to treat A7r5 cells. Calcification was detected by alizarin red staining and hematoxylin-eosin staining. The interaction of bone morphogenetic protein 2 with Runt-related transcription factor 2 was detected by immunofluorescence, and the expressions of bone morphogenetic protein 2, Runt-related transcription factor 2, and α-smooth muscle actin were detected by immunohistochemistry and western blot assay. RESULTS AND CONCLUSION: (1) Human carotid artery tissue staining revealed that compared with the non-calcification group, inflammatory cells increased and calcification positive rate increased in the calcification group (P < 0.05). Compared with the non-calcification group, the expressions of bone morphogenetic protein-2 and Runt-related transcription factor 2 were up-regulated, and the expression of α-smooth muscle actin was decreased in the calcification group (all P < 0.05). (2) The staining of mouse arterial specimens exhibited that, the positive rate of calcified area in the hyperhomocysteinemia group was significantly higher than that in the control group (P < 0.05); serum homocysteine level in the hyperhomocysteinemia group was significantly higher than that in the control group (P < 0.05). Compared with the control group, the expressions of bone morphogenetic protein-2 and Runt-related transcription factor 2 were up-regulated, and the expression of α-smooth muscle actin was decreased in the hyperhomocysteinemia group (all P < 0.05). (3) A7r5 cell culture analysis demonstrated that with the increase of homocysteine concentration gradient, the degree of calcification, the content of bone morphogenetic protein-2 and Runt-related transcription factor 2 protein in A7r5 cells increased (P < 0.05), and the content of α-smooth muscle actin protein decreased (P < 0.05). (4) The A7r5 cell culture analysis of overexpressed bone morphogenetic protein 2 showed that the calcification degree of the overexpressed bone morphogenetic protein 2 group was increased compared with the corresponding control group, the β-sodium glycerophosphate group, and the homocysteine group. RUNtrelated transcription factor 2 expression up-regulated (P < 0.05) and α-smooth muscle actin expression down-regulated (P < 0.05). (5) The expression of bone morphogenetic protein 2 increased in A7r5 cells cultured with homocysteine in calcified medium, and the expression of Runt-related transcription factor 2 increased with the increase of bone morphogenetic protein 2 expression. (6) The results confirm that bone morphogenetic protein-2 is a key target gene in the regulation of smooth muscle cell phenotypic transformation resulting in vascular calcification by hyperhomocysteinemia. Targeted regulation of bone morphogenetic protein-2 reduces hyperhomocysteinemia-induced vascular calcification. [ABSTRACT FROM AUTHOR]

Abstract Introduction Research has highlighted the role of runt-related transcription factor 2 (Runx2) in the development of osteoarthritis (OA); however, its causal association remains unclear. This study aimed to explore whether Runx2 expression is causally associated with OA and assess its therapeutic potential for OA. Methods Genetic proxy instruments for Runx2 expression were obtained from gene expression quantitative trait locus (eQTLs) study of eQTLGen Consortium (n = 31,684). Aggregated genome-wide association study (GWAS) data for OA (including all OA [177,517 cases and 649,173 controls], knee OA (KOA) [62,497 cases and 333,557 controls], and hip OA (HOA) [36,445 cases and 316,943 controls]) were extracted from the Genetics of Osteoarthritis Consortium. We integrated eQTLs data with OA GWAS data to estimate their causal association and to estimate the potential of Runx2 as a drug target in the treatment of OA using summary data-based Mendelian randomization (SMR) analysis. Furthermore, different OA GWAS data (including all OA [77,052 cases and 378,169 controls], KOA [24,955 cases and 378,169 controls], and HOA [15,704 cases and 378,169 controls]) derived from the GWAS Catalog database were used for replication study. Results SMR analysis showed that high expression levels of Runx2 were associated with an increased risk of all OA [odds ratio (OR) 1.044, 95% confidence interval (CI) 1.023–1.067; P = 5.03 × 10−5], KOA (OR 1.040, 95% CI 1.006–1.075; P = 0.021), and HOA (OR 1.067, 95% CI 1.022–1.113; P = 0.003). This suggests that Runx2 inhibitors may have promising potential for the treatment of OA. Notably, the causal effects of Runx2 with all OA (OR 1.053, 95% CI 1.027–1.079; P = 3.95 × 10−5) and KOA (OR 1.043, 95% CI 1.001–1.087; P = 0.045) were repeated in the replication study, but limited evidence supported the association of Runx2 expression levels with HOA (OR 1.045, 95% CI 0.993–1.101; P = 0.094). Conclusions Our analyses indicate a positive correlation between Runx2 expression and OA risk across all three phenotypes, suggesting the potential of Runx2 inhibitors in the treatment of OA and providing evidence from a genetic perspective.

Introduction: Research has highlighted the role of runt-related transcription factor 2 (Runx2) in the development of osteoarthritis (OA); however, its causal association remains unclear. This study aimed to explore whether Runx2 expression is causally associated with OA and assess its therapeutic potential for OA. Methods: Genetic proxy instruments for Runx2 expression were obtained from gene expression quantitative trait locus (eQTLs) study of eQTLGen Consortium (n = 31,684). Aggregated genome-wide association study (GWAS) data for OA (including all OA [177,517 cases and 649,173 controls], knee OA (KOA) [62,497 cases and 333,557 controls], and hip OA (HOA) [36,445 cases and 316,943 controls]) were extracted from the Genetics of Osteoarthritis Consortium. We integrated eQTLs data with OA GWAS data to estimate their causal association and to estimate the potential of Runx2 as a drug target in the treatment of OA using summary data-based Mendelian randomization (SMR) analysis. Furthermore, different OA GWAS data (including all OA [77,052 cases and 378,169 controls], KOA [24,955 cases and 378,169 controls], and HOA [15,704 cases and 378,169 controls]) derived from the GWAS Catalog database were used for replication study. Results: SMR analysis showed that high expression levels of Runx2 were associated with an increased risk of all OA [odds ratio (OR) 1.044, 95% confidence interval (CI) 1.023–1.067; P = 5.03 × 10−5], KOA (OR 1.040, 95% CI 1.006–1.075; P = 0.021), and HOA (OR 1.067, 95% CI 1.022–1.113; P = 0.003). This suggests that Runx2 inhibitors may have promising potential for the treatment of OA. Notably, the causal effects of Runx2 with all OA (OR 1.053, 95% CI 1.027–1.079; P = 3.95 × 10−5) and KOA (OR 1.043, 95% CI 1.001–1.087; P = 0.045) were repeated in the replication study, but limited evidence supported the association of Runx2 expression levels with HOA (OR 1.045, 95% CI 0.993–1.101; P = 0.094). Conclusions: Our analyses indicate a positive correlation between Runx2 expression and OA risk across all three phenotypes, suggesting the potential of Runx2 inhibitors in the treatment of OA and providing evidence from a genetic perspective. [ABSTRACT FROM AUTHOR]

Although various anti-osteoporosis drugs are available, the limitations of these therapies, including drug resistance and collateral responses, require the development of novel anti-osteoporosis agents. Rhizoma Drynariae displays a promising anti-osteoporosis effect, while the effective component and mechanism remain unclear. Here, we revealed the therapeutic potential of Rhizoma Drynariae-derived nanovesicles (RDNVs) for postmenopausal osteoporosis and demonstrated that RDNVs potentiated osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) by targeting estrogen receptor-alpha (ER α). RDNVs, a natural product isolated from fresh Rhizoma Drynariae root juice by differential ultracentrifugation, exhibited potent bone tissue-targeting activity and anti-osteoporosis efficacy in an ovariectomized mouse model. RDNVs, effectively internalized by hBMSCs, enhanced proliferation and ER α expression levels of hBMSC, and promoted osteogenic differentiation and bone formation. Mechanistically, via the ER α signaling pathway, RDNVs facilitated mRNA and protein expression of bone morphogenetic protein 2 and runt-related transcription factor 2 in hBMSCs, which are involved in regulating osteogenic differentiation. Further analysis revealed that naringin, existing in RDNVs, was the active component targeting ER α in the osteogenic effect. Taken together, our study identified that naringin in RDNVs displays exciting bone tissue-targeting activity to reverse osteoporosis by promoting hBMSCs proliferation and osteogenic differentiation through estrogen-like effects. The natural product Rhizoma Drynariae-derived nanovesicles isolated and purified from fresh Rhizoma Drynariae, have a high efficacy agonist of estrogen receptor α and a potential nano-drug platform for the prevention and treatment of postmenopausal osteoporosis. [Display omitted] [ABSTRACT FROM AUTHOR]

Cardiac fibrosis, characterized by the excessive deposition of extracellular matrix proteins, significantly contributes to the morbidity and mortality associated with cardiovascular diseases. This article explores the complex interplay between Runt-related transcription factor 2 (RUNX2), bone morphogenetic protein 2 (BMP2), and Notch signaling pathways in the pathogenesis of cardiac fibrosis. Each of these pathways plays a crucial role in the regulation of cellular functions and interactions that underpin fibrotic processes in the heart. Through a detailed review of current research, we highlight how the crosstalk among RUNX2, BMP2, and Notch not only facilitates our understanding of the fibrotic mechanisms but also points to potential biomolecular targets for intervention. This article delves into the regulatory networks, identifies key molecular mediators, and discusses the implications of these signaling pathways in cardiac structural remodeling. By synthesizing findings from recent studies, we provide insights into the cellular and molecular mechanisms that could guide future research directions, aiming to uncover new therapeutic strategies to manage and treat cardiac fibrosis effectively.

Abstract Background Bone defects remain a challenge today. In addition to osteogenic activation, the crucial role of angiogenesis has also gained attention. In particular, vascular endothelial growth factor (VEGF) is likely to play a significant role in bone regeneration, not only to restore blood supply but also to be directly involved in the osteogenic differentiation of mesenchymal stem cells. In this study, to produce additive angiogenic-osteogenic effects in the process of bone regeneration, VEGF and Runt-related transcription factor 2 (Runx2), an essential transcription factor for osteogenic differentiation, were coadministered with messenger RNAs (mRNAs) to bone defects in the rat mandible. Methods The mRNAs encoding VEGF or Runx2 were prepared via in vitro transcription (IVT). Osteogenic differentiation after mRNA transfection was evaluated using primary osteoblast-like cells, followed by an evaluation of the gene expression levels of osteogenic markers. The mRNAs were then administered to a bone defect prepared in the rat mandible using our original cationic polymer-based carrier, the polyplex nanomicelle. The bone regeneration was evaluated by micro-computerized tomography (μCT) imaging, and histologic analyses. Results Osteogenic markers such as osteocalcin (Ocn) and osteopontin (Opn) were significantly upregulated after mRNA transfection. VEGF mRNA was revealed to have a distinct osteoblastic function similar to that of Runx2 mRNA, and the combined use of the two mRNAs resulted in further upregulation of the markers. After in vivo administration into the bone defect, the two mRNAs induced significant enhancement of bone regeneration with increased bone mineralization. Histological analyses using antibodies against the Cluster of Differentiation 31 protein (CD31), alkaline phosphatase (ALP), or OCN revealed that the mRNAs induced the upregulation of osteogenic markers in the defect, together with increased vessel formation, leading to rapid bone formation. Conclusions These results demonstrate the feasibility of using mRNA medicines to introduce various therapeutic factors, including transcription factors, into target sites. This study provides valuable information for the development of mRNA therapeutics for tissue engineering.

We have previously demonstrated that auxiliary metformin therapy promotes healing of apical periodontitis. Here we aimed to investigate the effects of metformin on osteoblast differentiation and osteoclast formation in cultured cells and rat apical periodontitis. Murine pre-osteoblasts MC3T3-E1 and macrophages RAW264.7 were cultured under hypoxia (2% oxygen) or normoxia (21% oxygen) and stimulated with receptor activator of nuclear factor-κB ligand (RANKL) when indicated. Metformin was added to the cultures to evaluate its anti-hypoxic effects. Expressions of osteoblast differentiation regulator runt-related transcription factor 2 (RUNX2), RANKL, and osteoclast marker tartrate-resistant acid phosphatase (TRAP) were assessed by Western blot. Apical periodontitis was induced in mandibular first molars of 10 Sprague-Dawley rats. Root canal therapy with or without metformin supplement was performed. Periapical bone resorption was measured by micro-computed tomography. Immunohistochemistry was used to examine RUNX2, RANKL, and TRAP expressions. Hypoxia suppressed RUNX2 expression and enhanced RANKL synthesis in pre-osteoblasts. TRAP production increased in macrophages after hypoxia and/or RANKL stimulation. Metformin reversed hypoxia-induced RUNX2 suppression and RANKL synthesis in pre-osteoblasts. Metformin also inhibited hypoxia and RANKL-enhanced TRAP synthesis in macrophages. Intracanal metformin diminished bone loss in rat apical periodontitis. Comparing with vehicle control, cells lining bone surfaces in metformin-treated lesions had significantly stronger expression of RUNX2 and decreased synthesis of RANKL and TRAP. Alleviation of bone resorption by intracanal metformin was associated with enhanced osteoblast differentiation and diminished osteoclast formation in rat apical periodontitis. Our results endorsed the role of metformin as an effective medicament for inflammatory bone diseases. [ABSTRACT FROM AUTHOR]

Abstract Background Previous studies have demonstrated that Runt‐associated transcription factor 2 (RUNX2) serves as the main transcription factor for osteoblast differentiation and chondrocyte maturation. RUNX2 is related to a variety of tumors, particularly tumor invasion and metastasis, while the expression and molecular mechanisms of RUNX2 in clear cell renal cell carcinoma (ccRCC) keep to be determined. Stearyl CoA desaturase 1 (SCD1), an endoplasmic reticulum fatty acid desaturase, transfers saturated fatty acids to monounsaturated fatty acids, is expressed highly in numerous malignancies. Methods The Cancer Genome Atlas (TCGA) datebase and Western blot was used to analyzed the mRNA and protein levels of the target gene in ccRCC tissues and adjacent tissues. The proliferation ability of ccRCC cells was tested by colony forming and EdU assay. The migration ability of cells was detected by transwell assay. Immunoprecipitation was utilized to detect protein–protein interaction. Cycloheximide chase assay was used to measure the half‐life of SCD1 protein. Results In this study, the expressions of RUNX2 and SCD1 are increased in ccRCC tissues as well as ccRCC cell lines. Both RUNX2 and SCD1 could promote proliferation and migration in ccRCC cells. Furthermore, RUNX2 could physically interact with SCD1. In addition, the functional degradation and the inactivation of Wnt/β‐catenin signaling pathway triggered by the downregulation of RUNX2 could be partly offset by the overexpression of SCD1. Conclusion The findings indicate that the RUNX2/SCD1 axis may act as a potential therapeutic target via the Wnt/β‐catenin signaling pathway of ccRCC.

Long-term hypercaloric intake such as a high-fat diet (HFD) could act as negative regulators on bone remodeling, thereby inducing bone loss and bone microarchitecture destruction. Currently, food-derived natural compounds represent a promising strategy to attenuate HFD-induced bone loss. We previously prepared a whey protein hydrolysate (WPH) with osteogenic capacity. In this study, we continuously isolated and identified an osteogenic and antioxidant octapeptide TPEVDDA from WPH, which significantly promoted the alkaline phosphatase activities on MC3T3-E1 cells and exerted DPPH radical scavenging capacity. We then established an HFD-fed obese mice model with significantly imbalanced redox status and reduced bone mass and further evaluated the effects of different doses of WPH on ameliorating the HFD-induced bone loss and oxidative damages. Results showed that the administration of 2% and 4% WPH for 12 weeks significantly restored perirenal fat mass, improved serum lipid levels, reduced oxidative stress, and promoted the activity of antioxidant enzymes; meanwhile, WPH significantly preserved bone mass and bone mechanical properties, attenuated the degradation of trabecular microstructure, and regulated serum bone metabolism biomarkers. The protein levels of Runx2, Nrf2, and HO-1, as well as the phosphorylation level of GSK-3β in tibias, were notably activated by WPH. Overall, we found that the potential mechanism of WPH on ameliorating the HFD-induced bone loss mainly through its antioxidant and osteogenic capacity by activating Runx2 and GSK-3β/Nrf2 signaling pathway, demonstrating the potential of WPH to be used as a nutritional strategy for obesity and osteoporosis. [ABSTRACT FROM AUTHOR]

Objective To evaluate the effect of D-site binding protein （DBP） inhibition on vascular calcification in chronic uremic rats. Methods GSE146638 was downloaded from the public database （Gene Expression Omnibus， GEO）， and a total of 10 samples from rat of mRNA expression profiles were included in this study. DESeq2 was used to analyze the differentially expressed genes in the aortic smooth muscle between the control and chronic uremic rat model groups. RT-qPCR was employed to detect the expression levels of SLC22A2， ATF3， DBP and SMPD3. Male SD rats were randomly divided into four groups. In the blank group， no treatment was given. In the model group， chronic uremic rat models were established by nephrectomy of 2/3 of bilateral kidneys， and the surviving rats were fed with 8-week high phosphorus diet to induce vascular calcification. In the negative control lentivirus group （shNC group）， the rats were injected with negative control lentivirus for 4 weeks after establishing the vascular calcification uremic rat models. In the small interfering DBP group （shDBP group）， the rats were treated with siDBP lentivirus for 4 weeks after the establishment of vascular calcification uremic rat models. After corresponding interventions， the rat weight was measured， and serum creatinine， serum phosphorus， serum calcium and urea nitrogen levels were detected among four groups. Immunohistochemical staining and HE staining of the thoracic aorta tissues were used to evaluate the degree of injury. RT-qPCR and western blot were adopted to quantitatively measure the relative expression levels of Runt-related transcription factor 2 （RUNX2） and DBP. Results Compared with the control group， the rat weight was significantly less， whereas the serum creatinine levels were significantly higher in the model， shNC and shDBP groups （all P < 0.05）. In the model group， the thoracic aortic vascular wall was thickened， and the expression levels of RUNX2 and proliferating cell nuclear antigen （PCNA） were up-regulated. Compared with the model group， the serum creatinine level was declined， the expression levels of RUNX2 mRNA and protein in the thoracic aorta were down-regulated （all P < 0.05）， the expression level of PCNA in the thoracic aorta was down-regulated and the thoracic aortic vascular wall thickening was mitigated. Conclusion Inhibition of DBP may suppress vascular calcification and alleviate uremia-associated symptoms in chronic uremic rat models by down-regulating the expression levels of RUNX2 mRNA and protein in the aorta.

Abstract Background The pathogenesis of traumatic temporomandibular joint (TMJ) bony ankylosis remains unknown. This study aimed to explore the pathogenesis of traumatic TMJ bony ankylosis in a rat model. Methods Twenty-four 3-week-old male Sprague–Dawley rats were used in this study. Excision of the whole disc, the fibrocartilage damage of the condyle and glenoid fossa, and narrowed joint space were performed in the left TMJ of the operation group to induce TMJ bony ankylosis (experimental side). The right TMJ underwent a sham operation (sham side). The control group did not undergo any operations. At 1, 4, and 8 weeks postoperatively, rats of the operation group were sacrificed and TMJ complexes were evaluated by gross observation, Micro-CT, histological examinations, and immunofluorescence microscopy. Total RNA of TMJ complexes in the operation group were analyzed using RNA-seq. Results Gross observations revealed TMJ bony ankylosis on the experimental side. Micro-CT analysis demonstrated that compared to the sham side, the experimental side showed a larger volume of growth, and a considerable calcified bone callus formation in the narrowed joint space and on the rougher articular surfaces. Histological examinations indicated that endochondral ossification was observed on the experimental side, but not on the sham side. RNA-seq analysis and immunofluorescence revealed that Matrix metallopeptidase 13 (MMP13) and Runt-related transcription factor 2 (RUNX2) genes of endochondral ossification were significantly more downregulated on the experimental side than on the sham side. The primary pathways related to endochondral ossification were Parathyroid hormone synthesis, secretion and action, Relaxin signaling pathway, and IL-17 signaling pathway. Conclusions The present study provided an innovative and reliable rat model of TMJ bony ankylosis by compound trauma and narrowed joint space. Furthermore, we demonstrated the downregulation of MMP13 and RUNX2 in the process of endochondral ossification in TMJ bony ankylosis.

Successful bone regeneration on titanium (Ti) surfaces is a key process in dental implant treatment. Bone marrow mesenchymal stem cells (BMSCs) are fundamental cellular components of this process, and their early recruitment, proliferation, and differentiation into bone-forming osteoblasts are crucial. A proteoglycan (PG)-rich layer has been reported to exist between Ti surfaces and bones; however, the molecules that could potentially affect the formation of this layer remain unknown. Family with sequence similarity 20 member B (FAM20B) is a newly identified kinase that regulates the synthesis of glycosaminoglycans, an important component of the PG-rich layer. Because FAM20B is also closely associated with bone development, in this study, we examined the function of FAM20B in osteogenic differentiation of BMSCs on Ti surfaces. For this, BMSC cell lines with knocked down FAM20B (shBMSCs) were cultured on Ti surfaces. The results showed that the depletion of FAM20B reduced the formation of a PG-rich layer between the Ti surfaces and cells. The shBMSCs exhibited downregulated expression of osteogenic marker genes (ALP and OCN) and decreased mineral deposition. Moreover, shBMSCs reduced the molecular levels of p-ERK1/2, which plays an important role in MSC osteogenesis. The nuclear translocation of RUNX2, an important transcription factor for osteogenic differentiation, on the Ti surfaces is inhibited by the depletion of FAM20B in BMSCs. Moreover, the depletion of FAM20B reduced the transcriptional activity of RUNX2, which is important in regulating the expression of osteogenic genes. Bone healing and regeneration on implanted titanium surfaces is a cell–material interaction. Such an interaction is enabled by bone marrow mesenchymal stem cells (BMSCs), and their early recruitment, proliferation, and differentiation into bone-forming osteoblasts are essential for bone healing and osseointegration. In this study, we found that the family with sequence similarity 20-B influenced the formation of a proteoglycan rich layer between BMSCs and the titanium surface and regulated the differentiation of BMSCs into bone-forming osteoblasts. We believe that our study contributes significantly to the further exploration of bone healing and osseointegration mechanisms on implanted titanium surfaces. [Display omitted] [ABSTRACT FROM AUTHOR]

Background: Previous studies have demonstrated that Runt‐associated transcription factor 2 (RUNX2) serves as the main transcription factor for osteoblast differentiation and chondrocyte maturation. RUNX2 is related to a variety of tumors, particularly tumor invasion and metastasis, while the expression and molecular mechanisms of RUNX2 in clear cell renal cell carcinoma (ccRCC) keep to be determined. Stearyl CoA desaturase 1 (SCD1), an endoplasmic reticulum fatty acid desaturase, transfers saturated fatty acids to monounsaturated fatty acids, is expressed highly in numerous malignancies. Methods: The Cancer Genome Atlas (TCGA) datebase and Western blot was used to analyzed the mRNA and protein levels of the target gene in ccRCC tissues and adjacent tissues. The proliferation ability of ccRCC cells was tested by colony forming and EdU assay. The migration ability of cells was detected by transwell assay. Immunoprecipitation was utilized to detect protein–protein interaction. Cycloheximide chase assay was used to measure the half‐life of SCD1 protein. Results: In this study, the expressions of RUNX2 and SCD1 are increased in ccRCC tissues as well as ccRCC cell lines. Both RUNX2 and SCD1 could promote proliferation and migration in ccRCC cells. Furthermore, RUNX2 could physically interact with SCD1. In addition, the functional degradation and the inactivation of Wnt/β‐catenin signaling pathway triggered by the downregulation of RUNX2 could be partly offset by the overexpression of SCD1. Conclusion: The findings indicate that the RUNX2/SCD1 axis may act as a potential therapeutic target via the Wnt/β‐catenin signaling pathway of ccRCC. [ABSTRACT FROM AUTHOR]

Objective To investigate the effects of over expression and low expression of antisense transcripts of circular RNA cerebellar degeneration associated protein 1 (CDR1as) in Balb/C mouse bone marrow mesenchymal stem cells (BMSCs) on factors related to osteogenesis and angiogenesis. Methods BMSCs were cultured and identified in vitro. The lentiviral (LV) vector containing the overexpressed and silenced circRNA CDR1as genes and the control lentivirus were respectively transfected into mouse BMSCs, and stable cell lines were screened. The cells were divided into the circRNACDR1as over expression group and the over expression control group, and the CircRNACDR1as low expression group and the low expression control group. The components were stained with Alizarin Red S and alkaline phosphatase after 14 and 21 days of osteoinduction; qRT-PCR was used to detect the target genes circRNA CDR1as, osteogenic differentiation markers alkaline phosphatase (ALP), runt- related transcription factor 2 (RUNX2), osteocalcin (OCN), osteopontin (OPN), osterix(Osx), collagen I (COL-1), and the mRNA expression levels of vascular endothelial grown factor (VEGF) and angiogenin-1 (Ang-1). Results The results of alizarin red staining and alkaline phosphatase staining showed that the extracellular matrix calcium precipitation and ALP staining area of the over expression experimental group was greater than its control group, and those of the low expression experimental group was less than its control group. As the number of days of osteogenic induction increased, the calcium precipitation and ALP staining in each group also increased. RT-PCR results showed that the mRNA expression levels of circRNA CDR1as, ALP, RUNX2, OCN, OPN, OSX, COL-1, VEGF and Ang-1 in the over expression experimental group BMSCs were significantly increased (P

BACKGROUND: Exercise training can maintain bone mass and improve bone mineral density. However, its effect on senile osteoporosis and its possible mechanism are not clear. OBJECTIVE: To study the effects of exercise training on senile osteoporosis and to explore its possible molecular mechanism. METHODS: Sixteen Sprague-Dawley rats aged 24 months old were regarded as the model of senile osteoporosis and randomly divided into elderly model group (n=8) and exercise training group (n=8), while eight young Sprague-Dawley rats of 3 months old were randomly selected as young control group. All rats moved freely in their cages and those in the exercise training group underwent exercise training every day for 8 weeks. After 8 weeks of intervention, serum bone-specific alkaline phosphatase, type I procollagen amino-terminal peptide, tartrate-resistant acid phosphatase 5b and type I collagen aminoterminal peptide were detected using ELISA assay. Bone mineral density of the femur and the fourth lumbar vertebra were measured by dual energy X-ray absorptiometry, while bone microstructural parameters of the tibia and the fifth lumbar vertebra were determined by Micro-CT in each group. mRNA and protein expressions of bone morphogenetic protein 2, Runt-related transcription factor 2, Osterix, and peroxisome proliferator-activated receptor γ in bone marrow of the femur were detected by reverse transcriptase polymerase chain reaction and western blot. RESULTS AND CONCLUSION: Compared with the elderly model group, the serum levels of bone-specific alkaline phosphatase and type I procollagen aminoterminal peptide (P < 0.05), bone mineral density of the femur (P < 0.05) but not that of the fourth lumbar vertebra (P > 0.05), bone volume fraction and trabecular number of the tibia (P < 0.05), and the mRNA and protein expression of bone morphogenetic protein 2, Runt-related transcription factor 2, and Osterix in the femur (P < 0.05) were significantly increased in the elderly model group, while the serum levels of tartrate-resistant acid phosphatase 5b and type I collagen amino-terminal peptide (P < 0.05), trabecular separation of the tibia and the fifth lumbar vertebra (P < 0.05), and the mRNA and protein expression of peroxisome proliferator-activated receptor γ were significantly decreased (P < 0.05). In addition, exercise training could improve the bone volume fraction, trabecular number and trabecular thickness of the fifth lumbar vertebra in rats with senile osteoporosis, although the differences in changes were not always statistically significant (P > 0.05). The trabecular thickness of the tibia had no significant changes after exercise training (P > 0.05). To conclude, exercise training can significantly improve bone mineral density and bone microstructure, promote osteogenic differentiation, and inhibit adipogenic differentiation in aged osteoporotic rats. The mechanism may be related to the bone morphogenetic protein 2/Runt-related transcription factor 2/Osterix signaling pathway. [ABSTRACT FROM AUTHOR]

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Background: The pathogenesis of traumatic temporomandibular joint (TMJ) bony ankylosis remains unknown. This study aimed to explore the pathogenesis of traumatic TMJ bony ankylosis in a rat model. Methods: Twenty-four 3-week-old male Sprague–Dawley rats were used in this study. Excision of the whole disc, the fibrocartilage damage of the condyle and glenoid fossa, and narrowed joint space were performed in the left TMJ of the operation group to induce TMJ bony ankylosis (experimental side). The right TMJ underwent a sham operation (sham side). The control group did not undergo any operations. At 1, 4, and 8 weeks postoperatively, rats of the operation group were sacrificed and TMJ complexes were evaluated by gross observation, Micro-CT, histological examinations, and immunofluorescence microscopy. Total RNA of TMJ complexes in the operation group were analyzed using RNA-seq. Results: Gross observations revealed TMJ bony ankylosis on the experimental side. Micro-CT analysis demonstrated that compared to the sham side, the experimental side showed a larger volume of growth, and a considerable calcified bone callus formation in the narrowed joint space and on the rougher articular surfaces. Histological examinations indicated that endochondral ossification was observed on the experimental side, but not on the sham side. RNA-seq analysis and immunofluorescence revealed that Matrix metallopeptidase 13 (MMP13) and Runt-related transcription factor 2 (RUNX2) genes of endochondral ossification were significantly more downregulated on the experimental side than on the sham side. The primary pathways related to endochondral ossification were Parathyroid hormone synthesis, secretion and action, Relaxin signaling pathway, and IL-17 signaling pathway. Conclusions: The present study provided an innovative and reliable rat model of TMJ bony ankylosis by compound trauma and narrowed joint space. Furthermore, we demonstrated the downregulation of MMP13 and RUNX2 in the process of endochondral ossification in TMJ bony ankylosis. [ABSTRACT FROM AUTHOR]

Objective To evaluate the effect of D-site binding protein（ DBP） inhibition on vascular calcification in chronic uremic rats. Methods GSE146638 was downloaded from the public database （Gene Expression Omnibus， GEO）， and a total of 10 samples from rat of mRNA expression profiles were included in this study. DESeq2 was used to analyze the differentially expressed genes in the aortic smooth muscle between the control and chronic uremic rat model groups. RT-qPCR was employed to detect the expression levels of SLC22A2， ATF3， DBP and SMPD3. Male SD rats were randomly divided into four groups. In the blank group， no treatment was given. In the model group， chronic uremic rat models were established by nephrectomy of 2/3 of bilateral kidneys， and the surviving rats were fed with 8-week high phosphorus diet to induce vascular calcification. In the negative control lentivirus group （shNC group）， the rats were injected with negative control lentivirus for 4 weeks after establishing the vascular calcification uremic rat models. In the small interfering DBP group（ shDBP group）， the rats were treated with siDBP lentivirus for 4 weeks after the establishment of vascular calcification uremic rat models. After corresponding interventions， the rat weight was measured， and serum creatinine， serum phosphorus， serum calcium and urea nitrogen levels were detected among four groups. Immunohistochemical staining and HE staining of the thoracic aorta tissues were used to evaluate the degree of injury. RT-qPCR and western blot were adopted to quantitatively measure the relative expression levels of Runt-related transcription factor 2 （RUNX2） and DBP. Results Compared with the control group， the rat weight was significantly less， whereas the serum creatinine levels were significantly higher in the model， shNC and shDBP groups（ all P < 0.05）. In the model group， the thoracic aortic vascular wall was thickened， and the expression levels of RUNX2 and proliferating cell nuclear antigen （PCNA） were up-regulated. Compared with the model group， the serum creatinine level was declined， the expression levels of RUNX2 mRNA and protein in the thoracic aorta were down-regulated （all P < 0.05）， the expression level of PCNA in the thoracic aorta was down-regulated and the thoracic aortic vascular wall thickening was mitigated. Conclusion Inhibition of DBP may suppress vascular calcification and alleviate uremia-associated symptoms in chronic uremic rat models by down-regulating the expression levels of RUNX2 mRNA and protein in the aorta. [ABSTRACT FROM AUTHOR]

Objective To investigate the effect of silencing histone deacetylase 9 (HDAC9) expression on the proliferation and osteogenic differentiation of periodontal ligament stem cells (PDLSCs). Methods PDLSCs were isolated, cultured and identified in vitro. An siRNA construct specific for HDAC9 was transfected into PDLSCs (siHDAC9 group), and a nontargeting siRNA was used as a control (siNC group). The interference effect was determined by qRT-PCR. The cell cycle progression of PDLSCs was detected using flow cytometry. The proliferation activity of PDLSCs was detected via CCK-8 assay. Western blotting was used to detect the protein expression of proliferating cell nuclear antigen (PCNA). The mRNA expression of runt-related transcription factor 2 (RUNX2) and alkaline phosphatase (ALP) was investigated by qRT-PCR. The protein expression of RUNX2 was detected by western blotting. In addition, the formation of mineralized nodules was assessed by alizarin red staining. Results Compared with that in the siNC group, the mRNA expression of HDAC9 in the siHDAC9 group was lower (P < 0.01). Moreover, compared with those in the siNC group, the proliferation index (P

The modification of N6‐methyladenosine is involved in the progression of various cancers. This study aimed to clarify its regulatory mechanism in the pathogenesis of choroidal melanoma. Expression of methyltransferase‐like 14 in choroidal melanoma or normal choroidal tissues was determined by Western blot and immunohistochemistry. The impacts of methyltransferase‐like 14 on invasion and migration of choroidal melanoma cells were determined using functional and animal experiments. The interaction between methyltransferase‐like 14 and its downstream target was identified by methylated RNA immunoprecipitation and a dual‐luciferase reporter assay. Additionally, Wnt/β‐catenin signalling pathway was evaluated by Western blot. Methyltransferase‐like 14 was upregulated in choroidal melanoma compared to the normal choroidal tissues. Overexpression or knockdown of methyltransferase‐like 14 enhanced or inhibited the invasion and migration of choroidal melanoma cells, respectively, both in vivo and in vitro. Methyltransferase‐like 14 directly targeted downstream runt‐related transcription factor 2 mRNA, depending on N6‐methyladenosine. Additionally, the Wnt/β‐catenin signalling pathway was activated by methyltransferase‐like 14 in choroidal melanoma cells. Our study identified a novel RNA regulatory mechanism in which runt‐related transcription factor 2 was upregulated by enhanced expression of methyltransferase‐like 14 via N6‐methyladenosine modification, thus facilitating migration and invasion of choroidal melanoma cells. [ABSTRACT FROM AUTHOR]

A phenylalanine (Phe)-restricted diet is indispensable for individuals suffering from phenylketonuria (PKU). Our previous study reported a low-Phe-containing whey protein hydrolysate (LPH) prepared from a selected whey protein hydrolysate (TA2H). This study aimed to investigate the osteogenic activity of LPH and TA2H in MC3T3-E1 preosteoblast cells and explore the underlying mechanism. Results showed that the treatment of TA2H and LPH (at the final concentrations of 100–1000 μg/mL) had a stimulatory effect on the proliferation, differentiation, and mineralization of MC3T3-E1 cells. The LPH of 1000 μg/mL significantly increased cell proliferation (2.15- ± 0.11-fold) and alkaline phosphatase activity (1.22- ± 0.07-fold), promoted the protein and mRNA levels of runt-related transcription factor 2 (Runx2, 2.50- ± 0.14-fold and 2.97- ± 0.23-fold, respectively), enhanced the expression of differentiation biomarkers (type-I collagen, osteocalcin, and osteopontin), increased calcium deposition (1.56- ± 0.08-fold), and upregulated the ratio of osteoprotegerin/receptor activator of nuclear factor-κB ligand. The exploration of signaling pathways indicated that the activated p38-dependent Runx2 signaling contributed to the LPH-induced osteogenesis. These results provided evidence, for the first time, that a prepared low-Phe whey protein hydrolysate positively modulated the activity of osteoblasts through the p38/Runx2 pathway, thereby providing a new osteoinductive protein substitute to make functional PKU food. [ABSTRACT FROM AUTHOR]

Dieda ointment, a Chinese medicine, has been widely used in the clinical treatment of fracture after years of clinical research to prove that it has significant effects on healing injury and relieving pain, reinforcing tendons and setting bones. However, the mechanism of its effects on fracture healing is still unclear. The purpose of this study was to explore the mechanism of Dieda ointment in promoting fracture healing. The materials and methods explored were used to establish a fracture model of Sprague-Dawley rats, which was externally applied with Dieda ointment for 1, 2 and 4 w and imaging examination of the fracture site was performed. Histological examination was also performed. Western blotting and quantitative reverse transcription polymerase chain reaction were used to detect the expression of proteins and its messenger ribonucleic acid. We found that Dieda ointment can effectively accelerate callus formation at tibia fractures in rats and increase the messenger ribonucleic acid and protein expression levels of bone morphogenetic protein-2, suppressor of mothers against decapentaplegic 1, runt-related transcription factor 2 and Sp7 transcription factor, regulate bone formation and promote fracture healing. The study on the mechanism of Dieda ointment promoting fracture healing, not only provides theoretical and experimental basis for the subsequent study of Dieda ointment. [ABSTRACT FROM AUTHOR]

Calcific aortic valve disease (CAVD) is linked to high mortality. Melatonin inhibits nuclear factor-kappa B (NF-κB)/cyclic AMP response element-binding protein (CREB), contributing to CAVD progression. This study determined the role of melatonin/MT1/MT2 signaling in valvular interstitial cell (VIC) calcification. Western blotting and Alizarin red staining were used to analyze NF-κB/CREB/runt-related transcription factor 2 (Runx2) signaling in porcine VICs treated with an osteogenic (OST) medium without (control) or with melatonin for 5 days. Chromatin immunoprecipitation (ChIP) assay was used to analyze NF-κB's transcription regulation of NF-κB on the Runx2 promoter. OST medium-treated VICs exhibited a greater expression of NF-κB, CREB, and Runx2 than control VICs. Melatonin treatment downregulated the effects of the OST medium and reduced VIC calcification. The MT1/MT2 antagonist (Luzindole) and MT1 receptor neutralized antibody blocked the anticalcification effect of melatonin, but an MT2-specific inhibitor (4-P-PDOT) did not. Besides, the NF-κB inhibitor (SC75741) reduced OST medium-induced VIC calcification to a similar extent to melatonin at 10 nmol/L. The ChIP assay demonstrated that melatonin attenuated OST media increased NF-κB binding activity to the promoter region of Runx2. Activation of the melatonin/MT1-axis significantly reduced VIC calcification by targeting the NF-κB/CREB/Runx2 pathway. Targeting melatonin/MT1 signaling may be a potential therapeutic strategy for CAVD.

Objective To investigate the serum expression levels of vitamin K2 (VK2), runt related transcription factor 2 (Runx2) and their relationship with cardiovascular calcification in patients with stage Ⅲ-Ⅴ non dialysis chronic kidney disease (CKD). Methods A total of 149 patients with stage Ⅲ - Ⅴ CKD who did not receive dialysis were selected. According to the clinical stage, patients were divided into the stage Ⅲ group (58 cases), the stage Ⅳ group (52 cases) and the stage Ⅴ group (39 cases). In addition, according to the cardiovascular calcification, the patients were divided into the calcification group (42 cases) and the non calcification group (107 cases). The general data of all patients were collected. Serum levels of phosphorus, calcium, creatinine (Cr) and cystatin C (Cys C) were detected by automatic biochemical analyzer. The serum expressions of VK2 and Runx2 were detected by enzyme-linked immunosorbent assay kit. Results The levels of phosphorus, Cr, Cys C, Runx2 and cardiovascular calcification rate were increased gradually in the stage Ⅲ group, the stage Ⅳ group and the stage Ⅴ group, and levels of calcium and VK2 were decreased gradually, and the difference was statistically significant between groups (P＜0.05). The levels of phosphorus, Cys C and Runx2 were higher in the calcified group than those in the non calcified group, and the level of VK2 was lower than that in the non calcified group (P＜0.05). Multivariate analysis showed that the high expression levels of phosphorus and Runx2 were the risk factors of cardiovascular calcification in patients with CKD, while the high expression of VK2 was the protective factor of cardiovascular calcification in patients with CKD (P＜0.05). ROC analysis showed that serum levels of VK2, Runx2 and phosphorus had high evaluation value for patients with CKD complicated with cardiovascular calcification, and the areas under the curve were 0.843 (95%CI: 0.773-0.913), 0.819 (95%CI: 0.749-0.889) and 0.797 (95%CI: 0.726-0.868) respectively. Conclusion The serum level of VK2 is decreased with the increase of stage, and the level of Runx2 is increased with the increase of stage in patients with stage Ⅲ-Ⅴ non dialysis CKD. Both VK2 and Runx2 are closely related to the situation of cardiovascular calcification. [ABSTRACT FROM AUTHOR]

To study the effects of Zuoguiwan ( ZGW) on nourishing kidney essence, strengthening bone and marrow, and promotion of the proliferation of bone marrow mesenchymal stem cells ( BMSCs) and differentiation into osteoblasts. Methods The ZGW containing serum was prepared. 10% PBS DMEM group, osteogenic liquid group, 2. 5% serum of health rats with osteogenic liquid group, and 2. 5% serum containing ZGW osteogenic liquid group were set up. MTT method was used to observe the proliferation of BMSCs in each group. The differentiation of BMSCs into osteoblasts was tested using alkaline phosphatase calcium cobalt staining method . Alkaline phosphatase ( AKP) method was used to detect the content of AKP in cell culture medium. The expression of runt related transcription factor 2 ( Runx2 ), osteocalcin ( OPN), and alkaline phosphatase (ALP) protein in BMSCs were detected using Western blotting. Results Rapid proliferation of BMSCs was showed in 2. 5% serum containing ZGW osteogenic liquid group. The plateau stage appeared earlier. It promoted the secretion of AKP in BMSCs, increased the content of ALP, and up-regulated the protein expression levels of Runx2, ALP, and OPN in BMSCs cells. Conclusion The molecular mechanism of ZGW against osteoporosis may be related to the regulation of Runx2, activation of ALP and OPN protein expression, promotion of BMSCs proliferation and differentiation into osteoblasts. [ABSTRACT FROM AUTHOR]

Objective To explore the effect of calcitriol capsule (Cal) on tibial fracture healing in rats through bone morphogenetic protein-2 (BMP-2)/SMAD/Runt-related transcription factor 2 (Runx2) signaling pathway. Methods Fifty-two SD rats were randomly divided into control group (Cont), tibial fracture group, Cal group (gavage 10 ng/mL Cal for 4 consecutive weeks), and Cal + Noggin (BMP specific inhibitor) group (gavage 10 ng/mL Cal+percutaneous injection of 50 ng/mL Noggin for 4 consecutive weeks). X-ray, automatic biochemical analyzer, micro CT, HE staining and Western blotting were used to analyze the treatment effect and possible mechanism. Results After 4 weeks of the fractures, the fracture line in the tibial fracture group was obvious. The fracture line in the Cal group was almost invisible and there was a lot of callus at the broken end. The fracture healing in the Cal+Noggin group was poor. Bone mineral density, bone volume fraction, levels of ALP, OCN, PINP and the protein levels of BMP-2, Runx2, and p-SMAD1/5/8/SMAD1/5/8 in the tibial fracture group were higher than those in the Cont group (p<0.05), while trabecular separation was lower than that in the Cont Group (P<0. 05). Compared to those in the tibial fracture group. bone mineral density, bone volume fraction, levels of ALP, OCN, PINP, and the protein levels of BMP-2, Runx2, and p-SMAD1/5/8/SMAD1/5/8 in the Cal group increased, and trabecular separation decreased (P<0.05) Compared to those in the Cal group, bone mineral density, bone volume fraction, levels of ALP, OCN, PINP. and the protein levels of BMP-2, Runx2, and p-SMAD1/5/8/SMAD1/5/8 in the Cal + Noggin group decreased, and trabecular separation increased (P < 0.05). Conclusion Cal may improve bone mineral density and microstructure of new callus, and promote the healing of tibial fractures by activating BMP-2/SMAD/Runx2 pathway. [ABSTRACT FROM AUTHOR]

Objective To investigate the effects of casein kinase 2 interacting protein-1 (CKIP-1) on the osteogenic differentiation ability of human periodontal ligament stem cells (hPDLSCs). Methods The hPDLSCs were obtained by primary culture with periodontal ligament tissues that were collected from normal humans. Then, a lentiviral vector containing a CKIP-1-specific siRNA sequence was constructed, and the transcriptional level of CKIP-1 in hPDLSCs was downregulated after vector infection. The P4 cells were divided into four groups: the control group, negative control group (infected with a control vector), CKIP-siRNA group (infected by a CKIP-1 siRNA lentivirus) and CKIP-1 group (infected by a CKIP-1 overexpression virus). All of the cells were cultured under osteogenic induction for 21 days. Then, alizarin red staining and quantitative determination were performed to detect the osteogenic differentiation ability of the hPDLSCs. In addition, qPCR was used to detect the transcriptional level of osteogenesis-related regulatory factors, such as Runt-related transcription factor 2 (Runx2), alkaline phosphatase (ALP), osteocalcin (OCN), and receptor activator of nuclear factor kappa-B ligand (RANKL), and the osteogenesis-related regulatory factors of the bone morphogenetic protein (BMP) signaling pathway. Results There were no differences in the indexes between the negative control group and the control group (P > 0.05). Compared with the negative control group, the CKIP-siRNA group demonstrated more mineralized nodules (P < 0.05), significantly increased calcium salt deposition (P < 0.05), and increased mRNA levels of osteogenesis-related regulatory factors, such as Runx2 , ALP, OCN, and RANKL, and the osteogenesis-related regulatory factors of BMP signaling pathway (P < 0.05). Conclusion Downregulation of CKIP-1 could promote the osteogenic differentiation of hPDLSCs, which is related to the transcription level of osteogenic-related regulatory factors.

Abstract Background Circaea mollis Sieb. & Zucc. has been used as a traditional herbal medicine in Hani Ethnopharmacy and possesses anti-arthritic activities. This study aimed to investigate the effect of Circaea mollis Siebold & Zucc on postmenopausal osteoporosis. Methods For in vitro study, MCF7 breast cancer cells and MC3T3-E1 pre-osteoblast cells were utilized to estimate estrogenic and osteogenic activity. Osteoblastic markers were measured by western blot and real-time PCR. For in vivo study, female mature C57BL/6 mice were ovariectomized and oral administrated with 10 mg/kg and 40 mg/kg of EECM respectively. Results EtOH extract of Circaea mollis Siebold & Zucc. (EECM) increased alkaline phosphatase activity and osteoblast marker levels at day 7 during differentiation of mouse preosteoblasts. EECM reduced osteoclast differentiation and bone resorption in an osteoblast-osteoclast primary co-culture system. In ovariectomized mice, EECM prevented the decrease in bone mineral density and recovered OSX and Runx2 via BMP2/4, Smad1/5/9 and p38. Conclusions The results suggest that EECM may be effective in preventing bone loss, offering a promising alternative for the nutritional management of postmenopausal osteoporosis.

To explore the relationship between serum runt-related transcription factor 2 (Runx2) mRNA expression and arterial calcification in patients with uremia. Methods A total of 78 uremic patients who underwent autologous arteriovenous fistula plasty and resection and reconstruction in our hospital were selected as the research objects, and blood vessel tissue, serum samples and clinical data of patients were collected. The calcification score were based on silver nitrate results, the patients were divided into the calcified group (n=23) and the non-calcified group (n=55). The expression level of Runx2 in blood vessel tissue was detected by immunohistochemistry, and the expression level of Runx2 mRNA in serum was detected by real-time fluorescence quantitative method. The changes of clinical indicators were compared between the two groups. The factors affecting vascular calcification of each indicator were analyzed by linear regression, and the predictive ability of serum Runx2 mRNA on vascular calcification was evaluated by ROC curve. Results The results of silver nitrate staining showed that compared with the non-calcified group, the black deposits were significantly increased in the calcified group. The immunohistochemical results showed that the Runx2 score was significantly increased in the calcified group (P＜0.05). Compared with the non-calcified group, the age, dialysis age, blood phosphorus, iPTH and serum Runx2 mRNA expression levels were significantly increased in the calcified group (P＜0.05), and serum albumin and blood calcium were significantly decreased (P＜0.05). Spearman correlation analysis showed that age, dialysis age, blood phosphorus, iPTH, serum Runx2 mRNA, Runx2 score were positively correlated with calcification score, and serum albumin and blood calcium were negatively correlated with calcification score (P＜0.05). The results of multiple linear regression analysis showed that the elevated serum Runx2 mRNA expression levels, long dialysis age and hypocalcemia were independent risk factors for arterial vascular calcification in patients with uremia (P＜0.05). ROC curve analysis showed that the area under the ROC curve (AUC) of serum Runx2 mRNA combined with dialysis age and blood calcium to diagnose vascular calcification was 0.934. Conclusion Serum Runx2 mRNA expression is an independent risk factor for arterial vascular calcification in uremic patients, and which may be a biomarker for the diagnosis and prediction of arterial vascular calcification in uremic patients. [ABSTRACT FROM AUTHOR]

Fracture healing is related to osteogenic differentiation and mineralization. Recently, due to the unwanted side effects and clinical limitations of existing treatments, various natural product-based chemical studies have been actively conducted. Albiflorin is a major ingredient in Paeonia lactiflora, and this study investigated its ability to promote osteogenic differentiation and fracture healing. To demonstrate the effects of albiflorin on osteoblast differentiation and calcified nodules, alizarin red S staining and von Kossa staining were used in MC3T3-E1 cells. In addition, BMP-2/Smad and Wnt/β-catenin mechanisms known as osteoblast differentiation mechanisms were analyzed through RT-PCR and western blot. To investigate the effects of albiflorin on fracture healing, fractures were induced using a chainsaw in the femur of Sprague Dawley rats, and then albiflorin was intraperitoneally administered. After 1, 2, and 3 weeks, bone microstructure was analyzed using micro-CT. In addition, histological analysis was performed by staining the fractured tissue, and the expression of osteogenic markers in serum was measured. The results demonstrated that albiflorin promoted osteoblastogenesis and the expression of RUNX2 by activating BMP-2/Smad and Wnt/β-catenin signaling in MC3T3-E1 cells. In addition, albiflorin upregulated the expression of various osteogenic genes, such as alkaline phosphatase, OCN, bone sialoprotein, OPN, and OSN. In the femur fracture model, micro-CT analysis showed that albiflorin played a positive role in the formation of callus in the early stage of fracture recovery, and histological examination proved to induce the expression of osteogenic genes in femur tissue. In addition, the expression of bone-related genes in serum was also increased. This suggests that albiflorin promotes osteogenesis, bone calcification and bone formation, thereby promoting the healing of fractures in rats.

Fracture healing is related to osteogenic differentiation and mineralization. Recently, due to the unwanted side effects and clinical limitations of existing treatments, various natural product-based chemical studies have been actively conducted. Albiflorin is a major ingredient in Paeonia lactiflora , and this study investigated its ability to promote osteogenic differentiation and fracture healing. To demonstrate the effects of albiflorin on osteoblast differentiation and calcified nodules, alizarin red S staining and von Kossa staining were used in MC3T3-E1 cells. In addition, BMP-2/Smad and Wnt/β-catenin mechanisms known as osteoblast differentiation mechanisms were analyzed through RT-PCR and western blot. To investigate the effects of albiflorin on fracture healing, fractures were induced using a chainsaw in the femur of Sprague Dawley rats, and then albiflorin was intraperitoneally administered. After 1, 2, and 3 weeks, bone microstructure was analyzed using micro-CT. In addition, histological analysis was performed by staining the fractured tissue, and the expression of osteogenic markers in serum was measured. The results demonstrated that albiflorin promoted osteoblastogenesis and the expression of RUNX2 by activating BMP-2/Smad and Wnt/β-catenin signaling in MC3T3-E1 cells. In addition, albiflorin upregulated the expression of various osteogenic genes, such as alkaline phosphatase, OCN, bone sialoprotein, OPN, and OSN. In the femur fracture model, micro-CT analysis showed that albiflorin played a positive role in the formation of callus in the early stage of fracture recovery, and histological examination proved to induce the expression of osteogenic genes in femur tissue. In addition, the expression of bone-related genes in serum was also increased. This suggests that albiflorin promotes osteogenesis, bone calcification and bone formation, thereby promoting the healing of fractures in rats. [ABSTRACT FROM AUTHOR]

Stromal derived factor 1 (SDF1) has been shown to be involved in the pathogenesis of pulmonary artery hypertension (PAH). However, the detailed molecular mechanisms remain unclear. To address this, we utilized primary cultured rat pulmonary artery smooth muscle cells (PASMCs) and monocrotaline (MCT)-induced PAH rat models to investigate the mechanisms of SDF1 driving PASMCs proliferation and pulmonary arterial remodeling. SDF1 increased runt-related transcription factor 2 (Runx2) acetylation by Calmodulin (CaM)-dependent protein kinase II (CaMKII)-dependent HDAC4 cytoplasmic translocation, elevation of Runx2 acetylation conferred its resistance to proteasome-mediated degradation. The accumulation of Runx2 further upregulated osteopontin (OPN) expression, finally leading to PASMCs proliferation. Blocking SDF1, suppression of CaMKII, inhibition the nuclear export of HDAC4 or silencing Runx2 attenuated pulmonary arterial remodeling and prevented PAH development in MCT-induced PAH rat models. Our study provides novel sights for SDF1 induction of PASMCs proliferation and suggests that targeting SDF1/CaMKII/HDAC4/Runx2 axis has potential value in the management of PAH. [ABSTRACT FROM AUTHOR]

Objective:To observe the effect of Jiangu granule on the expression of miR-141, Dlx5, Msx2 and Runx2 in bone tissue of ovariectomized mice, and to explore the mechanism of Jiangu granule in preventing and treating postmenopausal osteoporosis.Methods:A total of 30 C57 mice were randomly divided into sham operation group, saline group and Jiangu granule group, with 10 cases in each group. Fat around ovary of the sham operation group was removed, but not ovary, bilateral ovaries of the saline group and Jiangu granule group were removed surgically, and administrate respectively by normal saline and Jiangu granule.After eight weeks of intervention, the tibial morphology was observed by micro-CT, the expression of microRNA-141, Dlx5, Msx2 and Runx2 in bone tissue were detected by real-time PCR, protein contents of Dlx5, Msx2 and Runx2 in bone tissue were detected by Western blot.Results:Compared with the saline group, the number, width, length, shape and distribution of trabeculae in the Jiangu granule group recovered partially, and the density and connectivity of trabeculae increased, which could not be restored to the level of the sham operation group, The area of cortical bone increased, the bone marrow cavity decreased, and basically restored to the level of the sham operation group; BMC, BMD, BV/TV, BS/TV, Tb.N increased significantly; Tb.Sp, SMI reduced significantly;the relative amount of microRNA-141 and Msx2 in bone tissue decreased significantly;the relative contents of Runx2 and Dlx5 in bone tissue increased significantly, with statistically significant differences (P

Vascular calcification (VC) is an urgent worldwide health issue with no available medical treatment. It is an active cell-driven process by osteogenic differentiation of vascular cells with complex mechanisms. The AMP-activated protein kinase (AMPK) serves as the master sensor of cellular energy status. Accumulating evidence reveals the vital role of AMPK in VC progression. AMPK is involved in VC in various ways, including inhibiting runt-related transcription factor 2 signaling pathways, triggering autophagy, attenuating endoplasmic reticulum stress and dynamic-related protein 1-mediated mitochondrial fission, and activating endothelial nitric oxide synthase. AMPK activators, like metformin, are associated with reduced calcification deposits in certain groups of patients, indicating that AMPK is a potential therapeutic target for VC.

Background: Calcific aortic valve disease is associated with ageing and high mortality. However, no effective pharmacological treatment has been developed. Vascular endothelial growth factor (VEGF) and its receptor are overexpressed in the calcified aortic valve tissue. However, the role of VEGF in calcific aortic valve disease pathogenesis and its underlying mechanisms remain unclear. Materials and methods: Runt-related transcription factor 2 expression and calciumrelated signalling were investigated in porcine valvular interstitial cells with or without human VEGF-A recombinant protein (VEGF165, 1-100 ng/mL) treatment and/or calmodulin-dependent kinase II (CaMKII) inhibitor (KN93, 10 µmol/L) and inositol triphosphate receptor inhibitor (2-aminoethyldiphenyl borate, 30 µmol/L) for 5 days. Results: VEGF165-treated cells had higher Runt-related transcription factor 2 expression and CaMKII/ adenosine 3',5'-monophosphate response element-binding protein (CREB) signalling activation than did control cells. KN93 reduced Runt-related transcription factor 2 expression and CREB phosphorylation in VEGF165-treated cells. The 2-aminoethyldiphenyl borate also reduced Runt-related transcription factor 2 expression in VICs treated with VEGF165. Conclusion: VEGF upregulated Runt-related transcription factor 2 expression in VICs by activating the IP3R/CaMKII/CREB signalling pathway. [ABSTRACT FROM AUTHOR]

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Although various anti-osteoporosis drugs are available, the limitations of these therapies, including drug resistance and collateral responses, require the development of novel anti-osteoporosis agents. Rhizoma Drynariae displays a promising anti-osteoporosis effect, while the effective component and mechanism remain unclear. Here, we revealed the therapeutic potential of Rhizoma Drynariae-derived nanovesicles (RDNVs) for postmenopausal osteoporosis and demonstrated that RDNVs potentiated osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) by targeting estrogen receptor-alpha (ER α ). RDNVs, a natural product isolated from fresh Rhizoma Drynariae root juice by differential ultracentrifugation, exhibited potent bone tissue-targeting activity and anti-osteoporosis efficacy in an ovariectomized mouse model. RDNVs, effectively internalized by hBMSCs, enhanced proliferation and ER α expression levels of hBMSC, and promoted osteogenic differentiation and bone formation. Mechanistically, via the ER α signaling pathway, RDNVs facilitated mRNA and protein expression of bone morphogenetic protein 2 and runt-related transcription factor 2 in hBMSCs, which are involved in regulating osteogenic differentiation. Further analysis revealed that naringin, existing in RDNVs, was the active component targeting ER α in the osteogenic effect. Taken together, our study identified that naringin in RDNVs displays exciting bone tissue-targeting activity to reverse osteoporosis by promoting hBMSCs proliferation and osteogenic differentiation through estrogen-like effects., Competing Interests: The authors have no conflicts of interest to declare., (© 2024 The Authors.)

Background: Circaea mollis Sieb. & Zucc. has been used as a traditional herbal medicine in Hani Ethnopharmacy and possesses anti-arthritic activities. This study aimed to investigate the effect of Circaea mollis Siebold & Zucc on postmenopausal osteoporosis. Methods: For in vitro study, MCF7 breast cancer cells and MC3T3-E1 pre-osteoblast cells were utilized to estimate estrogenic and osteogenic activity. Osteoblastic markers were measured by western blot and real-time PCR. For in vivo study, female mature C57BL/6 mice were ovariectomized and oral administrated with 10 mg/kg and 40 mg/kg of EECM respectively. Results: EtOH extract of Circaea mollis Siebold & Zucc. (EECM) increased alkaline phosphatase activity and osteoblast marker levels at day 7 during differentiation of mouse preosteoblasts. EECM reduced osteoclast differentiation and bone resorption in an osteoblast-osteoclast primary co-culture system. In ovariectomized mice, EECM prevented the decrease in bone mineral density and recovered OSX and Runx2 via BMP2/4, Smad1/5/9 and p38. Conclusions: The results suggest that EECM may be effective in preventing bone loss, offering a promising alternative for the nutritional management of postmenopausal osteoporosis. [ABSTRACT FROM AUTHOR]

Objective This study aimed to investigate the effect of cAMP ⁃ responsive ⁃ element ⁃ binding protein (CREB) overexpression on the differentiation of human stem cells from the apical papilla (hSCAPs), stimulated by trans⁃ forming growth factor⁃ beta (TGF⁃β1). Methods Cells were isolated from human immature third molars via enzymatic digestion. Four experimental groups were set up: ①a control group, receiving normal mineralization inducer (α⁃MEM, 10% FBS, 10 mmol/L β ⁃ sodium glycerophosphate, 50 μg/mL vitamin C, 10 nmol/L dexamethasone); ② a TGF ⁃ β1 group, receiving normal mineralization inducer and 5 μg/mL TGF⁃β1; ③ a TGF⁃β1+LV⁃empty group, receiving normal mineralization inducer and the transfected empty virus vector with 5 μg/mL TGF ⁃ β1; and ④ a TGF ⁃ β1+ ov ⁃CREB group, receiving normal mineralization inducer and the transfected CREB ⁃ overexpressing viral vector, with 5 μg/mL TGF⁃β1. The transfected cells were cultured in odontogenic medium in the presence or absence of TGF⁃β1 for 2 weeks. Alizarin red staining was used to detect mineralized nodules, and the mRNA expression of the mineralization genes runt⁃ related transcription factor 2 (RUNX2), dentin sialophosphoprotein (DSPP) and alkaline phosphatase (ALP) was mea⁃ sured by qPCR. Results Compared with the control group (1.12 ± 0.11), TGF⁃β1 inhibited the deposition of calcium minerals (0.67 ± 0.12) (P < 0.05) via hSCAPs and inhibited the mRNA expression of RUNX2 (0.60 ± 0.03), DSPP (0.43 ± 0.12) and ALP (0.69 ± 0.05) (P < 0.05). In contrast, overexpression of CREB attenuated the effect of TGF⁃β1 on hS⁃ CAPs, resulting in the development of a high number of mineralized nodules (1.27 ± 0.10) (P < 0.01) and increased RNA levels of RUNX2 (1.33 ± 0.07), DSPP (1.32 ± 0.11) and ALP (1.26 ± 0.03) (P＜0.05) compared with those in the TGF⁃β1 group. Conclusion Overexpressed CREB promotes odontogenic differentiation of hSCAPs by interfering with TGF⁃β1.

The neuropeptide Y (NPY) system is considered one of the primary neural signaling pathways. NPY, produced by osteoblasts and other peripheral tissues, is known to inhibit biological functions of osteoblasts. However, until recently, little was known of the autocrine mechanism by which NPY is regulated. To investigate this mechanism, overexpression plasmids and small interfering RNA (siRNA) targeting NPY were transfected into the MC3T3-E1 cell line to observe its effects on osteogenesis. NPY overexpression was found to markedly enhance the osteogenic ability of MC3T3-E1 cells by an autocrine mechanism, coincident with the upregulation of osterix and runt-related transcription factor 2 (Runx2). Furthermore, NPY increased the activities of alkaline phosphatase (ALP) and osteocalcin (OCN) by upregulating their osteoblastic expression in vitro (as well as that of osterix and Runx2). Following transfection with NPY-siRNA, the osteoblastic ability of MC3T3-E1 cells was markedly decreased, and NPY deficiency inhibited the protein expression of osterix, Runx2, OCN and ALP in primary osteoblasts. Collectively, these results indicated that NPY played an important role in osteoblast differentiation by regulating the osterix and Runx2 pathways. [ABSTRACT FROM AUTHOR]

Background: A recent study reported a novel long non-coding RNA (lncRNA) E2F-mediated cell proliferation enhancing lncRNA (EPEL, human chromosome 4, intergenic region) plays an oncogenic role in lung cancer.Aims: We aimed to investigate the role of lncRNA EPEL in gastric cancer.Methods: Gene expression was analyzed by RT-qPCR and western blot. Survival analysis was performed by comparing survival curves. Cell proliferation, migration, and invasion were analyzed by CCK-8 and Transwell assays.Results: We found that lncRNA EPEL and Runt-related transcription factor 2 (RUNX2) were both upregulated in gastric cancer. EPEL and RUNX2 were positively correlated in tumor. Patients with high expression level of lncRNA EPEL showed poor survival. LncRNA EPEL and RUNX2 overexpression promoted, while lncRNA EPEL siRNA silencing inhibited the migration, proliferation, and invasion of gastric cancers. In addition, RUNX2 overexpression completely rescued the inhibited cancer cell migration, proliferation, and invasion caused by lncRNA EPEL siRNA silencing. Consistently, EPEL overexpression resulted in upregulated RUNX2 expression, while RUNX2 overexpression did not affect lncRNA EPEL expression.Conclusions: Therefore, lncRNA EPEL may regulate cancer cell behaviors and affect prognosis of gastric cancer by interacting with RUNX2. [ABSTRACT FROM AUTHOR]

The probable positive effects of photobiomodulation therapy (PBMT) and oxytocin (OT) treatments together or alone were evaluated on cell viability along with the changes in the gene expression of Osteocalcin (OC), Osteoprotegerin (OPG), and Runt-related transcription factor 2 (Runx2) levels of sham (healthy)-Bone marrow mesenchymal stem cell(BMMSC) and ovariectomy-induced osteoporosis (OVX)-BMMSC. BMMSC was harvested from healthy and OVX rats and was cultured in osteogenic induction medium (OIM). There were five groups of BMMSCs: (1) sham -BMMSCs; (2) control -OVX-BMMSCs; (3) OT-treated-OVX-BMMSCs; (4) PBMT-treated-OVX-BMMSCs, and (5) OT + PBMT-OVX-BMMSCs. In all 5 groups, BMMSC viability and proliferation as well as gene expression of OC, OPG, and RUNX2 were evaluated. PBMT and PBMT + OT treatments showed a promising effect on the increased viability of OVX-BMMSC (ANOVA test; LSD test, p = 0.01, p = 0.002). The results of gene expression analysis revealed that the sham- BMMSCs responded optimally to OT treatment. It was also found that OVX-BMMSCs responded optimally to PBMT + OT and PBMT treatments at early and middle stages of osteogenic induction process. Nevertheless, they responded optimally to PBMT + OT and OT especially at the late stage of osteogenic induction process. PBMT and PBMT + OT treatments significantly increased viability of OVX-BMMSC in OIM in vitro. Both PBMT and PBMT + OT treatments could promote mineralization of OVX-BMMSC in the culture medium at early and middle stages of osteogenic induction process. Both OT and PBMT + OT treatments could promote mineralization of OVX-BMMSC in vitro at late stages of osteogenic induction process. [ABSTRACT FROM AUTHOR]

Objectives: To determine glucose transporter 1 (GLUT1) and runt‐related transcription factor 2 (RUNX2) expression during reparative dentinogenesis after pulpotomy with mineral trioxide aggregate (MTA) capping. Subjects and methods: Eight‐week‐old male Wistar rats were used. Pulp of the upper left first molar was exposed and capped with MTA. The upper right first molar of the same animal was used as a control. After collecting molars at various time points, GLUT1, RUNX2 and mammalian target of rapamycin (MTOR) were examined by immunohistochemistry. mRNA levels of Slc2a1 (encoding GLUT1), Runx2, Nestin and Mtor were determined by real‐time PCR. Results: Pulp exhibited progressive formation of reparative dentine lined with GLUT1‐ and MTOR‐immunoreactive odontoblast‐like cells at 5 days after pulpotomy. RUNX2 was detected in nuclei of most pulp tissue cells at day 5 after pulpotomy. Double immunofluorescence staining revealed GLUT1 immunoreactivity on odontoblast‐like cells positive for Nestin or RUNX2, 5 days after pulpotomy. Slc2a1, Runx2, Nestin and Mtor mRNA levels were significantly upregulated on days 3–5 after pulpotomy. Conclusions: After rat molar pulpotomy, dental pulp induced formation of reparative dentine with colocalization of GLUT1 and Nestin or RUNX2. Moreover, mRNA levels of Slc2a1, Runx2, Nestin and Mtor were significantly upregulated in pulpotomized dental pulp. [ABSTRACT FROM AUTHOR]

Osteoporosis (OP) is an age-related bone disease occurring worldwide. Osteoporotic fracture is one of the leading causes of disability and death in elderly patients. MicroRNAs (miRNAs/miRs) are key molecular regulatory factors in bone remodeling processes. The present study investigated the expression and mechanism of miR-135b-5p in patients with osteoporosis. The present results suggested that miR-135b-5p was upregulated in bone tissue fragments of patients with osteoporosis compared with the control patients. MC3T3-E1 cells were used to perform osteogenic differentiation induction. Reverse transcription-quantitative PCR and western blot assay were used to detect the mRNA and protein expression levels of the osteogenic markers osteocalcin (OC), Osterix and alkaline phosphatase (ALP). A specific kit was used for detecting ALP activity. The present results indicated that the mRNA expression levels of OC, Osterix and ALP significantly increased on the 7 and 14th day after osteogenic differentiation induction compared with the control group. Protein expression levels of OC, Osterix and ALP also increased on the 7 and 14th day after induction. ALP assay showed that ALP activity was significantly increased on the 7 and 14th day after induction. In addition, the present study found that miR-135b-5p was downregulated in MC3T3-E1 cells 7 and 14 days after osteogenic differentiation induction. The results of TargetScan analysis and dual luciferase reporter gene assay indicated that runt-related transcription factor 2 (RUNX2) was a direct target gene of miR-135b-5p. RUNX2 was upregulated in MC3T3-E1 cells on the 7 and 14th day after induction. Moreover, the present study found that compared with the osteogenic differentiation induction group, miR-135b-5p mimic significantly decreased OC, Osterix and ALP expression, and reduced ALP activity in MC3T3-E1 cells. However, these reductions were reversed following overexpression of RUNX2. The present results showed that miR-135b-5p mimic significantly reduced cell viability in MC3T3-E1 cells and induced cell apoptosis, and these effects were significantly reversed following RUNX2 overexpression. In summary, the present results suggested that miR-135-5p participated in the occurrence and development of osteoporosis via inhibition of osteogenic differentiation and osteoblast growth by targeting RUNX2. The present study suggested a novel potential target that may faciliate the treatment of osteoporosis, and further study is required to examine this possibility. [ABSTRACT FROM AUTHOR]

Osteosarcoma (OS) is the most common form of bone malignancy in children and adolescents. MicroRNAs (miRNAs) have been associated with the development and progression of OS. In the present study, reverse transcription-quantitative PCR, western blotting, Cell Counting Kit-8, luciferase and Transwell assays were performed to investigate the biological function of microRNA-150 (miR-150) in OS. The results revealed that miR-150 was significantly downregulated in OS cell lines (HOS, SAOS2, MG-63 and U2OS) in comparison with the normal osteoblast cells (hFOB1.19). Overexpression of miR-150 significantly inhibited cell proliferation in OS cells. miR-150 could sensitize OS cells to chemotherapy treatment of doxorubicin. Runt-related transcription factor 2 (RUNX2) was identified as a target gene of miR-150. RUNX2 knockdown exhibited similar inhibitory effects on both OS cell proliferation and chemotherapy sensitivity. Restoration of RUNX2 reversed the biological function of miR-150. Finally, miR-150 overexpression and RUNX2 knockdown enhanced caspase-3 cleavage. Taken together, the present study established a novel molecular mechanism, in that miR-150 plays tumor suppressor and chemoprotective roles by targeting RUNX2 in OS, indicating that miR-150 may be a potential therapeutic target for OS therapy in the future. [ABSTRACT FROM AUTHOR]

Aim: Glucose fluctuations may be responsible for, or further the onset of arterial hypertension, but the exact mechanisms remain unclear. The purpose of this study was to investigate the mechanisms behind and related to aortic fibrosis and aortic stiffening induced by glucose fluctuations. Methods: Sprague-Dawley rats were injected with streptozotocin (STZ) and randomly divided into three treatment groups: controlled STZ-induced diabetes (C-STZ); uncontrolled STZ-induced diabetes (U-STZ); and STZ-induced diabetes with glucose fluctuations (STZ-GF). After 3 weeks, rat blood pressure (BP) was tested, and aortic fibrosis was detected by using the Masson trichrome staining technique. Levels of p38 mitogen-activated protein kinase (p38 MAPK), runt-related transcription factor 2 (Runx2), collagen type 1 (collagen I), and NADPH oxidases were determined by Western blot.Rat vascular smooth muscle cells in vitro were used to explore underlying mechanisms. Results: The systolic BP of diabetic rats in the C-STZ, U-STZ, and STZ-GF groups was 127.67 ± 6.53, 150.03 ± 5.24, and 171.63 ± 3.53 mm Hg, respectively (p< 0.05). The mean BP of diabetic rats in the three groups was 91.20 ± 10.07, 117.29 ± 4.28, and 140.58 ± 2.14 mm Hg, respectively (p< 0.05). The diastolic BP of diabetic rats in the three groups was 73.20 ± 12.63, 101.93 ± 5.79, and 125.37 ± 4.62 mm Hg, respectively (p< 0.05). The ratios of fibrosis areas in the aortas of the three groups were 11.85 ± 1.23, 29.00 ± 0.87, and 48.36 ± 0.55, respectively (p< 0.05). The expressions of p38 MAPK, Runx2, and collagen I were significantly increased in the STZ-GF group. In vitro, applications of inhibitors of reactive oxygen species (ROS) and p38 MAPK successfully reversed glucose fluctuations that would have possibly induced aortic fibrosis. Conclusions: Blood glucose fluctuations aggravate aortic fibrosis via affecting the ROS/p38 MAPK /Runx2 signaling pathway. [ABSTRACT FROM AUTHOR]

The role of long non-coding RNA heart and neural crest derivatives expressed 2-antisense RNA 1 (lncRNA HAND2-AS1) in tumor suppression has been reported in a number of cancer types, while its functionality in triple-negative breast cancer (TNBC) remains unclear. The present study aimed to investigate the involvement of lncRNA HAND2-AS1 in TNBC using different methodologies. HAND2-AS1 and RUNX2 expression was analyzed using reverse transcription-quantitative PCR and western blot analysis. Diagnostic analysis was performed using receiver operating characteristic curve. Overexpression experiments were performed to analyze the interaction between HAND2-AS1 and RUNX2 while the Cell Counting Kit-8 assay was performed to analyze cell proliferation. In patients with early-stage TNBC, the expression level of lncRNA HAND2-AS1 was downregulated, whilst runt-related transcription factor 2 (RUNX2) mRNA was upregulated in tumor tissues, compared with paired healthy tissues. Expression levels of lncRNA HAND2-AS1 and RUNX2 mRNA were inversely correlated in tumor tissues, but not in paired healthy tissues. Decreased plasma expression levels of lncRNA HAND2-AS1 were observed in TNBC patients compared with those in healthy controls, and the downregulation of lncRNA HAND2-AS1 distinguished patients with TNBC from healthy controls. lncRNA HAND2-AS1 overexpression inhibited RUNX2 expression, whilst RUNX2 overexpression did not significantly affect lncRNA HAND2-AS1 expression in the MDA-MB-231 and BT-20 cell lines. lncRNA HAND2-AS1 overexpression resulted in the inhibition of cell proliferation, while RUNX2 overexpression promoted the proliferation of TNBC cells. RUNX2 overexpression partially reversed the effects of lncRNA HAND2-AS1 overexpression on cancer cells. Therefore lncRNA HAND2-AS1 may inhibit the proliferation of cancer cells by inhibiting RUNX2 expression in TNBC. [ABSTRACT FROM AUTHOR]