Thioredoxin1 is a target to attenuate diabetes­induced RPE cell dysfunction in human ARPE19 cells by alleviating oxidative stress.

Diabetes­induced cell dysfunction of the retinal pigment epithelium (RPE) contributes to the initiation and progression of diabetic retinopathy (DR). Thioredoxin 1 (Trx1) plays a key role in DR. However, the effect and mechanism of Trx1 on diabetes­induced cell dysfunction of the RPE is not fully understood during DR. In the present study, the effect of Trx1 on this process and its related mechanism were investigated. A Trx1 overexpression cell line, ARPE19­Trx1/LacZ, was constructed and treated with or without high glucose (HG). Flow cytometry was used to analyze apoptosis of these cells and the mitochondrial membrane potential was analyzed using JC­1 staining solution. A DCFH­DA probe was also used to detect the reactive oxygen species (ROS) generation. Western blotting was used to examine the expression of related proteins in ARPE­19 cells after HG treatment. The results demonstrated that the RPE layer was damaged in clinical samples. ROS formation and RPE cell dysfunction increased after HG treatment in vitro. Besides, the expression of mitochondrial­mediated apoptosis related proteins (Bax, apoptosis­inducing factor, cytochrome C, Caspase3 and Caspase9) also increased; however, overexpression of Trx1 attenuated these changes and improved the function of ARPE19 cells. These results indicated that overexpression of Trx1 alleviated diabetes­induced RPE cell dysfunction in DR by attenuating oxidative stress.

Application of Organoids in Carcinogenesis Modeling and Tumor Vaccination.

Organoids well recapitulate organ-specific functions from their tissue of origin and remain fundamental aspects of organogenesis. Organoids are widely applied in biomedical research, drug discovery, and regenerative medicine. There are various cultivated organoid systems induced by adult stem cells and pluripotent stem cells, or directly derived from primary tissues. Researchers have drawn inspiration by combination of organoid technology and tissue engineering to produce organoids with more physiological relevance and suitable for translational medicine. This review describes the value of applying organoids for tumorigenesis modeling and tumor vaccination. We summarize the application of organoids in tumor precision medicine. Extant challenges that need to be conquered to make this technology be more feasible and precise are discussed.

Organoid culture system for patient-derived lung metastatic osteosarcoma.

Osteosarcoma (OS) is the most common primary bone malignancy with high rates of recurrence and metastasis. OS often spreads to lungs, an optimized model for studying lung metastatic OS cells may help develop potential therapies for patients with lung metastasis. Here we firstly report an organoid culture system for lung metastatic OS tissues. We provided a fully described formula that was required for establishing lung metastatic OS organoids (OSOs). Using this protocol, the lung OSOs were able to be maintained and serially propagated for at least six months; the OSOs can also be generated from cryopreserved patient samples without damaging the morphology. The patient-derived lung OSOs retained the cellular morphology and expression of OS markers (Vimentin and Sox9) that recapitulate the histological features of the human OS. The microenvironment of primary lung metastatic OSOs preserved a similar T cell distribution with the human lung OS lesions; this provided a possible condition to explore how OS cells may react to immunotherapy. OSOs established from this protocol can be further utilized for studying various aspects of OS biology (e.g., tumorigenesis and drug screen/discovery) for precision medicine.

Ghrelin promotes the osteogenic differentiation of rMSCs via miR-206 and the ERK1/2 pathway.

OBJECTIVE: Mesenchymal stem cells (MSCs) can differentiate into chondroblasts, adipocytes, or osteoblasts under appropriate stimulation. Ghrelin, an endogenous ligand for the growth hormone secretagogue receptor (GHSR), stimulates growth hormone (GH) secretion and exerts both orexigenic and adipogenic effects. The ERK1/2 signaling pathway is known to trigger osteogenic differentiation of rabbit bone marrow-derived mesenchymal stromal cells. In the present study, the function of miR-206 in the ghrelin-mediated osteogenic differentiation of rabbit bone marrow-derived mesenchymal stromal cells (rMSCs) was explored. METHODS: The expression of miR-206 was detected by qPCR, and phosphorylated ERK1/2 and the protein expression levels of ALP, RUNX2, and Osterix were assessed by western blotting. RESULTS: Ghrelin inhibited the expression of miR-206 to promote the osteogenic differentiation of rMSCs. Moreover, ghrelin increased the phosphorylation of ERK1/2, while overexpression of miR-206 suppressed ERK1/2 phosphorylation, indicating that miR-206 can regulate the ERK1/2 pathway. Further, inhibition of ERK1/2 had no influence on miR-206 expression; however, the phosphorylation of ERK1/2 was decreased, and the protein expression levels of ALP, RUNX2, and Osterix were downregulated. CONCLUSIONS: Ghrelin promotes the osteogenic differentiation of rMSCs via miR-206 and the ERK1/2 pathway.

LncRNA PCAT6 Accelerates the Progression and Chemoresistance of Cervical Cancer Through Up-Regulating ZEB1 by Sponging miR-543.

BACKGROUND: Cervical cancer (CC) is a common cancer with a poor prognosis due to the chemoresistance of CC cells to cisplatin. This study aimed to investigate the biological significance of lncRNA prostate cancer-associated transcript 6 (PCAT6) in the carcinogenesis of CC. MATERIALS AND METHODS: Quantitative real-time polymerase chain reaction (qRT-PCR) was carried out to measure the abundance of PCAT6, miR-543 and zinc finger E-box binding protein 1 (ZEB1) in CC tissues and cells. The combination between miR-543 and lncRNA PCAT6 or ZEB1 was predicted by Starbase and was verified by dual-luciferase reporter assay, RNA-pull down assay and RNA immunoprecipitation (RIP) assay. Cell proliferation and chemoresistance to cisplatin were detected by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cell apoptosis and metastasis were determined by flow cytometry, Western blot and transwell migration and invasion assays. RESULTS: The abundance of ZEB1 protein was measured by Western blot assay. Murine xenograft model was established to confirm the function of lncRNA PCAT6 in vivo. The abundance of lncRNA PCAT6 was enhanced in CC tissues and cells compared with that in corresponding normal tissues and normal cervical epithelial cells Ect1/E6E7. MiR-543 was a target of PCAT6 and was negatively regulated by PCAT6. PCAT6 accelerated the proliferation, metastasis and the chemoresistance of CC cells to cisplatin while suppressed the apoptosis of CC cells. The overexpression of PCAT6 reversed the inhibitory effects of miR-543 accumulation on the proliferation, metastasis and chemoresistance of CC cells to cisplatin and the promoting impact on the apoptosis of CC cells. ZEB1 was a direct target of miR-543, and it functioned as the downstream gene of PCAT6/miR-543 to exert its oncogenic role in CC. PCAT6 promoted the growth of murine xenograft tumor through miR-543/ZEB1 axis in vivo. CONCLUSION: LncRNA PCAT6 facilitated the proliferation, metastasis and chemoresistance of CC cells to cisplatin while impeded the apoptosis of CC cells via PCAT6/miR-543/ZEB1 axis. PCAT6/miR-543/ZEB1 axis might be a promising target for CC therapy.

Sesamin Promotes Osteoblastic Differentiation and Protects Rats from Osteoporosis.

BACKGROUND Osteoporosis is a common osteopathy, resulting in fractures, especially in elder people. Sesamin has many pharmacological effects, including supplying calcium. However, how sesamin might prevent osteoporosis is still under study. MATERIAL AND METHODS Bone marrow stromal cells (BMSCs) extracted from rat femur were induced for osteoblastic differentiation. Cell proliferation, alkaline phosphatase (ALP), osterix (OSX), SRY-box 9 (SOX9), runt-related transcription factor 2 (RUNX2), osteocalcin (OCN), ß-catenin, low density lipoprotein receptor-related protein 5 (LRP5), and glycogen synthase kinase-3ß (GSK-3ß) levels in BMSCs were detected in the presence or absence of sesamin (1 µM or 10 µM). In addition, FH535 (1 µM) was used to silence Wnt/ß-catenin in vitro. Ovariectomized (OVX) rats were established and intragastrically administrated sesamin (80 mg/kg), and then the rat bones were analyzed by micro-computed tomography. Osteocalcin and collagen type I were measured in the rat femurs. RESULTS Sesamin had no influence on BMSC proliferation. Higher sesamin concentration promoted Wnt/ß-catenin activity and enhanced more expressions of ALP, OSX, SOX9, RUNX2, and OCN, gradually and significantly (P<0.05). Silencing Wnt/ß-catenin weakened the enhancement on RUNX2 and OCN expression. Sesamin (80 mg/kg) promoted bone structure in ovariectomized rats, and significantly enhanced osteocalcin and collage type I expression (P<0.05). CONCLUSIONS Sesamin promoted osteoblastic differentiation of rat BMSCs by regulating the Wnt/ß-catenin pathway, and improved rat bone structure. Sesamin could have therapeutic and preventive effects on osteoporosis.

Clinical significance of bromodomain-containing protein 7 and its association with tumor progression in prostate cancer.

Prostate cancer (PCa) is a common malignancy in males. The current study assessed the clinical significance of bromodomain-containing protein 7 (BRD7) and its association with PCa tumor progression. Serum and tissue expression levels of BRD7 were analyzed by reverse transcription-quantitative polymerase chain reaction. Receiver operating characteristic (ROC) analysis was used to evaluate the diagnostic value of BRD7. Kaplan-Meier survival analysis and Cox regression analysis were performed to assess the prognostic performance of BRD7. The association of BRD7 with cell behavior was investigated by transfection with a pcDNA3.1-BRD7 vector. The results revealed that serum and tissue BRD7 expression levels were significantly decreased in PCa samples compared with normal controls (P<0.001). BRD7 expression was significantly associated with the pathological stage (P=0.037), lymph node metastasis (P=0.009) and TNM stage (P=0.010). An area under the ROC curve of 0.864 was obtained, with a sensitivity and specificity of 77.0 and 83.3%, respectively. Low BRD7 expression was significantly associated with a shorter survival time in both overall survival analysis (P=0.003) and cancer-specific survival analysis (P=0.029). Furthermore, BRD7 appeared to serve as an independent prognostic factor for PCa. The proliferation, migration and invasion of PCa cells were suppressed by BRD7 overexpression. In summary, downregulation of BRD7 in PCa may be involved in tumor progression and serve as an effective diagnostic and prognostic biomarker.

Inhibition of osteoclastogenesis for periprosthetic osteolysis therapy through the suppression of p38 signaling by fraxetin.

Periprosthetic osteolysis belongs to osteolytic diseases, which often occur due to an imbalance between osteoclast and osteoblast number or activity. Fraxetin, a natural plant extract, inhibits osteoblast apoptosis and has therapeutic potential for treating osteolytic diseases. However, data pertaining to the effects of fraxetin on osteoclasts are limited. In the present study, it was demonstrated that the inhibition of osteoclastogenesis by fraxetin had an important role on the therapy of titanium particle­induced osteolysis in vivo. In addition, fraxetin was demonstrated to suppress receptor activator of nuclear factor­κB ligand (RANKL)­mediated osteoclast differentiation and bone resorption in vitro in a dose­dependent manner. Fraxetin inhibited osteoclast differentiation and function through the suppression of p38 signaling and subsequently, the suppression of osteoclast­specific gene expression, including tartrate­resistant acid phosphatase, nuclear factor of activated T­cells, cytoplasmic 1, and cathepsin K. In conclusion, fraxetin administration may have potential as a treatment method for periprosthetic osteolysis and other osteolytic diseases.

Effects of the 1, 4-dihydropyridine L-type calcium channel blocker benidipine on bone marrow stromal cells.

Osteoporosis (OP) often increases the risk of bone fracture and other complications and is a major clinical problem. Previous studies have found that high blood pressure is associated with bone formation abnormalities, resulting in increased calcium loss. We have investigated the effect of the antihypertensive drug benidipine on bone marrow stromal cell (BMSC) differentiation into osteoblasts and bone formation under osteoporotic conditions. We used a combination of in vitro and in vivo approaches to test the hypothesis that benidipine promotes murine BMSC differentiation into osteoblasts. Alkaline phosphatase (ALP), osteocalcin (OCN), runt-related transcription factor 2 (RUNX2), ß-catenin, and low-density lipoprotein receptor-related protein 5 (LRP5) protein expression was evaluated in primary femoral BMSCs from C57/BL6 mice cultured under osteogenic conditions for 2 weeks to examine the effects of benidipine. An ovariectomized (OVX) mouse model was used to investigate the effect of benidipine treatment for 3 months in vivo. We found that ALP, OCN, and RUNX2 expression was up-regulated and WNT/ß-catenin signaling was enhanced in vitro and in vivo. In OVX mice that were intragastrically administered benidipine, bone parameters (trabecular thickness, bone mineral density, and trabecular number) in the distal femoral metaphysis were significantly increased compared with control OVX mice. Consistently, benidipine promoted BMSC differentiation into osteoblasts and protected against bone loss in OVX mice. Therefore, benidipine might be a suitable candidate for the treatment of patients with postmenopausal osteoporosis and hypertension.