Myeloma extracellular vesicle-derived RAGE increases inflammatory responses and myotube atrophy in multiple myeloma through activation of the TLR4/NF-κB p65 pathway.

Sarcopenia manifests as muscle atrophy and loss that is complicated with malignancy. This study explored the mechanism of extracellular vesicles (EVs) in multiple myeloma (MM) with sarcopenia. SP2/0 conditioned medium (CM) was collected to isolate SP2/0-EVs. C2C12 cells were incubated with SP2/0 CM or SP2/0-EVs. ROS, TNF-α, IL-6, MuRF1 and MyHC levels were detected by DCF-DA fluorescent probe, ELISA, and Western blot. GW4869 was used to inhibit EV secretion in SP2/0 to confirm its effect on muscle atrophy. Serum was collected from MM patients with or without sarcopenia to detect RAGE mRNA expression. SP2/0 cells were transfected with RAGE siRNA and C2C12 cells were treated with the isolated si-RAGE-EVs or/and TLR4 agonist. SP2/0 tumor-bearing mouse model was established. Healthy mice and SP2/0-tumor bearing mice were treated with SP2/0-EVs or si-RAGE-EVs. SP2/0 CM or SP2/0-EVs stimulated ROS, inflammatory responses, and myotube atrophy in C2C12 cells. GW4869 blocked EV secretion and the effects of SP2/0 CM. RAGE mRNA expression in serum EVs was increased in MM&Sarcopenia patients and RAGE knockdown in SP2/0-EVs partially nullified SP2/0-EVs' effects. SP2/0-EVs activated the TLR4/NF-κB p65 pathway by translocating RAGE. SP2/0-EVs-derived RAGE elevated ROS production, inflammation, and myotube atrophy in C2C12 cells and caused muscle loss in SP2/0 tumor-bearing mice by activating the TLR4/NF-κB p65 pathway. SP2/0-EVs partially recapitulated muscle loss in healthy mice. SP2/0-EVs-derived RAGE increased ROS production, inflammation, and myotube atrophy in MM through TLR4/NF-κB p65 pathway activation.

SIRT5 promote malignant advancement of chordoma by regulating the desuccinylation of c-myc.

Chordoma is a relatively rare and locally aggressive malignant tumor. Sirtuin (SIRT)5 plays pivotal roles in various tumors, but the role of SIRT5 in chordoma has not been found. This study was performed to investigate the regulatory effects of SIRT5 on cell proliferation, migration, and invasion and the underlying mechanism in chordoma. A xenograft tumor mouse model was established to assess tumor growth. Reverse transcription-quantitative polymerase chain reaction was used to analyze the mRNA levels of SIRT5 and c-myc. The effects of SIRT5 and c-myc on cell proliferation, migration, and invasion of chordoma cells were detected by cell counting kit-8, colony formation, and Transwell assays. The interaction between SIRT5 and c-myc was evaluated by co-immunoprecipitation (IP) assay. The succinylation of c-myc was analyzed by IP and Western blot. The results showed that SIRT5 expression was upregulated in chordoma tissues and cells. SIRT5 interacted with c-myc to inhibit the succinylation of c-myc at K369 site in human embryonic kidney (HEK)-293T cells. Silencing of SIRT5 suppressed the cell proliferation, migration, and invasion of chordoma cells, while the results were reversed after c-myc overexpression. Moreover, silencing SIRT5 suppressed tumor growth in mice. These findings suggested that SIRT5 promoted the malignant advancement of chordoma by regulating the desuccinylation of c-myc.

Analysis of the efficacy of separation surgery for severe neurological compression in multiple myeloma: a retrospective analysis of 35 cases.

PURPOSE: To investigate the effectiveness and safety of separation surgery for Epidural Spinal Cord Compression (ESCC) graded ≥ 2 in patients with Multiple Myeloma (MM), analyze factors influencing surgical outcomes, and develop a preliminary treatment decision framework for these patients. METHODS: A retrospective analysis was conducted on clinical data from 35 MM patients who underwent separation surgery for ESCC graded ≥ 2 between 2013 and 2018. Patient data, including baseline information, surgical details, complications, and pre-operative as well as one-month post-operative efficacy evaluation indicators were recorded. Statistical analysis was performed on pre-operative and post-operative efficacy indicators to determine if there were significant improvements (p < 0.05). Ordered logistic regression was utilized to assess factors associated with an unfavorable post-operative quality of life outcome. RESULTS: Compared to pre-operative values, at one-month post-surgery, patients showed significant improvements in Frankel Score Classification (4 vs 5, p < 0.05), Karnofsky Performance Score (30 vs 70, p < 0.05), and Visual Analogue Scale (8 vs 3, p < 0.05). Complications occurred in 7 cases (20%). The number of segments with ESCC (OR = 0.171, p < 0.05) and pre-operative chemotherapy (OR = 5.202, p = 0.05) were identified as independent factors influencing patient outcomes. Patients with more than two vertebral segments with ESCC exhibited significantly worse post-operative conditions. CONCLUSIONS: Separation surgery effectively alleviates pain, improves neurological function, and enhances the quality of life in patients with ESCC graded ≥ 2 due to MM.

Which factors are associated with adverse prognosis in multiple myeloma patients after surgery? - preliminary establishment and validation of the nomogram.

BACKGROUND: To investigate the prognosis of patients with Multiple Myeloma (MM) after surgery, analyze the risk factors leading to adverse postoperative outcomes, and establish a nomogram. METHODS: Clinical data from 154 patients with MM who underwent surgery at our institution between 2007 and 2019 were retrospectively analyzed. Assessing and comparing patients' pain levels, quality of life, and functional status before and after surgery (P < 0.05) were considered statistically significant. The Kaplan-Meier survival curve was used to estimate the median survival time. Adverse postoperative outcomes were defined as worsened symptoms, lesion recurrence, complication grade ≥ 2, or a postoperative survival period < 1 year. Logistic regression analysis was used to determine the prognostic factors. Based on the logistic regression results, a nomogram predictive model was developed and calibrated. RESULTS: Postoperative pain was significantly alleviated in patients with MM, and there were significant improvements in the quality of life and functional status (P < 0.05). The median postoperative survival was 41 months. Forty-nine patients (31.8%) experienced adverse postoperative outcomes. Multivariate logistic regression analysis identified patient age, duration of MM, International Staging System, preoperative Karnofsky Performance Status, and Hb < 90 g/L as independent factors influencing patient prognosis. Based on these results, a nomogram was constructed, with a C-index of 0.812. The calibration curve demonstrated similarity between the predicted and actual survival curves. Decision curve analysis favored the predictive value of the model at high-risk thresholds from 10% to-69%. CONCLUSION: This study developed a nomogram risk prediction model to assist in providing quantifiable assessment indicators for preoperative evaluation of surgical risk.

Implication of different frailty criteria in older people with atrial fibrillation: a prospective cohort study.

BACKGROUND: the prevalence of physical and multidimensional frailty and their prognostic impact on clinical outcomes in patients with atrial fibrillation (AF) is unclear. OBJECTIVE: to evaluated frailty in a cohort of patients with AF according to different criteria, and studied the prevalence and its prognostic impact on clinical outcomes. METHODS: in this multicenter prospective cohort, 197 inpatients ≥ 65 years old with AF were recruited from September 2018 to April 2019.We used Fried Frailty phenotype (Fried) to assess physical frailty, and comprehensive geriatric assessment-frailty index (CGA-FI) to assess multidimensional frailty. The primary outcome was a composite of all-cause mortality or rehospitalization. RESULTS: the prevalence of frailty was determined as 34.5% by Fried, 42.6% by CGA-FI. Malnutrition and ≥ 7 medications were independently associated with frailty. Kaplan-Meier survival curve showed that the presence of frailty by CGA-FI had significantly lower all-cause mortality or rehospitalization survival rate (log-rank P = 0.04) within 1 year. Multivariate Cox regression adjusted for age and sex showed that the frailty by CGA-FI was significantly associated with the risk of all-cause mortality or rehospitalization within 1 year (HR 1.79, 95% CI 1.10-2.90). However, those associations were absent with the physical frailty. After broader multivariate adjustment, those associations were no longer statistically significant for both types of frailty. CONCLUSIONS: in older people with AF, Multidimensional frailty is more significantly associated with a composite of all-cause mortality or rehospitalization within 1 year than physical frailty, but these association are attenuated after multivariate adjustment. CLINICAL TRIAL REGISTRATION: ChiCTR1800017204; date of registration: 07/18/2018.

Overexpressed VDAC1 in breast cancer as a novel prognostic biomarker and correlates with immune infiltrates.

BACKGROUND: More and more evidence suggests that cancer is a mitochondrial metabolic disease recently and mitochondria dysfunction is critical to tumorigenesis. As a gatekeeper of mitochondria, the voltage-dependent anion channel 1 (VDAC1) is associated with the development of breast cancer (BC). However, its potential mechanism and clinical significance remain unclear; thus, in this research, we aimed to explore it. METHODS: VDAC1 expression in BC tissues and normal tissues was obtained from The Cancer Genome Atlas (TCGA) and validated by datasets from the gene expression omnibus (GEO) database. Then, the relationships between VDAC1 expression and clinicopathological features were analyzed. Receiver operating characteristics (ROC) curves were used to identify the diagnostic value of VDAC1. The prognostic value was evaluated by Kaplan-Meier survival curves and Cox regression analysis. VDAC1 with its co-expression genes were subjected to enrichment analysis to explore potential mechanisms in BC and the protein-protein interaction (PPI) network was constructed. At last, the association between VDAC1 expression and infiltration levels of immune cell infiltration by various methods, as well as their corresponding markers, was analyzed. We also analyzed the correction between VDAC1 expression and eight immune checkpoint genes and the tumor immune dysfunction and exclusion (TIDE) scores of each BC sample in TCGA were calculated and the differences between high and low VDAC1 expression groups were analyzed. RESULTS: VDAC1 expression was remarkably elevated in BC (p < 0.001), and high expression of VDAC1 was associated with the positive expression of ER (p = 0.004), PR (p = 0.033), and HER2 (p = 0.001). ROC analysis suggested that VDAC1 had diagnosed value in BC. The Kaplan-Meier analysis suggested that higher expression of VDAC1 was associated with shorter overall survival (OS), and further Cox regression analysis revealed that VDAC1 was an independent factor of unfavorable prognosis in BC patients. Enrichment analysis of VDAC1 and its co-expression suggested that VDAC1 was related to the regulation of mitochondrial energy metabolism and protein modification, and the HIF-1 singing pathway might be the potential mechanism in BC. Notably, we found that VDAC1 expression was infiltration levels of most types of immune cells, as well as the expression of marker genes of immune cells. The ICGs PDCD1, CTLA4, LAG3, SIGLEC15, and TIGIT were negatively corrected with VDAC1 expression in BC. TIDE scores between the low and high expression groups showed no difference. CONCLUSION: Overexpressed VDAC1 in BC could be severed as a novel biomarker for diagnosis and VDAC1 was an independent factor for adverse prognosis prediction. Our study revealed that VDAC1 might inhibit tumor immunity and might be a novel therapeutic target in BC.

Silencing of UCA1 Protects Against MPP+-Induced Cytotoxicity in SK-N-SH Cells via Modulating KCTD20 Expression by Sponging miR-423-5p.

Parkinson's disease (PD) is the second most prevalent neurodegenerative disorder. Long noncoding RNA (lncRNA) urothelial carcinoma-associated 1 (UCA1) has been implicated in PD development. Nevertheless, little insight has been gained on the mechanisms of UCA1 in PD pathogenesis. The levels of UCA1, miR-423-5p and potassium channel tetramerization domain containing 20 (KCTD20) were assessed by qRT-PCR and western blot. Cell viability was gauged by the CCK-8 assay, and cell apoptosis was detected by flow cytometry. Targeted relationships among UCA1, miR-423-5p and KCTD20 were verified by dual-luciferase reporter and RNA immunoprecipitation assays. Our data showed that MPP+ induced UCA1 expression in SK-N-SH cells. UCA1 silencing protected against MPP+-evoked cytotoxicity in SK-N-SH cells. UCA1 functioned as a miR-423-5p sponge, and the protective impact of UCA1 silencing on MPP+-evoked cytotoxicity was mediated by miR-423-5p. KCTD20 was a direct target of miR-423-5p, and miR-423-5p overexpression mitigated MPP+-triggered cell injury by down-regulating KCTD20. Furthermore, UCA1 regulated KCTD20 expression by acting as a sponge of miR-423-5p in SK-N-SH cells. Our study first identified that the silencing of UCA1 protected SK-N-SH cells from MPP+-evoked cytotoxicity at least in part by targeting the miR-423-5p/KCTD20 axis.

NONO promotes hepatocellular carcinoma progression by enhancing fatty acids biosynthesis through interacting with ACLY mRNA.

BACKGROUND: Dysregulation of fatty acid (FA) metabolism is involved in hepatocellular carcinoma (HCC) development. Non-POU domain-containing octamer binding protein (NONO), known as the component of nuclear paraspeckles, has recently been found to promote HCC progression. In this study, we investigated the functions of NONO in regulating de novo FA synthesis and its underling mechanism during HCC development. METHODS: The roles of NONO in HCC development by applying gene function loss analysis in HCC cells were detected by quantitative real-time polymerase chain reaction, cell proliferation, and cell invasion assays. The underlying mechanism of NONO in HCC development was examined by western blotting, subcellular fractionation, RNA-binding protein immunoprecipitation-sequencing, chromatin immunoprecipitation, co-immunoprecipitation and mass spectrometry. The effect of NONO on tumorigenesis in vivo was performed with a subcutaneous xenograft mouse model of HCC. RESULTS: NONO promotes HCC progression by interacting with and increasing ATP-citrate lyase (ACLY) mRNA to enhance FA biosynthesis. Furthermore, NONO promotes ACLY expression through enhancing nuclear ACLY mRNA stability in Diethylnitrosamine-stimulated HCC cells, not related to nuclear paraspeckles. Moreover, we find that NONO/SFPQ (Splicing factor proline and glutamine rich) heterodimer is essential for NONO interacting with ACLY mRNA in DEN stimulated HCC cells. In addition, NONO, Insulin like growth factor 2 mRNA binding protein 1 (IGF2BP1) and ACLY expressions contribute HCC development in mice and are related to poor survival. CONCLUSION: NONO promotes HCC progression by enhancing FA biosynthesis through interacting with ACLY mRNA and provide a novel potential target for HCC therapy.

What intervention regimen is most effective prevention for Portal venous system thrombosis after splenectomy in cirrhotics patients with Portal hypertension? Systematic review and network meta-analysis.

Portal venous system thrombosis (PVST) is a life-threatening complication after splenectomy in cirrhotics patients with portal hypertension, while the application of intervention regimen may prevent the incidence of PVST. The aim of this network meta-analysis was to determine the most appropriate intervention regimen and application time. Several electronic databases were searched up to December 2019. We estimated summary odds ratios (OR) using pairwise and network meta-analyses with random effects for the outcome of occurrence of PVST. This work was registered with PROSPERO (CRD42019161406). The analysis was based on 19 researches, which included 1853 patients. The results drawn from the data in standard meta-analysis indicated that the application of intervention was better than no intervention use, and early application of interventions was better than delayed application in preventing the occurrence of PVST. Subsequent network meta-analysis was performed to determine the most suitable intervention regimen used early post-operation. For separate mono-therapy drug, alprostadil, antithrombin III, low molecular dextran were significantly more efficacious than others. However, mono-therapy analysis was not so close to clinical application. In the follow-up network meta-analysis, low molecular dextran combined with low molecular weight heparin exhibited the largest effect on the preventing the incidence of PVST (0.12, 0.03-0.49), followed by antithrombin III (0.12, 0.04-0.41) with low molecular dextran (0.14, 0.05-0.41). We could draw the conclusion that early application of low molecular weight heparin combined with low molecular dextran seems to be the most satisfactory treatment to prevent the incidence of PVST for patients with cirrhotic portal hypertension after splenectomy.

Adverse clinical events caused by pacemaker battery depletion: two case reports.

BACKGROUND: The clinical symptoms and adverse events caused by pacemaker battery depletion are not uncommon, but it is easy to miss or misdiagnose them clinically. To raise the level of awareness towards this clinical situation, we report two cases. CASE PRESENTATION: We described two cases of pacemaker battery depletion. Case 1 was an 83-year-old male manifesting chest pain and dyspnea. Automatic reprogramming after pacemaker battery depletion resulted in pacemaker syndrome. While case 2 was an 80-year-old female with complete atrioventricular heart block and torsade de pointes, due to complete depletion of pacemaker battery. In addition, we introduce a method that can easily identify the depletion of the pacemaker battery, which has clinical promotion value of a certain degree. CONCLUSIONS: Those cases emphasize that serious morbidity can arise from pacemaker battery depletion, even in the early stages. Therefore, early detection and diagnosis is especially important.

Effects of ticagrelor on the pharmacokinetics of rivaroxaban in rats.

CONTEXT: Rivaroxaban and ticagrelor are two common drugs for the treatment of atrial fibrillation and acute coronary syndrome. However, the drug-drug interaction between them is still unknown. OBJECTIVE: To investigate the effects of ticagrelor on the pharmacokinetics of rivaroxaban in rats both in vivo and in vitro. MATERIALS AND METHODS: A sensitive and reliable UPLC-MS/MS method was developed for the determination of rivaroxaban in rat plasma. Ten Sprague-Dawley rats were randomly divided into ticagrelor pre-treated group (10 mg/kg/day for 14 days) and control group. The pharmacokinetics of orally administered rivaroxaban (10 mg/kg, single dose) with or without ticagrelor pre-treatment was investigated with developed UPLC-MS/MS method. Additionally, Sprague-Dawley rat liver microsomes were also used to investigate the drug-drug interaction between these two drugs in vitro. RESULTS: The C max (221.34 ± 53.33 vs. 691.18 ± 238.31 ng/mL) and the AUC(0-t) (1060.97 ± 291.21 vs. 3483.03 ± 753.83 µg·h/L) of rivaroxaban increased significantly (p < 0.05) with ticagrelor pre-treatment. The MRT(0-∞) of rivaroxaban increased from 4.41 ± 0.79 to 5.97 ± 1.11 h, while the intrinsic clearance decreased from 9.93 ± 2.55 to 2.89 ± 0.63 L/h/kg (both p < 0.05) after pre-treated with ticagrelor. Enzyme kinetic study indicated that ticagrelor decreased rivaroxaban metabolic clearance with the IC50 value of 14.04 µmol/L. CONCLUSIONS: Our in vivo and in vitro results demonstrated that there is a drug-drug interaction between ticagrelor and rivaroxaban in rats. Further studies need to be carried out to verify whether similar interactions truly apply in humans and whether these interactions have clinical significance.

Cysteinyl leukotriene receptor type 1 antagonist montelukast protects against injury of blood-brain barrier.

The blood-brain barrier (BBB) is formed by tightly connected cerebrovascular endothelial cells. Injury of human brain endothelial cells can cause disruption of the BBB and severe injury to brain tissue. Signals mediated cysteinyl leukotrienes (cysLTs) and their receptors are involved in a variety of pathological conditions. In the current study, our results show that oxygen glucose-deprivation/reoxygenation (OGD/R) induced the expression of leukotriene receptor type 1 (cysLT1R) in brain endothelial cells. Blockage of cysLT1R by its specific antagonist montelukast suppressed OGD/R-induced altered permeability of the human brain endothelial cell (EC) monolayer. Mechanistically, montelukast treatment reversed OGD/R-induced reduction of the tight junction proteins occludin and zonula occludens-1 (ZO-1). Montelukast also ameliorated OGD/R-induced reduction of inhibitors of matrix metalloproteinases (TIMPs), such as TIMP-1 and TIMP-2. On the other hand, montelukast suppressed the expression and production of matrix metalloproteinases (MMPs) and cytokines including MMP-2, MMP-9, interleukin 1ß (IL-1ß), tumor necrosis factor-α (TNF-α), and interleukin 6 (IL-6). Using a murine middle cerebral artery occlusion brain injury model, we demonstrated that the administration of montelukast improved the surgery-induced brain injury and protected against disruption of brain endothelial junction proteins such as occludin and ZO-1. Collectively, our data suggest that montelukast might confer protective roles against injury in brain endothelial cells.

Long noncoding RNA PVT1 inhibits interferon-α mediated therapy for hepatocellular carcinoma cells by interacting with signal transducer and activator of transcription 1.

Long noncoding RNA (LncRNA) PVT1 has recently been reported to be involved in the development of hepatocellular carcinoma (HCC) and hsigh expression of oncogenic PVT1 is associated with poor prognosis of HCC. Interferon-α (IFN-α) has been used in clinic for HCC therapy. However, whether PVT1 is involved in the IFN-α therapy for HCC is completely unknown. Our study found that high PVT1 expression in HCC cells is associated with high unmethylation in PVT1 promoter region. IFN-α treatment further increases PVT1 expression in HCC cells by enhancing H3K4me3 modification on the promoter. Furthermore, PVT1 knockdown enhances IFN-α-induced HCC cell apoptosis by promoting phosphorylation of signal transducer and activator of transcription 1 (STAT1) and upregulating IFN-stimulated genes expression. Moreover, PVT1 specifically interacts with STAT1 in HCC cells. Taken together, these results for the first time indicate that IFN-α treatment promotes oncogenic PVT1 expression in HCC cells, which interacts with STAT1 to inhibit IFN-α signaling, ultimately blocking IFN-α-induced cells apoptosis, suggesting that lncRNA PVT1 may be a potential target to improve IFN-α-mediated HCC immunotherapies.

Abnormal brain white matter in patients with right trigeminal neuralgia: a diffusion tensor imaging study.

BACKGROUND: Idiopathic or classical trigeminal neuralgia (TN) is a chronic painful condition characterized by intermittent pain attacks. Enough evidence demonstrates classical TN is related to neurovascular compression (NVC) at the trigeminal root entry zone (REZ), but white matter change secondary to TN are not totally known. METHODS: Visual Analogue Scale (VAS) and diffusion tensor imaging were performed on 29 patients with right TN and 35 healthy individuals. Voxel-wise analyses were performed with TBSS using multiple diffusion metrics, including fractional anisotropy (FA), mean diffusivity (MD), axial diffusivity (AD) and radial diffusivity (RD). Group differences in these parameters were compared between right TN patients and controls using TBSS and correlations between the white matter change and disease duration and VAS in right TN patients were assessed. Multiple comparison correction were applied to test significant correlations. RESULTS: The right TN patients showed significantly lower FA and higher RD in most left white matter (P < 0.05, FWE corrected). Moreover, negative correlations were observed between disease duration and the FA values of left corona radiata, genu of corpus callosum, left external capsule and left cerebral peduncle, and between VAS and the FA values of left corona radiata, left external capsule and left cerebral peduncle (P < 0.05). Positive correlations were observed for disease duration and the RD values of left corona radiata, right external capsule, left fornix cerebri and left cerebral peduncle, and for VAS and the RD values of left corona radiata and left external capsule (P < 0.05). However, once Bonferroni corrections were applied, these correlations were not statistically significant. CONCLUSION: These findings suggest that TN selectively impairs widespread white matter, especially contralateral hemisphere, which may be the hallmark of disease severity in TN patients.

Epigenetic silencing of the Wnt antagonist APCDD1 by promoter DNA hyper-methylation contributes to osteosarcoma cell invasion and metastasis.

Osteosarcoma (OS) is the most common type of bone tumor in children and adults. However, the molecular mechanism underlying OS tumorigenesis remains unclear. Here, we report that the expression of APCDD1, a Wnt antagonist, was reduced in OS tissues and cells compared to adjacent normal tissue and osteoblast cells, respectively. Mechanistically, this was due to increased levels of methylation in the promoter region of the APCDD1 gene. Consistently, the DNA methyltransferase inhibitor 5-AZA-dC, reduced DNA methylation in the APCDD1 promoter, and restored APCDD1 expression in OS tissue and cells. Moreover, DNMT3a, but not DNMT1 or DNMT3b, was the major DNA methyltransferase that facilitated hyper-methylation of DNA in the APCDD1 promoter, thus reducing APCDD1 mRNA levels in OS tissues. Importantly, ectopic expression of APCDD1 suppressed activity of the Wnt/ß-Catenin signaling pathway in OS cells and inhibited their invasion and reversed their EMT-like properties, while depletion of APCDD1 promoted invasion and metastasis of osteosarcoma cells in vitro and in vivo. Thus, we have provided the first evidence that APCDD1 expression is epigenetically silenced in OS, which may facilitate invasion and metastasis of OS cells.

The p16/miR-217/EGR1 pathway modulates age-related tenogenic differentiation in tendon stem/progenitor cells.

Previous studies have shown that the differentiation potential declines with the age of progenitor cells and is linked to altered levels of senescence markers. The purpose of this study was to test whether senescence marker p16 affects age-related tenogenic differentiation in tendon stem/progenitor cells (TSPCs). Young and aged TSPCs were isolated from young/healthy and aged/degenerated human Achilles tendons, respectively. Cellular aging and capacity for tenogenic differentiation were examined. The results showed that the tenogenic differentiation capacity of TSPCs significantly decreases with advancing age. TSPCs from elderly donors showed upregulation of senescence-associated ß-galactosidase and p16 and concurrently a decrease in Type I collagen concentration and in the expressions of tendon-related markers: Scx, Tnmd, Bgn, Dcn, Col1, and Col3. Overexpression of p16 significantly inhibited tenogenic differentiation of young TSPCs. Analysis of the mechanism revealed that this effect is mediated by microRNA-217 and its target EGR1. These results indicated that p16 inhibits tenogenic differentiation of TSPCs via microRNA signaling pathways, which may serve as a potential target for the prevention or treatment in the future.

Enhanced tenogenic differentiation and tendon-like tissue formation by CHIP overexpression in tendon-derived stem cells.

The carboxyl terminus of Hsc70-interacting protein (CHIP, also known as STUB1) plays critical roles in the proliferation and differentiation of many types of cells. The potential function of CHIP in tendon-derived stem cells (TDSCs) remains largely unknown at present. Here, we investigated the effects of CHIP on tenogenic differentiation of TDSCs via lentivirus-mediated overexpression. Forced expression of CHIP induced morphological changes and significantly enhanced cell proliferation, as well as tendon differentiation in vitro. Upon stimulation with differentiation induction medium, CHIP-overexpressing TDSCs displayed significant inhibition of differentiation into osteogenic and adipogenic lineages. Subsequent implantation of TDSCs overexpressing CHIP with collagen sponges into nude mice induced a marked increase in ectopic tendon formation in vivo, compared with the control group. Our findings collectively suggest that CHIP is an important contributory factor to tenogenic tissue formation.

EGR1 induces tenogenic differentiation of tendon stem cells and promotes rabbit rotator cuff repair.

BACKGROUND/AIMS: The rate of healing failure after surgical repair of chronic rotator cuff tears is considerably high. The aim of this study was to investigate the function of the zinc finger transcription factor early growth response 1 (EGR1) in the differentiation of tendon stem cells (TSCs) and in tendon formation, healing, and tendon tear repair using an animal model of rotator cuff repair. METHODS: Tenocyte, adipocyte, osteocyte, and chondrocyte differentiation as well as the expression of related genes were determined in EGR1-overexpressing TSCs (EGR1-TSCs) using tissue-specific staining, immunofluorescence staining, quantitative PCR, and western blotting. A rabbit rotator cuff repair model was established, and TSCs and EGR1-TSCs in a fibrin glue carrier were applied onto repair sites. The rabbits were sacrificed 8 weeks after repair operation, and tissues were histologically evaluated and tenocyte-related gene expression was determined. RESULTS: EGR1 induced tenogenic differentiation of TSCs and inhibited non-tenocyte differentiation of TSCs. Furthermore, EGR1 promoted tendon repair in a rabbit model of rotator cuff injury. The BMP12/Smad1/5/8 signaling pathway was involved in EGR1-induced tenogenic differentiation and rotator cuff tendon repair. CONCLUSION: EGR1 plays a key role in tendon formation, healing, and repair through BMP12/Smad1/5/8 pathway. EGR1-TSCs is a promising treatment for rotator cuff tendon repair surgeries.

CTGF positively regulates BMP12 induced tenogenic differentiation of tendon stem cells and signaling.

BACKGROUND/AIMS: Disordered differentiation of tendon stem cells (TSCs) during repair of injured tendon can result in the pathogenesis of chronic tendinopathy. Understanding tenocyte differentiation may provide new therapeutic insights for the prevention and treatment of chronic tendinopathy. The aim of our study was to determine if CTGF exerts a similar effect on BMP12-driven differentiation of rat TSCs. METHODS: In overexpressing and RNA interference CTGF TSCs, tenogenic differentitation and the expression of related genes were determined by immunofluorescence staining, quantitative PCR, and western blotting, with or without BMP12 treatment. The interaction in vitro between CTGF and BMP12 was detected by Chemical crosslinking assay. RESULTS: Our results showed that BMP12 effectively increased the expression of the tenocyte lineage markers scleraxis (Scx) and tenomodulin (Tnmd) at both mRNA and protein levels. Over-expression of CTGF from a lentiviral vector increased the expression of Scx and Tnmd as well as tendon proteins type I collagen (ColI) and tenascin-C (Tn-C) in TSCs compared to non-treated control cells with or without simultaneous BMP12 stimulation. Knockdown of CTGF expression decreased the expression of Scx, Tnmd, ColI and Tn-C compared to control cells. Chemical crosslinking experiments demonstrated a direct interaction between CTGF and BMP12. CONCLUSION: In conclusion, BMP12 plays a crucial role in tenogenesis via the Smad1/5/8 pathway, and CTGF positively promotes this effect.

The role of Pin1 protein in aging of human tendon stem/progenitor cells.

Aging of tendon stem/progenitor cells (TSPCs) can lead to tissue degeneration and subsequent injury. However, the molecular mechanisms controlling TSPC aging are not completely understood. In the present study, we investigated the role of Pin1 in aging of human TSPCs. Pin1 mRNA and protein expression levels were significantly decreased during prolonged in vitro culture of human TSPCs. Furthermore, overexpression of Pin1 delayed the progression of cellular senescence, as confirmed by downregulation of senescence-associated ß-galactosidase, increased telomerase activity and decreased levels of the senescence marker, p16(INK4A). Conversely, Pin1 siRNA transfection promoted senescence in TSPCs. In addition, miR-140-5p regulated Pin1 expression at the translational level via directly targeting its 3'UTR. Our results collectively demonstrate that Pin1 acts as an important regulator of TSPC aging.