Downregulation of Claudin5 promotes malignant progression and radioresistance through Beclin1-mediated autophagy in esophageal squamous cell carcinoma.

BACKGROUND: Esophageal squamous cell carcinoma (ESCC) is a highly prevalent and aggressive cancer with poor treatment outcomes. Despite the critical role of tight junction proteins in tumorigenesis, the involvement of Claudin5 in ESCC remains poorly understood. Thus, this study aimed to investigate the role of Claudin5 in ESCC malignant progression and radioresistance, as well as the underlying regulatory mechanisms. METHODS: The expression of Claudin5 was evaluated in esophageal cancer tissue using both public databases and 123 clinical samples. CCK-8, transwell invasion, wound healing and clonogenic survival assays were used to examine the proliferation, invasion, migration and radiosensitivity of ESCC cells in vitro. Xenograft and animal lung metastasis experiments were conducted to examine the impact of Claudin5 on tumor growth and lung metastasis in vivo. The effect of Claudin5 on autophagy was detected via transmission electron microscopy, western blotting and autophagy flux. Immunohistochemical staining was used to detect Claudin5 expression in ESCC patient samples. The statistical difference was assessed with Student t test or one-way ANOVA. The correlation between Claudin5 expression and radiotherapy response rate was performed by the Chi-square test. The significance of Kaplan-Meier curves was evaluated by the Logrank test. RESULTS: Claudin5 expression was downregulated in ESCC tissues. Downregulation of Claudin5 promoted ESCC cell proliferation, invasion, and migration both in vitro and in vivo. Downregulation of Claudin5 decreased the radiosensitivity of ESCC cells. Moreover, downregulation of Claudin5 promoted autophagy and the expression of Beclin1. Beclin1 knockdown reversed the effect of Claudin5 downregulation on autophagy induction and the promotion of ESCC cell malignant progression and radioresistance. Additionally, low expression of Claudin5 in ESCC cancer tissues was associated with poor radiotherapy response and prognosis. CONCLUSIONS: In summary, these findings suggest that downregulation of Claudin5 promotes ESCC malignant progression and radioresistance via Beclin1-autophagy activation and may serve as a promising biomarker for predicting radiotherapy response and patient outcome in ESCC.

Identification of a new way to induce differentiation of dermal fibroblasts into vascular endothelial cells.

BACKGROUND: Human dermal fibroblasts (HDFs) have the potential to differentiate into vascular endothelial cells (VECs), but their differentiation rate is low and the mechanism involved is not clear. The small molecule pathway controls the phenotype of fibroblasts by activating cellular signaling pathways, which is a more convenient method in the differentiation strategy of HDFs into VECs. METHODS: In this study, HDFs were treated with the different doses of CPP ((E)-4-(4-(4-(7-(diethylamino)-2-oxo-2H-chromene-3-carbonyl) piperazin-1-yl) styryl)-1-methylpyridin-1-ium iodide), and the mRNA and protein levels of HDFs were detected by qPCR, Western blot, flow cytometry and immunofluorescent staining. The matrigel assays, acetylated-LDL uptake and angiogenesis assays of chick embryo chorioallantoic membrane (CAM) and hindlimb ischemia model of nude mice were performed to evaluate the functions of VECs derived from HDFs. RESULTS: Here, we report that the small chemical molecule, CPP, can effectively induce HDFs to differentiate into VECs. First, we observed the morphological changes of HDFS treated with CPP. Flow cytometry, Western blot and qRT-PCR analyses showed that CPP effectively decreased the level of the HDFs-marker Vimentin and increased levels of the VEC-markers CD31, CD133, TEK, ERG, vWF, KDR and CDH5. Detection of the percentage of CD31-positive cells by immunofluorescent staining confirmed that CPP can effectively induce HDFs to differentiate into VECs. The results of Matrigel assays, DiI-ac-LDL uptake, angiogenesis assays on CAM and hindlimb ischemia model of nude mice showed that CPP-induced HDFs have the functions of VECs in vitro and in vivo. Western blot and qRT-PCR analysis showed that CPP induces HDFs to differentiate into VECs by promoting the expression of pro-angiogenic factors (VEGF, FGF-2 and PDGF-BB). CONCLUSIONS: Our data suggest that the small chemical molecule CPP efficiently induces the differentiation of HDFs into VECs. Simultaneously, this new inducer provides a potential to develop new approaches to restore vascular function for the treatment of ischemic vascular diseases.

Integrated Analysis of miRNAs and Gene Expression Profiles Reveals Potential Biomarkers for Osteoarthritis.

Purpose: Currently, the early diagnosis and treatment of osteoarthritis (OA) remain a challenge. In the present study, we attempted to explore potential biomarkers for the diagnosis and treatment of OA. Methods: The differentially expressed genes (DEGs) were identified based on three mRNA datasets of synovial tissues for OA patients and normal controls downloaded from the Gene Expression Omnibus (GEO) database. Furthermore, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were used for evaluating gene function related categories. Then, miRNA sequencing was performed for differentially expressed miRNAs' identification. Finally, weighted gene co-expression network analysis (WGCNA) was performed for genes detected by the three mRNA datasets and a competing endogenous RNA (ceRNA) network with DEGs and differentially expressed microRNAs (miRNAs) was constructed for central genes identification. In addition, the relationship between central gene expression and immune infiltration was analyzed, and the candidate agents for OA were predicted based on the Connectivity Map database. Quantitative RT-PCR (qRT-PCR), Western blotting analysis, and immunofluorescent staining were performed to validate the expression levels of differentially expressed miRNAs and differentially expressed target genes in normal and OA tissues and chondrocytes. MiRNA-mRNA network was also validated in chondrocytes in vitro. Results: A total of 259 DEGs and 26 differentially expressed miRNAs were identified, among which 94 miRNA-mRNA interactions were predicted. The brown module in WGCNA was most closely correlated with the clinical traits of OA. After overlapping the brown module genes with miRNA-mRNA pairs, 27 miRNA-mRNA pairs were obtained. A ceRNA network was constructed with 5505 lncRNA-miRNA-mRNA interactions. B-cell translocation gene 2(BTG2), Abelson-related gene (ABL2), and vascular endothelial growth factor A (VEGFA) were identified to be the central genes with good predictive performance, which were significantly correlated with immune cell infiltration in OA, reflected by declined activated dendritic cells (aDCs), and elevated contents of B cells, macrophages, neutrophils, and T helper cells. Anisomycin, MG-132, thapsigargin, and lycorine were predicted to be the potential candidate agents for OA intervention. In vitro, the expression levels of differentially expressed miRNAs and biomarkers identified in the present study were consistent with the results obtained in normal or OA knee cartilage tissues and chondrocytes. Furthermore, BTG2 was identified to be negatively regulated by miR-125a-5p. Conclusion: BTG2, ABL2, and VEGFA can be regarded as potential predictive and treatment biomarkers for OA, which might guide the clinical therapy of OA.

ZBM-H-induced activation of GRP78 ATPase promotes apoptosis via annexin A7 in A549 lung cancer cells.

Hypochlorous acid (HOCl) is an essential signal for the regulation of cancer cell fate, including autophagy and apoptosis. HOCl regulated autophagy by affecting the oxidation modification of glucose-regulated protein 78 (GRP78) and the activity of GRP78 ATPase. The mechanism of GRP78 ATPase in cell apoptosis has however not yet been clarified. Here we reported that ZBM-H, as a probe of HOCl, was able to directly bind to GRP78 in the presence or absence of ATP. Following ZBM-H treatment, the interaction between GRP78 and annexin A7 (ANXA7) was promoted, and this was accompanied by increased phosphorylation of integrin ß4 (ITGB4). In addition, ZBM-H enhanced the phosphorylation of ANXA7. ABO, an inhibitor of ANXA7, inhibited ZBM-H-induced ITGB4 phosphorylation and apoptosis, while ANXA7 activator SEC had opposite effect. Collectively, these data provide new evidence for the mechanism by which ZBM-H-induced activation of GRP78 ATPase regulates apoptosis of A549 lung cancer cells.

Locking Plate Alone or in Combination with Cannulated Screws for Hoffa Fractures: A Retrospective Study.

OBJECTIVE: To determine the efficacy of distal femur condyle locking plate (DFCLP) alone or in combination with cannulated screws for Hoffa fractures. METHODS: In this study, between May 2014 and February 2019, 13 patients between 26 and 64 years with isolated Hoffa fractures were enrolled during the study period and retrospectively analyzed. All patients underwent open reduction and internal fixation by DFCLP alone or in combination with cannulated screws followed by early active rehabilitation postoperatively. The primary outcome was evaluated using range of movement (ROM), Knee Society Score (KSS), International Knee Documentation Committee (IKDC) scoring system, and the fracture healing time of the patients during the 24-month follow-up period. Postoperative complications were also used to assess the patients' conditions. RESULTS: A total of 13 patients completed the 24-month follow-up assessment and achieved bone re-union at Hoffa fracture sites. The average follow-up period was 24.5 months (ranging from 24 to 28 months). Six patients were treated by DFCLP in combination with cannulated screws and the remaining seven patients were treated by DFCLP alone. The mean ROM was determined as 119° (ranging from 100° to 130°). The mean KSS score was 87.9 (ranging from 80 to 92 points), with 11 patients evaluated as excellent, two as good, and zero bad cases. The mean IKDC score was 84.2 (ranging from 74.7 to 89.7 points), with 10 evaluated as excellent, three as good, and zero bad cases. The mean IKDC score was 83.3 for patients with medial Hoffa fractures and 84.4 for those with lateral Hoffa fractures. The average time to healing was 3.5 months (ranging from 3 to 4 months), and at month 3, the fracture healing was evident in seven patients (54%), and at month 4, fracture healing was seen in six patients (46%). It is worth mentioning that two patients suffered from knee joint stiffness and osteoarthritis during the 24 months follow-up. Eleven patients (84.6%) achieved satisfactory knee joint function through early postoperative rehabilitation. CONCLUSION: In patients with Hoffa fractures, treatment with DFCLP alone or in combination with cannulated screws followed by early active rehabilitation resulted in great stability and satisfactory functional outcomes after 24 months. Our findings may provide surgeons with a new way to treat Hoffa fractures.

Closing-wedge distal femoral osteotomy combined with medial patellofemoral ligament reconstruction for recurrent patellar dislocation with genu valgum.

BACKGROUND: Medial patellofemoral ligament reconstruction (MPFLR) is a well-established procedure for addressing recurrent patellar dislocation (RPD) in young patients. However, despite being a promising procedure for RPD with genu valgum, there is a scarcity of reports on simultaneous MPFLR and closing-wedge distal femoral osteotomy (CWDFO). The purpose of the present study was to observe and analyse the clinical and imaging findings of CWDFO combined with MPFLR for RPD with genu valgum. METHODS: From May 2015 to April 2018, 25 patients with RPD and genu valgum were surgically treated in our department. Anteroposterior long-leg, weight-bearing, lower-extremity radiographs, lateral radiographs and computed tomography (CT) scans of the patellofemoral joint were obtained, and the anatomical femorotibial angle (aFTA), mechanical lateral distal femoral angle (mLDFA), weight-bearing line rate (WBLR), patellar height, patellar lateral shift (PLS) and tibial tubercle-trochlear groove (TT-TG) distance were analysed. Validated knee scores, such as the Kujala, Lysholm, visual analogue scale (VAS) scores and Tegner socres, were evaluated preoperatively and 2 years postoperatively. RESULTS: 25 patients, with an average age of 19.8 years (14-27), were evaluated. During the 2-year follow-up period, all patients were able to achieve a better sports level without any problems, with no recurrence of patellar instability. Compared with preoperation, the aFTA, mLDFA, WBLR and PLS showed statistically significant improvement following the procedure (p < 0.001). Meanwhile, no significant differences in the Insall index and TT-TG distance were found. The mean Kujala score, average Lysholm score, VAS score and Tegner socres showed significant postoperative improvement. CONCLUSIONS: CWDFO combined with MPFLR is a suitable treatment for RPD with genu valgum, and can lead to significant improvement in the clinical and imaging findings of the knee in the short term.

Effects of medial meniscal slope and medial posterior tibial slope on the locations of meniscal tears: A retrospective observational study.

The aim of this study was to analyze the relationship between medial posterior tibial slope (MPTS) and medial meniscus slope (MMS) with the location of meniscal lesions. We hypothesize that meniscuses with greater MPTS and MMS are more likely to have lesions in posterior horn.A total of 292 patients underwent arthroscopic surgery between January 2014 to September 2019 due to knee osteoarthritis (OA) and meniscal lesions were reviewed. Based upon the location of meniscal tears, patients were categorized as group B (tears in posterior horn) and group A (other sites). MPTS and MMS were measured from magnetic resonance imaging (MRI) slices. Osteoarthritis grade was evaluated in anteroposterior radiographs by the criteria defined by Kellgeren and Lawrence. Demographic data, OA grade, MPTS, and MMS for the 2 groups were compared and analyzed.The group A had 29 (39%) male and 45 (61%) female subjects with a mean age of 57.07 ±â€Š6.79 years. Group B consists of 74 (34%) male and 144 (66%) female subjects with a mean age of 58.90 ±â€Š7.594 years. (P = .067 and P = .458 for age and sex, respectively). In group A, 31 knees (42%) were determined to be Kellgren-Lawrence grade one, 32 knees (43%) grade two, and 11 knees (15%) grade three. In group B, 86 knees (39%) were categorized in grade one, 85 knees (39%) in grade two, and 47 knees (26%) in grade three (P = .085). The mean MPTS was 5.06 ±â€Š2.11 degree for group A and 6.15 ±â€Š2.37 degree for group B (P = .001). The mean MMS for group A was lower than group B (1.38 ±â€Š2.12 degree vs 3.14 ±â€Š2.92 degree; P < .000)This study demonstrated that increased MPTS and MMS may be considered as the risk factors for medial meniscal posterior horn tears.

Second-look arthroscopic findings after medial open-wedge high tibial osteotomy combined with all-inside repair of medial meniscus posterior root tears.

PURPOSE: To examine the results of medial open-wedge high tibial osteotomy (MOWHTO) combined with all-inside repair focusing on medial meniscus posterior root tears (MMPRT) by second-look arthroscopy and determine the clinical relevance of the findings. METHODS: From June 2015 to June 2017, 27 consecutive patients underwent MOWHTO and meniscal treatment for MMPRT using all-inside repair. All were available for second-look arthroscopy evaluation at the time of plate removal. Cartilage regeneration in the medial condyles and healing status of the MMPRT were assessed at the time of second-look arthroscopy. Clinical outcomes were evaluated by the Hospital for Special Surgery (HSS) scores and the Lysholm score scale. The potential factors affecting the healing of MMPRT, including age, body mass index (BMI), weight-bearing line rate (WBLR) and femorotibial angle (FTA) were discussed. RESULTS: There were 11 (41%) cases with complete healing (group A) and 16 (59%) cases with lax healing (group B). A complete cartilage coverage of chondral lesions was observed at second-look arthroscopy. Improvements of HSS in both groups were 41.25 ± 6.18 and 38.82 ± 5.43, while improvements in Lysholm score seen in both groups were 30.40 ± 4.84 and 32.65 ± 4.11. There was no significant difference in the postoperative clinical results between the two groups. In group A, the age (months), BMI, WBLR, and FTA are 54.61 ± 2.53, 23.58 ± 2.39, 66.10 ± 2.86% and 169.00 ± 2.62°, respectively, while in group B, they are 55.16 ± 4.92, 29.58 ± 3.91, 57.51 ± 5.49% and 175.21 ± 2.87°, respectively. A significant difference has been identified between two groups except for age. CONCLUSION: As seen in this retrospective study, a higher healing rate of MMPRT using all-inside repair and regeneration of degenerated articular cartilage in the medial condyles after MOWHTO can be expected. Healing of the MMPRT was not related to a better clinical outcome. Moreover, it can be inferred that BMI, WBLR and FTA may affect the healing status of MMPRT.

[Prognostic significance of neuroendocrine differentiation in non-small cell lung cancer patients' survival].

BACKGROUND: To observe the relationship between non-small cell lung cancer with neuroendocrine differentiation (NSCLC-NE) and patients' postoperative survival. METHODS: During April 1997 to April 1999, ninety-eight patients with lung cancer who were surgical treated were enrolled in this study. The tumor specimens were stained by NE markers: neuron specific enolase (NSE) and synaptophysin (SY). The NE marker reactions were divided as +, ++ and +++ scale groups. The same specimens were also examined by electron microscope for the specific neuroendocrine granule. All enrolled patients were followed up to 36 months, and the maximum follow-up time was 60 months. The Cox proportional hazard model multivariate analysis was applied to observe the relationship between the NSCLC-NE and the patients' postoperative survival. RESULTS: Ninety-one cases were diagnosed as NSCLC. In these patients, 63.7% (58/91) were positive for NE stain. Among them, 59.3% (54/91) were NSE-positive, and 24.1%(22/91) were SY-positive. The specific NE granules were found by the electron microscope examination in 33.0%(30/91). Combination of NE marker stain reaction and electronic microscope examination results, 48.4% (44/91) were considered as NSCLC-NE. The Cox proportional hazard model multivariate analysis showed that the NSCLC-NE patients' survival was significantly reduced (P=0.048). The following factors were related to NSCLC-NE patients' survival: lung cancer cell differentiation (P=0.006), stages of the cancer (P=0.001), and the NE markers reaction (P=0.054). CONCLUSIONS: NSCLC-NE is significantly related to the cancer cell differentiation and the patients' postoperative survival. The NE markers should be applied clinically as one of prognostic factors to evaluate the postoperative survival for NSCLC patients.