Tenascin-C promotes endochondral ossification and fracture healing through Hedgehog and Hippo signaling.

Fractures are frequent and severe musculoskeletal injuries. This study aimed to investigate the function of tenascin-C (TNC) in regulating chondrogenic during fracture healing and elucidate the underlying molecular mechanisms. A well-established femur fracture model in male C57BL/6J mice was used to transect the middle diaphysis of the femur. To identify the essential role of TNC, shTNC lentiviruses or TNC protein were administered in the animal model. Micro-CT analysis, histologic analysis, immunostaining assays, and gene expression analysis were employed to investigate the effect of TNC during fracture healing. An in vitro mesenchymal stem cell culture system was developed to investigate the role and molecular mechanism of TNC in regulating chondrogenesis. TNC expression was induced at the inflammatory phase and peaked at the cartilaginous callus phase during fracture healing. Knockdown of TNC expression in callus results in decreased callus formation and impaired fracture healing. Conversely, administration of exogenous TNC promoted chondrogenic differentiation, cartilage template formation and ultimately improved fracture healing. Both the Hedgehog and Hippo signaling pathways were found to be involved in the pro-chondrogenic function of TNC. Our observations demonstrate that TNC is a crucial factor responsible for endochondral ossification in fracture healing and provide a potential therapeutic strategy for promoting fracture healing.

Intermittent administration of PTH for the treatment of inflammatory bone loss does not enhance entheseal pathological new bone formation.

OBJECTIVE: To investigate the effect of intermittent parathyroid hormone (iPTH) administration on pathological new bone formation during treatment of ankylosing spondylitis-related osteoporosis. METHODS: Animal models with pathological bone formation caused by hypothetical AS pathogenesis received treatment with iPTH. We determined the effects of iPTH on bone loss and the formation of pathological new bone with micro-computed tomography (micro-CT) and histological examination. In addition, the tamoxifen-inducible conditional knockout mice (CAGGCre-ERTM; PTHflox/flox, PTH-/-) was established to delete PTH and investigate the effect of endogenous PTH on pathological new bone formation. RESULTS: iPTH treatment significantly improved trabecular bone mass in the modified collagen-induced arthritis (m-CIA) model and unbalanced mechanical loading models. Meanwhile, iPTH treatment did not enhance pathological new bone formation in all types of animal models. Endogenous PTH deficiency had no effects on pathological new bone formation in unbalanced mechanical loading models. CONCLUSION: Experimental animal models of AS treated with iPTH show improvement in trabecular bone density, but not entheseal pathological bone formation，indicating it may be a potential treatment for inflammatory bone loss does in AS.

Allicin attenuates calcium oxalate crystal deposition in the rat kidney by regulating gap junction function.

Renal calculus is a global common urological disease that is closely related to crystal adhesion and renal tubular epithelial cell impairment. Gap junctions (GJs) and their components (connexins and Cxs) are involved in various pathophysiology processes, but their roles in renal calculi progression are not well defined. Our previous RNA microarray analysis suggests that GJs are one of the key predicted pathways involved in the renal calcium oxalate (CaOx) crystal rat model. In the current study, we found that the Cx43 and Cx32 expression and the GJ function decreased significantly after stimulation with CaOx or sodium oxalate (NaOx) in NRK-52E, MDCK, and HK-2 cells, and Cx43 expression also decreased in renal tissues in renal CaOx crystal model rats. Inhibition of Cx43 in NRK-52E cells by small interference RNA significantly increased the CD44 and androgen receptor expression, and the adhesion between CaOx crystals and cells, which were consistent with the function of GJ inhibitors. On the other hand, after GJ function and Cx43 expression were increased by allicin, diallyl disulfide, or diallyl trisulfide, the impairment of NRK-52E cells by NaOx or other GJ inhibitors and the adhesion between CaOx crystals and renal cells decreased significantly. Furthermore, allicin also increased Cx43 expression and inhibited crystal deposition in rat kidneys. Taken together, our results provide a basis that GJs and Cx43 may participate in renal CaOx stone progression and that allicin, together with its analogues, could be potential drugs for renal calculus precaution.

Cell death-related molecules and biomarkers for renal cell carcinoma targeted therapy.

Renal cell carcinoma (RCC) is not sensitive to conventional radio- and chemotherapies and is at least partially resistant to impairments in cell death-related signaling pathways. The hallmarks of RCC formation include diverse signaling pathways, such as maintenance of proliferation, cell death resistance, angiogenesis induction, immune destruction avoidance, and DNA repair. RCC diagnosed during the early stage has the possibility of cure with surgery. For metastatic RCC (mRCC), molecular targeted therapy, especially antiangiogenic therapy (e.g., tyrosine kinase inhibitors, TKIs, such as sunitinib), is one of the main partially effective therapeutics. Various forms of cell death that may be associated with the resistance to targeted therapy because of the crosstalk between targeted therapy and cell death resistance pathways were originally defined and differentiated into apoptosis, necroptosis, pyroptosis, ferroptosis and autophagic cell death based on cellular morphology. Particularly, as a new form of cell death, T cell-induced cell death by immune checkpoint inhibitors expands the treatment options beyond the current targeted therapy. Here, we provide an overview of cell death-related molecules and biomarkers for the progression, prognosis and treatment of mRCC by targeted therapy, with a focus on apoptosis and T cell-induced cell death, as well as other forms of cell death.

Early and rapid prediction of postoperative infections following percutaneous nephrolithotomy in patients with complex kidney stones.

OBJECTIVES: To obtain more accurate and rapid predictors of postoperative infections following percutaneous nephrolithotomy (PCNL) in patients with complex kidney stones, and provide evidence for early prevention and treatment of postoperative infections. PATIENTS AND METHODS: A total of 802 patients with complex kidney stones who underwent PCNL, from September 2016 to September 2017, were recruited. Urine tests, urine cultures (UCs) and stone cultures (SCs) were performed, and the perioperative data were prospectively recorded. RESULTS: In all, 19 (2.4%) patients developed postoperative urosepsis. A multivariate logistic regression analysis revealed that an operating time of ≥100 min, urine test results with both positive urine white blood cells (WBC+) and positive urine nitrite (WBC+NIT+), positive UCs (UC+), and positive SCs (SC+) were independent risk factors of urosepsis. The incidence of postoperative urosepsis was higher in patients with WBC+NIT+ (10%) or patients with both UC+ and SC+ (UC+SC+; 8.3%) than in patients with negative urine test results or negative cultures (P < 0.01). Preoperative WBC+NIT+ was predictive of UC+SC+, with an accuracy of >90%. The main pathogens found in kidney stones were Escherichia coli (44%), Proteus mirabilis (14%) and Staphylococcus (7.4%); whilst the main pathogens found in urine were E. coli (54%), Enterococcus (9.4%) and P. mirabilis (7.6%). The incidence of E. coli was more frequent in the group with urosepsis than in the group without urosepsis (P < 0.05). CONCLUSIONS: WBC+NIT+ in preoperative urine tests could be considered as an early and rapid predictor of UC+SC+ and postoperative urosepsis. Urosepsis following PCNL was strongly associated with E. coli infections in patients with complex kidney stones.

Endourologic strategies for a minimally invasive management of urinary tract stones in patients with urinary diversion.

OBJECTIVE: To present our experience in minimally invasive management of urinary tract stones in patients with urinary diversion. MATERIALS AND METHODS: We retrospectively reviewed 26 patients with urinary tract stones after cystectomy and urinary diversion. The types of urinary diversion were ileal conduit, colon conduit, ileal orthotopic neobladder in 19, 4, and 3 patients, respectively. At postoperative days 2, a plain KUB and urinary ultrasonography were performed in order to assess stone fragmentation or hydronephrosis. According to postoperative imaging, stone free rate (SFR) was defined as complete absence of fragments or residual stones less than 4mm. RESULTS: 19 patients were treated with minimally invasive percutaneous lithotripsy (MPCNL) and 2 patients required second-look MPCNL. Anterograde flexible ureteroscopy was performed in 2 patients, while in 2 patients a combined anterograde and retrograde approach was required. Three reservoir stones were treated by transurethral neo-bladder lithotripsy. Postoperative significant complications occurred in 2 patients (7.7%). The highest percentage of stone composition was struvite, as a result of chronic urinary tract infection (UTI). SFR was 88.5% (23 of 26). CONCLUSIONS: Our experience showed that MPCNL is a safe and effective treatment modality with little morbidity for renal and upper ureteral stones in patients with urinary diversion. For middle and lower ureteral stones, an anterograde approach could be also considered as a first line treatment, but a combined anterograde and retrograde approach was required when the anterograde access alone cannot provide acceptable results.

Connexin43 is associated with the progression of clear cell renal carcinoma and is regulated by tangeretin to sygergize with tyrosine kinase inhibitors.

BACKGROUND: The roles of Connexin43 (Cx43) in clear cell renal cell carcinoma (ccRCC) microenviroment remains to be poorly defined. METHODS: The expression profile, prognosis and immune analysis of Cx43 in various cancers, particularly in ccRCC were performed using TCGA database, and various biological function assays were applied to explore the physiological role of Cx43 and tangeretin in ccRCC. Western blot were applied to examine the protein expression and Kunming mice were used to evaluate preliminary safety or anti-tumor activity of tangeretin and sunitinib. RESULTS: Compared with the normal group, higher expression levels of Cx43 in ccRCC, and distinct associations between Cx43 expression and ccRCC prognosis or immune infiltration, were found. Notably, the expression of Cx43 was found to be highly correlated with that of receptor tyrosine kinases (RTKs), particularly with VEGFR1, VEGFR2 and VEGFR3. The expression of Cx43 and EGFR was also found to be higher in ccRCC than that in the para-cancerous specimens. Knocking down Cx43 expression decreased RCC cell viability, cell migration, p-EGFR, MMP-9 and survivin expression. Using 14 Chinese medicine monomers, tangeretin was screened and found to inhibit tumor cell viability and Cx43 expression. Tangeretin also enhanced the sensitivity of RCC cells to tyrosine kinase inhibitors (TKIs) sunitinib and sorafenib. However, the same concentration of tangeretin exerted a less prominent effect on normal renal cell viability. CONCLUSIONS: Cx43 is strongly associated with RTK expression and ccRCC progression, while tangeretin can inhibit RCC cell malignancy by inhibiting Cx43 expression and enhance the sensitivity of RCC cells to TKIs.

CXCL12/CXCR4-Rac1-mediated migration of osteogenic precursor cells contributes to pathological new bone formation in ankylosing spondylitis.

Ankylosing spondylitis (AS) is a chronic inflammatory disease characterized by inflammatory back pain and spinal ankylosis due to pathological new bone formation. Here, we identified CXCL12 as a critical contributor to pathological new bone formation through recruitment of osteogenic precursor cells (OPCs). CXCL12 was found highly expressed in the regions that would potentially develop pathological new bone. OPCs were recruited to the regions where CXCL12 was up-regulated. Inhibition of CXCL12/CXCR4 axis with AMD3100 or conditional knockout of CXCR4 attenuated OPCs migration and subsequent pathological new bone formation in animal models of AS. By contrast, a genetically engineered animal model with CXCL12 overexpression developed a joint ankylosis phenotype. Furthermore, Rac1 was found essential for OPCs migration and pathological new bone formation. These findings ravel the novel role of CXCL12 in AS and indicate a potential strategy for targeting the CXCL12/CXCR4-Rac1 axis to prevent progression of axial skeleton ankylosis.

BRD4 promotes heterotopic ossification through upregulation of LncRNA MANCR.

AIMS: Acquired heterotopic ossification (HO) is a debilitating disease characterized by abnormal extraskeletal bone formation within soft-tissues after injury. The exact pathogenesis of HO remains unknown. It was reported that BRD4 may contribute to osteoblastic differentiation. The current study aims to determine the role of BRD4 in the pathogenesis of HO and whether it could be a potential target for HO therapy. METHODS: Achilles tendon puncture (ATP) mouse model was performed on ten-week-old male C57BL/6J mice. One week after ATP procedure, the mice were given different treatments (e.g. JQ1, shMancr). Achilles tendon samples were collected five weeks after treatment for RNA-seq and real-time quantitative polymerase chain reaction (RT-qPCR) analysis; the legs were removed for micro-CT imaging and subsequent histology. Human bone marrow mesenchymal stem cells (hBMSCs) were isolated and purified bone marrow collected during surgeries by using density gradient centrifugation. After a series of interventions such as knockdown or overexpressing BRD4, Alizarin red staining, RT-qPCR, and Western Blot (Runx2, alkaline phosphatase (ALP), Osx) were performed on hBMSCs. RESULTS: Overexpression of BRD4 enhanced while inhibition of Brd4 suppressed the osteogenic differentiation of hBMSCs in vitro. Overexpression of Brd4 increased the expression of mitotically associated long non-coding RNA (Mancr). Downregulation of Mancr suppressed the osteoinductive effect of BRD4. In vivo, inhibition of BRD4 by JQ1 significantly attenuated pathological bone formation in the ATP model (p = 0.001). CONCLUSION: BRD4 was found to be upregulated in HO and Brd4-Mancr-Runx2 signalling was involved in the modulation of new bone formation in HO. Cite this article: Bone Joint Res 2021;10(10):668-676.

The characteristic and relationship of Escherichia coli isolated from urine and stones in patients with calcium oxalate stones.

Previous clinical studies have shown that Escherichia coli (E. coli) predominated in urine and stone culture from calcium oxalate (CaOx) stone disease. The characteristic and relationship between E. coli isolated from urine cultures (EUC) and stone cultures (ESC) are compared. 83 E. coli (33 EUC and 50 ESC, respectively) from 66 CaOx stone patients were recruited in the study. E. coli in urine and stones from those patients were assessed by antimicrobial susceptibility test, genotyping and phylogenetic grouping. Furthermore, whole genome sequencing and comparative genomic analysis in paired ESC and EUC isolated strains from eight patients were carried out. The E. coli strains from ESC and EUC were not only multidrug resistant (MDR), but also had the similar pattern of resistant genes. The dominant phylogenetic group was B2, which was found in 54.0% of the ESC samples and 69.7% of the EUC samples, respectively. The virulence genes of E. coli, which isolated from stones and urine in the same patients, were highly homologous and largely consistent. Meanwhile, these E. coli strains were located in the same clade originated from a common ancestor. ESC and EUC isolated from patients with CaOx stones had a high prevalence of phylogenetic groups B2. Bacterial strains isolated from urine and stones in the same patient had consistent antimicrobial susceptibility profiles, genotyping, phylogenetic groups, virulence and resistance genes, also with high sequence co-linearity and close relationships.

Safety of Surgery in benign Prostatic Hyperplasia Patients on Antiplatelet or Anticoagulant Therapy: A Systematic Review and Meta-Analysis.

PURPOSE: The management strategies of anticoagulant (AC) or antiplatelet (AP) therapy in the preoperative period of benign prostatic hyperplasia (BPH) is still controversial. Therefore, a meta-analysis to systematically evaluate the surgical safety for BPH patients on AC or AP therapy was performed. MATERIALS AND METHODS: The protocol for the review is available on PROSPERO (CRD42018105800). A literature search was performed by using MEDLINE, Web of Science, PubMed, Cochrane library, and Embase. Summarized odds ratios (OR), mean difference (MD) and 95% confidence intervals (CI) were used to assess the difference in outcomes. RESULTS: We identified 13 trials with a total of 3767 patients. An intragroup significant difference was found in bleeding complications and blood transfusions when undergoing transurethral resection of the prostate (TURP). For laser surgery, the intragroup significant difference was found in the result of blood transfusion. Bridging therapy would not cause a higher risk of bleeding complications and blood transfusion during the perioperative period. Besides, no difference existed in operation time, catheterization time, hospitalization, and thromboembolic events. CONCLUSION: Patients with BPH on perioperative AC/AP therapy would have a risk of postoperative hemorrhage after TURP or laser treatments. To reduce the risk of hemorrhage, bridging therapy could be a good choice.

Comparison of Supracostal and Infracostal Access For Percutaneous Nephrolithotomy: A Systematic Review and Meta-Analysis.

PURPOSE: In this meta-analysis, we aimed to compared efficacy and safety of supracostal and infracostal access for percutaneous nephrolithotomy (PCNL). MATERIALS AND METHODS: We included eligible studies from PubMed, EMBASE, Cochrane Library, Web of Science and China National Knowledge Infrastructure. Literature searching, quality assessment and data extraction were performed by two independent reviewers. Data were analyzed by RevMan software. Binary and continuous variables were calculated as odds ratios (OR) and mean difference (MD). RESULTS: Two prospective comparative studies and seven retrospective observational studies were included in the meta-analysis, which contained 1,024 cases of supracostal access and 1,249 cases of infracostal access for PCNL. The supracostal access resulted in a significant reduced mean hemoglobin (95% CI: 0.26-3.46, MD = 1.86 g/L, P = .02) and a higher incidence of hydrothorax (95% CI: 4.77-22.95: OR = 10.47, P < .00001) compared to infracostal access. However, there no difference between supracostal and infracostal access regarding additional procedures (95% CI: 0.70-1.69, OR = 1.09, P = .71), stone-free rate (95% CI: 0.80-1.72, OR = 1.18, P = .41), length of hospital stay (95% CI: -0.03-0.37, MD = 0.17 day, P = .10), and occurrence of fever (95% CI: 0.95-2.03, OR = 1.39, P = .09) and blood transfusion (95% CI: 0.45-1.70, OR = 0.88, P = .70). No publication bias was identified in the study. CONCLUSION: Supracostal access was effective, but not as safe as infracostal access PCNL due to a higher risk of reduced hemoglobin and hydrothorax. Therefore, infracostal access should be the preferred safe and effective approach recommended for PCNL. When a supracostal puncture is performed, essential precautions to avoid hemoglobin loss and hydrothorax should be used.

LncRNA-miRNA-mRNA expression variation profile in the urine of calcium oxalate stone patients.

BACKGROUND: To explore long-non-coding RNA (lncRNA), microRNA (miRNA) and messenger RNA (mRNA) expression profiles and their biological functions in the urine samples in calcium oxalate (CaOx) patients. METHODS: Five CaOx kidney stone patients were recruited in CaOx stone group and six healthy people were included as control group, whose midstream morning urine was collected before the patients were given any medicine on admission. After total RNA was extracted from urine, microarray of miRNA, mRNA and lncRNA were applied to explore their expression variation. Gene ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were performed to reveal the gene functions of the dysregulated lncRNA-associated competing endogenous RNA (ceRNA) network. Quantitative real-time PCR were performed on HK-2 cells treated with sodium oxalate (NaOx) to further screen out the differentially expression profiles of these RNAs. RESULTS: A total of nine miRNAs, 883 mRNAs and 1002 lncRNAs were differentially expressed in urine of CaOx patients compared with normal population. GO analysis revealed that most of mRNAs from ceRNA network were enriched in terms of respiratory burst, regulation of mitophagy, and protein kinase regulator activity. KEGG pathway analysis of these genes related to ceRNA network highlight their critical role in pentose phosphate pathway, glyoxylate and dicarboxylate metabolism, and Janus kinase/signal transducer and activator of transcription (JAK-STAT) signaling pathway. Five miRNAs (miR-6796-3p, miR-30d-5p, miR-3192-3p, miR-518b and miR-6776-3p), four mRNAs (NT5E, CDH4, CLEC14A, CCNL1) and six lncRNAs (lnc-TIGD1L2-3, lnc-KIN-1, lnc-FAM72B-4, lnc-EVI5L-1, lnc-SERPINI1-2, lnc-MB-6) from the HK-2 cells induced by NaOx were consistent with the expression changes of microarray results. CONCLUSION: The differential expressed miRNAs, mRNAs and lncRNAs may be associated with numerous variations of the signaling pathways or regulation of metabolism and kinase activity, providing potential biomarkers for early diagnosis of urolithiasis and new basis for further research of urolithiasis mechanism.

Use of the ureteral access sheath during ureteroscopy: A systematic review and meta-analysis.

The debate still rages on for the usefulness of ureteral access sheath (UAS). Therefore, a meta-analysis to discuss the effects of applying UAS during ureteroscopy was performed. The protocol for the review is available on PROSPERO (CRD42017052327). A literature search was conducted up to November, 2017 using the Web of science, PUBMED, EMBASE and Cochrane Library. The quality of articles was assessed by the Jadad scale and Newcastle Ottawa Scale (NOS). Egger's test and the trim-and-fill method were used to evaluate publication bias. Effect sizes were calculated by pooled odds ratio (ORs) and mean differences (MDs). Sensitivity analyses and subgroup analyses were performed to explore the origin of heterogeneity. Eight trials with a total of 3099 patients and 3127 procedures were identified. Results showed no significant difference in stone-free rate (SFR) (OR = 0.83, 95% CI 0.52-1.33, P = 0.45), intraoperative complications (OR = 1.16, 95% CI 0.81-7.69, P = 0.88), operative time (MD = 4.09, 95% CI -15.08-23.26, P = 0.68) and hospitalization duration (MD = -0.13, 95% CI -0.32-0.06, P = 0.18). However, the incidence of postoperative complications was higher in UAS group (OR = 1.46, 95% CI 1.06-2.00, P = 0.02). Evidence from meta-analysis indicated that the use of UAS during ureteroscopy did not manifest advantages. However, given the intrinsic restrictions of the quality of selected articles, more randomized controlled trials (RCTs) are warranted to update the findings of this analysis.

Endourologic strategies for a minimally invasive management of urinary tract stones in patients with urinary diversion

ABSTRACT Objective To present our experience in minimally invasive management of urinary tract stones in patients with urinary diversion. Materials and Methods We retrospectively reviewed 26 patients with urinary tract stones after cystectomy and urinary diversion. The types of urinary diversion were ileal conduit, colon conduit, ileal orthotopic neobladder in 19, 4, and 3 patients, respectively. At postoperative days 2, a plain KUB and urinary ultrasonography were performed in order to assess stone fragmentation or hydronephrosis. According to postoperative imaging, stone free rate (SFR) was defined as complete absence of fragments or residual stones less than 4mm. Results 19 patients were treated with minimally invasive percutaneous lithotripsy (MPCNL) and 2 patients required second-look MPCNL. Anterograde flexible ureteroscopy was performed in 2 patients, while in 2 patients a combined anterograde and retrograde approach was required. Three reservoir stones were treated by transurethral neo-bladder lithotripsy. Postoperative significant complications occurred in 2 patients (7.7%). The highest percentage of stone composition was struvite, as a result of chronic urinary tract infection (UTI). SFR was 88.5% (23 of 26). Conclusions Our experience showed that MPCNL is a safe and effective treatment modality with little morbidity for renal and upper ureteral stones in patients with urinary diversion. For middle and lower ureteral stones, an anterograde approach could be also considered as a first line treatment, but a combined anterograde and retrograde approach was required when the anterograde access alone cannot provide acceptable results.