Dual roles of BK Polyomavirus in promoting urothelial carcinoma progression via regulating CLDN1.

Uncontrolled productive infection of BK polyomaviruses (BKV) in immunocompromised patients was reported to result in serious diseases, especially renourinary malignancies. However, the mechanism of BKV as a role of human carcinogen is still unknown. In this study, we showed that there is a significant association between BKV infection and metastasis of urothelial carcinoma (UCA). BKV-infected tumor tissues exhibit invasive histologic phenomena with vascular invasion and myometrial invasion. Then we identified that BKV promotes UCA invasion in a mode of dual regulation of tumor cells (TCs) invasion and endothelial cells (ECs) adhesion by encoding miRNAs. In cancer cells, BKV-B1-miR-5p promotes cell motility and invasiveness by directly targeting CLDN1. Moreover, exosomal-BKV-B1-miR-3p derived from BK-infected BC cells would be transferred to ECs and increase its adhesion to tumor cells by switching on the CLDN1 enhancer, which subsequently destroyed endothelial monolayers and increased permeability. In a human urothelial cancer metastasis mouse model, BK-inoculated cells exhibited higher incidence of vascular leakage and liver colonization. However, the vascular leakage and liver metastasis could be reduced when knocking down miRNAs in BK-inoculated cells. Our research delineates the bifunctional impact of BKV-encoded microRNAs on the expression of CLDN1 within both TCs and ECs, which orchestrates the establishment of a pre-metastatic niche in UCA.

Phenotypic and genetic characterization of hypervirulent Klebsiella pneumoniae in patients with liver abscess and ventilator-associated pneumonia.

Ventilator-associated pneumonia (VAP) and pyogenic liver abscess (PLA) due to Klebsiella pneumoniae infection can trigger life-threatening malignant consequences, however, there are few studies on the strain-associated clinical pathogenic mechanisms between VAP and PLA. A total of 266 patients consist of 129 VAP and 137 PLA were included for analysis in this study. We conducted a comprehensive survey for the two groups of K. pneumoniae isolates, including phenotypic experiments, clinical epidemiology, genomic analysis, and instrumental analysis, i.e., to obtain the genomic differential profile of K. pneumoniae strains responsible for two distinct infection outcomes. We found that PLA group had a propensity for specific underlying diseases, especially diabetes and cholelithiasis. The resistance level of VAP was significantly higher than that of PLA (78.57% vs. 36%, P < 0.001), while the virulence results were opposite. There were also some differences in key signaling pathways of biochemical processes between the two groups. The combination of iucA, rmpA, hypermucoviscous phenotype, and ST23 presented in K. pneumoniae infection is more important and highly prudent for timely treatment. The present study may contribute a benchmark for the K. pneumoniae clinical screening, epidemiological surveillance, and effective therapeutic strategies.

Immune escape of SARS-CoV-2 variants to therapeutic monoclonal antibodies: a system review and meta-analysis.

BACKGROUND: Omicron's high transmissibility and variability present new difficulties for COVID-19 vaccination prevention and therapy. In this article, we analyzed the sensitivity of vaccine-induced antibodies as well as the effect of booster vaccinations against Omicron sublineages. METHODS: We looked for Randomized Controlled Trials and cohort studies that reported the COVID-19 vaccines against Omicron sublineages up to 28 July 2022 through PubMed, the Cochrane Library, EMBASE, and Web of Science. Quantitative synthesis was carried out using Stata 16.0 and RevMa5.3, then the serum NT50 and antibody sensitivity to neutralize Omicron sublineages were assessed before and after booster vaccination. This study was registered with PROSPERO number CRD42022350477. RESULTS: This meta-analysis included 2138 patients from 20 studies, and the booster vaccination against Omicron sublineages showed a significant difference compared to 2 dosage: BA.1/BA.1.1 (SMD = 0.80, 95% CI: 0.75-0.85, P = 0.00), BA.2/BA.2.12.1 (SMD = 0.77, 95% CI: 0.69-0.85, P = 0.00), BA.3 (SMD = 0.91, 95% CI: 0.83-1.0, P = 0.00), and BA.4/5 (SMD = 0.77, 95% CI: 0.60-0.94, P = 0.00). The sensitivity of vaccines-induced antibodies decreased by at least 5-folds after booster vaccination, particularly in the case of BA.4/5 which had the most notable decline in vaccine effectiveness. CONCLUSION: After the booster vaccination, the NT50 and the neutralization ability of vaccine-induced antibodies increased, but the susceptibility of antibodies decreased compared with the control virus, which may be a clue for future Omicron sublineages prevention.

Characterization of Phage Resistance and Their Impacts on Bacterial Fitness in Pseudomonas aeruginosa.

The emergence and spread of antibiotic resistance pose serious environmental and health challenges. Attention has been drawn to phage therapy as an alternative approach to combat antibiotic resistance with immense potential. However, one of the obstacles to phage therapy is phage resistance, and it can be acquired through genetic mutations, followed by consequences of phenotypic variations. Therefore, understanding the mechanisms underlying phage-host interactions will provide us with greater detail on how to optimize phage therapy. In this study, three lytic phages (phipa2, phipa4, and phipa10) were isolated to investigate phage resistance and the potential fitness trade-offs in Pseudomonas aeruginosa. Specifically, in phage-resistant mutants phipa2-R and phipa4-R, mutations in conferring resistance occurred in genes pilT and pilB, both essential for type IV pili (T4P) biosynthesis. In the phage-resistant mutant phipa10-R, a large chromosomal deletion of ~294 kb, including the hmgA (homogentisate 1,2-dioxygenase) and galU (UTP-glucose-1-phosphate uridylyltransferase) genes, was observed and conferred phage phipa10 resistance. Further, we show examples of associated trade-offs in these phage-resistant mutations, e.g., impaired motility, reduced biofilm formation, and increased antibiotic susceptibility. Collectively, our study sheds light on resistance-mediated genetic mutations and their pleiotropic phenotypes, further emphasizing the impressive complexity and diversity of phage-host interactions and the challenges they pose when controlling bacterial diseases in this important pathogen. IMPORTANCE Battling phage resistance is one of the main challenges faced by phage therapy. To overcome this challenge, detailed information about the mechanisms of phage-host interactions is required to understand the bacterial evolutionary processes. In this study, we identified mutations in key steps of type IV pili (T4P) and O-antigen biosynthesis leading to phage resistance and provided new evidence on how phage predation contributed toward host phenotypes and fitness variations. Together, our results add further fundamental knowledge on phage-host interactions and how they regulate different aspects of Pseudomonas cell behaviors.

Low molecular weight heparin reduces arterial blood lactic acid content and increases estimated glomerular filtration rate in patients with moderate Covid-19 pneumonia.

BACKGROUND: Coronavirus disease 2019 (Covid-19) remains a serious health threat worldwide. We aimed to investigate whether low molecular weight heparin (LMWH) can promote organ function recovery in moderate Covid-19 pneumonia patients. METHODS: We initiated an LMWH protocol in Covid-19 patients with increased D-dimer, body mass index >30 kg/m2 or a history of diabetes from January 18, 2020 at Shanghai Public Health Clinical Center. In this retrospective study, we assigned moderate Covid- 19 pneumonia patients admitted between January 18th and April 18, 2020 receiving the LMWH protocol to the LMWH group. Moderate patients who met the inclusion criteria but did not receive LMWH protocol were included in the control group by 1:2 propensity score matching. General clinical information, indicators for renal function, arterial blood gas analyses, arterial blood lactic acid content (mmol/L), and coagulation indexes at 0 day, 3 days, 7 days, and 11 days after admission were recorded and compared between the two groups. RESULTS: There were 41 patients in the LMWH group and 82 patients in the control group. General information in both groups were similar. Compared to the control group, the arterial blood lactic acid content (mmol/L) at day 11 (1.3 [1.1, 1.7] vs. 1.2 [0.9, 1.3], P = 0.016) was reduced in the LMWH group. The estimated glomerular filtration rate (eGFR) in the LMWH group was higher than that in the control group at day 7 (108.54 [89.11, 128.17] vs. 116.85 [103.39, 133.47], P = 0.039) and day 11 (113.74 [94.49, 126.34] vs. 128.31 [112.75, 144, 12], P  = 0.003). The serum creatinine levels (Scr) in the LMWH group were lower than that in the control group at day 7 (62.13 [51.47, 77.64] vs. 55.49 [49.50, 65.75], P = 0.038) and day 11 (63.35 [50.17, 75.73] vs. 51.62 [44.62, 61.24], P = 0.005). CONCLUSIONS: LMWH treatment can reduce arterial blood lactic acid levels and improve eGFR in moderate Covid-19 pneumonia patients. Randomized controlled trials are warranted to further investigate this issue. TRIAL REGISTRATION: ChiCTR.org.cn, ChiCTR2000034796.

Isolation and Characterization of Novel Phages Targeting Pathogenic Klebsiella pneumoniae.

Klebsiella pneumoniae is a dominant cause of community-acquired and nosocomial infections, specifically among immunocompromised individuals. The increasing occurrence of multidrug-resistant (MDR) isolates has significantly impacted the effectiveness of antimicrobial agents. As antibiotic resistance is becoming increasingly prevalent worldwide, the use of bacteriophages to treat pathogenic bacterial infections has recently gained attention. Elucidating the details of phage-bacteria interactions will provide insights into phage biology and the better development of phage therapy. In this study, a total of 22 K. pneumoniae isolates were assessed for their genetic and phenotypic relatedness by multi-locus sequence typing (MLST), endonuclease S1 nuclease pulsed-field gel electrophoresis (S1-PFGE), and in vitro antibiotic susceptibility testing. In addition, the beta-lactamase gene (blaKPC) was characterized to determine the spread and outbreak of K. pneumoniae carbapenemase (KPC)-producing enterobacterial pathogens. Using these ST11 carbapenem-resistant K. pneumoniae isolates, three phages (NL_ZS_1, NL_ZS_2, and NL_ZS_3) from the family of Podoviridae were isolated and characterized to evaluate the application of lytic phages against the MDR K. pneumoniae isolates. In vitro inhibition assays with three phages and K. pneumoniae strain ZS15 demonstrated the strong lytic potential of the phages, however, followed by the rapid growth of phage-resistant and phage-sensitive mutants, suggesting several anti-phage mechanisms had developed in the host populations. Together, this data adds more comprehensive knowledge to known phage biology and further emphasizes their complexity and future challenges to overcome prior to using phages for controlling this important MDR bacterium.

Interactions between the Prophage 919TP and Its Vibrio cholerae Host: Implications of gmd Mutation for Phage Resistance, Cell Auto-Aggregation, and Motility.

Prophage 919TP is widely distributed among Vibrio cholera and is induced to produce free φ919TP phage particles. However, the interactions between prophage φ919TP, the induced phage particle, and its host remain unknown. In particular, phage resistance mechanisms and potential fitness trade-offs, resulting from phage resistance, are unresolved. In this study, we examined a prophage 919TP-deleted variant of V. cholerae and its interaction with a modified lytic variant of the induced prophage (φ919TP cI-). Specifically, the phage-resistant mutant was isolated by challenging a prophage-deleted variant with lytic phage φ919TP cI-. Further, the comparative genomic analysis of wild-type and φ919TP cI--resistant mutant predicted that phage φ919TP cI- selects for phage-resistant mutants harboring a mutation in key steps of lipopolysaccharide (LPS) O-antigen biosynthesis, causing a single-base-pair deletion in gene gmd. Our study showed that the gmd-mediated O-antigen defect can cause pleiotropic phenotypes, e.g., cell autoaggregation and reduced swarming motility, emphasizing the role of phage-driven diversification in V. cholerae. The developed approach assists in the identification of genetic determinants of host specificity and is used to explore the molecular mechanism underlying phage-host interactions. Our findings contribute to the understanding of prophage-facilitated horizontal gene transfer and emphasize the potential for developing new strategies to optimize the use of phages in bacterial pathogen control.

Intracellular Low Iron Exerts Anti-BK Polyomavirus Effect by Inhibiting the Protein Synthesis of Exogenous Genes.

BK polyomavirus (BKPyV) is a small double-stranded DNA virus and ubiquitous human pathogen that particularly affects immunocompromised individuals. Antiviral therapy for BKPyV is urgently needed. Intracellular irons have an important role in many viral infections, yet its contribution to BKPyV and replication has not been explored. In this study, we explored the interaction between BKPyV infection and intracellular iron and the inhibitory effect of iron depletion on BKPyV infection. By creating a low-intracellular-iron environment, we demonstrated that the iron-chelating-induced iron depletion inhibits BKPyV infection in primary renal tubular epithelial cells (RPTECs) and urinary bladder cancer cells (TCCSUP cells). Iron depletion exerts an inhibitory effect after BKPyV enters the nucleus, which might be due to the inhibition of the protein synthesis of exogenous genes in iron-depleted cells. Further exploration of the target proteins of iron-regulating viral infection could potentially be used to develop new strategies for urgently needed anti-BKPyV therapies. IMPORTANCE BKPyV poses a serious threat to the health of immunocompromised patients, and there are currently no curative drugs. Understanding the relationship between the virus and intracellular environment contributes to the discovery of antiviral targets. We demonstrate here that BKPyV is inhibited in cells with a low-iron environment. We also find that iron-chelating-induced iron depletion inhibits viral and exogenous protein synthesis. Further exploration of the target proteins of iron regulation could have great potential in developing new drugs against BKPyV and other viruses.

BK polyomavirus infection promotes growth and aggressiveness in bladder cancer.

BACKGROUND: Recent studies have confirmed the integration of the BK polyomavirus (BKPyV) gene into the cellular genome of urothelial carcinomas in transplant recipients, further confirming the correlation between BKPyV and urothelial carcinomas after transplantation. However, the role BKPyV infections play in the biological function of bladder cancer remains unclear. METHODS: We developed a BKPyV-infected bladder cancer cell model and a mice tumor model to discuss the role of BKPyV infections. RESULTS: Our research proves that BKPyV infections promote the proliferation, invasion and migration of bladder cancer cells, while the activation of ß-catenin signaling pathway is one of its mediation mechanisms. CONCLUSIONS: We first described BKPyV infection promotes the proliferation, invasion and migration of bladder cancer. We verified the role of ß-catenin signaling pathway and Epithelial-Mesenchymal Transition effect in BKPyV-infected bladder cancer. These results provide meaningful information towards the diagnosis and treatment of clinical bladder cancer.

High-mobility group box 1 protein antagonizes the immunosuppressive capacity and therapeutic effect of mesenchymal stem cells in acute kidney injury.

BACKGROUND: Kidney ischemia reperfusion injury (IRI) is a common cause of acute kidney injury and an unavoidable consequence of kidney transplantation and still lacks specific therapeutics. Recently, mesenchymal stem cell (MSC) has been emerging as a promising cell-based therapy for IRI in the context of transplantation. MSC negatively regulates the secretion of pro-inflammatory as well as the activation of immune cells during IRI through its unique immunosuppressive property. METHODS: We employed mice kidney IRI model and MSC cell line to monitor the IRI related checkpoints. siRNAs were utilized to knock down the potential key factors for mechanistic analysis. Statistical analysis was performed by using one-way ANOVA with Tukey's post hoc procedure by SPSS. RESULTS: The expression of high-mobility group box 1 protein (HMGB1) is increased in the acute phase as well as the recovery stage of IRI. Importantly, the HMGB1 upregulation is correlated with the injury severity. HMGB1 diminishes the MSC induced immunosuppressive capacity in the presence of pro-inflammatory cytokines in vitro. Toll like receptor 4 (TLR4)-mediated inducible nitric oxide synthase (iNOS) inhibition contributes to the negative effect of HMGB1 on MSCs. HMGB1-TLR4 signaling inhibition augments the therapeutic efficacy of MSCs in mice renal IRI model. CONCLUSIONS: These findings demonstrate that HMGB1 plays a crucial role in shaping the immunoregulatory property of MSCs within the microenvironments, providing novel insights into the crosstalk between MSCs and microenvironment components, suggesting HMGB1 signals as a promising target to improve MSC-based therapy.

Non-active antibiotic and bacteriophage synergism to successfully treat recurrent urinary tract infection caused by extensively drug-resistant Klebsiella pneumoniae.

We report a case of a 63-year-old female patient who developed a recurrent urinary tract infection (UTI) with extensively drug-resistant Klebsiella pneumoniae (ERKp). In the initial two rounds of phage therapy, phage resistant mutants developed within days. Although ERKp strains were completely resistant to sulfamethoxazole-trimethoprim, the combination of sulfamethoxazole-trimethoprim with the phage cocktail inhibited the emergence of phage resistant mutant in vitro, and the UTI of patient was successfully cured by this combination. Thus, we propose that non-active antibiotic and bacteriophage synergism (NABS) might be an alternative strategy in personalized phage therapy.

Clinical progression of patients with COVID-19 in Shanghai, China.

BACKGROUND: Studies on the 2019 novel coronavirus disease (COVID-19) have generally been limited to the description of the epidemiology and initial clinical characteristics. We investigated the temporal progression in patients with COVID-19. METHODS: In this retrospective, single-center study, we included confirmed cases of COVID-19 from Jan 20 to Feb 6, 2020 in Shanghai. Final date of follow-up was February 25, 2020. RESULTS: Of the 249 patients enrolled, the median age was 51 years old, and 126 (50.6%) were male. The duration from onset of symptoms to hospitalization was 4(2-7) days in symptomatic patients. Fever was occurred in 235(94.3%) patients. A total of 215 (86.3%) patients had been discharged after 16(12-20) days hospitalization. The estimated median duration of fever in all the patients with fever was 10 days (95 confidential intervals [CIs]: 8-11 days) after onset of symptoms. Patients who were transferred to intensive care units (ICU) had significantly longer duration of fever as compared to those not in ICU (31 days v.s. 9 days after onset of symptoms, respectively, P <0.0001). Radiological aggravation of initial image was observed in 163 (65.7%) patients on day 7 after onset of symptoms. 154(94.5%) of these patients showed radiological improvement on day 14. The median duration to negative reverse-transcriptase PCR tests of upper respiratory tract samples was 11 days (95 CIs: 10-12 days). Viral clearance was more likely to be delayed in patients in ICU than those not in ICU (P <0.0001). In multivariate logistical analysis, age (Odds ratio [OR] = 1.06) and CD4 T cell count (OR = 0.55 per 100 cells/ul increase) were independently associated with ICU admission. CONCLUSIONS: The majority of COVID-19 cases are mild. The clinical progression pattern suggests that early control of viral replication and application of host-directed therapy in later stage is essential to improve the prognosis of CVOID-19.

Mast cell infiltration associated with loss of interstitial cells of Cajal and neuronal degeneration in achalasia.

BACKGROUND: Achalasia is a motility disorder of unknown etiology. Previous studies supported the hypothesis that autoimmune-mediated inflammatory responses produce inhibitory neuronal degeneration. This study was designed to explore the role of mast cells in achalasia. METHODS: We collected information from 116 patients with achalasia who underwent peroral endoscopic myotomy between December 2016 and May 2017. Lower esophageal sphincter (LES) muscle biopsy was performed in all patients with achalasia, as well as 20 control subjects. The number of mast cells, interstitial cells of Cajal (ICCs), nNOS-positive cells, and S-100-positive cells in the LES were evaluated by immunohistochemistry. Pathological and clinical data were compared between groups. KEY RESULTS: Compared with controls, the LES of patients with achalasia had significantly fewer ICCs, nNOS-positive cells, and S-100-positive cells and a higher number of mast cells (all P < 0.001). Furthermore, the increased mast cell infiltration was significantly associated with decreased ICCs, nNOS-positive cells, and S-100-positive cells in patients with achalasia (all P < 0.05). Clinically, the number of strongly positive mast cells was highest in patients with type I achalasia and lowest in those with type III achalasia (P < 0.001). In addition, patients with a history of autoimmune disease or viral infection had greater mast cell infiltration in the LES muscle (P = 0.040). CONCLUSIONS & INFERENCES: In patients with achalasia, mast cell infiltration in the LES muscle is increased, in association with loss of ICCs and neuronal degeneration. Mast cells may thereby play a crucial role in the development of achalasia.

Transplantation of Telocytes Attenuates Unilateral Ureter Obstruction-Induced Renal Fibrosis in Rats.

BACKGROUND/AIMS: Previous studies imply that telocytes may have a protective effect on fibrosis in various organs, including the liver, colon, and heart. The effect of telocytes on renal fibrosis remains unknown. Herein, this study was designed to investigate the effect of telocytes on renal fibrosis and the potential mechanisms involved. METHODS: In a unilateral ureteral obstruction (UUO)-induced renal fibrosis model, telocytes were injected via the tail vein every other day for 10 days. The degree of renal damage and fibrosis was determined using histological assessment. The expression of collagen I, fibronectin, epithelial-mesenchymal transition markers, and Smad2/3 phosphorylation was examined by western blot analyses. Real-time PCR and enzyme-linked immunosorbent assay were performed in vivo to detect the levels of transforming growth factor (TGF)-ß1 and various growth factors. RESULTS: Telocytes attenuated renal fibrosis, as evidenced by reduced interstitial collagen accumulation, decreased expression of fibronectin and collagen I, upregulation of E-cadherin, and downregulation of α-smooth muscle actin. Furthermore, telocytes decreased serum TGF-ß1 levels, suppressed Smad2/3 phosphorylation, and increased the expression of hepatocyte growth factor (HGF) in rat kidney tissue following UUO. Blockage of HGF counteracted the protective effect of telocytes on UUO-treated kidneys. Through the detection of HGF mRNA levels in vitro, we found that telocytes had no effect on HGF expression compared with renal fibroblasts. CONCLUSION: Telocytes attenuated UUO-induced renal fibrosis in rats, likely through enhancing the expression of HGF in an indirect manner.

Protective effects of cyclic helix B peptide on aristolochic acid induced acute kidney injury.

BACKGROUND: Aristolochic acid (AA) injuries remain a serious condition associated with acute renal dysfunction. Herein, the effect and mechanism of a novel tissue protective peptide, cyclic helical B-peptide (CHBP) derived from erythropoietin, were investigated in a mice model. METHODS: Mice were randomly divided into four groups, receiving the following treatments (1: saline; 2: AA 10mg/kg; 3: AA 10mg/kg +CHBP 4nmol/kg; 4: AA 10mg/kg +CHBP 8nmol/kg). RESULTS: Blood urea nitrogen and serum creatinine was increased by AA but decreased by CHBP in a dose-dependent fashion. CHBP also significantly improved renal tubular injury and inflammatory infiltration, which was gradually increased by AA. Apoptotic cells, infiltrating inflammatory cells, and active caspase-3+ cells were greatly reduced by CHBP. In addition, CHBP inhibited caspase-3, 9 and improved bcl-2, bcl-xl protein expression in vivo. CONCLUSION: Taken together, we demonstrated, for the first time, that CHBP effectively improved renal function and tissue damage caused by AA, which maybe through reducing caspase-3 activation, apoptosis, and inflammation.

Differential microRNA expression in aristolochic acid-induced upper urothelial tract cancers ex vivo.

Aristolochic acid (AA) is a carcinogenic, mutagenic and nephrotoxic compound commonly isolated from members of the plant family of Aristolochiaceae (such as Aristolochia and Asarum) and used in Chinese herbal medicine. Use of AA and AA­containing plants causes chronic kidney disease (CKD) and upper urinary tract carcinoma (UUC); however, the underlying mechanism remains to be defined. miRNAs regulate a number of biological processes, including cell proliferation, differentiation and metabolism. This study explored differentially expressed miRNAs between AA­induced upper urothelial tract cancer (AAN­UUC) and non­AAN­UUC tissues. Patients with AAN­UUC and non­AAN­UUC (n=20/group) were recruited in the present study. Five tissue samples from each group were used for miRNA microarray profiling and the rest of the tissue samples were subjected to reverse transcription-quantitative polymerase chain reaction analysis including seven selected miRNAs for confirmation. A total of 29 miRNAs were differentially expressed between AAN­UUC and non­AAN­UUC tissues (P<0.05). TargenScan and Gene ontology analyses predicted the functions and targeted genes of these differentially expressed miRNAs, i.e. Akt3, FGFR3, PSEN1, VEGFa and AR. Subsequently, expression of the selected differentially expressed miRNAs (Hsa­miR­4795­5p, Hsa­miR­488, Hsa­miR­4784, Hsa­miR­330, Hsa­miR­3916, Hsa­miR­4274 and Hsa­miR­181c) was validated in another set of tissue samples. A total of 29 miRNAs were identified to be differentially expressed between AAN­UUC and non­AAN­UUC tissues and these miRNA target genes in FGFR3 and Akt pathways, which regulate cell growth and tumor progression, respectively.

Downregulation of homeodomain-interacting protein kinase-2 contributes to bladder cancer metastasis by regulating Wnt signaling.

Homeodomain-interacting protein kinase-2 (Hipk2) has been shown to have important regulatory roles in cancer biology, such as cancer cell proliferation, cell cycle, and cell invasion. However, the contributions of Hipk2 to bladder cancer metastasis remain largely unknown. In the current study, we assayed the expression level of Hipk2 in bladder cancer tissues by real-time PCR, and defined its biological functions. We found that Hipk2 levels were downregulated in most bladder cancer tissues compared with adjacent normal tissues, and Hipk2 levels were remarkably decreased in metastasized tumor tissues when compared with primary tumors. SiRNA-mediated Hipk2 silencing increased bladder cancer cell invasion. Hipk2 knockdown resulted in decrease of E-cadherin expression and increase of N-cadherin and fibronectin expression, indicated that epithelial-mesenchymal transition (EMT) was induced. We further demonstrated that Hipk2 knockdown induced Wnt signaling activation and ß-catenin nuclear localization. Finally, we confirmed that Hipk2 inhibition promoted EMT and subsequent cell invasion, at least in part by activating Wnt signaling. These data suggest an important role of Hipk2 in regulating metastasis of bladder cancer and implicate the potential application of Hipk2 in bladder cancer therapy.

Impact of HLA antibodies on graft survival in long-term renal recipients with functional grafts.

OBJECTIVES: Accumulating evidence supports the hypothesis that HLA antibodies play an important role in early renal allograft injury. However, the effects of HLA antibodies on long-term graft survival are still poorly understood. In this study, we examined the impact of HLA antibodies on graft survival in long-term renal recipients with functional grafts for 10 years. MATERIAL AND METHODS: In this retrospective study, long-term renal recipients were defined as kidney transplant recipients who had normally functioning renal grafts (serum creatinine level <2.0 mg/dl) for 10 years. Posttransplant serum samples from a total of 92 long-term renal allograft recipients on cyclosporine-based triple maintenance drug therapy (121.6 ± 0.886 months) were screened for the specificities of anti-HLA antibodies. The results of HLA antibodies before the transplantation, assessed using the same test method, were compared with those after their transplantations. Moreover, these 92 patients who received cadaveric renal transplant between January 2000 and December 2002 were followed up for about 10 years (range 107-135 months). RESULTS: 27 patients had HLA-I antibodies and 16 patients had HLA-II antibodies before the transplantation, whereas 12 patients had HLA-I antibodies and 18 patients had HLA-II antibodies after the transplantation. Moreover, the types of HLA antibodies were different from those found before the transplantation. In these renal recipients with functioning renal grafts, the estimated glomerular filtration rate was 66.52 ± 14.52 ml/min/1.73 m(2) in the HLA antibody-positive group (n = 23) and 69.09 ± 25.54 ml/min/1.73 m(2) negative in the HLA antibody-negative group (n = 69, p > 0.05). Three patients (3.26%) (3 out of 92) had donor-specific anti-HLA antibodies (DSA). The frequency of DSA in this study was lower than that in the general Chinese Han renal recipient population. CONCLUSIONS: We find that all HLA antibodies in the long-term renal grafts are newly formed after the transplantation. The HLA antibody status has little impact on the renal graft function in the long-term renal recipients.

Downregulation of GAS5 promotes bladder cancer cell proliferation, partly by regulating CDK6.

UNLABELLED: Long non-coding RNAs (lncRNAs) play important roles in diverse biological processes, such as transcriptional regulation, cell growth and tumorigenesis. However, little is known about whether lncRNA-GAS5 (growth arrest-specific 5) regulates bladder cancer progression. In the present study, we found that the GAS5 expression is commonly downregulated in bladder cancer cell lines and human specimens. Knockdown of GAS5 promotes bladder cancer cell proliferation, whereas forced expression of GAS5 suppresses cell proliferation. We further demonstrated that knockdown of GAS5 increases CDK6 mRNA and protein levels in bladder cancer cells. Expectedly, GAS5 inhibition induces a significant decrease in G0/G1 phase and an obvious increase in S phase. Gain-of-function and loss-of-function studies showed that GAS5 inhibits bladder cancer cell proliferation, at least in part, by regulating CDK6 expression. CONCLUSIONS: Downregulated GAS5 promotes bladder cancer cell proliferation, partly by regulating CDK6, and thus may be helpful in the development of effective treatment strategies against bladder cancer.

Knockdown of regulator of cullins-1 (ROC1) expression induces bladder cancer cell cycle arrest at the G2 phase and senescence.

Regulator of Cullins-1 (ROC1) is a key subunit in the Cullin-RING ligase (CRL) protein complex. Overexpression of ROC1 protein is associated with tumor progression and poor prognosis of non-muscle invasive bladder transitional cell carcinoma (NMIBC). This study was designed to assess the effects of ROC1 knockdown in bladder cancer cells and to determine the potential mechanisms involved. A total of 112 bladder cancer tissue specimens were recruited for immunohistochemical analyses of ROC1 overexpression. Bladder cancer cell lines were used to knockdown ROC1 expression using ROC1 siRNA. Our data showed that ROC1 knockdown remarkably inhibited bladder cancer cell growth, arrested cells at the G2 phase of the cell cycle, and induced the p53-dependent cell senescence. Molecularly, G2 arrest was associated with upregulation of p21, p27, cyclin B1, and Cdc2 proteins. ROC1 knockdown induced-senescence functioned through p53/p21 pathway. Knockdown of p21 expression partially rescued ROC1 knockdown-induced growth inhibition in cancer cells. Furthermore, nude mouse xenograft analyses confirmed these in vitro data. In conclusion, data from the current study indicate that ROC1 plays an essential role in bladder cancer progression and could serve as a novel anticancer target for bladder transitional cell carcinoma (BTCC).