The intracellular domain of UNC5B facilities proliferation and metastasis of bladder cancer cells.

The intracellular domain of UNC5B contains both death domain and caspase-3 cleavage site, and is regarded as a functional domain that mediates apoptosis. However, in our previous studies, we found that the death domain of UNC5B in bladder cancer cells could not be activated to promote apoptosis. In this study, different UNC5B truncates (residue 399-945, residue 412-945) were created to explore whether the caspase-3 cleavage site (site 412), as another potential functional domain of its intracellular portion, could be activated to induce apoptosis in bladder cancer cells. Using mass spectrometry, we acquired a comprehensive and detailed identification of differentially expressed proteins by overexpressing UNC5B and its truncates. Protein-protein-interaction (PPI) network analysis was also applied to investigate the aggregation of related proteins and predict the functional changes. EDU assay, apoptosis, xenograft tumour implantation, migration, invasion and tumour metastasis were performed to comprehensively identify the effects of UNC5B truncates on bladder cancer cells. We demonstrate that the intracellular domain of UNC5B promotes cell proliferation in vitro and tumour formation in vivo, by binding to a large number of ribosomal proteins. The overexpression of intracellular domain also facilitates cells to migrate, invade and metastasize by interacting with fibronectin, beta-catenin and vimentin. In addition, we reveal that overexpressing the intracellular domain of UNC5B cannot bind or activate cleaved caspase-3 to trigger apoptosis in bladder cancer cells.

UNC5B mediates G2/M phase arrest of bladder cancer cells by binding to CDC14A and P53.

UNC5B is a known tumor suppressor gene in a variety of cancers. As a transmembrane protein, UNC5B also induces apoptosis in a P53-dependent manner. In this study, we demonstrate that UNC5B inhibits proliferation through G2/M phase arrest by mass spectrometry and bioinformatics analysis in bladder cancer cells. By combing with CDC14A and P53, UNC5B dephosphorylated P53 at Ser-315 site. This dephosphorylation facilitated G2/M phase arrest by reducing the expression of cyclin B1 and increasing the expression of p-CDK1, thus inhibiting tumor proliferation. Knockdown of CDC14A suppressed the G2/M phase arrest induced by UNC5B in vitro, and eliminated the inhibitory effect of UNC5B on tumor proliferation in vivo. Our results show that UNC5B-mediated cell cycle arrest may act as a potential treatment for bladder cancer.

Functional and morphological alterations of the urinary bladder in type 2 diabetic FVB(db/db) mice.

AIMS: Diabetic bladder dysfunction (DBD) has been extensively studied in animal models of type 1 diabetes. We aimed to examine the functional and morphological alterations of the urinary bladder in a type 2 diabetes model, FVB(db/db) mice. METHODS: FVB(db/db) mice and age-matched FVB/NJ control mice were tested at either 12, 24 or 52weeks of age. Body weight, blood glucose and glycated hemoglobin (HbA1c) levels were measured. Bladder function was assessed by measurement of 24-h urination behavior and conscious cystometry. Bladder was harvested for Masson's Trichrome staining and morphometric analysis. RESULTS: The body weights of FVB(db/db) mice were twice as those of FVB/NJ control mice. The blood glucose and HbA1c levels were higher in FVB(db/db) mice at 12 and 24weeks, but not at 52weeks. A significant increase in the mean volume per void, but decrease in the voiding frequency, in FVB(db/db) mice was observed. Cystometry evaluation showed increased bladder capacity, voided volume, and peak micturition pressure in FVB(db/db) mice compared with FVB/NJ mice. Morphometric analysis revealed a significant increase in the areas of detrusor muscle and urothelium in FVB(db/db) mice. In addition, some FVB(db/db) mice, especially males at 12 and 24weeks, showed small-volume voiding during 24-h urination behavior measurement, and detrusor overactivity in the cystometry measurement. CONCLUSIONS: The FVB(db/db) mouse, displaying DBD characterized by not only increased bladder capacity, void volume, and micturition pressure, but also bladder overactivity, is a useful model to further investigate the mechanisms of type 2 diabetes-related bladder dysfunction.

Bladder function in mice with inducible smooth muscle-specific deletion of the manganese superoxide dismutase gene.

Manganese superoxide dismutase (MnSOD) is considered a critical component of the antioxidant systems that protect against oxidative damage. We are interested in the role of oxidative stress in bladder detrusor smooth muscle (SM) in different disease states. In this study, we generated an inducible, SM-specific Sod2(-/-) mouse model to investigate the effects of MnSOD depletion on the function of the bladder. We crossbred floxed Sod2 (Sod2(lox/lox)) mice with mice containing heterozygous knock-in of a gene encoding a tamoxifen-activated Cre recombinase in the SM22α promoter locus [SM-CreER(T2)(ki)(Cre/+)]. We obtained Sod2(lox/lox),SM-CreER(T2)(ki)(Cre/+) mice and injected 8-wk-old males with 4-hydroxytamoxifen to induce Cre-mediated excision of the floxed Sod2 allele. Twelve weeks later, SM-specific deletion of Sod2 and depletion of MnSOD were confirmed by polymerase chain reaction, immunoblotting, and immunohistochemistry. SM-specific Sod2(-/-) mice exhibited normal growth with no gross abnormalities. A significant increase in nitrotyrosine concentration was found in bladder SM tissue of SM-specific Sod2(-/-) mice compared with both wild-type mice and Sod2(+/+), SM-CreER(T2)(ki)(Cre/+) mice treated with 4-hydroxytamoxifen. Assessment of 24-h micturition in SM-specific Sod2(-/-) mice revealed significantly higher voiding frequency compared with both wild-type and SM-specific Cre controls. Conscious cystometry revealed significantly shorter intercontraction intervals and lower functional bladder capacity in SM-specific Sod2(-/-) mice compared with wild-type mice. This novel model can be used for exploring the mechanistic role of oxidative stress in organs rich in SM in different pathological conditions.

Short-term diabetes- and diuresis-induced alterations of the bladder are mostly reversible in rats.

OBJECTIVES: To determine whether diabetes mellitus- and diuresis-induced alterations in the bladder can be reversed in rats. METHODS: Male Sprague-Dawley rats were randomly distributed into eight groups (n = 16 per group): 3 weeks and 11 weeks age-matched controls, 3 weeks and 11 weeks after streptozotocin-induced diabetes mellitus, 3 weeks after diabetes mellitus induction then treated with insulin for 8 weeks, 3 weeks and 11 weeks after 5% sucrose-induced diuresis, and 3 weeks after 5% sucrose-induced diuresis followed by removal of 5% sucrose for 8 weeks. Bodyweight, blood glucose and glycated hemoglobin A1c were monitored. At the designated time-points, 24-h urinary habits were examined, and cystometry was carried out in half of the animals. The bladders from the remaining animals were harvested for histological examination, and quantification of smooth muscle, urothelium and collagen components. RESULTS: Insulin treatment reversed hyperglycemia and polyuria in diabetic animals successfully, which was shown by normalization of blood glucose, glycated hemoglobin A1c and 24-h urinary habits. Subsequently, bodyweight, bladder weight and percentage change of bladder components (smooth muscle, collagen, urothelium) in total bladder cross-sectional area were reversed to almost normal levels, and the bladder dysfunction was mostly reversed by 8 weeks of glycemic control, seen in the cystometry study. Similar alterations and reversed effects were seen in diuretic rats without and with 5% sucrose removal, respectively. CONCLUSIONS: Short-term (3-week induction) diabetes- and polyuria-induced functional and morphological alterations of the bladder can mostly be reversed in rats.

Successful induction of stress urinary incontinence in mice by vaginal distension does not depend on the estrous cycle.

OBJECTIVE: To determine if the performance of vaginal distension (VD) in different estrous cycle phases affects the successful induction of stress urinary incontinence in mice. METHODS: Female virgin C57BL/6 mice were distributed into 4 groups according to their estrous cycle phase: proestrus, estrus, metestrus, or diestrus. The estrous cycle was staged by examining vaginal smears, and the method was corroborated by histologic examination of the vagina in a subset of each group. Each group was divided into 4 subgroups for measurement of 24-hour micturition behavior and the leak point pressure (LPP) 4 or 20 days after VD or sham VD. RESULTS: Voiding events increased and mean void volume decreased 4 days, but not 20 days after VD, and the LPP was decreased 4 days, but not 20 days after VD compared with the corresponding sham group, regardless of the estrous cycle phase when VD was performed. There were no differences in 24-hour micturition behavior or LPP among the 4 different estrous cycle phases either in sham or VD group. CONCLUSION: Successful induction of reversible stress urinary incontinence in mice is not dependent on the estrous cycle.

Roles of polyuria and hyperglycemia in bladder dysfunction in diabetes.

PURPOSE: Diabetes mellitus causes diabetic bladder dysfunction. We identified the pathogenic roles of polyuria and hyperglycemia in diabetic bladder dysfunction in rats. MATERIALS AND METHODS: A total of 72 female Sprague-Dawley® rats were divided into 6 groups, including age matched controls, and rats with sham urinary diversion, urinary diversion, streptozotocin induced diabetes mellitus after sham urinary diversion, streptozotocin induced diabetes mellitus after urinary diversion and 5% sucrose induced diuresis after sham urinary diversion. Urinary diversion was performed by ureterovaginostomy 10 days before diabetes mellitus induction. Animals were evaluated 20 weeks after diabetes mellitus or diuresis induction. We measured 24-hour drinking and voiding volumes, and cystometry. Bladders were harvested to quantify smooth muscle, urothelium and collagen. We measured nitrotyrosine and Mn superoxide dismutase in the bladder. RESULTS: Diabetes and diuresis caused increases in drinking and voiding volume, and bladder weight. Bladder weight decreased in the urinary diversion group and the urinary diversion plus diabetes group. The intercontractile interval, voided volume and compliance increased in the diuresis and diabetes groups, decreased in the urinary diversion group and further decreased in the urinary diversion plus diabetes group. Total cross-sectional tissue, smooth muscle and urothelium areas increased in the diuresis and diabetes groups, and decreased in the urinary diversion and urinary diversion plus diabetes groups. As a percent of total tissue area, collagen decreased in the diuresis and diabetes groups, and increased in the urinary diversion and urinary diversion plus diabetes groups. Smooth muscle and urothelium decreased in the urinary diversion and urinary diversion plus diabetes groups. Nitrotyrosine and Mn superoxide dismutase increased in rats with diabetes and urinary diversion plus diabetes. CONCLUSIONS: Polyuria induced bladder hypertrophy, while hyperglycemia induced substantial oxidative stress in the bladder, which may have a pathogenic role in late stage diabetic bladder dysfunction.

[A case report of pharynx leiomyosarcoma].

The patient was hospitalized for foreign body sensation at pharynx persisted for two months. The patient noticed that the left pharyngeal palate was elevated two months ago and was treated as pharyngitis without success. Because of no eating difficulty, loss of voice or sore throat, the patient did not seek for further treatment. Recently the foreign body sensation at pharynx was worsened and the elevation at left pharyngeal palate enlarged and affected pronunciation and speech. A bulge about 4 cm x 5 cm in size was found around left soft palate and tonsil, relatively hard in texture; The mucosal membrane of the bulge was intact with slight hyperemia. The bulge was not movable and exhibited no tenderness to touch and no bleeding to press. Left tonsil was swelling of degree I degrees, grayish white and unsmooth on the surface. Uvula was deviated slight to the right. The soft palate movement was not satisfactory. No swelling on right tonsil. The CT indicated a soft tissue mass at left parapharyngeal space, about 3.4 cm x 5.4 cm in size. leiomyosarcoma (pharynx).

[Analyses of differential expression of Homeobox genes between lingual squamaous cell carcinoma and normal mucosa].

OBJECTIVE: To investigate the differently expressed Homeobox genes between lingual squmaous cell carcinoma and normal mucosa. METHODS: Seven paired specimens including lingual squmaous cell carcinoma and its surrounding normal tissue were obtained from 7 patients. Customized Oligo microarray which contains numerous probes of 232 human Homeobox genes was used to analyse the results. All datas were scanned by Agilent scanner and differentiately expressed genes were sorted out. RESULTS: Homeobox gene NANOG was found up-regulated in 5 samples. PHTF2 was found down-regulated in 7 samples, and CRX, PITX1, OTEX was found down-regulated in 5 samples. CONCLUSION: As the key gene to cellular proliferation and differentiation, Homeobox genes is closely releverant to the oncogenesis of lingual squmaous cell carcinoma.