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Introduction: The clinical-histologic correlation in diabetic nephropathy is not completely known. Methods: We analyzed nephrectomy specimens from 90 patients with diabetes and diverse degrees of proteinuria and glomerular filtration rate (GFR). Results: Thirty-six (40%) subjects had normoalbuminuria, 33 (37%) microalbuminuria, and 21 (23%) non-nephrotic proteinuria. Mean estimated GFR (eGFR) was 65±23 (40% <60 ml/min per 1.73 m2). About 170 glomeruli per patient were analyzed, and all samples included vascular tissue. Six subjects (7%) were classified in diabetic nephropathy class I, 61 (68%) in class II-a, 13 (14%) in class II-b, 9 (10%) class III, and 1 (1%) in class IV. Eighty percent to 90% of those with normoalbuminuria or microalbuminuria were classified in class II-a or II-b and <10% in class III; 52% of those with proteinuria were in class II-a, 15% in class II-b, and 19% in class III. Nodular sclerosis (57%) and mesangial expansion (15%) were more frequent in cases with proteinuria than in normoalbuminuria (28% and 8%; P = 0.028 and 0.017). About 20% to 30% of all cases, regardless the level of albuminuria or proteinuria or the histologic class had tubular atrophy, interstitial fibrosis, or inflammation in >10% to 20% of the sample. Moderate hyalinosis and arteriolar sclerosis were observed in 80% to 100% of cases with normoalbuminuria, microalbuminuria, proteinuria, as well as in class I, II, or III. Conclusions: Weak correspondence between analytical parameters and kidney histology was found. Thus, disease may progress undetected from the early clinical stages of the disease. Finally, vascular damage was a very common finding, which highlights the role of ischemic intrarenal disease in diabetes.

Introduction: The clinical-histologic correlation in diabetic nephropathy is not completely known. Methods: We analyzed nephrectomy specimens from 90 patients with diabetes and diverse degrees of proteinuria and glomerular filtration rate (GFR). Results: Thirty-six (40%) subjects had normoalbuminuria, 33 (37%) microalbuminuria, and 21 (23%) non-nephrotic proteinuria. Mean estimated GFR (eGFR) was 65±23 (40% <60 ml/min per 1.73 m2). About 170 glomeruli per patient were analyzed, and all samples included vascular tissue. Six subjects (7%) were classified in diabetic nephropathy class I, 61 (68%) in class II-a, 13 (14%) in class II-b, 9 (10%) class III, and 1 (1%) in class IV. Eighty percent to 90% of those with normoalbuminuria or microalbuminuria were classified in class II-a or II-b and <10% in class III; 52% of those with proteinuria were in class II-a, 15% in class II-b, and 19% in class III. Nodular sclerosis (57%) and mesangial expansion (15%) were more frequent in cases with proteinuria than in normoalbuminuria (28% and 8%; P = 0.028 and 0.017). About 20% to 30% of all cases, regardless the level of albuminuria or proteinuria or the histologic class had tubular atrophy, interstitial fibrosis, or inflammation in >10% to 20% of the sample. Moderate hyalinosis and arteriolar sclerosis were observed in 80% to 100% of cases with normoalbuminuria, microalbuminuria, proteinuria, as well as in class I, II, or III. Conclusions: Weak correspondence between analytical parameters and kidney histology was found. Thus, disease may progress undetected from the early clinical stages of the disease. Finally, vascular damage was a very common finding, which highlights the role of ischemic intrarenal disease in diabetes.

The inhibitory effects demonstrated by activation of cannabinoid receptors (CB) on cancer proliferation and migration may also play critical roles in controlling bladder cancer (BC). CB expression on human normal and BC specimens was tested by immunohistochemistry. Human BC cells RT4 and RT112 were challenged with CB agonists and assessed for proliferation, apoptosis, and motility. Cellular sphingolipids (SL) constitution and metabolism were evaluated after metabolic labelling. CB1-2 were detected in BC specimens, but only CB2 was more expressed in the tumour. Both cell lines expressed similar CB2. Exposure to CB2 agonists inhibited BC growth, down-modulated Akt, induced caspase 3-activation and modified SL metabolism. Baseline SL analysis in cell lines showed differences linked to unique migratory behaviours and cytoskeletal re-arrangements. CB2 activation changed the SL composition of more aggressive RT112 cells by reducing (p amp;lt; 0.01) Gb3 ganglioside (-50 +/- 3%) and sphingosine 1-phosphate (S1P, -40 +/- 4%), which ended up to reduction in cell motility (-46 +/- 5%) with inhibition of p-SRC. CB2-selective antagonists, gene silencing and an inhibitor of SL biosynthesis partially prevented CB2 agonist-induced effects on cell viability and motility. CB2 activation led to ceramide-mediated BC cell apoptosis independently of SL constitutive composition, which instead was modulated by CB2 agonists to reduce cell motility., Funding Agencies|Lions Club Milano ai Cenacoli

The inhibitory effects demonstrated by activation of cannabinoid receptors (CB) on cancer proliferation and migration may also play critical roles in controlling bladder cancer (BC). CB expression on human normal and BC specimens was tested by immunohistochemistry. Human BC cells RT4 and RT112 were challenged with CB agonists and assessed for proliferation, apoptosis, and motility. Cellular sphingolipids (SL) constitution and metabolism were evaluated after metabolic labelling. CB1-2 were detected in BC specimens, but only CB2 was more expressed in the tumour. Both cell lines expressed similar CB2. Exposure to CB2 agonists inhibited BC growth, down-modulated Akt, induced caspase 3-activation and modified SL metabolism. Baseline SL analysis in cell lines showed differences linked to unique migratory behaviours and cytoskeletal re-arrangements. CB2 activation changed the SL composition of more aggressive RT112 cells by reducing (p amp;lt; 0.01) Gb3 ganglioside (-50 +/- 3%) and sphingosine 1-phosphate (S1P, -40 +/- 4%), which ended up to reduction in cell motility (-46 +/- 5%) with inhibition of p-SRC. CB2-selective antagonists, gene silencing and an inhibitor of SL biosynthesis partially prevented CB2 agonist-induced effects on cell viability and motility. CB2 activation led to ceramide-mediated BC cell apoptosis independently of SL constitutive composition, which instead was modulated by CB2 agonists to reduce cell motility., Funding Agencies|Lions Club Milano ai Cenacoli

The inhibitory effects demonstrated by activation of cannabinoid receptors (CB) on cancer proliferation and migration may also play critical roles in controlling bladder cancer (BC). CB expression on human normal and BC specimens was tested by immunohistochemistry. Human BC cells RT4 and RT112 were challenged with CB agonists and assessed for proliferation, apoptosis, and motility. Cellular sphingolipids (SL) constitution and metabolism were evaluated after metabolic labelling. CB1-2 were detected in BC specimens, but only CB2 was more expressed in the tumour. Both cell lines expressed similar CB2. Exposure to CB2 agonists inhibited BC growth, down-modulated Akt, induced caspase 3-activation and modified SL metabolism. Baseline SL analysis in cell lines showed differences linked to unique migratory behaviours and cytoskeletal re-arrangements. CB2 activation changed the SL composition of more aggressive RT112 cells by reducing (p amp;lt; 0.01) Gb3 ganglioside (-50 +/- 3%) and sphingosine 1-phosphate (S1P, -40 +/- 4%), which ended up to reduction in cell motility (-46 +/- 5%) with inhibition of p-SRC. CB2-selective antagonists, gene silencing and an inhibitor of SL biosynthesis partially prevented CB2 agonist-induced effects on cell viability and motility. CB2 activation led to ceramide-mediated BC cell apoptosis independently of SL constitutive composition, which instead was modulated by CB2 agonists to reduce cell motility., Funding Agencies|Lions Club Milano ai Cenacoli

The inhibitory effects demonstrated by activation of cannabinoid receptors (CB) on cancer proliferation and migration may also play critical roles in controlling bladder cancer (BC). CB expression on human normal and BC specimens was tested by immunohistochemistry. Human BC cells RT4 and RT112 were challenged with CB agonists and assessed for proliferation, apoptosis, and motility. Cellular sphingolipids (SL) constitution and metabolism were evaluated after metabolic labelling. CB1-2 were detected in BC specimens, but only CB2 was more expressed in the tumour. Both cell lines expressed similar CB2. Exposure to CB2 agonists inhibited BC growth, down-modulated Akt, induced caspase 3-activation and modified SL metabolism. Baseline SL analysis in cell lines showed differences linked to unique migratory behaviours and cytoskeletal re-arrangements. CB2 activation changed the SL composition of more aggressive RT112 cells by reducing (p amp;lt; 0.01) Gb3 ganglioside (-50 +/- 3%) and sphingosine 1-phosphate (S1P, -40 +/- 4%), which ended up to reduction in cell motility (-46 +/- 5%) with inhibition of p-SRC. CB2-selective antagonists, gene silencing and an inhibitor of SL biosynthesis partially prevented CB2 agonist-induced effects on cell viability and motility. CB2 activation led to ceramide-mediated BC cell apoptosis independently of SL constitutive composition, which instead was modulated by CB2 agonists to reduce cell motility., Funding Agencies|Lions Club Milano ai Cenacoli

The inhibitory effects demonstrated by activation of cannabinoid receptors (CB) on cancer proliferation and migration may also play critical roles in controlling bladder cancer (BC). CB expression on human normal and BC specimens was tested by immunohistochemistry. Human BC cells RT4 and RT112 were challenged with CB agonists and assessed for proliferation, apoptosis, and motility. Cellular sphingolipids (SL) constitution and metabolism were evaluated after metabolic labelling. CB1-2 were detected in BC specimens, but only CB2 was more expressed in the tumour. Both cell lines expressed similar CB2. Exposure to CB2 agonists inhibited BC growth, down-modulated Akt, induced caspase 3-activation and modified SL metabolism. Baseline SL analysis in cell lines showed differences linked to unique migratory behaviours and cytoskeletal re-arrangements. CB2 activation changed the SL composition of more aggressive RT112 cells by reducing (p amp;lt; 0.01) Gb3 ganglioside (-50 +/- 3%) and sphingosine 1-phosphate (S1P, -40 +/- 4%), which ended up to reduction in cell motility (-46 +/- 5%) with inhibition of p-SRC. CB2-selective antagonists, gene silencing and an inhibitor of SL biosynthesis partially prevented CB2 agonist-induced effects on cell viability and motility. CB2 activation led to ceramide-mediated BC cell apoptosis independently of SL constitutive composition, which instead was modulated by CB2 agonists to reduce cell motility., Funding Agencies|Lions Club Milano ai Cenacoli

The transient receptor potential melastin 8 ion channel (TRPM8) is implicated in bladder sensing but limited information on TRPM8 antagonists in bladder overactivity is available. This study characterizes a new TRPM8-selective antagonist (DFL23448 [5-(2-ethyl-2H-tetrazol-5-yl)-2-(3-fluorophenyl)-1,3-thiazol-4-ol]) and evaluates it in cold-induced behavioral tests and tests on bladder function and experimental bladder overactivity in vivo in rats. DFL23448 displayed IC50 values of 10 and 21 nM in hTRPM8 human embryonic kidney 293 cells activated by Cooling Agent 10 or cold, but it had limited activity (IC50 > 10 mu M) at transient receptor potential vanilloids TRPV1, TRPA1, or TRPV4 or at various G protein-coupled receptors. In rats, DFL23448 administered intravenously or orally had a half-life of 37 minutes or 4.9 hours, respectively. DLF23448 (10 mg/kg i.v.) reduced icilin-induced "wet dog-like" shakes in rats. Intravesical DFL23448 (10 mg/l), but not vehicle, increased micturition intervals, micturition volume, and bladder capacity. During bladder overactivity by intravesical prostaglandin E-2 (PGE(2)), vehicle controls exhibited reductions in micturition intervals, micturition volumes, and bladder capacity by 37%-39%, whereas the same parameters only decreased by 12%-15% (P < 0.05-0.01 versus vehicle) in DFL23448-treated rats. In vehicle-treated rats, but not in DFL23448-treated rats, intravesical PGE(2) increased bladder pressures. Intravenous DFL23448 at 10 mg/kg, but not 1 mg/kg DFL23448 or vehicle, increased micturition intervals, micturition volumes, and bladder capacity. During bladder overactivity by intravesical PGE(2), micturition intervals, micturition volumes, and bladder capacity decreased in vehicle- and 1 mg/kg DFL23448-treated rats, but not in 10 mg/kg DFL23448-treated rats. Bladder pressures increased less in rats treated with DFL23448 10 mg/kg than in vehicle-or 1 mg/kg DFL23448-treated rats. DFL23448 (10 mg/kg i.v.), but not vehicl, Funding Agencies|Dompe Pharmaceuticals; Gester Foundation

Mesenchymal stem cells (MSC) as vehicles of therapeutic genes represent a unique tool to activate drugs within a neoplastic mass due to their property to home and engraft into tumours. In particular, MSC expressing the cytosine deaminase:: uracil phosphoribosyltransferase (CD-MSC) have been previously demonstrated to inhibit growth of subcutaneous prostate cancer xenografts thanks to their ability to convert the non-toxic 5-fluorocytosine into the antineoplastic 5-fluorouracil. Since both the immune system and the tumour microenvironment play a crucial role in directing cancer progression, in order to advance towards clinical applications, we tested the therapeutic potential of this approach on animal models that develop autochthonous prostate cancer and preserve an intact immune system. As cell vectors, we employed adipose-tissue and bone-marrow MSC. CD-MSC toxicity on murine prostate cancer cells and tumour tropism were verified in vitro and ex-vivo before starting the preclinical studies. Magnetic Resonance Imaging was utilised to follow orthotopic tumour progression. We demonstrated that intravenous injections of CD-MSC cells, followed by intraperitoneal administration of 5-fluorocytosine, caused tumour regression in the transgenic adenocarcinoma of the mouse prostate (TRAMP) model, which develops aggressive and spontaneous prostate cancer. These results add new insights to the therapeutic potential of specifically engineered MSC in prostate cancer disease., Funding Agencies|Sanofi; Associazione Amici di URI onlus; Slovak League against Cancer

Purpose: We assessed whether spinal inhibition of the cannabinoid degrading enzyme FAAH would have urodynamic effects in normal rats and rats with bladder overactivity induced by partial urethral obstruction or prostaglandin E2. We also determined the expression of FAAH, and the cannabinoid receptors CB1 and CB2 in the sacral spinal cord. Materials and Methods: We used 44 rats for functional (cystometry) and Western blot experiments. The FAAH inhibitor oleoyl ethyl amide (3 to 300 nmol) was administered intrathecally (subarachnoidally) or intravenously. The expression of FAAH and CB1/CB2 receptors was determined by Western blot. Results: Oleoyl ethyl amide given intrathecally affected micturition in normal rats and rats with bladder overactivity but effects were more pronounced in the latter. In normal rats oleoyl ethyl amide only decreased micturition frequency, while it decreased frequency and bladder pressures in rats with bladder overactivity. Intravenous oleoyl ethyl amide (3 to 300 nmol) had no urodynamic effect. FAAH and CB1/CB2 receptors were expressed in the rat sacral spinal cord. The expression of CB1/CB2 receptors but not FAAH was higher in obstructed than in normal rats. Conclusions: FAAH inhibition in the sacral spinal cord by oleoyl ethyl amide resulted in urodynamic effects in normal rats and rats with bladder overactivity. The spinal endocannabinoid system may be involved in normal micturition control and it appears altered when there is bladder overactivity.

BACKGROUND: Information on autonomic neurapraxia in female urogenital surgery is scarce, and a model to study it is not available. OBJECTIVE: To develop a model to study the impact of autonomic neurapraxia on bladder function in female rats, as well as to assess the effects of corticosteroid therapy on the recovery of bladder function in this model. DESIGN, SETTING, AND PARTICIPANTS: Female Sprague-Dawley rats were subjected to bilateral pelvic nerve crush (PNC) and perioperatively treated with betamethasone or vehicle. Bladder function and morphology of bladder tissue were evaluated and compared with sham-operated rats. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Western blot, immunohistochemistry, organ bath experiments, and cystometry. RESULTS AND LIMITATIONS: Sham-operated rats exhibited regular micturitions without nonvoiding contractions (NVCs). Crush of all nerve branches of the pelvic plexus or PNC resulted in overflow incontinence and/or NVCs. Betamethasone treatment improved recovery of regular micturitions (87.5% compared with 27% for vehicle; p<0.05), reduced lowest bladder pressure (8 ± 2 cm H(2)O compared with 21 ± 5 cm H(2)O for vehicle; p<0.05), and reduced the amplitude of NVCs but had no effect on NVC frequency in PNC rats. Compared with vehicle, betamethasone-treated PNC rats had less CD68 (a macrophage marker) in the pelvic plexus and bladder tissue. Isolated bladder from betamethasone-treated PNC rats exhibited better nerve-induced contractions, contained more cholinergic and sensory nerves, and expressed lower amounts of collagen III than bladder tissue from vehicle-treated rats. CONCLUSIONS: PNC causes autonomic neurapraxia and functional and morphologic changes of isolated bladder tissue that can be recorded as bladder dysfunction during awake cystometry in female rats. Perioperative systemic betamethasone treatment reduced macrophage contents of the pelvic plexus and bladder, partially counteracted changes in the bladder tissue, and, Funding Agencies|Swedish Medical Research Council||Gester Foundation||Urological Research Institute