Nociception and pain in humans lacking a functional TRPV1 channel.

BACKGROUNDChronic pain is a debilitating illness with currently limited therapy, in part due to difficulties in translating treatments derived from animal models to patients. The transient receptor potential vanilloid 1 (TRPV1) channel is associated with noxious heat detection and inflammatory pain, and reports of adverse effects in human trials have hindered extensive efforts in the clinical development of TRPV1 antagonists as novel pain relievers.METHODSWe examined 2 affected individuals (A1 and A2) carrying a homozygous missense mutation in TRPV1, rendering the channel nonfunctional. Biochemical and functional assays were used to analyze the mutant channel. To identify possible phenotypes of the affected individuals, we performed psychophysical and medical examinations.RESULTSWe demonstrated that diverse TRPV1 activators, acting at different sites of the channel protein, were unable to open the cloned mutant channel. This finding was not a consequence of impairment in the expression, cellular trafficking, or assembly of protein subunits. The affected individuals were insensitive to application of capsaicin to the mouth and skin and did not demonstrate aversive behavior toward capsaicin. Furthermore, quantitative sensory testing of A1 revealed an elevated heat-pain threshold but also, surprisingly, an elevated cold-pain threshold and extensive neurogenic inflammatory, flare, and pain responses following application of the TRPA1 channel activator mustard oil.CONCLUSIONOur study provides direct evidence in humans for pain-related functional changes linked to TRPV1, which is a prime target in the development of pain relievers.FUNDINGSupported by the Israel Science Foundation (368/19); Teva's National Network of Excellence in Neuroscience grant (no. 0394886) and Teva's National Network of Excellence in Neuroscience postdoctoral fellowship.

The effect of poly (ADP-ribose) polymerase inhibition on aminoglycoside-induced acute tubular necrosis in rats.

INTRODUCTION: Aminoglycosides (AG) cause nephrotoxicity in 10 - 20% of patients. One of the mechanisms is by generating reactive oxygen species (ROS), leading to DNA destruction and activation of poly(ADPribose) polymerase (PARP) causing necrotic tubular cell death. PARP inhibition on gentamicin-induced nephrotoxicity was studied. METHODS: 19 female Wistar-Kyoto rats divided into 3 groups: control (3 rats receiving no treatment); gentamicin-treated group (8 rats); and 8 rats treated with gentamicin combined with 3-aminobenzamide (3 AB). Kidney functions, protein, and gentamicin levels as well as urinary trypsin inhibitory activity (TIA) were measured. Tissue microscopic examination and immunohistochemical study for proliferative cell nuclear antigen (PCNA) were determined. The effect of PARP inhibitor on the bactericidal activity of gentamicin was also assessed. RESULTS: The following results were statistically significant: urea (mg/dL) 39.9 ± 5.86, 88.3 ± 50.3, and 48.5 ± 12.7 (p = 0.048); serum creatinine (mg/dL): 0.6 ± 0.26, 1.05 ± 0.7, 0.6 ± 0.06 (p = 0.043); proteinuria (mg/24-hours): 7.27 ± 3.65, 41.2 ± 18.1, and 17.6 ± 13.9 (p = 0.050); the number of tubular macronuclei (per 10 mm2): 18.33 ± 16.07, 218 ± 101.8, 41.7 ± 36.2 (p = 0.012); the number of dilated tubes (per 10 mm2): 61.67 ± 12.58, 276.3 ± 112.7, 140.0 ± 90.9 (p = 0.04); and the number of PCNA positive nuclei (per 10 mm2): 223.3 ± 95.69, 3,585 ± 2,215.3, 626.7 ± 236.9 (p = 0.034) in the control, gentamicin, and gentamicin+3AB-treated groups, respectively. The following biochemical and histologic parameters were also examined, however, they showed no statistically significant difference: TIA (p = 0.055), mitoses (p = 0.14), mononuclear infiltrate (p = 0.188), and intratubular cast formation (p = 0.084). No effect on bactericidal activity was observed. CONCLUSION: This study illustrates that PARP inhibitor significantly attenuates gentamicin-induced nephrotoxicity in rats with no effect on the bactericidal activity.

The Effect of Cyclooxygenase-2 Expression in Uterine Carcinosarcoma on Survival: A Reassessment Based on Mature Data.

OBJECTIVE: To reassess the effect cyclooxygenase-2 (COX-2) expression in carcinosarcoma on survival based on mature 5-year survival data. METHOD: A comparison of 5-year survival of 27 patients with carcinosarcoma according to the presence of COX-2 immunohistochemical staining and staining score was performed. RESULTS: The 5-year survival of those with positive and negative COX-2 staining was statistically not different. However, there was a clear trend for more favorable 5-year survival in patients with a high staining score than in those with a low score, and the difference was of borderline significance (38.5% vs 7.1%; P = 0.06). CONCLUSION: In view of the role of COX-2 in carcinogenesis, our finding that COX-2 expression may confer a better survival in patients with carcinosarcoma is intriguing. Larger studies are indicated to elucidate the effect of COX-2 expression on survival in patients with carcinosarcoma because this may have therapeutic implications.

CA125 expression in the tissue of uterine leiomyosarcoma.

UNLABELLED: Background: Elevated serum levels of the epithelial marker CA125 are occasionally observed in leiomyosarcoma (LMS) patients. OBJECTIVES: To assess the immunohistochemical expression of this marker in the tissue of LMS. METHODS: The consecutive unselected records of all patients with LMS diagnosed during the period 1995-2012 were located and abstracted. After verification of the diagnosis, 4 µm unstained slides were prepared from each case for immunohistochemical staining for CA125. Sections of ovarian carcinoma known to express CA125 were used as positive controls. RESULTS: We located 17 LMS patients from the period under study. Bleeding was the presenting symptom in 9 patients; the diagnosis was established prior to treatment in 11 patients. The tumor was in an advanced stage in 6 patients, and in 7 unstaged patients it was grossly confined to the uterus. Ten patients died within 14 months after the diagnosis. Serum CA125 levels prior to treatment were assessed in only 8 patients and were above normal limits (> 35 U/ml) in 3 of them. Two of the three with elevated serum levels were in stage III, and the third was an unstaged apparent stage I patient. None of the LMS tissue specimens demonstrated immunohistochemical expression of CA125. CONCLUSIONS: CA125 was not immunohistochemically expressed in the tissue of any LMS tumors examined by us. The origin of elevated serum CA125 in some of these tumors is therefore not in its tissue and remains unknown.

Effect of atorvastatin on IgA nephropathy in the rat.

BACKGROUND: IgA nephropathy (IgAN) is the most common chronic glomerulonephritis in humans and is a major cause of end-stage kidney disease worldwide. There is no agreement on the exact underlying mechanism or therapeutic intervention for this disorder. Mesangial proliferation typifies the renal histopathology in IgAN. Statin drugs, as prenylationinhibitors, have been shown to have an antiproliferative effect on renal mesangial cells and to reduce IgAN-associated glomerulusclerosis and proteinuria. The aim of this study is to examine the effect of atorvastatin on kidney function, proteinuria and kidney histology changes in IgANinduced rats. METHODS: IgAN was induced in Wistar-Kyoto rats by bovine γ-globulin (BGG). Four groups of rats were treated in metabolic cages: 1) control; 2) atorvastatin (2 mg/kg body weight/day through nasogastric tube) - treated rats; 3) IgAN-rats; 4) IgAN-rats treated with atorvastatin. Urine volume, urine protein excretion, blood urea and creatinine concentrations in addition to creatinine clearance were examined every 14 days, throughout the duration of the study (56 days). All kidneys from sacrificed rats were examined for histology including glomerular cell nuclei count and immunofluorescence. RESULTS: There were no differences in blood creatinine concentrations between the groups. Creatinine clearance was lower on the 42nd day and proteinuria was higher on Days 14, 42 and 56, in rats in Group 3 compared to all others; additionally, histology examination revealed a higher glomerular cell nuclei count in this group. Immunofluorescence was equally positive for IgA in mesangial cells in the kidneys from rats of Groups 2, 3 and 4. CONCLUSIONS: Atorvastatin attenuates kidney-function impairment, proteinuria and mesangial cell proliferation in BGG model of IgANinduced rats.

COX-2 expression in nonepithelial ovarian malignancies.

The aim of the study was to assess the expression of cyclooxygenases (COX)-2 in nonepithelial ovarian malignancies.COX-2 immunohistochemical staining was performed on newly prepared deparaffinized slides from formalin-fixed, paraffin-embedded archival tissue blocks of unselected nonepithelial ovarian malignancies diagnosed between January 1993 and October 2009 after reconfirmation of the diagnosis. Staining was assessed according to intensity of staining and the proportion of stained cells. Staining of more than 10% of the cells was considered positive.During the study period, 26 histologically proven nonepithelial ovarian malignancies were diagnosed. Of them, 16 were granulosa cell tumors and 10 were germ cell tumors (4 dysgerminomas and 6 immature teratomas). Five (31.2%) granulosa cell tumors had positive immunohistochemical COX-2 staining. Positive staining was observed only in 1 immature teratoma and in none of the dysgerminomas.Our data seem to indicate that COX-2 expression by immunohistochemical methods is not frequent in nonepithelial ovarian malignancies.

COX-2 expression in uterine carcinosarcoma.

OBJECTIVE: To assess cyclooxygenase-2 (COX-2) expression in the carcinomatous and sarcomatous elements of uterine carcinosarcoma. DESIGN: Retrospective study. SETTING: Medical center. SAMPLE: Twenty-seven consecutive uterine specimens with carcinosarcoma. METHODS: COX-2 expression assessment by immunohistochemistry was performed on deparaffinized sections of paraffin-embedded tissue blocks. Analysis of all stained tissue sections was done by means of light microscopy counting 10 high power fields with a minimum of 1,000 cells per slide. Analysis of the stained tissue sections was done by pathologists blinded to the clinical data and the follow-up course of the malignancy. MAIN OUTCOME MEASURES: Staining of more than 10% of the cells was considered positive. Staining intensity was graded on a 0-3 scale. A scoring index was calculated by multiplying the intensity grade by the percentage of stained cells and considered low when it was < or =1 and high when it was >1. COX-2 expression was associated with prognostic factors and survival. RESULTS: COX-2 immunohistochemical staining was positive in 74% of the carcinomatous component of carcinosarcoma and a high staining score was observed in 48.2%. Staining parameters were significantly lower in the sarcomatous component. No significant association between COX-2 expression with stage, grade, depth of myometrial invasion, lymphvascular involvement or with survival was observed. CONCLUSIONS: COX-2 is expressed in a high percentage of uterine carcinosarcoma, but does not seem to be associated with prognostic factors or survival.

Is Her-2/neu expressed in nonepithelial ovarian malignancies?

OBJECTIVE: The objective of this study was to assess the expression of Her-2/neu in nonepithelial ovarian malignancies. STUDY DESIGN: Formalin-fixed paraffin-embedded archival tissue blocks of 20 unselected nonepithelial ovarian malignancies (12 granulosa cell tumors and 8 germ cell tumors) diagnosed between 1993 and 2005 were immunohistochemically stained for Her-2/neu. RESULTS: Immunohistochemical staining for Her-2/neu was not present in any of these nonepithelial malignancies examined. CONCLUSION: Our limited sample size does not allow a generalized conclusion concerning the lack of Her-2/neu expression in nonepithelial ovarian malignancies, but it adds information with regard to the expression of this oncogene in these rare neoplasms and seems to indicate that it is not a frequent occurrence.

Assessment of Her-2/neu expression in hydatidiform moles for prediction of subsequent gestational trophoblastic neoplasia.

OBJECTIVE: The aim of the present study was to asses the ability of Her-2/neu immunohistochemical staining of the molar tissue to predict the risk of developing gestational trophoblastic neoplasia (GTN). METHODS: Sections prepared from 33 consecutive formalin-fixed paraffin-embedded archival reconfirmed hydatidiform mole tissue blocks were immunohistochemically stained for Her-2/neu. The staining was scored according to the subjectively evaluated intensity of staining and the proportion of stained villous cytotrophoblastic cells. Clinical data were abstracted from medical files. RESULTS: 23 patients had a complete and 10 a partial mole. Nine patients (27.3%) were diagnosed with GTN [7 of 23 patients with a complete mole (30.4%) and 2 of the 10 (20.0%) with a partial mole]. A positive immunohistochemical Her-2/neu stain was found in 6 (18.2%) of the patients with hydatidiform mole (3 with a complete mole). The rate of Her-2/neu expression was somewhat higher in moles with subsequent GTN than in moles with an uneventful course (22.2% vs. 16.6%, respectively). The difference did not reach significance (Fisher's Exact Test, P=0.55) possibly due to the small number of cases (power of <5%). The sensitivity and specificity of Her-2/neu expression for prediction of GTN was 22.2% and 83.3%, respectively, and the positive and negative predictive value 33.3% and 74.1%, respectively. CONCLUSION: While the specificity of Her-2/neu immunohistochemical staining for prediction of GTN is relatively high, the low sensitivity and low positive and negative predictive value precludes its practical clinical use for prediction of post-molar GTN. The quest for a precise predictor of post-molar GTN should continue.

Asymptomatic or minimally symptomatic hyperCKemia: histopathologic correlates.

BACKGROUND: Persistent creatine kinase elevation is occasionally encountered in subjects without any clinical manifestation of a neuromuscular disorder or any condition known to be associated with increased serum CK levels. It is still unresolved whether extensive investigations and specifically a muscle biopsy should be performed in clinically normal individuals with elevated CK levels. OBJECTIVE: To study the muscle pathology of patients with asymptomatic or minimally symptomatic hyperCKemia. METHODS: The clinical and laboratory data of patients with persistent hyperCKemia and normal neurologic examination were reviewed and their muscle biopsies evaluated. RESULTS: The study group included 40 patients aged 7-67 years; the male to female ratio was 3:1. Nineteen patients were completely asymptomatic, 20 had mild non-specific myalgia, and 1 had muscle cramps. Electromyography was performed in 27 patients and showed myopathic changes in 7 (26%). Abnormal muscle biopsy findings (e.g., increased variation in fiber size, increased number of central nuclei, and occasional degenerating fibers) were detected in 22 of the 40 patients (55%). No fat or glycogen accumulation was detected. Immunohistochemistry demonstrated abnormal dystrophin staining in 3 patients (8%), resembling the pathologic changes of Becker muscular dystrophy. No abnormal findings were detected on immunohistochemical staining for merosin, dysferlin, caveolin 3, or alpha and gamma sarcoglycans. The EMG findings did not correlate with the pathologic findings. CONCLUSIONS: Abnormal muscle biopsies were found in 55% of patients with asymptomatic or minimally symptomatic hyperCKemia. Specific diagnosis of muscular dystrophy, however, was possible in only 8% of the patients.

Expression of c-kit in uterine carcinosarcoma.

OBJECTIVES: The use of tyrosine kinase inhibitors (TKIs) has resulted in successful treatment of KIT-positive neoplasms. Carcinosarcoma is a very aggressive neoplasm. Consequently, c-kit expression may have significant clinical implications for this tumor. The purpose of the present study was to assess c-kit expression in the carcinomatous and sarcomatous element of carcinosarcoma to identify if KIT represents a therapeutic target for treatment of this neoplasm. METHODS: Immunohistochemical staining for c-KIT was performed on paraffin-embedded tissue blocks of 20 consecutive uterine specimens with carcinosarcoma, 40 with endometrial carcinoma, and 12 with atrophic endometrium. Two pathologists assessed the scoring index of staining. RESULTS: In the stromal element of carcinosarcoma, no immunohistochemically c-KIT stained cells were observed and in the epithelial element, a low scoring index of staining was present. In endometrial endometrioid carcinoma, the scoring index was high in only 15% of the specimens. In the atrophic endometrium, a low scoring index was seen in all specimens. CONCLUSIONS: Our results seem to suggest that c-kit probably plays no major role in the pathogenesis of the majority of these tumors while it may be involved in the pathogenesis of some endometrial carcinomas. In view of the multiple molecular targets that are activated by imatinib mesylate, no definitive conclusions can be drawn with regard its effectiveness in carcinosarcomas based on the present study. Further studies of c-kit expression in larger series of carcinosarcomas in order to settle the controversial issues with regard to frequency of expression, prognostic, and clinical value are warranted.