High-performance liquid chromatography method for urinary trans,trans-muconic acid. Application to environmental exposure to benzene

This report describes a specific and precise high-performance liquid chromatography (HPLC) method for the quantification of trans,trans-muconic acid in human urine. The procedure involved a highly efficient Bond-Elut SAX extraction with 20% acetic acid elution. The HPLC analysis used a sodium acetate/methanol mobile phase with a C18 reverse phase column and UV detection at 265 nm. The recovery, precision, linearity, and limits of detection and quantification of the method were determined. Mean absolute recoveries were between 97% and 115%. The calibration curve showed a correlation coefficient of 0.9955 and the limit of detection was determined to be 10.8 microg/L. The method is suitable for evaluation of occupational and environmental benzene exposure in humans. The study of urinary trans,trans-muconic acid of two populations of children to evaluate environmental benzene exposure is presented.     

Neurospheres enriched in cancer stem-like cells are highly effective in eliciting a dendritic cell-mediated immune response against malignant gliomas

Cancer stem-like cells (CSC) could be a novel target for cancer therapy, including dendritic cell (DC) immunotherapy. To address this, we developed experiments aimed at DC targeting of neurospheres (NS) from GL261 glioma cells because neurospheres can be enriched in CSC. We obtained murine neurospheres by growing GL261 cells in epidermal growth factor/basic fibroblast growth factor without serum. GL261-NS recapitulated important features of glioblastoma CSC and expressed higher levels of radial glia stem cell markers than GL261 cells growing under standard conditions (GL261 adherent cells, GL261-AC), as assessed by DNA microarray and real-time PCR. GL261-NS brain gliomas were highly infiltrating and more rapidly lethal than GL261-AC, as evidenced by survival analysis (P < 0.0001), magnetic resonance imaging and histology. DC from the bone marrow of syngeneic mice were then used for immunotherapy of GL261-NS and GL261-AC tumors. Strikingly, DC loaded with GL261-NS (DC-NS) cured 80% and 60% of GL261-AC and GL261-NS tumors, respectively (P < 0.0001), whereas DC-AC cured only 50% of GL261-AC tumors (P = 0.0022) and none of the GL261-NS tumors. GL261-NS expressed higher levels of MHC and costimulatory molecules (CD80 and CD86) than GL261-AC; the JAM assay indicated that DC-NS splenocytes had higher lytic activity than DC-AC splenocytes on both GL261-NS and GL261-AC, and immunohistochemistry showed that DC-NS vaccination was associated with robust tumor infiltration by CD8+ and CD4+ T lymphocytes. These findings suggest that DC targeting of CSC provides a higher level of protection against GL261 gliomas, a finding with potential implications for the design of clinical trials based on DC vaccination.     

Interrelations between fat distribution, muscle lipid content, adipocytokines, and insulin resistance: effect of moderate weight loss in older women

BACKGROUND: Interrelations between fat distribution, muscle lipid infiltration, adipocytokines, insulin resistance, and moderate weight loss have not been investigated in obese older subjects. OBJECTIVE: The objective was to evaluate relations between fat distribution, muscle lipid content, adipocytokines, and insulin resistance in older women and the effects of moderate weight loss. DESIGN: In 35 healthy women aged 58-83 y, body mass index, waist circumference, sagittal abdominal diameter (SAD), and body composition measured by dual-energy X-ray absorptiometry were evaluated. A midthigh single computed tomography scan was performed to determine subcutaneous adipose tissue (AT), intermuscular AT (IAT), muscular tissue, and muscle lipid infiltration, evaluated as low-density lean tissue. Metabolic variables, insulin resistance measured by homeostasis model assessment, adiponectin, leptin, and high-sensitivity C-reactive protein were measured in all subjects and after weight loss in a subgroup of 15 obese women. RESULTS: Waist circumference and SAD were positively correlated with leptin and insulin resistance and negatively correlated with adiponectin. Adiponectin was associated negatively with insulin resistance and positively with HDL cholesterol, whereas leptin was positively associated with insulin resistance and triacylglycerols. Midthigh subcutaneous AT was associated with insulin resistance and leptin, whereas IAT was associated with triacylglycerols. Stepwise regression with insulin resistance as the dependent variable and body mass index, SAD, triacylglycerols, HDL cholesterol, adiponectin, leptin, high-sensitivity C-reactive protein, and midthigh subcutaneous AT as independent variables showed that SAD entered the regression first (R(2) = 0.492) followed by adiponectin (R(2) = 0.63). After moderate weight loss, midthigh subcutaneous AT, IAT, low-density lean tissue, leptin, and insulin resistance decreased significantly; no significant changes in adiponectin were observed. CONCLUSIONS: Fat distribution indexes and adiponectin are independently associated with insulin resistance. Even in older women, moderate weight loss improves body fat distribution, muscle lipid infiltration, and insulin resistance. Moderate weight loss results in a significant decrease in leptin but no changes in adiponectin.     

Stimulation of prostaglandin EP2 receptors prevents NMDA-induced excitotoxicity

Prostaglandin E(2) (PGE(2)) plays an important role in inflammation and neurologic disorders. The neuromodulatory effects of PGE(2) are mediated through regulation of four G-protein-coupled receptors known as EP1, EP2, EP3, and EP4. The goal of the current study was to determine whether EP2 receptor activation protects neurons from acute NMDA-mediated excitotoxicity. To examine the effects of EP2 activation, mice were given an injection of the EP2 receptor-selective agonist butaprost (K (i) = 110 nM for EP2 receptor; K (i) > 10,000 for other prostaglandin receptors) in the cerebral ventricle and then an injection of NMDA in the right striatum. After 48 h, a significant reduction in NMDA-induced lesion volume was observed in groups pretreated with butaprost (1-300 nmol/L), with maximal protection at 100 nmol/L (p < 0.001). To determine if EP2-activated protection was specific to neurons, mouse neuronal cultures were treated with butaprost, and cell viability was analyzed after 24 h of NMDA excitotoxicity. The results showed that butaprost significantly increased neuron survival in a dose-dependent fashion. Furthermore, treatment of primary neurons with butaprost significantly increased cAMP levels (p < 0.001). Together, these data reveal that EP2 receptor stimulation mediates neuroprotection against NMDA excitotoxicity both in vivo and in vitro and that butaprost can limit acute brain damage. Development and testing of specific PGE(2) receptor mimetics could lead to a decrease in side effects associated with anti-inflammatory drugs and could help to fight acute and/or chronic neurologic disorders.     

Use of a new hemicellulose dressing (Veloderm) for the treatment of split-thickness skin graft donor sites A within-patient controlled study

A multi-centre, open, within-patient controlled study was performed on 23 adult burnt patients to investigate the effectiveness, safety and tolerability of Veloderm^® in comparison with Algisite M™ and Jaloskin^® in split-thickness skin graft donor site care. The areas dressed with Veloderm^® completely healed within 10–13 days in a significant higher proportion than the other two dressings (47.6% for Veloderm^® versus 26.3% for Algisite M™ and 10% for Jaloskin^®, P < 0.03), showing during the whole study less incidence of exudates and of peri-lesional erythema. The aesthetic outcome of the treated lesions after healing was significantly better for Veloderm^® (P = 0.0016). Veloderm^® and Jaloskin^® required very few renewals of the medication during the first week of treatment, while Algisite M™ needed several multiple re-dressings. Veloderm^® was judged better than the other two treatments as far as the acceptability (P < 0.001), ease of use (P < 0.001) and efficacy (P < 0.00001). Both pain during application or at removal of dressings and local infections were negligible with all treatments. No scars were formed in any skin donor site. In conclusion Veloderm^® is a safe and effective dressing for the re-epithelialization of the skin graft donor sites: it showed higher activity than the other two compared dressings.     

Antisperm antibodies in prepubertal boys with cryptorchidism

The presence of antisperm antibodies in male individuals before puberty is controversial due to the lack of finally differentiated male germ cells. It was questioned whether the pathologic conditions of the male gonad may influence antisperm antibody formation in individuals before puberty. Sera samples of 76 individuals and 10 healthy boys with testicular failure (mainly uni- or bilateral cryptorchidism) were examined by means of indirect immunobead-binding test (IDIBT). The presence of antisperm antibodies was found in 3.95% of the studied subjects. Antibodies recognizing antigenic determinants present on the surface of mature sperm cells may be produced before puberty in individuals suffering from cryptorchidism or the other gonadal disorders. Antisperm antibodies that did develop in a minority of the studied male population may be proof for individual predispositions to autoimmune reactions.     

Surgical strategies in cervico-mediastinal goiters

AIM: We retrospectively studied clinical picture, diagnosis, treatment and complications of patients with cervico-mediastinal goiter who underwent surgery. METHODS: Sixty-three patients underwent surgical treatment for cervico-mediastinal goiter in the last 10 years. Thirty-two patients (50.8%) had cervico-mediastinal goiter, 24 patients (33.3%) had mediastino-cervical goiter and 7 patients (11.1%) had mediastinal goiter. Forty-seven cases were prevascular goiters and 16 were retrovascular goiters. We performed total thyroidectomy in 25 patients, subtotal thyroidectomy in 31 patients and in 7 cases the resection of residual goiter. In 50 patients we used a cervical approach, in 12 patients the cervical incision was combined with median sternotomy (6 in total) and in 1 patient with transverse sternotomy. RESULTS: Three patients (4.7%) died in the postoperative period (2 cardio-respiratory failure and 1 pulmonary embolism). The histologic study revelead 5 (7.9%) carcinomas. Postoperative complications were: dyspnea in 7 cases (11.1%), transient vocal cord paralysis in 5 patients (7.9%), temporary hypoparathyroidism in 6 patients (9.5%) and kidney failure in 1 case (1.6%). CONCLUSIONS: Cervicotomy is the approach of choice but in some limited cases (carcinoma, thyroiditis, retrovascular goiter, ectopic goiter) the sternotomy should be performed. Postoperative mortality and morbidity is very low, independent of surgical techniques.     

The utility of HPV DNA triage in the management of cytological AGC

OBJECTIVE: Given the demonstrated utility of human papilloma virus DNA triage in the management of atypical squamous cells of undetermined significance, this study sought to evaluate the potential role of human papilloma virus DNA testing in the evaluation and management of cytological atypical glandular cells. STUDY DESIGN: Following institutional review board approval, 28 women presenting with cytological atypical glandular cells underwent repeat thin-prep cytology, Hybrid Capture 2 human papilloma virus DNA testing, colposcopic evaluation, Fisher electrosurgical conization, and endometrial sampling. Human papilloma virus test results in each patient were then correlated with histologic lesions, if present. RESULTS: Sixteen of the 28 study patients had pathologic lesions (11/28 high-grade squamous intraepithelial lesion, 3/28 low-grade squamous intraepithelial lesion, 1/28 adenocarcinoma in situ, 1/28 simple endometrial hyperplasia). Human papilloma virus DNA testing was available in 24 of 28 subjects (86%). The sensitivity of human papilloma virus positivity to predict the presence of cervical intraepithelial neoplasia was 100% (confidence interval 77% to 100%), specificity 64% (confidence interval 35% to 85%), positive predictive value 76%, and negative predictive value 100%. Women who tested human papilloma virus positive were 12 times more likely to have cervical intraepithelial neoplasia than women who were human papilloma virus negative (Fisher P<.001). Human papilloma virus positivity was not predictive of endometrial pathology; women who were human papilloma virus positive were less likely to have endometrial pathology than were women who were human papilloma virus negative (risk ratio 1.6, 95% confidence interval 0.01-1.7). CONCLUSION: Atypical glandular cells can represent a variety of lesions. The majority of the lesions will be squamous intraepithelial lesions of the cervix (50%), with high-grade squamous intraepithelial lesion present in 40% of subjects. Human papilloma virus DNA testing is a sensitive test for the presence of squamous intraepithelial lesion, with excellent negative predictive value for the absence of squamous intraepithelial lesion. The results of this study suggest human papilloma virus DNA testing might be an effective screening test in the initial evaluation and management of cytological atypical glandular cells.