Cigarette smoking is associated with an altered vaginal tract metabolomic profile.

Cigarette smoking has been associated with both the diagnosis of bacterial vaginosis (BV) and a vaginal microbiota lacking protective Lactobacillus spp. As the mechanism linking smoking with vaginal microbiota and BV is unclear, we sought to compare the vaginal metabolomes of smokers and non-smokers (17 smokers/19 non-smokers). Metabolomic profiles were determined by gas and liquid chromatography mass spectrometry in a cross-sectional study. Analysis of the 16S rRNA gene populations revealed samples clustered into three community state types (CSTs) ---- CST-I (L. crispatus-dominated), CST-III (L. iners-dominated) or CST-IV (low-Lactobacillus). We identified 607 metabolites, including 12 that differed significantly (q-value < 0.05) between smokers and non-smokers. Nicotine, and the breakdown metabolites cotinine and hydroxycotinine were substantially higher in smokers, as expected. Among women categorized to CST-IV, biogenic amines, including agmatine, cadaverine, putrescine, tryptamine and tyramine were substantially higher in smokers, while dipeptides were lower in smokers. These biogenic amines are known to affect the virulence of infective pathogens and contribute to vaginal malodor. Our data suggest that cigarette smoking is associated with differences in important vaginal metabolites, and women who smoke, and particularly women who are also depauperate for Lactobacillus spp., may have increased susceptibilities to urogenital infections and increased malodor.

Experience with intensity-modulated radiotherapy in the treatment of head and neck cancer.

PURPOSE: Studies using intensity-modulated radiation therapy (IMRT) in the treatment of head and neck tumors have shown to decrease acute and late radiation toxicity. However, the high conformity of this technique can increase the risk of recurrence due to geographic miss. The aim of this study was to analyze whether the results of IMRT met the theoretical expectations concerning treatment efficacy. METHODS: From a total of 185 patients (152 males and 33 females, mean age 58±10.36 years) 176 were evaluable and were studied. Eighty-nine (48.1%) patients had surgical treatment and 50 of them were scheduled for concomitant cisplatin chemotherapy. Irradiation was performed using IMRT, a sliding window with 9 fields in a Varian 2100 C/D linear accelerator, X-ray beam, 6 MeV. The prescribed dose in the planning treatment volume (PTV1), i.e., the area of the primary tumor and nodal area, was 66 Gy/2.2 Gy-70 Gy/2.12 Gy. In the PTV2 (the area at high risk) the dose was 60Gy/2 Gy-59.4 Gy/ 1.8 Gy, and in the PTV 3 (the area treated with prophylactic irradiation) the prescribed dose was 54 Gy/1.8 Gy/50.4 Gy/1.53 Gy. RESULTS: The 3-year overall survival (OS) and relapse-free survival (RFS) of IMRT-treated patients, most of whom were in stages III and IV (158 out of 177), were 50 and 57%, respectively. Using postoperative radiotherapy/chemoradiotherapy 3-year locoregioncal control was achieved in 75% of the cases as compared with 35% in non-operated patients. CONCLUSIONS: The worst outcomes were found in oral cavity and hypopharyngeal tumors, and the best in laryngeal and oropharyngeal tumors. Better results were found in surgically treated patients, and in lower disease stages. Despite the high conformity of dose distribution and efforts to spare healthy tissues, most cases of locoregional relapse occurred in areas receiving the full radiation dose. If dividing relapses into cases of persistence and local recurrence, the former predominated.

PET/CT significance for planning radiotherapy of head and neck cancer.

The combination of positron emission tomography and computed tomography (PET/CT) offers metabolic mapping in addition to anatomic information of the primary lesion, nodal and distant metastases in patients with head and neck tumors, and may be therefore beneficial for radiotherapy planning. The aim of our study was to evaluate benefits of combined PET and CT imaging for staging and target volume delineation in this group of patients.Fifty three patients (40 men and 13 women) with confirmed advanced, inoperable or non-radically operated head and neck cancer were assessed based on the results of PET/CT as well as standard diagnostic examinations. All patients were subsequently treated with intensity modulated radiotherapy (IMRT) with simultaneous integrated boost (SIB) of 6 MV X-rays. There was an agreement between the standard examinations results and results of PET/CT in 30 cases. In 23 cases there was disagreement either in tumor size, nodal involvement or presence of distant metastases. Results of the tumor size assessment differed significantly in 5 cases. There was no agreement found in nodal involvement in 10 cases. The cancer confirmed by standard examination was not found by PET/CT in 2 cases; 3 PET/CT positive findings were not confirmed by standard examinations. In 3 patients PET-CT revealed new distant metastatic disease. Based on PET/CT assessment we changed treatment strategy and applied potentially curative dose of radiotherapy to previously undiscovered regions in 9 patients. We decided to change the treatment intent in 3 cases and only palliative treatment was applied. Based on our experience and the literature review, PET/CT may be considerable contribution to the standard diagnostic procedures in approximately one third of cases.

Autophagy is essential to suppress cell stress and to allow BCR-Abl-mediated leukemogenesis.

Hematopoietic cells normally require cell extrinsic signals to maintain metabolism and survival. In contrast, cancer cells can express constitutively active oncogenic kinases such as BCR-Abl that promote these processes independent of extrinsic growth factors. When cells receive insufficient growth signals or when oncogenic kinases are inhibited, glucose metabolism decreases and the self-digestive process of autophagy is elevated to degrade bulk cytoplasm and organelles. Although autophagy has been proposed to provide a cell-intrinsic nutrient supply for mitochondrial oxidative metabolism and to maintain cellular homeostasis through degradation of damaged organelles or protein aggregates, its acute role in growth factor deprivation or inhibition of oncogenic kinases remains poorly understood. We therefore developed a growth factor-dependent hematopoietic cell culture model in which autophagy can be acutely disrupted through conditional Cre-mediated excision of the autophagy-essential gene Atg3. Treated cells rapidly lost their ability to perform autophagy and underwent cell cycle arrest and apoptosis. Although Atg3 was essential for optimal upregulation of mitochondrial oxidative pathways in growth factor withdrawal, this metabolic contribution of autophagy did not appear critical for cell survival, as provision of exogenous pyruvate or lipids could not completely rescue Atg3 deficiency. Instead, autophagy suppressed a stress response that otherwise led to p53 phosphorylation and upregulation of p21 and the pro-apoptotic Bcl-2 family protein Puma. Importantly, BCR-Abl-expressing cells had low basal levels of autophagy, but were highly dependent on this process, and rapidly underwent apoptosis upon disruption of autophagy through Atg3 deletion or treatment with chemical autophagy inhibitors. This dependence on autophagy extended in vivo, as Atg3 deletion also prevented BCR-Abl-mediated leukemogenesis in a cell transfer model. Together these data demonstrate a critical role for autophagy to mitigate cell stress, and that cells expressing the oncogenic kinase BCR-Abl appear particularly dependent on autophagy for cell survival and leukemogenesis.

The relationship of intracellular free calcium activity to amylase secretion in substance P- and isoprenaline-stimulated rat parotid acini.

The relationship between intracellular free calcium activity ([Ca2+]i) and stimulated amylase secretion was investigated in rat parotid acini by measuring the effects of substance P methyl ester and isoprenaline on quin2 fluorescence and amylase release. Although both of these drugs evoked concentration-dependent increases in [Ca2+]i and amylase release, the tachykinin had a greater effect on [Ca2+]i while the catecholamine was the more effective secretagogue. Whereas isoprenaline exerted equipotent effects on amylase secretion and [Ca2+]i, the dose-response relationship for stimulation of secretion by substance P was dissociated by three orders of magnitude to the right of that for elevation of [Ca2+]i by this peptide. It is concluded that these data do not support the hypothesis that substance P-stimulated amylase secretion is mediated solely through an increase in [Ca2+]i and that other second-messengers may be involved in mediation of this secretory response.

Description of an automated assay for measurement of alpha-amylase in vitro from rat parotid gland slices.

1. A modified continuous assay of alpha-amylase is detailed and its usefulness in measuring amylase secretion from rat parotid salivary gland described. 2. The alpha-amylase secretion from the rat parotid gland can be evoked by acetylcholine, substance P and isoprenaline. 3. The secretion of amylase was greatest following isoprenaline stimulation and was of a slower time course than with the other two agonists.

The role of dopamine in salivation in the rat parotid gland.

The ability of dopamine to evoke secretion from the rat parotid gland has been investigated. The actions of dopamine were investigated alone and during carbachol mediated secretion. Dopamine increased flow rate and protein secretion in both experimental models. The increase in protein secretion is mediated via beta 1-adrenoceptors whereas the increase in flow rate may be due to activation of specific dopamine receptor.

Disopyramide fatality: case report and GC/FID analysis.

This case involves massive ingestion of disopyramide by a 19-year-old male, found dead, not having had the benefit of medical intervention. Quantitation of the drug was accomplished by gas chromatography using a flame ionization detector (GC/FID). Identification of the compound was done by thin layer chromatography (TLC) and ultraviolet spectrophotometry (UVS). The blood concentration of disopyramide was 15 times the usual therapeutic level and was easily detected by FID.

Utilization of thin-layer chromatography and enzyme immunoassay systems to screen and confirm the presence of morphine, codeine, phencyclidine and benzoylecgonine.

Thin-layer chromatography (TLC) and enzyme immunoassay (EMIT) were coordinated to produce unequivocal identification of certain substances. Samples were extracted and subjected to TLC. After development, "positive" spots were scraped from the plate, buffered, centrifuged, and subjected to EMIT. Interferences were not encountered.

Propoxyphene-associated deaths: methods, post-mortem levels in blood and liver.

Gas chromatographic methodology for the determination of d-propoxyphene in blood and liver samples is given. The method results in a clean extract and sharp response peaks. Quantitation is based on peak height ratios of propoxyphene and pyrroliphene. Post-mortem levels in blood range from 0.1-0.7 mg/dl; in liver from 1.1-18.9 mg/100 gm. The highest blood level has the lowest blood-liver ratio, the rest being widely variable. The incidence of propoxyphene in accidental and suicidal deaths appears to be increasing steadily.