ReProComet: a new in vitro method to assess DNA damage in mammalian sperm.

The increasing request of chemical safety assessment demands for the validation of alternative methods to reduce the resort to animal experimentation. Methods that evaluate reproductive toxicity are among those requiring the largest use of animals. Presently, no validated in vitro alternative exists for the assessment of reproductive toxicity. Mammalian sperm are sensitive targets of DNA-reactive chemicals, which form premutagenic adducts. Here, we propose a new method based on comet assay to detect DNA damage induced by potential germ cell mutagens in bull sperm available from assisted reproduction practices. In somatic cells, chemical-induced adducts can be revealed by comet assay that detects DNA breaks produced during adduct repair. Mature sperm, however, are devoid of repair enzymes, and adducts are processed only after fertilization. For this reason, comet assay is not sensitive to detect DNA lesions induced in sperm by most chemicals. To overcome such limitation, we developed a modified comet assay based on the addition of a protein extract from HeLa cells to agarose-embedded sperm on microscopic slides. To test the method, sperm were treated in vitro with methyl methanesulfonate (MMS) or melphalan (MLP) and comet assay was conducted both with and without protein supplementation. No effect of MMS or MLP was detected without protein supplementation; on the contrary, a clear-cut dose-dependent effect was measured after addition of the cell extract. These results represent a proof of concept of a novel in vitro mutagenicity test on sperm that could offer a promising approach to complement previously validated in vivo germ cell genotoxicity assays.     

Asymmetric dimethyl-arginine (ADMA) response to inflammation in acute infections.

BACKGROUND AND METHODS: The endogenous inhibitor of nitric oxide synthase (NOs) asymmetrical dimethyl-arginine (ADMA) has been implicated as a possible modulator of inducible NOs during acute inflammation. We examined the evolution in the plasma concentration of ADMA measured at the clinical outset of acute inflammation and after its resolution in a series of 17 patients with acute bacterial infections. RESULTS: During the acute phase of inflammation/infection, patients displayed very high levels of C-reactive protein (CRP), interleukin-6 (IL-6), procalcitonin and nitrotyrosine. Simultaneous plasma ADMA concentration was similar to that in healthy subjects while symmetric dimethyl-arginine (SDMA) levels were substantially increased and directly related with creatinine. When infection resolved, ADMA rose from 0.62 +/- 0.23 to 0.80 +/- 0.18 micromol/l (+29%, P = 0.01) while SDMA remained unmodified. ADMA changes were independent on concomitant risk factor changes and inversely related with baseline systolic and diastolic pressure. Changes in the ADMA/SDMA ratio were compatible with the hypothesis that inflammatory cytokines activate ADMA degradation. CONCLUSIONS: Resolution of acute inflammation is characterized by an increase in the plasma concentration of ADMA. The results imply that ADMA suppression may actually serve to stimulate NO synthesis or that in this situation plasma ADMA levels may not reflect the inhibitory potential of this methylarginine at the cellular level.     

Synergistic antiproliferative activity of quercetin and cisplatin on ovarian cancer cell growth

It has been demonstrated that the flavonoid quercetin (3,3',4',5-7-pentahydroxyflavone) (Q) inhibits the growth of several cancer cell lines and that the antiproliferative activity of this substance is mediated by a so-called type II estrogen binding site (type II EBS). We investigated the effects of quercetin and cisplatin (CDDP) alone and in combination on the proliferation of the ovarian cancer cell line OVCA 433. Both drugs exhibited a dose-related growth inhibition in a range of concentrations between 0.01 and 2.5 microM and 0.01 and 2.5 micrograms/ml for Q and CDDP respectively. The combination of the two drugs resulted in a synergistic antiproliferative activity. Two other flavonoids tested, i.e., rutin (3-rhamnosylglucoside of quercetin) and hesperidin [7-b rutinoside of hesperetin (3'-5-3-hydroxy-4-methoxyflavone)] were ineffective both alone and in combination with CDDP. Since both rutin and hesperidin do not bind to type II EBS it can be hypothesized that Q synergizes with CDDP by acting through an interaction with these binding sites.     

Food cravings in relation to body mass index, restraint and estradiol levels: a repeated measures study in healthy women.

The study considered the nature and extent of cravings in 108 healthy women between the ages of 20 and 37 who were tested at four time points over a 2-year period. There was substantial consistency over the four widely separated time points (3 months-1 year) in the types of foods craved, with chocolate and ice cream highest on the list, followed by fatty and spicy foods, and sweets. Women with a higher body mass index reported more consistent cravings for salty foods, especially those with high flavor intensity. There were no significant relationships between dietary restraint and the number, frequency or types of cravings. There were also no strong relationships between estradiol levels and the number, frequency or types of cravings women reported in the whole sample. The data suggest that women have a stable core of foods for which they experience cravings, relatively independent of estradiol levels, BMI or degree of dietary restraint.     

Prevalence, clinical characteristics, quality of life, and prognosis of patients with congestive heart failure and isolated left ventricular diastolic dysfunction.

Prevalence of isolated left ventricular (LV) diastolic dysfunction has been reported to be as high as one-third of all heart failure (HF) cases, with an increasing prevalence in the elderly population. However, there is a paucity of prospective data about the prevalence and prognosis of isolated LV diastolic dysfunction in an unselected population of patients hospitalized with HF. Therefore, we prospectively evaluated 179 consecutive patients discharged from our hospital with HF to assess the prevalence of systolic versus diastolic LV dysfunction among patients hospitalized with HF and to compare their demographics, clinical features, self-perceived quality of life (QOL), and 6-month readmission rate and mortality. Among them, 133 (59% men, median age 74 years) showed in sinus rhythm and had no significant primary valvular disease. LV diastolic dysfunction was diagnosed on the basis of the European Study Group on Diastolic HF echocardiographic criteria. QOL was assessed at hospital discharge and 6-month follow-up visit using the Minnesota Living with HF questionnaire. Survival of patients with HF was compared with that of age- and sex-matched general population. In all, 29 patients (22%) had isolated LV diastolic dysfunction and 102 (78%) had prevalent LV systolic dysfunction (ie, LV ejection fraction 相似文献   

Significance of cyclin D1 expression in meningiomas: a preliminary study.

Forty-four evaluable patients with intracranial meningiomas were assessed for the expression of the cell-cycle regulator cyclin D1 and of proteins involved in proliferation and apoptosis such as PCNA, MIB-1, p53 and bcl-2. Analyses were carried out by western blot and immunohistochemistry after immediate processing of fresh tumor specimens. By western blot, expression of cyclin D1 significantly correlated with p53 (p=0.02) and with proliferative activity, as assessed by PCNA expression (p=0.0009). By immunohistochemistry, a significant relationship between cyclin D1 and the proliferation marker MIB-1 was confirmed (p=0.05), whereas significance with bcl-2 expression was not found (p=0.01). Moreover, although the association with tumor grade appeared of borderline statistical significance (p=0.07), all the grade II/III meningiomas showed increased expression of cyclin D1 and high proliferative activity. In conclusion, data from this preliminary study seem to suggest a potential value of the combined expression of cyclin D1 and proliferation indicators in defining subgroups of meningiomas with a more aggressive biological behavior.     

Clinical epidemiology and survival of progressive multifocal leukoencephalopathy in the era of highly active antiretroviral therapy: Data from the Italian Registry Investigative Neuro AIDS (IRINA)

Human immunodeficiency virus (HIV)-associated progressive multifocal leukoencephalopathy (PML) remains a relevant clinical problem even in the era of highly active antiretroviral therapy (HAART). Aims of the study were to analyze clinical and treatment-related features and the survival probability of PML patients observed within the Italian Registry Investigative Neuro AIDS (IRINA) during a 29-month period of HAART. Intravenous drug use, the presence of focal signs, and the involvement of white matter at neuroradiology increased the risk of having PML. A reduced probability of PML was observed when meningeal signs were reported. Patients starting HAART at PML diagnosis and previously naïve for antiretrovirals showed significantly higher 1-year probability of survival (.58), compared to those continuing HAART (.24), or never receiving HAART (.00). Higher CD4 cell count were associated with a higher survival probability (.45). At multivariate analysis, a younger age, higher CD4, starting HAART at PML diagnosis, the absence of previous acquired immunodeficiency syndrome (AIDS)-defining events, and the absence of a severe neurologic impairment were all associated with a reduced hazard of death. The use of cidofovir showed a trend towards a reduced risk of death.