Paolo B. DePetrillo

Serum 25-Hydroxyvitamin D and the Incidence of Acute Viral Respiratory Tract Infections in Healthy Adults

Background Declining serum concentrations of 25-hydroxyvitamin D seen in the fall and winter as distance increases from the equator may be a factor in the seasonal increased prevalence of influenza and other viral infections. This study was done to determine if serum 25-hydroxyvitamin D concentrations correlated with the incidence of acute viral respiratory tract infections. Methodology/Findings In this prospective cohort study serial monthly concentrations of 25-hydroxyvitamin D were measured over the fall and winter 2009–2010 in 198 healthy adults, blinded to the nature of the substance being measured. The participants were evaluated for the development of any acute respiratory tract infections by investigators blinded to the 25-hydroxyvitamin D concentrations. The incidence of infection in participants with different concentrations of vitamin D was determined. One hundred ninety-five (98.5%) of the enrolled participants completed the study. Light skin pigmentation, lean body mass, and supplementation with vitamin D were found to correlate with higher concentrations of 25-hydroxyvitamin D. Concentrations of 38 ng/ml or more were associated with a significant (p<0.0001) two-fold reduction in the risk of developing acute respiratory tract infections and with a marked reduction in the percentages of days ill. Conclusions/Significance Maintenance of a 25-hydroxyvitamin D serum concentration of 38 ng/ml or higher should significantly reduce the incidence of acute viral respiratory tract infections and the burden of illness caused thereby, at least during the fall and winter in temperate zones. The findings of the present study provide direction for and call for future interventional studies examining the efficacy of vitamin D supplementation in reducing the incidence and severity of specific viral infections, including influenza, in the general population and in subpopulations with lower 25-hydroxyvitamin D concentrations, such as pregnant women, dark skinned individuals, and the obese.

Determining the Hurst exponent of fractal time series and its application to electrocardiographic analysis

An alternative regression-based method for estimating the Hurst coefficient of a fractal time series is proposed. A formal mathematical description of the methodology is presented. The geometric relationship of the algorithm to the family of self-similar fractal curves is outlined. The computational structure of the algorithm is optimal for generation of real-time estimates of H. We show that the method can be applied to biologically-derived time series such as the cardiac interbeat interval and we obtain estimates of H from several diverse electrocardiographic data sets.

Serotonin transporter promoter polymorphism and monoamine oxidase type A VNTR allelic variants together influence alcohol binge drinking risk in young women.

The short allelic variant of the serotonin transporter protein promoter polymorphism (5HTTLPR) appears to influence binge drinking in college students. Both monoamine oxidase type A (MAOA) and the serotonin transporter protein are involved in the processing of serotonin, and allelic variants are both associated with differences in the efficiency of expression. We hypothesized that a significant gene × gene interaction would further stratify the risk of binge drinking in this population. Participants were college students (n = 412) who completed the College Alcohol Study, used to measure binge drinking behaviors. Genomic DNA was extracted from saliva for PCR based genotyping. The risk function for binge drinking was modeled using logistic regression, with final model fit P < 0.0005. This model was valid only for Caucasian females (n = 223), but the power to detect sex and ethnic effects was small. Young Caucasian women carrying higher expression MAOA VNTR alleles homozygous for the short allelic variant of the 5HTTLPR demonstrated the highest rate of binge drinking by self‐report, odds ratio (genotype odds: population odds) and 95% confidence intervals, 3.11 (1.14–18.10). Individuals carrying higher expression MAOA VNTR alleles carrying at least one long 5HTTLPR allelic variant had the lowest risk of binge drinking 0.46 (0.28–0.71). These results support the hypothesis that binge drinking behavior in young adulthood may be influenced by neurobiological differences in serotonergic function conferred by functional polymorphisms in genes involved in serotonin processing.

Ritonavir inhibition of calcium-activated neutral proteases.

Calpains (EC 3.4.22.17) are intracellular calcium-activated cysteine proteases that mediate tissue injury following post-ischemic and post-traumatic stress. Both human HIV protease and calpains share a similar secondary structure, where the active site is flanked by hydrophobic regions. The present study demonstrates that ritonavir, a hydrophobic HIV protease inhibitor, also inhibits calpain activity. In PC12 cell extracts assayed for calpain at maximal activity (2mM calcium), ritonavir exhibited competitive inhibition with a K(i) of 11+/-7.0 microM. Experiments with purified enzymes showed inhibition for both m- and mu-calpain isoforms (m-calpain, K(i)=9.2+/-1.2 microM; mu-calpain, K(i)=5.9+/-1.4 microM). Ritonavir also inhibited calcium-stimulated calpain activity in PC12 cells in situ. These results suggest that ritonavir or analogues of the drug should be investigated as cytoprotective agents in conditions where cell death or injury is mediated via calpain activation.

Initial ethanol exposure results in decreased heart rate variability in ethanol-naive rhesus monkeys.

Ethanols effects on heart rate variability may contribute to the increased cardiac disease and mortality observed in alcoholics. We assessed cardiac response to ethanol in seven previously ethanol-naive monkeys given a standard dose of ethanol, or saline. Ethanol exposure reduced cardiac signal complexity [mean+/-S.D. (ethanol: Hurst parameter=0.39+/-0.02; saline: Hurst parameter=0.32+/-0.06)] and increased the spectral exponent (ethanol: beta=1.36+/-0.35; saline: beta=1.12+/-0.35) when compared to saline, while heart rate itself was unaffected (saline: interbeat interval=303.57+/-24.57; ethanol: interbeat interval=308.14+/-20.45). Taken together with data that show autonomic disregulation in alcoholics, these findings provide further evidence of deleterious ethanol effects on cardiac signal dynamics.

Differential effects of MK801 and lorazepam on heart rate variability in adolescent rhesus monkeys (Macaca Mulatta)

Previous research shows that ketamine significantly alters cardiac signal regulation in rhesus monkeys, however relatively little is known about the mechanism for this effect. In the study reported here the relative contributions of NMDA receptor activation on cardiac signal dynamics were determined by administering a specific NMDA antagonist, MK801, to rhesus monkeys. The general effects of sedation were assessed by measuring cardiac response to lorazepam, a sedative drug without direct NMDA receptor activity. Electrocardiographic signal dynamics were examined before and after I.V. administration of either MK801 (0.16 mg/kg) or lorazepam (0.48 mg/kg). Inter-beat interval time series data were analyzed in the frequency domain after Fourier transform, and a nonlinear measure of autocorrelation, the Hurst exponent (H), was derived. After MK801 administration, log [HF /Total power] increased post-infusion (M = 1.11, SD = 0.45) compared with pre-infusion values [M = −0.19, SD = 0.32, F(1,4) = 19.49, P = 0.01] while H decreased, mean pre versus post 0.52+/−S.D. 0.10 versus 0.01+/− 0.05, P = 0.0002. Lorazepam administration did not significantly alter heart rate variability measures obtained in the frequency or nonlinear domains. To our knowledge, this is the first study that has defined the effects of peripherally administered MK801 on cardiovascular dynamics in primates and establishes that peripheral administration of NMDA antagonists result in large increases the high-frequency components of cardiac rhythm and increased heart rate variability compatible with MK801-associated increases in parasympathetic outflow.

Calcium-Activated Neutral Protease Activity Is Decreased in PC12 Cells After Ethanol Exposure

Abstract: Calcium‐activated neutral protease activity was determined in PC12 cells exposed to ethanol for 96 h using a fluorescence‐based assay with N‐succinyl‐Leu‐Tyr 7‐amido‐4‐methylcoumarin as the substrate. Stimulated activity was measured at high (1,400 µM) or low (140 µM) Ca2+ concentrations in the presence of 20 µM ionomycin. Kinetic parameters were derived by fitting a model relating fluorescence intensity to time: Ft = Ffinal*(1 −e−kobst). Cell extracts were subjected to nondenaturing gel electrophoresis and casein zymography with quantification of the activity of the two calpain isoforms. Exposure to ethanol significantly decreased whole cell calpain activity measured by kobs beginning at 20 mM, to 27.8% of control at 1,400 µM Ca2+ and 29.2% of control at 140 µM Ca2+ in the presence of 20 µM ionomycin. No changes in μ‐calpain or m‐calpain activities were found in cell extracts from cells exposed to 20 mM ethanol, whereas at 40 and 80 mM ethanol, significant decreases in both μ‐calpain and m‐calpain activities were discovered.

Inhibition of rat PC12 cell calpain activity by glutathione, oxidized glutathione and nitric oxide

Calpain, a calcium activated neutral protease, is involved in mediating neurotoxicity resulting from conditions of oxidative stress and free radical formation, such as hypoxia and ischemia. Nitric oxide (NO) may also be involved in modulating the cytotoxic effects of oxidative stress. We investigated the roles of reduced glutathione (GSH), oxidized glutathione (GSSG), and NO in modulating calpain activity in PC12 cells. Cell extracts were treated with GSSG, GSH, or the NO-donor S-nitroso-N-acetylpenicillamine. Calpain activity was determined by means of a fluorescent assay. Non-linear regression analysis was used to determine the type of inhibition (competitive, uncompetitive, or non-competitive). GSH displayed uncompetitive inhibition, with K(i)=7.0+/-2.0 mM (Mean+/-SEM) while GSSG exhibited competitive inhibition with K(i)=2.5+/-0.3 mM. NO was an irreversible inhibitor of calpain activity. These results suggest that both GSH and GSSG may be important physiological modulators of calpain activity.

Ritonavir protects hippocampal neurons against oxidative stress-induced apoptosis.

Oxidative stress plays an important role in many neurodegenerative conditions including Alzheimers disease and Parkinsons disease. 4-Hydroxynonenal (HNE), a lipid-soluble aldehydic product of membrane peroxidation, has been known to decrease neuronal survival by impairing Na+, K+, and -ATPase activity. HNE also increases neuronal vulnerability to excitotoxic injury and disrupts homeostasis by activating proteases which mediate the destruction of cellular protein and structure. The present study demonstrated that the hydrophobic HIV protease inhibitor, ritonavir inhibited HNE-mediated apoptosis in hippocampal primary neurons. In neurons exposed to oxidative stress induced by HNE (1 microM), ritonavir at 100 pM increased cell survival and completely abolished the apoptotic effects of HNE (P < 0.01). Ritonavir and its analogues might have useful cytoprotective effects for use in limiting the natural course of tissue injury after conditions where oxidative stress plays a role.

Sex differences in total body water in adolescent rhesus macaques estimated by ethanol dilution

Abstract:  Non‐human primates are widely used in research, yet relatively few studies have addressed potential pharmacokinetic differences between males and females. The present study examined the relationship between total body water, sex, age, and weight in the rhesus macaque (Macaca mulatta). Ethanol‐naïve, adolescent rhesus macaques (n = 119) were administered ethanol (males, 2.1 g/kg; females, 2.0 g/kg) intravenously, and blood samples for blood ethanol concentration obtained at 5, 10, and 60 minutes following the end of the infusion. Non‐linear regression was used to compare and contrast a series of pharmacokinetic models examining the relationship between weight, sex, age, Vd and zero‐order elimination rate. Vd (mean ± SEM) for male rhesus was 0.771 ± 0.008 l/kg and for females was 0.730 ± 0.008 l/kg, different at P < 0.00001. There were no sex differences in the rate of zero‐order ethanol elimination, estimated to be 0.0032 ± 0.0004 g/kg/minute. The data reported here may be useful in designing and interpreting pharmacokinetic studies using rhesus monkeys.