DESIGN AND SYNTHESIS OF NOVEL SYNTHETIC ANTIOXIDANTS FOR THE TREATMENT OF OXIDATIVE STRESS RELATED DISEASES

Free radicals play an important role in many physiological processes that occur in the human body such as cellular defense responses to infectious agents and a variety of cellular signaling pathways. While at low concentrations free radicals are involved in many significant metabolic reactions, high levels of free radicals can have deleterious effects on biomolecules like proteins, lipids, and DNA. Many physiological disorders such as diabetes, ageing, neurodegenerative diseases, and ischemia-reperfusion (I/R) injury are associated with oxidative stress.1 In particular, the deleterious effects caused by I/R injury developed during organ transplantation, cardiac infarct, and stroke have become the main cause of death in the United States and Europe.1,2 In this context, we synthesized and characterized a series of novel indole-amino acid conjugates as potential antioxidants for I/R injury. The synthesis of indole-phenol conjugate compounds is also discussed. Phenolic derivatives such as caffeic acid, butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), resveratrol, and its analogues are known for their significant antioxidative properties. A series of resveratrol analogues have been designed and synthesized as potential antioxidants. The radical scavenging mechanisms for potential antioxidants and assays for the in vitro evaluation of antioxidant activities are also discussed.

The moderating effect of personal mastery and the relations between stress and Plasminogen Activator Inhibitor-1 (PAI-1) antigen

OBJECTIVE: This study tested whether feelings of personal control over one's life circumstances (i.e., personal mastery) would attenuate the relations between stress (i.e., negative life events and caregiving distress) and Plasminogen Activator Inhibitor (PAI)-1 antigen, an inhibitor of fibrinolysis implicated in the development of cardiovascular disease. DESIGN: Seventy-one spousal dementia caregivers were assessed for plasma levels of PAI-1 antigen, negative life events, caregiver distress, and feelings of personal mastery. Regression analysis was used to determine if personal mastery moderated the relations between stress (i.e., life stress and caregiving distress) and PAI-1 antigen levels. MAIN OUTCOME MEASURE: Plasminogen activator inhibitor (PAI)-1 antigen in plasma. RESULTS: After controlling for other factors associated with PAI-1 antigen levels, negative life events were positively associated with plasma PAI-1 antigen concentrations in participants low in personal mastery (beta = .31; p = .050) but not in individuals high in personal mastery (beta = .22; p = .184). The moderating effect of mastery on the relations between caregiving distress and PAI-1 antigen did not reach statistical significance (p = .091). CONCLUSIONS: These data suggest that mastery may protect individuals from some of the alterations in hemostatic factors that have been linked to cardiovascular risk.

Measures of endothelial dysfunction in plasma of patients with posttraumatic stress disorder

Posttraumatic stress disorder (PTSD) confers an increased cardiovascular risk. In 14 otherwise healthy patients with PTSD and in 14 age- and gender-matched non-PTSD controls, we investigated whether the categorical diagnosis of PTSD and severity of PTSD symptom clusters (i.e. re-experiencing, avoidance, arousal, and overall score) would be associated with plasma concentrations of three markers of endothelial dysfunction [soluble tissue factor (sTF), von Willebrand factor (VWF), and soluble intercellular adhesion molecule (sICAM)-1]. Compared with controls, patients had significantly higher sTF; this difference became nonsignificant when controlling for psychological distress. VWF and sICAM-1 levels were not significantly different between patients and controls. In the entire sample virtually all PTSD symptom clusters correlated significantly and positively with sTF and VWF but not with sICAM-1. The correlation between symptoms of re-experiencing and sTF was significantly different between patients and controls. Controlling for symptoms of anxiety and depression (i.e. psychological distress) rendered most associations between PTSD symptom clusters and sTF nonsignificant, whereas controlling for age retained significance of associations with VWF. Posttraumatic stress showed a continuous relationship with sTF and VWF, with the former relationship being partly affected by psychological distress. This suggests one mechanism by which posttraumatic stress could contribute to atherosclerosis.

Autonomic stress responses elicited by watching a live broadcast soccer game: a pilot study

Aim: Increased rates of hospitalization due to cardiovascular events have been reported during phases of World Soccer Championships (WSC). The purpose of this pilot study was to explore acute psychological and physiological effects of watching a live broadcast soccer game during the WSC 2006. Methods: Seven male supporters (age: M=24; SD=2.7) of the Swiss National Soccer Team watched a game of their team in a controlled laboratory setting. Heart rate (HR), heart rate variability (HRV), salivary cortisol, alpha-amylase (sAA), and testosterone concentrations, as well as several mood ratings were captured repeatedly before, during, and after the game. Results: Subjects reported feeling stressed, and HR and sAA activity showed an increase during the game. In contrast, HRV, cortisol and testosterone were unaffected. Conclusion: Watching a sports competition seems to specifically affect the sympathetic nervous system, which can be measured by sensitive electrocardiographic and salivary markers.

No evidence for an association of posttraumatic stress disorder with circulating levels of CRP and IL-18 in a population-based study.

Several studies have shown associations of posttraumatic stress disorder (PTSD) with the development of cardiometabolic diseases. The underlying psychopathological mechanisms, including potential links to inflammatory processes, have been discussed but remain elusive. Therefore, the aim of the present study was to evaluate the association of PTSD symptoms with the inflammatory biomarkers C-reactive protein (CRP) and interleukin-18 (IL-18). The study population consisted of 3012 participants aged 32-81years drawn from the population-based KORA F4 study conducted in 2006-08 in the Augsburg region (Southern Germany). PTSD symptoms were measured by the Impact of Event Scale, the Posttraumatic Diagnostic Scale and interview data and classified as no, partial or full PTSD. The associations of PTSD with CRP and IL-18 concentrations were estimated by multiple regression analyses with adjustments for age, sex and cardiometabolic risk factors. Linear regression analyses showed no significant association between PTSD and CRP or IL-18 concentration: adjusted for age and sex, the geometric mean concentrations in participants with full PTSD was for CRP 9% lower and for IL-18 1% higher than in participants with no PTSD (p values 0.53 and 0.89). However, further analyses indicated that individuals with partial PTSD had an increased chance of belonging to the highest quartile of the IL-18 concentration. No significant association was observed for any of the three subscales intrusion, avoidance or hyperarousal with CRP or IL-18 concentration. This large, population-based study could not find an association of full PTSD with CRP and IL-18 concentrations. Further research is needed to analyse these relationships.

Satellite-Derived Variability in Chlorophyll, Wind Stress, Sea Surface Height, and Temperature in the Northern California Current System

Satellite-derived data provide the temporal means and seasonal and nonseasonal variability of four physical and biological parameters off Oregon and Washington ( 41 degrees - 48.5 degrees N). Eight years of data ( 1998 - 2005) are available for surface chlorophyll concentrations, sea surface temperature ( SST), and sea surface height, while six years of data ( 2000 - 2005) are available for surface wind stress. Strong cross-shelf and alongshore variability is apparent in the temporal mean and seasonal climatology of all four variables. Two latitudinal regions are identified and separated at 44 degrees - 46 degrees N, where the coastal ocean experiences a change in the direction of the mean alongshore wind stress, is influenced by topographic features, and has differing exposure to the Columbia River Plume. All these factors may play a part in defining the distinct regimes in the northern and southern regions. Nonseasonal signals account for similar to 60 - 75% of the dynamical variables. An empirical orthogonal function analysis shows stronger intra-annual variability for alongshore wind, coastal SST, and surface chlorophyll, with stronger interannual variability for surface height. Interannual variability can be caused by distant forcing from equatorial and basin-scale changes in circulation, or by more localized changes in regional winds, all of which can be found in the time series. Correlations are mostly as expected for upwelling systems on intra-annual timescales. Correlations of the interannual timescales are complicated by residual quasi-annual signals created by changes in the timing and strength of the seasonal cycles. Examination of the interannual time series, however, provides a convincing picture of the covariability of chlorophyll, surface temperature, and surface height, with some evidence of regional wind forcing.

Social support and oxytocin interact to suppress cortisol and subjective responses to psychosocial stress

BACKGROUND The presence of social support has been associated with decreased stress responsiveness. Recent animal studies suggest that the neuropeptide oxytocin is implicated both in prosocial behavior and in the central nervous control of neuroendocrine responses to stress. This study was designed to determine the effects of social support and oxytocin on cortisol, mood, and anxiety responses to psychosocial stress in humans. METHODS In a placebo-controlled, double-blind study, 37 healthy men were exposed to the Trier Social Stress Test. All participants were randomly assigned to receive intranasal oxytocin (24 IU) or placebo 50 min before stress, and either social support from their best friend during the preparation period or no social support. RESULTS Salivary free cortisol levels were suppressed by social support in response to stress. Comparisons of pre- and poststress anxiety levels revealed an anxiolytic effect of oxytocin. More importantly, the combination of oxytocin and social support exhibited the lowest cortisol concentrations as well as increased calmness and decreased anxiety during stress. CONCLUSIONS Oxytocin seems to enhance the buffering effect of social support on stress responsiveness. These results concur with data from animal research suggesting an important role of oxytocin as an underlying biological mechanism for stress-protective effects of positive social interactions.

Influence of Chilling Stress on the Intercellular Distribution of Assimilatory Sulfate Reduction and Thiols in Zea mays

Abstract: The effect of chilling on the intercellular distribution of mRNAs for enzymes of assimilatory sulfate reduction, the activity of adenosine 5′-phosphosulfate reductase (APR), and the level of glutathione was analysed in leaves and roots of maize (Zea mays L). At 25 °C the mRNAs for APR, ATP sulfurylase, and sulfite reductase accumulated in bundle-sheath only, whereas the mRNA for O-acetylserine sulfhydrylase was also detected in mesophyll cells. Glutathione was predominantly detected in mesophyll cells; however, oxidized glutathione was equally distributed between the two cell types. Chilling at 12 °C induced oxidative stress which resulted in increased concentrations of oxidized glutathione in both cell types and a prominent increase of APR mRNA and activity in bundle-sheath cells. After chilling, mRNAs for APR and sulfite reductase, as well as low APR activity, were detected in mesophyll cells. In roots, APR mRNA and activity were at higher levels in root tips than in the mature root and were greatly increased after chilling. These results demonstrate that chilling stress affected the levels and the intercellular distribution of mRNAs for enzymes of sulfate assimilation.

The Effect of Heavy Metal Stress on Avian Proximal Tubule Urate Secretion

In both humans and birds, urate is an important antioxidant when maintained at normal plasma concentrations. Though human kidneys primarily reabsorb filtered urate, while those of birds perform mostly secretion, both maintain urate levels at ~300microM. The importance of maintaining urate levels within the homeostatic range was observed when the study of several prominent diseases revealed an association with hyperuricemia. This study examined the effect of elevated zinc concentration on avian urate secretion. Here, acute exposure of chicken proximal tubule epithelial cells (cPTCs) to zinc stress had no effect on urate secretion, but prolonged zinc-induced cellular stress inhibited active transepithelial urate secretion with no change in Mrp4 expression, glucose transport, or transepithelial resistance. Moreover, zinc had no effect on urate transport by isolated brush border membrane vesicles, suggesting involvement of a more complex cellular stress adaptation. Previous work has demonstrated that AMP-activated protein kinase (AMPK), a critical metabolic regulator, conserves energy during cellular stress by shutting down ATP-utilizing processes and activating ATP-generating processes. Pharmacological activation of AMPK by AICAR produced decreased urate secretion by cPTCs similar to the effect seen with prolonged exposure to zinc, while the AMPK inhibitor Compound C prevented both AICAR and zinc inhibition of urate secretion, suggesting a stress induced mechanism of regulation. Supported by NSF. IACUC #A08-046.

Female-specific regulation of cytochrome P450 3As and their response to xenobiotic stress

Cytochrome P450 3As (CYP3As) are phase I enzymes responsible for metabolizing more than 50% of clinical drugs. Recent studies have revealed that expression of CYP3As is two-fold higher in women than in men leading to a faster metabolic clearance of therapeutic drugs in women. In this study, we analyzed the female specific rat CYP3A isoform, CYP3A9. We evaluated the effects of progesterone and estrogen on CYP3A9 regulation and showed a distinct role for estrogen in mediating female dominance of CYP3A9. We also observed changes in CYP3A9 expression at various stages of pregnancy which correlates well with varying physiological estradiol concentrations. In addition, by the in vitro data shows that estradiol mediated induction can be abrogated with estrogen receptor antagonist ICI182,780. We also identified three novel murine CYP3A isoforms CYP3A13, CYP3A41 and CYP3A44 and characterized their genomic structures and expression profiles. CYP3A41 and CYP3A44 show female specific expression but surprisingly this female dominance is not mediated via estrogen. Control male mice did not exhibit any CYP3A41 mRNA levels but showed minimal levels of CYP3A44. In order to gain insights into the governance ofαthe female specific genes, the hepatic regulation of CYP3A41 and CYP3A44 by the xeno-sensors PXR and CAR was examined. In female mice, pregnenolone-16α-carboxynitrile, suppressed CYP3A41 and CYP3A44 mRNA levels in PXR−/− background whereas dexamethasone-dependent suppression of CYP3A41 was mediated by PXR. In addition, phenobarbital challenge in PXR−/− revealed up-regulation of both CYP3A44, CYP3A41 levels only in males. No role for CAR was seen in the regulation of either CYP3A41 or CYP3A44 gene expression in female mice. Interestingly, PXR and CAR ligands induced male CYP3A44 levels in a receptor dependent fashion. This increase of CYP3A44 transcript in male mice is in contrast to the response seen in female mice, which clearly indicates an additional layer of regulation. Our findings suggest that gender plays a strategic role in directing the CAR/PXR mediated effects of CYP3A44/CYP3A41. This implies that differential regulation of female specific CYP3A isoforms may be the key to explain some of the gender differences observed in clearance of certain therapeutics like antidepressants and analgesics. ^

Salivary cortisol, psychological stress and depressive symptoms among patients undergoing colon cancer screenings

As the second leading cause of cancer-related deaths in the United States, colon cancer has a high cure rate if detected early by a colonoscopy (U.S. Cancer Statistics Working Group, 2007). However, more than 41 million at-risk Americans are not properly receiving colonoscopy screenings according to the recommendations of the Center for Disease Control. This study provides insight into the physiological and psychological benefits of the colonoscopy procedure over and above cancer detection and prevention. Thirty-six patients receiving colonoscopic screening at the University of Connecticut Health Center participated in this study. A questionnaire battery that assessed perceived stress, depressive symptoms, colon cancer related worry, and social support, and optional saliva sampling was completed 2 weeks prior to and post colonoscopy. It was hypothesized that salivary cortisol concentrations, perceived stress, and self-reported depressive symptoms would show significant decreases from pre to post colonoscopy, and that these variables would all be positively correlated with one another. Results showed significant, positive correlations between depressive symptoms and both salivary cortisol (r (34)= .348, p< .05) and perceived stress (r (34)= .635, p< .01). Morning salivary cortisol levels decreased significantly from pre to post colonoscopy to levels below the population mean (t (16)=-3.711, p<. 01). No such differences were observed in either perceived stress or depressive symptoms. These results indicate that by decreasing cortisol concentrations to levels below that of the population mean, the colonoscopy provided physiological health benefits to patients beyond cancer screening. From a health psychology standpoint, this may encourage some of the 41 million Americans not receiving proper colon cancer screenings to adopt this potentially life-saving health behavior.

Effects of salinity, temperature and cadmium stress on cadmium -binding protein in the grass shrimp, {\it Palaemonetes pugio}

The combined effects of salinity, temperature and cadmium stress on survival and adaptation through cadmium-binding protein (CdBP) accumulation were studied in the grass shrimp, Palaemonetes pugio. In 96-hour bioassays, shrimp were exposed to zero or one of three levels of cadmium, under one of six different salinity (15, 25, or 35$\perthous$) and temperature (20 or 30$\sp\circ$C) regimes. CdBP concentrations were quantified in survivors from the 24 exposure groups. Salinity and temperature did not affect survivorship unless the shrimp were also exposed to cadmium. Grass shrimp were most sensitive to cadmium at low salinity-high temperature, and least sensitive at high salinity-low temperature. The incidence of cadmium-associated black lesions in gill tissue was influenced by salinity and temperature stress. P. pugio produced a 10,000 dalton metallothionein-like CdBP when exposed to at least 0.1 mg Cd$\sp{2+}$/L for 96 hours. Accumulation of CdBP was increased with increases in the exposure cadmium level, increases in temperature and decreases in salinity, independently and in conjunction with one another. Maximum CdBP concentrations occurred in grass shrimp that survived the salinity-temperature-cadmium conditions creating maximum stress as measured by highest mortality, not necessarily in shrimp exposed to the highest cadmium levels. The potential utility of this method as a monitor of physiological stress in estuarine biota inhabiting metal-polluted environments is discussed. ^

Concentrations of metals and POPs in liver tissue of wild sea bird chicks from the Norwegian arctic region

Lysosomal membrane stability, lipofuscin (LF), malondialdehyde (MDA), neutral lipid (NL) levels, as well as halogenated organic compounds (HOCs), Cr, Cd, Pb and Fe concentrations were analyzed in liver of black-legged kittiwake (BK), herring gull (HG), and northern fulmar (NF) chicks. There were significant species differences in the levels of NL, LF and lysosomal membrane stability. These parameters were not associated with the respective HOC concentrations. LF accumulation was associated with increasing Cr, Cd and Pb concentrations. HG presented the lowest lysosomal membrane stability and the highest. LF and NL levels, which indicated impaired lysosomes in HG compared to NF and BK. Lipid peroxidation was associated with HOC and Fe2+ levels. Specific HOCs showed positive and significant correlations with MDA levels in HG. The study indicates that contaminant exposure can affect lysosomal and lipid associated parameters in seabird chicks even at low exposure levels. These parameters may be suitable markers of contaminant induced stress in arctic seabirds.

(Table 1) Concentrations of persistant organic pollutants in brain and liver of glaucous gulls, Bjørnøya, Svalbard archipelago

Dead and dying glaucous gulls (Larus hyperboreus) were collected on Bjornoya in the Barents Sea in 2003, 2004 and 2005. Autopsies of the seabirds only explained a clear cause of death for three (14%) of the 21 birds. A total of 71% of the birds were emaciated. Liver and brain samples were analysed for organochlorine pesticides (OCPs), polychlorinated biphenyls (PCBs), polybrominated diphenyl ether (PBDEs), hexabromo-cyclododecanes (HBCDs) and mercury (Hg). High levels of OCPs, PCBs, PBDEs and alpha-HBCD were found in liver and brain. Compared to the dead and dying glaucous gulls found 1989, the congeners' composition tended to change toward more persistent compounds in the 2003-2005 samples. The brain levels of OCPs and PCBs did not differ between 1989 and 2003-2005, while the liver levels were significantly lower. The brain/liver ratio for PCB and PBDE significantly decreased with halogenations of the molecule, indicating a clear discrimination of highly halogenated PCBs and PBDEs entering the brain. There was further a clear negative correlation between contaminant concentrations and body condition. The brain levels were not as high as earlier published lethal levels of p,p'-DDE or PCB. However, more recent studies reported a range of sub-lethal OCP- and PCB-related effects in randomly sampled glaucous gulls. An additional elevation of pollutants due to emaciation may increase the stress of the already affected birds. The high brain levels of OCP, PCB and PBDE of present study might therefore have contributed to the death of weakened individuals of glaucous gull.

CO2-driven seawater acidification increases photochemical stress in a green alga

Increased CO2 and associated acidification in seawater, known as ocean acidification, decreases calcification of most marine calcifying organisms. However, there is little information available on how marine macroalgae would respond to the chemical changes caused by seawater acidification. We hypothesized that down-regulation of bicarbonate acquisition by algae under increased acidity and CO2 levels would lower the threshold above which photosynthetically active radiation (PAR) becomes excessive. Juveniles of Ulva prolifera derived from zoospores were grown at ambient (390 µatm) and elevated (1000 µatm) CO2 concentrations for 80 days before the hypothesis was tested. Here, the CO2-induced seawater acidification increased the quantum yield under low levels of light, but induced higher nonphotochemical quenching under high light. At the same time, the PAR level at which photosynthesis became saturated was decreased and the photosynthetic affinity for CO2 or inorganic carbon decreased in the high-CO2 grown plants. These findings indicated that ocean acidification, as an environmental stressor, can reduce the threshold above which PAR becomes excessive.