Alois Strasser

miR-17, miR-19b, miR-20a, and miR-106a are down-regulated in human aging.

Aging is a multifactorial process where deterioration of body functions is driven by stochastic damage while counteracted by distinct genetically encoded repair systems. To better understand the genetic component of aging, many studies have addressed the gene and protein expression profiles of various aging model systems engaging different organisms from yeast to human. The recently identified small non‐coding miRNAs are potent post‐transcriptional regulators that can modify the expression of up to several hundred target genes per single miRNA, similar to transcription factors. Increasing evidence shows that miRNAs contribute to the regulation of most if not all important physiological processes, including aging. However, so far the contribution of miRNAs to age‐related and senescence‐related changes in gene expression remains elusive. To address this question, we have selected four replicative cell aging models including endothelial cells, replicated CD8+ T cells, renal proximal tubular epithelial cells, and skin fibroblasts. Further included were three organismal aging models including foreskin, mesenchymal stem cells, and CD8+ T cell populations from old and young donors. Using locked nucleic acid‐based miRNA microarrays, we identified four commonly regulated miRNAs, miR‐17 down‐regulated in all seven; miR‐19b and miR‐20a, down‐regulated in six models; and miR‐106a down‐regulated in five models. Decrease in these miRNAs correlated with increased transcript levels of some established target genes, especially the cdk inhibitor p21/CDKN1A. These results establish miRNAs as novel markers of cell aging in humans.

L-arginine induces dopamine release from the striatum in vivo

Recent reports indicate that induction of nitric oxide (NO) evokes dopamine (DA) release from the striatum in vitro. In this study, we used L-arginine (L-Arg) to demonstrate the in vivo stimulation of DA release from the striatum of Mongolian gerbils using microdialysis. The content of DA in the striatal extracellular fluid (ECF) increased 7–15-fold in the presence of L-Arg in the perfusate as compared with that of the controls (DA level in drug-free perfusate varied from 0.050 ± 0.009 to 0.092 ± 0.023 pmol 10 μUl−1). Simultaneous perfusion of L-Arg with nitro-L-arginine (NLA), an inhibitor of nitric oxide synthase, markedly reduced the L-Arg effect on DA release from the striatum. The NLA-perfused animals contained DA levels significantly lower than those observed in the control striatal dialysate. These findings indicate for the first time that DA release in vivo can be induced by L-Arg, the precursor of NO. The data strongly suggest that NO may modulate striatal DA release.

Cerebral postischemic hypoperfusion is mediated by ETA receptors

Effect of ETA-receptor antagonist, BQ123, on postischemic hypoperfusion in the presence or absence of nitric oxide synthetase inhibitor, N omega-nitro-L-arginine (NLA), was investigated in Mongolian gerbils. BQ123 given prior to ischemia reversed the early incomplete recovery of cerebral blood flow observed with NLA without affecting the late postischemic hypoperfusion. Additional postischemic administration of BQ123 also reversed (P < 0.01) the late postischemic hypoperfusion seen in NLA-, N omega-nitro-D-arginine methyl ester- or Ringers-treated animals.

Immune modulation following immunization with polyvalent vaccines in dogs.

A decline in T-cell-mediated immunity and transient state of immunosuppression after immunization has been reported in dogs. Nevertheless, dogs are still routinely vaccinated with polyvalent live vaccines and severe disease does not generally occur. In order to investigate these effects on the canine immune system and to elucidate possible mechanisms we determined the following immune parameters in the blood of 33 clinically sound German shepherd dogs before and after standard vaccination with a polyvalent vaccine against distemper, parvovirus, viral hepatitis, leptospirosis, kennel cough and rabies: white and differential blood cell count, the serum concentrations and/or activities of IL-1, IL-2, IFN-gamma, TNF-alpha, neopterin and IgG, natural killer (NK) cell activity, bactericidal activity and complement hemolytic activity, lymphocyte proliferation test (LPT) and nitroblue tetrazolium test (NBT). Our major findings were that significant postvaccinal decreases in T-cell mitogenic response to PHA and in neutrophil function and neopterin serum concentration were accompanied by simultaneous increase in plasma IgG and hemolytic complement activity. This suggests a transient shift in the balance between cell-mediated and humoral (T(H)1/T(H)2) immunity rather than immunosuppression. These results do not imply that dogs should not receive live vaccines, as the response to vaccines just seems to create a state of altered homeostasis when immunization elicits protection by humoral and cell-mediated immunity. However, these recognized compromises of immune function should be considered and vaccines still be applied only in healthy animals and strictly according to the rules and regulations given by the manufacturer.

A simple method for the simultaneous separation of peripheral blood mononuclear and polymorphonuclear cells in the dog

A simple method for the simultaneous separation and purification of peripheral blood mononuclear cells (PBMC) and polymorphonuclear neutrophil cells (PMNC) was developed for comparative and functional studies in the immune system of the dog. Purity and cell viability were > 95%, yields were similar to those obtained by other techniques but without red blood cell contamination. Differential blood cell count studies of the isolated cells in blood samples of beagle dogs and German shepherd dogs demonstrated that the 1.077/1.119 double density centrifugation is an effective method of acquiring both highly purified blood mononuclear cells and polymorphonuclear cells as separate entities from the same sample. The interface between plasma and 1.077 contained an average 97% blood mononuclear cells vs. 3% polymorphonuclear cells, and the interface between 1.077 and 1.119 an average 96% polymorphonuclear cells vs. 4% blood mononuclear cells. These data indicate that Histopaque 1.077/1.119 double density gradient allows the purification and physical separation of lymphocytes and phagocytes from a blood sample in the dog, enabling the investigator to examine both cell types from the same sample simultaneously.

Modulation of striatal dopamine release in cerebral ischemia byL-arginine

The effect ofL-arginine, the precursor of nitric oxide, on ischemic dopamine release from the striatum was investigated in Mongolian gerbils subjected to bilateral carotid artery occlusion (15 min) alone or with reflow (2 h). Dopamine and its metabolites were measured in the striatal extracellular space dialysate after continuous perfusion (2 μl/min) of artificial extracellular fluid in the presence or absence of 15 mmol/literL- orD-arginine or 1 mmol/liter nitro-L-arginine.L-Arginine but notD-arginine increased the striatal content of dopamine in pre- and postischemia whereas it lowered the levels of dopamine and 3-methoxytyramine induced by ischemia. In contrast, nitro-L-arginine reduced the preischemic levels of dopamine and 3,4-dihydroxyphenyl-acetic acid, and had no effect on the ischemic release of dopamine. These findings indicate thatL-arginine stereospecifically modified the ischemic release and metabolism of dopamine. The data also suggest that the basal level of nitric oxide is not involved in dopamine release during ischemia but may participate in regulating dopamine release under physiological conditions.

Nitro-L-arginine augments the endothelin-1 content of cerebrospinal fluid induced by cerebral ischemia

The effect of nitro-L-arginine (NLA), inhibitor of NO synthase, on ET-1 content in cerebrospinal fluid (CSF) and on the vascular system was investigated in global ischemia/reperfusion of Mongolian gerbils. The results indicate that NLA induced a prolonged (2-3-fold) increase of ET-1 concentration above that seen in the CSF of untreated animals during ischemia/reperfusion. Both the transient and prolonged rise of ET-1 content observed in the CSF coincided with the reduction in the cerebral blood flow seen in untreated and NLA-treated gerbils, respectively, at the time of reperfusion.

Effects of feed contaminant deoxynivalenol on plasma cytokines and mRNA expression of immune genes in the intestine of broiler chickens.

An experiment was conducted to investigate the individual and combined effects of dietary deoxynivalenol (DON) and a microbial feed additive on plasma cytokine level and on the expression of immune relevant genes in jejunal tissues of broilers. A total of 40 broiler chicks were obtained from a commercial hatchery and divided randomly into four groups (10 birds per group). Birds were reared in battery cages from one day old for 5 weeks. The dietary groups were 1) control birds fed basal diet; 2) DON group fed basal diet contaminated with 10 mg DON/ kg feed; 3) DON + Mycofix group fed basal diet contaminated with 10 mg DON/ kg feed and supplemented with a commercial feed additive, Mycofix® Select (MS) (2.5 kg/ton of feed); 4) Mycofix group fed basal diet supplemented with MS (2.5 kg/ton of feed). At 35 days, the plasma levels of tumor necrosis factor alpha (TNF-α) and interleukin 8 (IL-8) were quantified by ELISA test kits. Furthermore, the mRNA expression of TNF-α, IL-8, IL-1β, interferon gamma (IFNγ), transforming growth factor beta receptor I (TGFBR1) and nuclear factor kappa-light-chain-enhancer of activated B cells 1 (NF-κβ1) in jejunum were quantified by qRT-PCR. The results showed that the plasma TNF-α decreased in response to DON, while in combination with MS, the effect of DON was reduced. DON down-regulated the relative gene expression of IL-1β, TGFBR1 and IFN-γ, and addition of MS to the DON contaminated diet compensates these effects on IL-1β, TGFBR1 but not for IFN-γ. Furthermore, supplementation of MS to either DON contaminated or control diet up-regulated the mRNA expression of NF-κβ1. In conclusion, DON has the potential to provoke and modulate immunological reactions of broilers and subsequently could increase their susceptibility to disease. The additive seemed to have almost as much of an effect as DON, albeit on different genes.

The Impact of Environment in Comparison with Moderate Physical Exercise and Dietary Restriction on BDNF in the Cerebral Parietotemporal Cortex of Aged Sprague-Dawley Rats

Background and Aims: Moderate physical exercise, dietary restriction as well as enriched environment in separate studies have been reported to delay some of the adverse effects of aging on brain function, parallel to an increase in brain-derived neurotrophic factor (BDNF). In order to elucidate these influences in a comparative setting, we examined the tissue concentrations of BDNF in the cerebral parietotemporal cortex of old Sprague-Dawley rats. Methods: Male Sprague-Dawley rats were divided randomly into six groups, living from 5 months (baseline group BL) up to 23 months of age as follows: voluntary running in wheels (RW), food restricted by feeding to pair weight with RW animals (PW), forced running on treadmills (TM), and sedentary controls with ad libitum access to food, either housed individually (S1) or in groups of 4 animals (S4). BDNF concentrations were determined by a commercially available ELISA. Results: We found higher BNDF concentrations in the 5 months old animals than in the 23 months old animals of group S1. The old sedentary group S4 showed significantly higher BNDF concentrations in comparison with the old individually caged groups RW, TM, PW and S1. Their BNDF concentrations were even higher than those of the young baseline group. Conclusions: Our data suggest that housing and social interactions have more influence on BDNF concentrations in the cerebral parietotemporal cortex of aging Sprague-Dawley rats than physical exercise and food restriction.

Antioxidant properties of the vasoactive endocannabinoid, 2-arachidonoyl glycerol (2-AG)

Reactive oxygen species (ROS) were shown to play a role in altering blood-brain barrier (BBB) permeability and formation of brain edema induced by trauma and/or ischemia. 2-arachidonoyl glycerol (2-AG), a novel, potent vasodilatory and cytoprotective endocannabinoid has been implicated to act as an antioxidative agent. This study examines: 1) the possible 2-AG modulation of BBB injury and edema formation induced by closed head injury (CHI); and 2) comparable effects between 2-AG and 4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl (TPL), a known antioxidant nitroxide on endothelial Ca2+ and cytoskeletal responses to H2O2 (ROS). 2-AG treatment reduced the CHI-induced increase in BBB permeability and brain edema. The endothelial H2O2-stimulated Ca2+ mobilization and cytoskeleton (vimentin) rearrangement was modified by either 2-AG or TPL. These findings provide evidence of 2-AG antioxidant activity and are consistent with the involvement of ROS in the pathomechanism of CHI-induced BBB injury and brain edema.