Interleukin-1, nitric oxide and reactive astrocytes

Recent work in many laboratories has revealed that cytokines are important mediators of inflammation, host defense, and tissue injury in a variety of neurological diseases. A role for astrocytes and microglia in these diseases has been considered pivotal, since both cell types readily produce and respond to cytokines in vitro and show morphologic and immunocytochemical evidence for activation in vivo. Although much of the work documenting these events has been generated in rodent systems, our laboratory has focused on human cell culture systems to define the nature of the activating signals for human microglia and astrocytes and their responses to activating cytokines and growth factors and evidence for activation. The results have shown that interleukin-1 (IL-1) is a potent activator of human astrocytes and induces cytokines such as tumor necrosis factor alpha and interleukin-6, and is a potent activator of nitric oxide generation in astrocytes. Astrocytes also promote microglial growth and differentiation through production of colony-stimulating factors, an activity that is enhanced following activation with IL-1. This review will summarize the human glia data generated in this and other laboratories and present hypotheses how glia may participate in certain human central nervous system diseases.     

Override of spontaneous respiratory pattern generator reduces cardiovascular parasympathetic influence

We investigated the effects of voluntary control of breathing on autonomic function in cardiovascular regulation. Variability in heart rate was compared between 5 min of spontaneous and controlled breathing. During controlled breathing, for 5 min, subjects voluntarily reproduced their own spontaneous breathing pattern (both rate and volume on a breath-by-breath basis). With the use of this experimental design, we could unmask the effects of voluntary override of the spontaneous respiratory pattern generator on autonomic function in cardiovascular regulation without the confounding effects of altered respiratory pattern. Results from 10 subjects showed that during voluntary control of breathing, mean values of heart rate and blood pressure increased, whereas fractal and spectral powers in heart rate in the respiratory frequency region decreased. End-tidal PCO2 was similar during spontaneous and controlled breathing. These results indicate that the act of voluntary control of breathing decreases the influence of the vagal component, which is the principal parasympathetic influence in cardiovascular regulation.     

Medicine taking in Southampton: a second look

1. A 1 in 200 sample of the Southampton electorate were sent a postal questionnaire in January 1993. Of the 756 adults surveyed, 400 (52.9%) returned completed questionnaires. One hundred and eighty-eight (47.0%) of the respondents had been prescribed a medicine within the previous month. 2. Compared with a survey 9 years earlier, medicine taking had increased amongst men (44.1% vs 33.7% NS) and drugs acting on the respiratory system were in more widespread use (19 vs 7 patients P < 0.05). 3. Patterns of storage of medicines were almost identical to those found in 1984. However, methods of disposal were significantly different with 34% of the respondents stating that they would return left-over medicines to the Doctor or Pharmacist compared with 17% in the previous study (P < 0.01). 4. Of those taking medicines 120 (63.8%) had received a manufacturers' information leaflet. Medicines used to treat disorders of the respiratory and cardiovascular systems were most likely to be accompanied by such a leaflet (74% and 70% respectively). 5. Those who received a leaflet were almost all satisfied by it. However, patient awareness of potential side effects remained poor with only 30% being aware of any which their medicine might cause. 6. Despite improvements in attitudes towards medicine taking over time patients awareness of potential adverse effects remains limited. Further research is necessary in order to determine how best to educate patients on this topic.     

Enhanced sensitivity of ubiquinone-deficient mutants of Saccharomyces cerevisiae to products of autoxidized polyunsaturated fatty acids

Coenzyme Q (ubiquinone or Q) plays a well known electron transport function in the respiratory chain, and recent evidence suggests that the reduced form of ubiquinone (QH2) may play a second role as a potent lipid-soluble antioxidant. To probe the function of QH2 as an antioxidant in vivo, we have made use of a Q-deficient strain of Saccharomyces cerevisiae harboring a deletion in the COQ3 gene [Clarke, C. F., Williams, W. & Teruya, J. H. (1991) J. Biol. Chem. 266, 16636-16644]. Q-deficient yeast and the wild-type parental strain were subjected to treatment with polyunsaturated fatty acids, which are prone to autoxidation and breakdown into toxic products. In this study we find that Q-deficient yeast are hypersensitive to the autoxidation products of linolenic acid and other polyunsaturated fatty acids. In contrast, the monounsaturated oleic acid, which is resistant to autoxidative breakdown, has no effect. The hypersensitivity of the coq3delta strains can be prevented by the presence of the COQ3 gene on a single copy plasmid, indicating that the sensitive phenotype results solely from the inability to produce Q. As a result of polyunsaturated fatty acid treatment, there is a marked elevation of lipid hydroperoxides in the coq3 mutant as compared with either wild-type or respiratory-deficient control strains. The hypersensitivity of the Q-deficient mutant can be rescued by the addition of butylated hydroxytoluene, alpha-tocopherol, or trolox, an aqueous soluble vitamin E analog. The results indicate that autoxidation products of polyunsaturated fatty acids mediate the cell killing and that QH2 plays an important role in vivo in protecting eukaryotic cells from these products.     

Expression of type II nitric oxide synthase in primary human astrocytes and microglia: role of IL-1beta and IL-1 receptor antagonist

In this work, we studied the expression of type II nitric oxide synthase (NOS) in primary cultures of human astrocytes and microglia. Cytokine-activated human fetal astrocytes expressed a 4.5-kb type II NOS mRNA that was first evident at 8 h, steadily increased through 48 h, and persisted through 72 h. The inducing signals for astrocyte NOS II mRNA expression were in the order IL-1beta + IFN-gamma > IL-1beta + TNF-alpha > IL-1beta. SDS-PAGE analysis of cytokine-stimulated astrocyte cultures revealed an approximately 130-kDa single NOS II band that was expressed strongly at 48 and 72 h (72 h > 48 h). Specific NOS II immunoreactivity was detected in cytokine-treated astrocytes, both in the cytosol and in a discrete paranuclear region, which corresponded to Golgi-like membranes on immunoelectron microscopy. In human microglia, cytokines and LPS failed to induce NOS II expression, while the same stimuli readily induced TNF-alpha expression. In cytokine-treated human astrocytes, neither NOS II mRNA/protein expression nor nitrite production was inhibited by TGF-beta, IL-4, or IL-10. In contrast, IL-1 receptor antagonist exerted near complete inhibition of NOS II mRNA and nitrite induction. Monocyte chemoattractant peptide-1 mRNA was induced in TGF-beta-treated astrocytes, demonstrating the presence of receptors for TGF-beta in astrocytes. These results confirm that in humans, cytokines stimulate astrocytes, but not microglia, to express NOS II belonging to the high output nitric oxide system similar to that found in rodent macrophages. They also show that the regulation of type II NOS expression in human glia differs significantly from that in rodent glia. A crucial role for the IL-1 pathway in the regulation of human astrocyte NOS II is shown, suggesting a potential role for IL-1 as a regulator of astrocyte activation in vivo.     

Gastric trichobezoar in a banner-tailed kangaroo rat (Dipodomys spectabilis)

A male, wild-caught kangaroo rat developed anorexia and wasting. The animal was euthanized and a gastric trichobezoar found at necropsy. The paucity of information regarding the clinical medicine of this species is a hindrance to those charged with the care of kangaroo rats. Gastric trichobezoar should be considered as a differential diagnosis in cases of anorexia in kangaroo rats.     

Autoxidation of ubiquinol-6 is independent of superoxide dismutase

Ubiquinone (Q) is an essential, lipid soluble, redox component of the mitochondrial respiratory chain. Much evidence suggests that ubiquinol (QH2) functions as an effective antioxidant in a number of membrane and biological systems by preventing peroxidative damage to lipids. It has been proposed that superoxide dismutase (SOD) may protect QH2 form autoxidation by acting either directly as a superoxide-semiquinone oxidoreductase or indirectly by scavenging superoxide. In this study, such an interaction between QH2 and SOD was tested by monitoring the fluorescence of cis-parinaric acid (cPN) incorporated phosphatidylcholine (PC) liposomes. Q6H2 was found to prevent both fluorescence decay and generation of lipid peroxides (LOOH) when peroxidation was initiated by the lipid-soluble azo initiator DAMP, dimethyl 2,2'-azobis (2-methylpropionate), while Q6 or SOD alone had no inhibitory effect. Addition of either SOD or catalase to Q6H2-containing liposomes had little effect on the rate of peroxidation even when incubated in 100% O2. Hence, the autoxidation of QH2 is a competing reaction that reduces the effectiveness of QH2 as an antioxidant and was not slowed by either SOD or catalase. The in vivo interaction of SOD and QH2 was also tested by employing yeast mutant strains harboring deletions in either CuZnSOD and/or MnSOD. The sod mutant yeast strains contained the same percent Q6H2 per cell as wild-type cells. These results indicate that the autoxidation of QH2 is independent of SOD.