The heat shock/oxidative stress connection

Involvement of free-radical oxidations in the aging process has been a topic of interest since Harman's original contribution. Because of the close association between aging and Alzheimer disease (AD) and the qualitative similarity in the neuropathology of both conditions, it has been proposed by many investigators that oxidative stress may be important in AD. If such modality of injury was indeed involved, one should expect to find markers of oxidation and heat shock (since free radicals are key mediators of heat-shock induction) in brains of patients with AD. In fact, several studies documented abnormal expression of antioxidant enzymes and heat-shock proteins (HSP) along with other markers of oxidation in AD brains. We showed that abnormally expressed antioxidant enzymes are topographically associated with senile plaques and neurofibrillary tangles, and that the activity of these enzymes is (contrary to what one would expect) markedly reduced. These findings have recently been confirmed by other investigators. Despite a large amount of evidence that suggests an association between oxidative stress and the pathogenesis of AD, it is not yet known whether oxidative stress is a cause or consequence of the disorder. Future research efforts regarding the oxidative stress hypothesis of AD should include attempts, at generating AD pathology by oxidative means in laboratory animals, determining the role and integrity of the heat-shock response in AD, as well as that of various antioxidant systems, growth factors, and hormones with antioxidant and neuroprotective properties.     

Human choriogonadotropin-induced coupling of receptor and Gs protein and the effect of hormone deglycosylation

The detergent-soluble extract of rat ovary plasma membranes contained a Gs protein of about 100 kDa as shown by its elution behavior on a Bio Gel A-1.5m column. However, the cell membranes exposed to hCG (37° C, 15 min) contained in addition a higher molecular weight Gs protein complex of 300 kDa comprised of human chorionic gonadotropin (hCG) receptor (hCGR) and Gs. The complex bound with an affinity column of GTP-Sepharose and could be released with Gpp(NH)p and GTP inhibited this binding. The presence of the hCGR in the complex was shown by its binding to ¹²⁵I-hCG. Furthermore, GTP inhibited the binding of hCG to the complex. These results indicate the presence of hCGR and Gs protein complex in the hCG-treated membranes. hCGR and Gs protein were individually purified and reconstituted into phospholipid vesicles. The protein-phospholipid vesicles showed saturation kinetics of binding of ¹²⁵I-hCG and ³H-Gpp(NH)p. Incubation of phospholipid vesicles with hCG resulted in a 2–3-fold increase in the binding of ³H-Gpp(NH)p and GTPase activity. Activation of Gs protein was dependent on the length of incubation and the hormone concentration. Deglycosylated hCG was about 10 times less potent than hCG suggesting a role of carbohydrates of hCG in inducing hCG-Gs protein interactions. The data with the in vitro reconstitution system rule out the involvement of a carbohydrate-binding lectin in the function of the hormone.     

The effects of interferon-beta on phorbol ester or calcium ionophore-induced intercellular adhesion molecule-I expression in epidermal carcinoma cells.

Keratinocyte intercellular adhesion molecule (ICAM)-I expression is induced by interferon (IFN)-gamma. It has been previously reported that IFN-beta suppresses IFN-gamma-induced ICAM-I expression in A431 cells, a human squamous cell carcinoma cell line. In this study, the suppression mechanisms were investigated at the post second messenger level. Both 12-O-tetradecanoylphorbol-13-acetate (TPA) and calcium ionophore (A23187) induce ICAM-I expression in A431 cells. ICAM-I expression induced by either was not suppressed with cotreatment with IFN-beta. Furthermore, IFN-beta did not inhibit the translocation of protein kinase C (PKC) by TPA. It appears that the pathways involved in ICAM-I expression induced by activation of PKC or increased in intracellular Ca++ are not affected by IFN-beta.     

锌离子对大鼠海马突触体Ca^2＋—Mg^2＋ATP酶活性及蛋白合成的影响

行为实验己经证明，锌过多或缺锌均可影响脑功能。锌作为体内重要的微量元素，影响多种酶的活性及蛋白质和核酸的台成。本实验通过体外分离大鼠脑海马突触体，观察不同浓度锌离子对Ca2 －Mg2 ATP酶的活性和3H－Leu掺入突触蛋白合成的影响．结果表明：1．锌离子浓度在25μmol／L时增加该酶的活性（＜0．01），并促进3H－Leur掺入蛋白质的合成（＜0．05）。2．锌离子在50,100，200μmol／L的较高浓度时对Co2 －M2 ATP酶的活性有显著的抑制作用（分别为：P＜0.05．P＜0．01，P＜0.01），仅200μmol／L对3H—Leu掺入突触蛋白合成有抑制作用。本研究提示：适量的锌对突触体功能的维持是必要的，但剂量过高则起相反作用。     

Influence of continuous infusion of interleukin-1 alpha on the core protein and the core protein fragments of the small proteoglycan decorin in cartilage.

Decorin, a collagen-binding small proteoglycan, is considered to have a specific function in the organization or stability of the collagen network. Therefore, alteration of its molecular properties may be of pathophysiological relevance during the development of cartilage damage. It is shown here that normal cartilage from rabbit knee-joint contains glycosaminoglycan chain-bearing core protein fragments of 39, 23, and 18 kDa, each one amounting to approximately 5-6% of the intact decorin core protein. Continuous infusion of human recombinant interleukin-1 alpha for 14 days (200 ng/day) into a knee-joint led in condylar cartilage to a reduction in the amount of intact core protein from 2 micrograms/mg wet tissue to about 1.1 micrograms/mg. The increase in its quantity found after infusion of heat-inactivated interleukin-1 was not statistically significant. The concentration of all three core protein fragments became reduced to a similar extent as the intact core protein under the influence of the cytokine, and additional fragments were not found. Surprisingly, there was a much smaller response to interleukin-1-treatment in patellar cartilage.     

Maintenance and synthesis of proteins for an anucleate axon

The anucleate (distal) segment of a crayfish medial giant axon (MGA) remains intact for months in vivo after severing the axon from its cell body, a phenomenon referred to as long-term survival (LTS). We collected axoplasm from chronic anucleate MGAs by perfusing 2-cm lengths of axons with an intracellular saline. This axoperfusate was analyzed by SDS-PAGE and silver stained. Axoperfusate proteins from intact MGAs and from chronic anucleate MGAs exhibiting LTS for up to 6 months were the same. Furthermore, immunoreactive levels of actin and β-tubulin were similar in axoperfusates from intact and chronic anucleate MGAs. This maintenance of proteins in chronic anucleate MGAs must be due to a lack of protein degradation and/or to local protein synthesis by a source other than the cell body. To investigate local protein synthesis in vitro, we added [³⁵S]-methionine to the extracellular saline surrounding intact and chronic anucleate MGAs. After 4- to 6-h incubations, radiolabelled proteins were detected in axoperfusates analyzed by SDS-PAGE and fluorography. The similarity between radiolabelled proteins in axoperfusates and MGA glial sheaths indicated a glial origin for the radiolabelled axoperfusate proteins. Various observations and control experiments suggested that glial-axonal protein transfer occurred by a physiological process. Glial-axonal protein transfer may contribute to the maintenance of proteins during LTS of chronic anucleate MGAs.     

Alcohol and sauna bathing: effects on cardiac rhythm, blood pressure, and serum electrolyte and cortisol concentrations.

The effect of heavy drinking and sauna bathing on cardiac rhythm, blood pressure, and serum electrolyte and cortisol concentrations was studied in 10 healthy male volunteers. Sauna bathing induced a comparable, significant increase in heart rate with and without alcohol consumption. During sauna bathing without alcohol, systolic blood pressure remained at the baseline level, whereas sauna and alcohol together decreased systolic blood pressure markedly from 136 +/- 4 to 113 +/- 3 mmHg (P less than 0.01). Neither sauna alone, nor sauna combined with alcohol intake, increased the frequency of premature ventricular complexes. Serum potassium, calcium and cortisol concentrations changed slightly during sauna, but alcohol consumption did not contribute further to this. In conclusion, sauna bathing, even in combination with heavy drinking, does not appear to provoke cardiac arrhythmias in healthy young men. However, the risk of hypotension is increased when sauna bathing is combined with alcohol consumption.