PGE1 abolishes the mitochondrial-independent cell death pathway induced by D-galactosamine in primary culture of rat hepatocytes

Background and Aim:  PGE₁ reduces in vivo and in vitro D-galactosamine (D-GalN)-induced cell death in hepatocytes. The present study was undertaken to elucidate the intracellular pathway by which D-GalN induces cell death in cultured hepatocytes. In addition, we evaluated if PGE₁ was able to modulate different parameters related to D-GalN-induced apoptosis in cultured rat hepatocytes.
Methods:  Hepatocytes were isolated from male Wistar rats (225–275 g) by the classical collagenase procedure. PGE₁ (1 µM) was administered 2 h before D-GalN (5 mM) in primary culture of rat hepatocytes. Apoptosis was determined by DNA fragmentation and caspase-3, -6, -8 and -9 activation in hepatocytes. Caspase activation was evaluated by the detection of the related cleaved product and its associated activity. Cell necrosis was determined by the measurement of lactate dehydrogenase (LDH) activity in culture medium. To elucidate the role of mitochondria, we measured neutral (nSMase) and acid (aSMase) sphingomyelinase, as well as the expression of cytochrome c in mitochondria and cytoplasm fractions from D-GalN treated hepatocytes.
Results:  D-GalN induced caspase-3 activation and DNA fragmentation in hepatocytes. This apoptotic response was not associated with the activation of caspase-6, -8 or -9. The use of specific inhibitors confirmed that only caspase-3 was involved in D-GalN-induced apoptosis. D-GalN did not modify nSMase and aSMase activities, nor mitochondrial cytochrome c release in hepatocytes.
Conclusions:  D-GalN induced apoptosis through caspase-3 activation but without modification of the activity of caspase-6, -8, -9, SMases or cytochrome c release. PGE₁ appears to prevent D-GalN-induced apoptosis by a mitochondria-independent mechanism.     

Anti-mitochondrial antibodies in patients with dilated cardiomyopathy (anti-M7) are directed against flavoenzymes with covalently bound FAD

Anti-mitochondrial antibodies (anti-M7) in sera from patients with dilated cardiomyopathy and myocarditis recognize, besides mitochondrial antigens, bacterial sarcosine dehydrogenase. The common target antigen was identified as the covalently bound FAD of mitochondrial and bacterial flavoenzymes. Thus, anti-M7-positive serum reacted on Western blots exclusively with covalently flavinylated enzymes. The antigenic specificity of anti-M7 sera was reproduced by an antiserum raised in rabbits with 6-hydroxy- D -nicotine oxidase. The heart mitochondrial membrane antigen recognized by anti-M7 serum was identified as the flavoprotein subunit of succinate dehydrogenase, the antigens in rat liver mitochondrial matrix as the flavoenzymes dimethylglycine dehydrogenase and sarcosine dehydrogenase. Anti-M7 serum contained a specific anti-flavoenzyme antibody fraction. Nanomolar concentrations of FAD and riboflavin inhibited the immune reaction on Western blots and in ELISA, and incubation with FAD-agarose depleted the anti-M7 activity of the serum. N-terminally deleted dimethylglycine dehydrogenase proteins were only immunoprecipitated by anti-M7 sera when the FAD was covalently incorporated. An affinity constant (K_D) of 10^?8 M was established for the anti-flavoenzyme antibodies by competitive ELISA. Of patients with cardiomyopathy and myocarditis, 36% and 25%, respectively, were anti-flavoenzyme-positive by Western blot and ELISA, but only two of 15 patients with other heart diseases and none of 50 healthy controls.     

维吾尔族和汉族弱精子症病人精子mtDNA4977bp缺失相关性研究

目的探讨维吾尔族和汉族弱精子症病人精子mtDNA~(4977bp)缺失的相关性。方法本文采用聚合酶链反应(PCR)技术对60例标本分两组,维、汉各30例,其中精子活力正常标本(对照组)和弱精子症患者(观察组)各15例进行了精子线粒体DNA(mtDNA)缺失的研究。结果发现15例做内对照的维族精子活力正常人中仅有3例(20%)有mtDNA~(4977bp)缺失；而15例维吾尔族弱精子症患者中有10例(67%) mtDNA~(4977bp)缺失；两组4977bp缺失频率比较有显著性差异(P＜0．05)。15例做内对照的汉族精子活力正常标本中无mtDNA~(4977bp)缺失；15例汉族弱精子症患者中有12例(80%)mtDNA~(4977bp)缺失。两组4977bp缺失频率比较差异有显著性(P＜0．05)。同时发现维吾尔族男性不育症与汉族男性不育症mtDNA~(4977bp)缺失频率无显著性差异。结论维吾尔族和汉族mtDNA~(4977bp)缺失在弱精子症的发病中可能起重要作用。     

Inflammaging as a Major Characteristic of Old People: Can It Be Prevented or Cured?

Widespread aging at the population level is a recent phenomenon that emerged in affluent societies. Inflammation is necessary to cope with damaging agents and is crucial for survival, particularly to cope with acute inflammation during our reproductive years. But chronic exposure to a variety of antigens, especially to some viruses such as cytomegalovirus, for a period much longer than that predicted by evolution, induces a chronic low-grade inflammatory status that contributes to age-associated morbidity and mortality. This condition carries the proposed name "inflammaging". Centenarians are unique in that, despite high levels of pro-inflammatory markers, they also exhibit anti-inflammatory markers that may delay disease onset. The key to successful aging and longevity is to decrease chronic inflammation without compromising an acute response when exposed to pathogens.     

Superoxide dismutases of muscle in mitochondrial encephalomyopathies

Lmmunohistochemical analyses were made of the superoxide dismutases (Mn-SOD and CuiZn-SOD) in biopsied muscles from 7 patients with mitochondrial encephalomyopathies that included mitochondrial encephalomyopathy, lactic acidosis and strokelike episodes (MELAS), and chronic progressive external ophthalmoplegia (CPEO). Mn-SOD mainly was present in the subsarcolemmal region, but it also was found in a coarsely granular, reticular, or diffuse pattern of staining within the muscle fibers. These Mn-SOD-positive fibers corresponded almost completely to the ragged-red fibers. The immunoreaction for CuiZn-SOD was weakly positive in some of the muscle fibers positive for Mn-SOD. In CPEO, Mn-SOD-positive fibers predominantly showed decreased cytochrome c oxidase (COX) activity. In MELAS, Mn-SOD-positive fibers tended to be stained deeply for COX although a few were COX-negative. These findings suggest that Mn-SOD-positive fibers can be used to make a differential diagnosis between CPEO and MELAS and that in mitochondrial encephalomyopathies Mn-SOD in the raggedred fibers may protect against oxidative stress. © John Wiley & Sons, Inc.     

Calcium transport and ATPase activity in mitochondria and fragments of sarcoplasmic reticulum of the heart and muscles of rabbits with hypercholesteremia

Keeping rabbits on a high-cholesterol diet (1 g/kg) for 3–7 months led to an increase in cholesterol concentration in the mitochondrial membranes and fragments of the sarcoplasmic reticulum (SPR) of the myocardium and skeletal muscles. Saturation of the membranes with cholesterol led to a decrease in efficiency of the Ca-pump of the SPR, as reflected in lowering of the Ca/ATP ratio and an increase in the outflow of Ca⁺⁺ from the SPR. Under these conditions the rate of accumulation of Ca⁺⁺ was higher in SPR than in the mitochondria. Activity of mitochondrial Mg⁺⁺-activated 2,4-DNP-ATPase was reduced in hypercholesteremia.Laboratory of Molecular Pathology and Biochemistry, Institute of General Pathology and Pathological Physiology, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR A. M. Chernukh.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 89, No. 3, pp. 292–294, March, 1980.     

Electron-cytochemical investigation of the brain mitochondria at various times after death

An electron-cytochemical investigation was made of oxidation of 3,3-diaminobenzidine (DAB) in the brain of rats and man at different times after death. The oxidation product of DAB was localized in the mitochondria, lipofuscin granules, and erythrocytes. Oxidation of DAB by rat and human brain mitochondria was shown to be only very slightly depressed even 2 days after death.Laboratory of Experimental Pathology and Pathomorphology of the Brain, Institute of Psychiatry, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR A. V. Snezhnevskii.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 81, No. 6, pp. 757–759, June, 1976.     

Liver and Heart Mitochondrial Succinate Dehydrogenase Activity of Newborn Rats in Anoxic Hypoxia and Starvation

Succinate dehydrogenase activity was determined in the liver and heart of newborn rats after 3 and 48 hours' exposure to anoxic hypoxia (10% O₂) and after 48 hours' starvation. Control determinations were made on newborn animals of corresponding ages, full term foetuses (21 days), infantile (1 and 2 weeks) and full grown animals. Hypoxia for 3 h had no influence on succinate dehydrogenase activity at all in either the heart or liver mitochondria of the newborn animals. After 48 h no difference was observed in the liver between the hypoxic animals and the starved controls of the same age, though starvation itself had resulted in a significant increase in activity, as much as 42%. When liver mitochondrial succinate dehydrogenase in normal mitochondria was activated by preincubation mitochondria with the substrate, the activity increase obtained was greater than that resulting from starvation. The increase in activity in the heart of the hypoxic or starved animals was not significant (< 10%).