Antioxidant enzyme systems in skeletal muscle atrophied by immobilization

To clarify the mechanism of oxidative stress in skeletal muscle atrophied by immobilization, we investigated the change of antioxidant enzyme activities in a typical slow red muscle, the soleus. Atrophied soleus muscles were collected from male Wistar rats (16 weeks old), one ankle joint of which had been immobilized in the fully extended position for 7 days. Also, soleus muscles were collected from intact age-matched rats as control. The activities of Mn-containing superoxide dismutase (Mn-SOD), Cu,Zn-containing superoxide dismutase (Cu,Zn-SOD), Se-dependent glutathione peroxidase (Se-GSHPx), glutathione S-transferase (GST), catalase, and glutathione reductase (GSSGRx) were measured. The activities of Cu,Zn-SOD, GST, and GSSGRx were significantly higher in atrophied muscles, while the others were unchanged. Increased Cu,Zn-SOD and unchanged Mn-SOD levels might reflect increased generation of superoxide anions in the cytoplasm rather than in the mitochondria. Owing to the enhancement of Cu,Zn-SOD and the unaltered Se-GSHPx and catalase activities, hydrogen peroxide is thought to be increased in the cytoplasm. Because there is also an increase of iron in the microsomes of atrophied muscles, the production of hydroxyl radicals, the most aggressive of radicals, might consequently be elevated.     

Antistressor effect of dalargin in rats with immobilization stress

Laboratory of Experimental Surgery, A. V. Vishnevskii Institute of Surgery, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR M. I. Kuzin.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 111, No. 6, pp. 617–619, June, 1991.     

Diversity of antisperm antibodies bound to sperm surface in male immunological infertility

PROBLEM: The presence of antisperm antibodies (ASA) in males can reduce fecundity, however, relationship between the two is disputed. This study was performed to investigate if there is diversity of ASA bound to sperm surface using immunobead test (IBT) combined with complement dependent sperm immobilization test (SIT). METHODS: The ASA bound to sperm surface were detected using the direct IBT (D-IBT) in 275 semen samples. In some cases with ASA detected by D-IBT, sperm immobilizing antibodies bound to sperm surface were also evaluated using direct SIT (D-SIT). RESULTS: The incidence of the immunoglobulin G (IgG), IgA, and IgM classes of ASA detected by D-IBT were 2.5, 1.8, and 0.4%, respectively. Totally, nine (3.3%) infertile men had ASA on the sperm surface. D-SIT was tested positive in four (66.7%) of six cases with ASA assessed by D-IBT. CONCLUSIONS: Some of the sperm-bound antibodies are associated with complement dependent sperm immobilizing antibodies, indicating that there exists a heterogeneity of sperm-bound antibodies. This result might be one of the reasons for the controversy about the relationship between ASA and immunological infertility in men.     

Influence of sperm immobilization on onset of Ca(2+) oscillations after ICSI

Sperm immobilization prior to intracytoplasmic sperm injection (ICSI) is thought to be necessary for efficient fertilization. A variety of methods of sperm immobilization (pipetting, squeezing and piezo application) are currently employed in ICSI. The effect of differences in immobilization method on the timing of initial Ca(2+) oscillations of oocytes in ICSI was investigated. Motile spermatozoa were immobilized in eosin Y solution using pipetting, squeezing and piezo application. Complete staining of the sperm head was achieved after 220.7, 42.2 and 5.0 s respectively. Oscillations after ICSI were measured fluorometrically for each method. The onset of Ca(2+) oscillations was observed at 4.8 to 80.4 min after ICSI. Ca(2+) oscillations developed earlier with the piezo method (14.4 +/- 6.4 min) than other methods (pipetting, 43.1 +/- 20.2 min, P < 0.01; squeezing, 18.4 +/- 3.8 min, P = NS). The piezo method produced the earliest staining of the sperm head and may have caused the most damage to the sperm membrane. A more rapid onset of Ca(2+) oscillations was also observed with the piezo method. The method of sperm immobilization may be important for the rapid release of sperm factors that initiate oocyte activation. This study also showed that Ca(2+) oscillations develop earlier in human oocytes treated by ICSI than indicated in previous reports.     

Correlation Between Regional Specificity of Antisperm Antibodies to the Spermatozoan Surface and Complement-Mediated Sperm Immobilization

ABSTRACT: Sera from men at risk for immunity to spermatozoa were screened for antisperm antibodies by immunobead binding following passive antibody transfer to antibody-free sperm of fertile donors. The percent motile sperm after incubation in diluted antibody positive serum in the presence of complement was compared with the regional distribution of immunoglobulins bound to the sperm surface. The extent of complement-mediated sperm immobilization varied with immunoglobulin class and with the location of antibody bound to the sperm surface. Tests utilizing complement-mediated immobilization of sperm are insensitive to the presence of antibodies of IgG and IgA classes that are directed against the head, the distal one-fifth of the sperm tail principal piece, or the tail end piece. A high degree of immobilization was found only when IgG binding occurred on the distal two-fifths to three-fifths of the principal piece of the tail or when IgM bound to the sperm tail end piece.     

Covalent immobilization of proteins onto photoactivated polystyrene microtiter plates for enzyme-linked immunosorbent assay procedures

Enhancement of the speed and sensitivity of an ELISA technique was achieved by doing it on a polystyrene microtiter plate preactivated by a simple photochemical reaction. Immobilization of Epicoccum nigrum antigen (allergenic antigen) or goat anti-rabbit IgG onto the photoactivated plates was found to occur in only 45 min with higher binding than that obtained through adsorption during the same period onto the untreated surface. Nearly 1.5-2-folds higher readings were obtained when the ELISA was carried out with the solid phase prepared on the photoactivated surface rather than on the untreated surface. Moreover, solid phases prepared on the activated surface could detect IgE (E. nigrum antibody) even at 1/50 (v/v) dilutions, whereas a solid phase prepared on the untreated surface failed to do so. Around three times higher ELISA values were obtained in the activated plate than the untreated plate when IgE was diluted to 1/5 (v/v). Such photoactivated surface could be of great importance in diagnostic tests involving the ELISA technique particularly to confirm false negative cases and for other immunoassays such as radioimmunoassay procedures.     

聚醚砜表面光固定脲酶的研究

我们利用含芳香叠氮基的光活性酯将脲酶光固定在聚醚砚（Polyether sulfone,PES）膜的表面。同时研究了紫外光辐照时间对固定化脲酶密度、固定化脲酶活性的影响，温度、PH值对自由脲酶、固定化脲酶相对活性的影响；并考察了固定化脲酶的相对活性随光活性酶浓度的变化、固定化脲酶的重复使用次数及储存稳定性等性质。结果表明，PES膜表面固定化脲酶的浓度为0.33mg/cm^2；当紫外光辐照时间为5分钟时，固定化脲酶的相对活性最高；固定化脲酶的最适Ph值和最适温度分别为7℃和50 ℃；在50℃时，连续使用12次后，固定化脲酶仍具有50%的催化活性；在4℃保存储存一个多月仍可保持80%的催化活性。     

The effect of immobilization on myotendinous junction: an ultrastructural,histochemical and immunohistochemical study

Kannus , P., Jozsa , L., Kvist , M., Lehto , M. & Järvinen , M. 1992. The effect of immobilization on myotendinous junction: an ultrastructural, histochemical and immunohistochemical study. Acta Physiol Scand 144 , 387–394. Received 28 April 1 991 , accepted 13 October 1991. ISSN 0001–6772. Tampere Research Station of Sports Medicine, UKK-Institute, and Department of Surgery, Tampere University Central Hospital, Tampere, Finland; Department of Morphology, National Institute of Traumatology, Budapest, Hungary; and Sports Medical Research Unit, Paavo Nurmi Center, University of Turku, Turku, Finland. The effect of immobilization on the myotendinous junction of the calf muscles in the rat was studied histochemically, immunohistochemically and morphometrically with a transmission electron microscope. After 3 weeks of immobilization, the contact area between the muscle cells and tendineal collagen fibres was reduced by almost 50% in both type I (slow-twitch) and type II (fast-twitch) muscle fibres. The terminal finger-like processes of the muscle cells became shallow and cylindrical or were completely atrophied. Their basal membranes were slightly thickened. Histochemically, the most remarkable alteration in the myotendinous junction was the marked decrease in the sulphate containing glyco-saminoglycans. In the basal lamina of the muscle fibres, the glycosaminoglycan and proteoglycan content was also reduced. Immunohistochemical analyses revealed that the amount of type III collagen was markedly increased on the myotendinous interface, but the amount and distribution of type I collagen was not affected by immobilization. These findings suggest that immobilization causes degenerative changes at the myotendinous junction, which, in turn, most likely decrease its tensile strength and may predispose it to rupture during activity.     

Development of a reversible bienzymatic system in immunoenzymatic technique

The use of specific captors solubilized with a ligand on to proteic membranes together with an automatic-computerized system is proposed for the determination of various haptens or antigens in biological and industrial fluids by an enzyme-linked immunoassay test. Two enzymes are used in this technique: the first enzyme for linking reversibly the immunocomplex to the insoluble matrix, the substrate of this enzyme being immobilized on the matrix; the second (beta-d-glucose oxidase) for labelling the antigen. Its activity is measured by fixing the immunocomplex gelatin membrane on to a pO(2) sensor. After incubation of the antigen labelled with glucose oxidase and the free antigen with specific antibodies linked with the first enzyme in a pre-established concentration, the reaction medium is introduced inside the continuous flow cell. O(2) consumption due to the enzyme reaction is measured in actual time when the electrode is in contact with the glucose standard solution. The signal is correlated by an injection of urea solution. The signal is processed with an automated microcomputer system.