[Treatment of postoperative ventral hernias with the use of mesh implants made of titanium thread]. / Lechenie posleoperatsionnykh ventral'nykh gryzh s ispol'zovaniem setchatykh implantov iz titanovoi niti.

AIM: Study of the results of the use of titanium thread mesh implants in the treatment of postoperative ventral hernias. METHODS: The study included 84 patients with postoperative ventral hernias. All performed open-access prosthetic hernioplasty. 2 groups were formed: the main group consisted of 32 patients who used a mesh implant made of titanium thread as an implant, the comparison group consisted of 52 patients whose hernioplasty was performed using a polypropylene mesh implant. There were no statistically significant differences between groups of patients by age, gender, average body mass index, risk class of anesthesia (ASA), size and location of hernias. RESULTS: The frequency of postoperative complications in the main group was 6.2%, in the comparison group - 3.8%. There were no statistically significant differences in this indicator between the groups. At the time of discharge from the hospital, the level of plasma C-reactive protein in patients of the main group was significantly lower than in patients of the comparison group. CONCLUSION: The use of titanium thread mesh implants in the treatment of postoperative ventral hernias is accompanied by a less inflammatory response of the body to the implant and does not lead to an increase in the frequency of postoperative complications.

[Titanium mesh implants in herniology]. / Titanovye setchatye implanty v gerniologii.

Literature review is devoted to the main implants used in hernia surgery and their disadvantages. Advisability of titanium mesh implants in surgery of anterior abdominal wall hernias is shown.

Identification of autophosphorylation sites in c-Yes purified from rat liver plasma membranes.

c-Yes was purified 322-fold from a rat liver plasma membrane fraction to a single 60-kDa band on SDS-PAGE. The purified protein contained essentially no phosphotyrosine residues and was autophosphorylated with Mg2+. ATP exclusively at tyrosine residues with a concomitant increase in the protein-tyrosine kinase activity. The autophosphorylated c-Yes was extensively digested by trypsin and the resultant two major phosphopeptides, peptides I and II, were purified by HPLC on a reversed-phase C-18 column. The amino acid sequence of peptide I was determined to be LIEDNEYTAR, which is identical with the sequence from Leu-418 through Arg-427 of mouse c-Yes, indicating that one of the autophosphorylation sites corresponds to Tyr-424 of the mouse c-Yes. After partial determination of the N-terminal sequence of 10 amino acid residues of peptide II, the 230 bp sequence of rat cDNA that encodes the N-terminal 76 amino acid residues of c-Yes covering peptide II, was determined. From the predicted amino acid sequence, the sequence of peptide II was assumed to be from Tyr-16 through Lys-46, YTPENPTEPVNTSAGHYGVEHATAATTSSTK. The purified c-Yes phosphorylated the tyrosine residue of synthetic peptides covering Tyr-32 and its surrounding sequence but did not phosphorylate peptides covering Tyr-16 and its surrounding sequence, suggesting that the other autophosphorylation site is Tyr-32.

Purification and characterization of a possible protooncogene fyn product, p59fyn, from a rat brain particulate fraction.

Four tyrosine-protein kinases that reacted with antibodies specific to p62c-yes, p60c-src, p60c-src+, and p59fyn, respectively, were solubilized from a rat brain particulate fraction and separated by casein-Toyopearl column chromatography. Possible p59fyn, with a pI of 6.5, was purified 490-fold as a single 59-kDa protein band on SDS-PAGE. The purified enzyme contained almost no phosphotyrosine residues but was autophosphorylated with Mg2+. ATP exclusively at tyrosine residues, with a concomitant increase in the kinase activity toward tyrosine-glutamate (1:4) copolymers. The rate of the copolymer phosphorylation was proportional to the square of the enzyme concentration, suggesting activation through intermolecular catalysis. In the presence of Mn2+, however, the reaction showed a first-order dependence on the enzyme concentration.

Purification and characterization of a rat liver membrane tyrosine-protein kinase, the possible protooncogene c-yes product, p60c-yes.

A tyrosine-protein kinase was purified more than 270-fold from the rat liver plasma membrane fraction by successive column chromatographies on Sephacryl S-300, wheat germ agglutinin-agarose, casein-Sepharose, and hydroxylapatite, followed by isoelectrofocusing electrophoresis. The enzyme with pI of 6.2 was a 60-kDa single polypeptide which represented 42% of total protein. The enzyme reacted quantitatively with a monoclonal antibody to the amino-terminal sequence (Cys-3 to Ser-66) specific to the human c-yes protein, but not with antibodies to the specific amino-terminal sequences of the c-src, fyn, and lck proteins. The purified enzyme contained almost no phosphotyrosine residue but was autophosphorylated with Mg.ATP exclusively at tyrosine residues with concomitant increase in the kinase activity. The rates of autophosphorylation of the enzyme and phosphorylation of tyrosine-glutamate (1:4) copolymers, catalyzed by the enzyme were proportional to the square of enzyme concentration, suggesting that p60c-yes undergoes autophosphorylation through intermolecular catalysis, resulting in stimulation of the enzyme activity. Although the enzyme reaction showed an essential requirement for Mg2+ or Mn2+ with optimal concentrations of 20 and 3 mM, respectively, autophosphorylation significantly activated the enzyme only in the presence of Mg2+. Autophosphorylation of the enzyme reduced the Km for tyrosine-glutamate copolymers and tubulin, but not for ATP, and increased the Vmax of copolymer and tubulin phosphorylation.

Metal ion binding sites of bacteriorhodopsin. Laser-induced lanthanide luminescence study.

Laser-excited luminescence lifetimes of lanthanide ions bound to bacteriorhodopsin have been measured in deionized membranes. The luminescence titration curve, as well as the binding curve of apomembrane (retinal-free) with Eu3+, has shown that the removal of the retinal does not significantly affect the affinity of Eu3+ for the two high affinity sites of bacteriorhodopsin. The D2O effects on decay rate constants indicate that Eu3+ bound to the high affinity sites of native membrane or apomembrane is coordinated by about six ligands in the first coordination sphere. Tb3+ is shown to be coordinated by four ligands. The data indicate that metal ions bind to the protein with a specific geometry. From intermetal energy transfer experiments using Eu3+-Pr3+, Tb3+-Ho3+, and Tb3+-Er3+, the distance between the two high affinity sites is estimated to be 7-8 A.

Effects of arginine modification on the photocycle of halorhodopsin.

Exhaustive reaction with phenylglyoxal removed 9 of the 12 arginine and 1 of the 2 lysine residues in detergent-solubilized halorhodopsin, without affecting the chromophore. The consequences of this extensive removal of positive charges on various chloride-binding equilibria and the photochemistry were evaluated. No significant effects were seen on the affinity of Site I to chloride and on the increase in the pKa of Schiff-base deprotonation, which is caused by the chloride binding at this site. No significant effects were seen on the affinity of Site II to chloride, either. However, the photocycle of the pigment was affected. Kinetic modeling of the observed changes in flash-induced absorption changes suggests that the modification increases the affinity of the main halorhodopsin photointermediate to chloride by about fourfold. If chloride translocation involves release of chloride from this intermediate during the transport cycle, the result might explain the observed partial inhibitory effects on chloride transport. Plausible models of chloride translocation include reversible binding of the anion by positively charged groups, strategically arranged in the protein. The results indicate that two of the three spectroscopically observable chloride-dependent equilibria do not depend on a large number of positively charged residues in the protein. To the extent that the unaffected equilibria represent association and dissociation which occur during chloride translocation, at least part of the chloride translocation might be accomplished with the participation of only a few positively charged residues.

Characterization of metal ion-binding sites in bacteriorhodopsin.

We have investigated the effects of the binding of various metal ions to cation-free bacteriorhodopsin ("blue membrane"). The following have been measured: shift of the absorption maximum from 603 to 558 nm (blue to purple transition), binding isotherms, the release of H+ upon binding, and the decay of the deprotonated intermediate of the photocycle, M412. We find that all cations of the lanthanide series, as well as the alkali and alkali earth metals earlier investigated, are able to bring about the absorption shift, whereas Hg2+ and Pt4+ are not. Sigmoidal spectroscopic titration curves and nonsigmoidal binding curves suggest that there are two high affinity sites for cations in bacteriorhodopsin. Binding to the site with the second highest affinity is responsible for the absorption shift. Divalent cation binding to blue membrane causes release of about six protons, whereas higher numbers of protons are released by trivalent cations, suggesting that the shift of absorption maximum involves proton release from carboxyl group(s). The metal ion bound to this site must be surrounded by carboxyl oxygen atoms acting together as a multidentate ligand with a specific geometry because multivalent ions are effective only when capable of octahedral coordination. Lanthanide ions dramatically inhibit M412 decay at pH above 6.3, an effect probably due to binding to lipid phosphoryl groups.

[Vessels of the macro- and microcirculatory bed of the heart in the early developmental stage of acute myocardial infarct viewed by angioroentgenology, histology and electron microscopy]. / Sosudy makro-mikrogemotsirkuliatornogo rusla serdtsa na rannei stadii razvitiia ostrogo infarkta miokarda v angiorentgenologicheskom, gistologicheskom i élektronno-mikroskopichesm osveshchenii.

A report is based on the investigation of the hearts of persons who died at an early ischemic (up to twenty-four hours) stage of the acute myocardial infarction studied under the light microscope (39 cases) and at the ultrastructural level (5 cases). An original technique of the combined filling of the macro- and microvascular bed was applied as well as the intravascular silver impregnation and special histologic methods for detection of the early lesions of cardiomyocytes were used in some cases. The interrelationship and sequence of lesions of the coronary arteries and vessels of the cardiac microcirculatory bed are shown. The lesions of microvessels bear mainly a secondary character and depend upon the duration of myocardial infarction, its dimensions and its deepness. The ultrastructural alterations of microvessels are characterized by the heterogeneity and dependence on the degree and character of the cardiomyocyte lesion. A morphological estimation of the compensatory-adaptive alterations in the cardiac vascular bed at the early stages of myocardial infarction development is given.

Evidence for a sulfhydryl group near the retinal-binding site of halorhodopsin.

Amino acid analysis of the halorhodopsin chromoprotein shows that this protein contains a cysteine residue. Such a residue is absent in bacteriorhodopsin. Low concentrations (micromolar) of HgCl2 inhibit light-dependent chloride transport by halorhodopsin in envelope vesicles prepared from Halobacterium halobium strain L-33 and increase the Km for chloride. The decay rate of the flash-induced absorption change of halorhodopsin, measured at 570 nm, is considerably slowed by HgCl2, and this effect is reversed at higher concentrations of chloride. In addition, the magnitude of the absorption changes is diminished by HgCl2. These effects of the mercurial are also seen with the purified, solubilized chromoprotein. Upon addition of HgCl2 to the chromoprotein at low chloride concentrations in the dark, a decrease of absorption at 580 nm and an increase at 380 nm occur, as well as a blue shift of the chromophore by about 20 nm. Sustained illumination of halorhodopsin results in a 410 nm photoproduct. The reconversion of this species to 580 nm in the dark is strongly inhibited by HgCl2. These results show that a thiol group is essential for the stability of the halorhodopsin chromophore and for its photochemical reactions and suggest that this group is in the vicinity of both the retinal Schiff's base and the chloride-binding site.

Halide binding by the purified halorhodopsin chromoprotein. I. Effects on the chromophore.

The halorhodopsin chromoprotein, a retinal-protein complex with an apparent molecular mass of 20 kilo-daltons, exhibits all of the halide-dependent effects found for the chromophore of functional halorhodopsin in cell envelope vesicles. With increasing halide concentration (a) an alkali-dependent 580/410 nm chromophore equilibrium (attributed to reversible deprotonation of the retinal Schiff's base) is shifted toward the 580-nm chromophore and (b) the flash-induced photocycle proceeds increasingly via P520, rather than via P660. The halide-binding site(s) responsible for these effects must reside, therefore, in the chromoprotein. Chloride and bromide are about equivalent, but iodide is much less effective in these effects and in being transported. Several other anions, i.e. thiocyanate, nitrate, phosphate, and acetate, affect the absorption maximum of the chromophore but do not allow the production of P520 upon flash illumination and are not transported. However, these ions appear to compete with chloride in the flash experiments. These observations suggest that binding of anions to a relatively nonspecific site affects the protonation state of the Schiff's base in the chromophore. Either this site directly or a more specific site, connected to the first one by a sequential pathway, is involved with the photocycle intermediates and with chloride transport by halorhodopsin.

Oxidation of reactive sulfhydryl groups of sarcoplasmic reticulum ATPase.

The role of reactive sulfhydryl groups of sarcoplasmic reticulum ATPase has been investigated. Incubation of ATPase with 17 mol o-iodosobenzoic acid per mol ATPase results in a 15% inhibition of Ca2+ uptake with only a 5% loss of ATPase activity. When ATPase is treated with 15 mol KMnO4 per mol ATPase, Ca2+ uptake is completely inhibited. From the measurement of remaining SH groups using 5,5'-dithiobis-(2-nitrobenzoic acid), it is found that the oxidation of approximately four SH groups per ATPase molecule with KMnO4 leads to a complete loss of Ca2+ uptake, while the oxidation of five SH groups per ATPase with o-iodosobenzoic acid results in only 15% inhibition of Ca2+ uptake. The results of amino acid analysis indicate that KMnO4 oxidizes the reactive SH groups to sulfonic acid groups. Among the five o-iodosobenzoic acid-reactive SH groups, at least one shows a distinct Ca2+ dependence. Addition of o-iodosobenzoic acid to the reaction medium containing KMnO4 does not increase the number of oxidized SH groups, indicating that both o-iodosobenzoic acid and KMnO4 oxidize the same SH groups of the enzyme. The different effects of two oxidizing agents on sarcoplasmic reticulum ATPase eliminate the possibility of direct involvement of SH group(s) in the ATPase reaction.

18O-probes of phosphoenzyme formation and cooperativity with sarcoplasmic reticulum ATPase.

ATP is known to produce unusual modulations of catalytic steps in its own hydrolysis by sarcoplasmic reticulum ATPase both in the micromolar range and in the millimolar range of ATP concentration. The nature of these modulations can be probed with 18O-exchange techniques. Particularly valuable are recent approaches based on measurement of the [18O]Pi species formed from highly 18O-labeled ATP. Such analyses show that as ATP is decreased in the micromolar range there is a much greater tendency for reversal of hydrolysis of the phosphoryl enzyme prior to release of Pi to the medium. No modulation of steps involved in oxygen exchange is seen in the millimolar concentration range. Ways are presented in which the modulation at lower ATP could result from interaction of ATP at independent catalytic sites.

Characterization of medium inorganic phosphate-water exchange catalyzed by sarcoplasmic reticulum vesicles.

Effects of temperature, Ca2+, and ATP on the extent and characteristics of the medium Pi in equilibrium HOH exchange catalyzed by sarcoplasmic reticulum ATPase are reported. Measurements of the patterns of [18O]Pi species formed from highly labeled [18O]Pi show that a single catalytic pathway is involved in the rapid medium Pi in equilibrium HOH exchange with Mg2+ present and the much slower exchange with both Mg2+ and Ca2+ present. A continued high rate of exchange is observed when ATP concentration is increased up to 5 mM even though the amount of phosphoenzyme formed from Pi is much greater at lower ATP concentrations. This result reveals that binding of ATP in some manner causes a pronounced increase in the rate constant for hydrolysis of the phosphoenzyme. During the rapid oxygen exchange in the absence of Ca2+ at 10 and 30 degrees C, the rate of Pi release from the enzyme-Pi complex is about 5-6 times greater than the rate of phosphoenzyme formation. Both Pi release and phosphoenzyme formation are much slower in the presence of Ca2+, with a greater relative tendency for phosphoenzyme formation, particularly at the lower temperature.

Modification of rabbit muscle phosphorylase b by a water-soluble carbodiimide.

Glycogen phosphorylase b from rabbit muscle was rapidly inactivated by incubation with 1-cyclohexyl-3-(2-morpholinyl-(4)-ethyl)carbodiimide metho-p-toluenesulfonate (CMC) at pH 5.1. The inactivation was pH-dependent and was not restored by treatment with hydroxylamine. The addition of glycine ethyl ester or N-(2,4-dinitrophenyl)-ethylenediamine (DNP-EDA) markedly increased the rate of inactivation. Of the various amino analogs of glucose tested, only glucosyl amine accelerated the inactivation, although they are all bound to the glucose 1-phosphate site of the enzyme. In the absence of amines, incorporation of about 3 mol of [metho-14C]CMC per protein monomer was observed on complete inactivation. In the presence of DNP-EDA, however, only 2 mol of [metho-14C]CMC and 1 mol of DNP-EDA were incorporated before the activity was completely lost. The treatment of phosphorylase b with CMC did not change the Km values of the enzyme for glucose 1-phosphate and AMP, in spite of the 56% inactivation. It is suggested that, in the phosphorylase-catalyzed reaction, an essential carboxyl group of the enzyme plays a role in the protonation of the glucosidic oxygen of glucose 1-phosphate.

Affinity of glucose analogs for alpha-glucan phosphorylases from rabbit muscle and potato tubers.

The action of phosphorylase b from rabbit muscle and potato phosphorylase was inhibited to various extents by several glucose analogs. Like glucose itself, all of the glucosidic oxygen-substituted analogs tested in kinetic experiments showed a nonlinear competitive inhibition for muscle phosphorylase b and a linear competitive one for potato phosphorylase. 5-Thio-D-glucose, one of the ring oxygen-substituted analogs, also inhibited the action of muscle phosphorylase b in the same manner, while the inhibition pattern of 5-amino-D-glucose (nojirimycin) was of a linear noncompetitive type. Since the conformation of 5-amino-D-glucose in aqueous solution is half-chair (Reese et al. (1971) Carbohyd. Res. 18, 381-388), the unusual kinetic behavior of the compound toward muscle phosphorylase b was supposed to be due to its half-chair conformation. In the glucosidic oxygen-substituted analogs, the affinity for both muscle phosphorylase b and potato phosphorylase decreased with decreasing order of magnitude of electronegativity of the glucosidic atom. The strong positive correlation between the affinity and the electronegativity suggests that D-glucose-1-P, the substrate, may bind to phosphorylase with the formation of a hydrogen bond between its glucosidic oxygen and a hydrogen donor of the enzyme.

Inhibition of alpha-glucan phosphorylase by alpha-D-glucopyranosyl fluoride.

Alpha-D-Glucopyranosyl fluoride was found to inhibit strongly the action of alpha-glucan phosphorylase b[EC 2.4.1.1] from rabbit muscle, and that of the enzyme from potato tubers rather weakly. The inhibition is highly specific, being competitive with respect to glucose 1-phosphate and noncompetitive with respect to polysaccharide, during polysaccharide synthesis. In the reverse process, it is competitive with respect to Pi. These results have been explained by assuming that the inhibitor binds to the glucose 1-phosphate site of the enzyme, occupying both subsites which normally bind the glucosyl and phosphate moities of the substrate, but does not directly interact with the polysaccharide site. Based on this assumption, the dissociation constants of the enzyme-inhibitor and enzyme-polysaccharide-inhibitor complexes have been evaluated (0.43 and 0.20 mM for the muscle enzyme, respectively; 24 and 23 mM for the potato enzyme, respectively). Glucosyl fluoride also acts as a noncompetitive inhibitor with respect to AMP. A high concentration of AMP causes an inhibitory effect on the action of the muscle enzyme, the effect being menifested in the presence of glucosyl fluoride.

Alpha-glucan phosphorylase from sweet potato: isolation and properties of the partially degraded enzyme.

Alpha-Glucan phosphorylase (EC 2.4.1.1.) was purified from sweet potato roots. Apparently homogeneous preparations obtained are partially degraded products from phosphorylase, as judged from the results of molecular weight determination, NH-2-termini analysis and pyridoxal-5'-P assay. Phosphorylase is shown to be degraded in the crude extract from sweet potato. The degradation is partly suppressed by EDTA and by salts and is accelerated by reducing agents. It is proposed that sweet potato phosphorylase in its intact form has a similar molecular structure and similar properties to the white potato enzyme. Both plant phosphorylases are preferentially cleaved by protease near the middle of their polypeptide chains without much loss of enzyme activity.