Effect of contrycal on activity of dehydrogenases and their isozymes in muscles and organs of animals with developing granulation tissue

The effect of contrycal on the state of the enzyme systems of the muscles, liver, kidneys, and heart was investigated in rats with developing granulation tissue. This protease inhibitor was found to stimulated lactate and malate dehydrogenase activity and also the isozyme spectrum of these enzymes. The action of the inhibitor was manifested as a change in the state of the enzyme systems both at the site of injury (granulations and underlying tissue) and in certain internal organs (liver and kidneys).Presented by Academician of the Academy of Medical Sciences of the USSR A. A. Vishnevskii [deceased].Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 82, No. 7, pp. 807–810, July, 1976.     

Effect of organosilicon compounds on protein biosynthesis in granulation and fibrous tissue

The effect of propoxysilatrane (POS) on biosynthesis of the principal biopolymers of connective tissue was studied. Administration of POS as 0.5 and 2.0% ointments in lanolin-petrolatum base cause stimulation of cell proliferation in granulation and fibrous tissues developing in open skin defects in albino rats. Stimulation of cell proliferation in these animals was shown to be accompanied by increased biosynthesis of collagen and noncollagen proteins. In concentrations of 10^–3–10^–4 M, POS caused intensification of collagen biosynthesis (the formation of peptidebound nondialyzable hydroxyproline-¹⁴C) in vitro in chick embryonic cartilage tissue. The silatranes are thus biologically active substances with a regulatory influence on the course of repair and proliferation in connective tissue.Laboratory of Biochemistry and Pharmacokinetics, Institute of Organic Synthesis, Academy of Sciences of the Latvian SSR, Riga. Laboratory of Experimental and Clinical Biochemistry, Institute of Traumatology and Orthopedics, Ministry of Health of the Latvian SSR, Riga. (Presented by Academician of the Academy of Medical Sciences of the USSR V. V. Zakusov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 87, No. 2, pp. 153–155, February, 1979.     

Oxidation-reduction processes during allergic reactions of the delayed type to microbial antigens

The effect of the general allergic reaction of delayed type (GARDT) to brucellosis antigen on glycolytic processes and activity of dehydrogenases of the Krebs' cycle was studied in the blood and organs of guinea pigs. Besides inhibition of the activity of the four dehydrogenases studied, inhibition of glycolysis connected with a decrease in the concentrations of lactate and pyruvate and inhibition of lactate dehydrogenase (LD) activity was observed. An increase in the content of anaerobic fractions and a decrease in the content of aerobic fractions were observed in the LD isozyme spectrum, together with excessiveness of the spectrum itself.Central Research Laboratory. Alma-Ata Medical Institute. (Presented by Academician of the Academy of Medical Sciences of the USSR A. D. Ado.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 86, No. 9, pp. 347–349, September, 1978.     

Changes in the isozyme spectrum of some enzymes of energy metabolism and in mitochondrial ultrastructure in virus-induced carcinogenesis

The dynamics of redistribution of the isozyme spectrum of lactate and malate dehydrogenases in rat embryonic fibroblasts (REF) in contact with type 12 human adenovirus was studied and compared with the dynamics of changes in the fine structure of the mitochondria of the same cell models. The action of the oncogenic virus was shown to produce changes in the isozyme spectra of lactate and malate dehydrogenases in both the nucleus and cytoplasm. These disturbances began after the first days of interaction between the REF culture and the oncogenic virus and were not connected with proliferative growth of the cell culture, but were due to the action of the virus on the cell. Comparison of the kinetics of changes in the enzymes studied with the results of quantitative analysis of the ultrastructure of the mitochondrial apparatus showed that the biochemical changes arise much sooner, virtually at the moment of infection.Laboratory of Virology, P. A. Gertsen Moscow Oncologic Research Institute. (Presented by Academician of the Academy of Medical Sciences of the USSR N. N. Zhukov-Verezhnikov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 84, No. 8, pp. 205–208, August, 1977.     

粥样硬化斑块中组织因子和组织因子途径抑制物的表达

目的 观察股动脉粥样硬化斑块中组织因子TF和组织因子途径抑制物TFPI的表达、分布.方法 采用免疫组化、双染组化方法检测股动脉粥样硬化斑块中TF和TFPI的表达和分布,RT-PCR检测TF mRNA和TFPI mRNA的表达.脐动脉作为对照.结果 脐动脉外膜表达少量TF和TFPI蛋白及其mRNA,而动脉粥样硬化斑块血管增生内膜大量表达TF和TFPI蛋白及其mRNA.结论 增生内膜中所有细胞类型及细胞间质都表达TF和TFPI.     

Caspase抑制剂对冻融卵巢组织的影响

目的研究Caspase抑制剂z-DEVD-fmk对冻融小鼠卵巢组织的影响。方法将15只小鼠随机分为3组：A组（新鲜对照组）、B组（添加z-DEVD-fmk组）和C组（未添加z-DEVD-fmk组）,B组和C组的卵巢组织进行玻璃化冷冻,一部分卵巢皮质组织进行组织学分析,另一部分卵巢皮质组织进行自体移植,手术后1个月后测血清雌二醇浓度和移植物增值细胞核抗原（PCNA）表达率。结果光镜观察发现冷冻组的始基卵泡和初级卵泡的形态正常率均明显低于对照组,B组的始基卵泡和初级卵泡的形态正常率与C组比较无明显差异。然而,B组血清雌二醇浓度高于C组,差异有统计学意义。同样,B组移植物卵细胞和间质细胞PCNA蛋白表达率高于C组。结论实验结果提示：Caspase抑制剂z-DEVD-fmk在卵巢组织冻融过程中具有保护作用。     

蛋白酶抑制剂对肥大细胞类胰蛋白酶分泌的影响

目的 探讨蛋白酶抑制剂和组胺对肥大细胞类胰蛋白酶分泌的影响。方法 扁桃体组织经酶消化后，细胞成份用全HBSS重新悬浮，肥大细胞激发和抑制剂作用的试验在37℃条件下完成。类胰蛋白酶水平用酶联免疫吸附试验（ELISA）方法测定。结果 鱼精蛋白具有刺激人类扁桃体肥大细胞释放类胰蛋白酶的作用，其分泌量可高达基础分泌量的5倍。高浓度TLCK和TPGK可抑制抗-IgE诱导的类胰蛋白酶释放，TLCK在有否预培养的情况下均能抑制钙离子导入剂（cal-cium ionophore,CI)诱导的类胰蛋白酶释放，而TPCK则只有在20min预培养后才能显示此作用。结论 TLCK和TPCK抑制IgE依赖性和非依赖性类胰蛋白酶释放提示具有胰蛋白酶和糜蛋白酶活性的酶参与肥大细胞的激活-分泌偶联过程。     

Effect of prolonged exposure of rats to high carbon dioxide concentrations

The lactate dehydrogenase (LD) isozyme spectrum of heart muscle, the heart rate (HR) and the ECG indices were studied during exposure to high concentrations of carbon dioxide, and the survival rate of rats also was determined in relation to the rate of rise of the carbon dioxide concentration in the atmosphere. Functional changes observed in cardiac activity during exposure to CO₂ in a concentration of 30% for 7.5 h were not accompanied by any disturbance of carbohydrate metabolism of the heart muscle. The most important condition of survival of rats exposed for a long time to an atmosphere with a high (up to 50%) CO₂ concentration is the rate at which the concentration rises.DeceasedInstitute for Medico-Biological Problems, Ministry of Health of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR S. S. Debov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 86, No. 9, pp. 285–287, September, 1978.     

Fallopian tube prolapse misdiagnosed as vault granulation tissue: a report of three cases

Prolapse of the fallopian tube into the vagina is an uncommon complication caused by either vaginal or abdominal hysterectomy. Recently, however, we encountered three cases with prolapse of the fallopian tube after abdominal hysterectomy. The patients presented with vaginal bleeding. A red hemorrhagic granular mass, misdiagnosed as vaginal granulation tissue both macroscopically and microscopically, was noted at the apex of vagina. Pathologically, one case was initially diagnosed as vaginal vault granulation tissue, but there were two recurrences after excision. Microscopically, the mass had a papillary or villous outer surface with a complex pattern of tubular and glandular structures, as well as acute and chronic inflammatory infiltrates in the fibrovascular stroma. A typical ciliated tubal type of epithelium was identified, and on immunohistochemical staining for cytokeratin, attenuated epithelial cells were detected. It is necessary to receive a pathologic confirmation by performing vaginal biopsy when fallopian tube prolapse is clinically suspected, thus preventing misdiagnosis.     

Expression of vascular endothelial growth factor in exuberant tracheal granulation tissue in children

Prolonged tracheotomy and endotracheal intubation often induce symptoms of airway obstruction and delay decannulation and extubation. Bronchoscopic examination of patients undergoing these treatments usually shows the presence of exuberant (pseudopapillary or nodular) granulation tissue occupying the airway lumen. An immunohistochemical analysis was undertaken of vascular endothelial growth factor (VEGF) expression in exuberant tracheal granulation tissue (n=17) obtained from children treated with prolonged tracheotomy or endotracheal intubation. Increased levels of VEGF protein and mRNA were expressed mainly by tracheal epithelial cells that migrated to cover the granulation tissue and partly by pericapillary macrophages in this tissue, whereas normal tracheal epithelium did not express VEGF. The VEGF expression level correlated significantly with the severity of the exuberant granulation tissue response (p=0·0018). As VEGF induces angiogenesis and vascular permeability, characteristics of granulation tissue, and plays a pivotal role in granulation tissue development, enhanced VEGF expression may be involved in the development of exuberant tracheal granulation tissue. Copyright © 1999 John Wiley & Sons, Ltd     

Wound healing: the role of gap junctional communication in rat granulation tissue maturation.

Granulation tissue maturation is dependent upon the orientation of collagen fibers and cell differentiation. Gap junctions are intercellular membrane gated channels that facilitate direct communication between cells known as gap junctional intercellular communication (GJIC). The hypothesis is that GJIC modulates the maturation of granulation tissue during wound repair. In vitro, GJIC optimizes fibroblast-populated collagen lattice contraction and influences cell morphology. It is reported that LiCl increases GJIC in cultured cardiac myocytes. Polyvinyl alcohol (PVA) sponge implants with central reservoirs were placed within separate subcutaneous pockets on the backs of adult male Sprague-Dawley rats. Each PVA implant received either 20 mM LiCl or saline injections on days 5, 7, and 10 after implantation. On day 11 implants were harvested and processed for light microscopy. By H&E staining LiCl-treated implants showed increased vascularization and decreased cell density compared to saline controls. Polarized light microscopy of Sirius red-stained specimens revealed more intense collagen fiber birefringence secondary to dense, parallel-organized collagen fiber bundles after LiCl treatment. This suggests that LiCl enhancement of GJIC between fibroblasts advances the maturation of granulation tissue. It is proposed that the degree of GJIC between granulation tissue fibroblasts influences both the quantity and the quality of granulation tissue deposited during the wound healing process.     

组织因子途径抑制因子/绿色荧光蛋白双顺反子真核表达载体构建及其在NIH 3T3细胞中表达

目的构建含人组织因子途径抑制因子(TFPI)和绿色荧光蛋白(GFP)基因的双顺反子真核表达载体,并验证其在NIH 3T3细胞中的表达,为血管再狭窄的防治提供一个具有示踪和治疗双重作用的有效载体。方法以含有全长cDNA的pIRES-TFPI为模板,多聚酶链反应(PCR)扩增TFPI全长cDNA,经酶切后插入到pIRES2-AcGFP1-Nuc载体中,构建成双顺反子真核表达载体,重组质粒经酶切图谱分析、PCR扩增及测序鉴定后命名为pIRES2-AcGFP1-Nuc-TFPI。将其转染NIH3T3细胞,采用荧光显微镜观察GFP在细胞中的表达,以RT-PCR检测TFPI在细胞内的表达。结果经酶切图谱分析、PCR扩增及DNA测序证实双顺反子真核表达载体构建正确;荧光显微镜可观察到细胞内GFP的表达;RT-PCR证实经TFPI基因转染的细胞内TFPI mRNA表达增高。结论成功构建了包括TFPI和GFP的真核双表达载体,并使其在NIH3T3细胞中顺利表达。     

Effect of neurogenic injury to the myocardium on activity of some enzymes of its energy metabolism

After electrical stimulation of the arch of the aorta in rabbits for 3 h exhaustion of the tissue noradrenalin (NA) reserves in the myocardium was accompanied by an increase in the activity of hexokinase (HK), lactate dehydrogenase (LD), and glucose-6 phosphate dehydrogenase (G6PD). Injection of L-Dopa after electrical stimulation prevented the fall of the NA level in the heart muscle and the change in activity of the above enzymes. The results confirm the important role of disturbances of mediator metabolism in mechanisms of development of metabolic and generative injuries.Laboratory of Experimental Pharmacology, Department of Pharmacology, Institute of Experimental Medicine, Academy of Medical Sciences of the USSR, Leningrad. (Presented by Academician of the Academy of Medical Sciences of the USSR S. V. Anichkov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 82, No. 10, pp. 1205–1206, October, 1976.     

幽门螺杆菌下调人胃黏膜分泌性白细胞蛋白酶抑制因子的表达

目的通过低剂量阿斯匹林的处理作为炎性模型,研究分析人胃和十二指肠黏膜分泌性白细胞蛋白酶抑制因子(SLPI)表达下调是否与幽门螺杆菌(Helicobacter pylori,Hp)定居或感染有关。方法选择20例健康志愿者分成Hp+和Hp-组,其中Hp+组成功进行了根除治疗。每组10人,每人口服阿司匹林100 mg/d。通过活检提取受试者不同部位胃黏膜的总RNA和蛋白,采用RT/real-time PCR检测SLPI的mRNA表达水平和单抗ELISA定量测定SLPI蛋白,统计分析SLPI基因表达的相关数据。结果与Hp-组比较,Hp+组胃窦黏膜SLPI表达水平显著降低,但Hp+组经抗生素根除治疗后其SLPI水平恢复至与Hp-组相当的水平。在低剂量阿司匹林处理下,所有受试者表现组织学观察到的活动性或慢性炎性征。各组在药物处理的不同时间虽有一定的差异,但与药物处理对照组(第0天)比较,SLPI的表达差异没有统计学意义[第1天:(77±880)pg/10μg蛋白;第3天:(941±149)pg/10μg蛋白;第7天:(763±363)pg/10μg蛋白]。阿司匹林处理的1周内以胃窦为主的胃炎持续存在(活动性:1.5~1.7;慢性:1.2~2.1),但与Hp-和Hpe(根除组)比较,Hp+组的胃窦黏膜SLPI蛋白表达仍显著降低。结论低剂量阿司匹林处理所致炎性反应对人胃黏膜SLPI的表达没有影响,胃窦黏膜SLPI基因表达的下调与Hp感染有关。     

Role of the inflammatory protein serine protease inhibitor Kazal in preventing cytolytic granule granzyme A-mediated apoptosis

Serine protease inhibitor Kazal (SPIK) is an inflammatory protein whose levels are elevated in numerous cancers. However, the role of this protein in cancer development is unknown. We have recently found that SPIK suppresses serine protease-dependent cell apoptosis. Here, we report that anti-SPIK antibodies can co-immmunoprecipitate serine protease granzyme A (GzmA), a cytolytic granule secreted by cytotoxic T lymphocytes and natural killer cells during immune surveillance, and that SPIK suppresses GzmA-induced cell apoptosis. Deletion studies show that the C3-C4 region of SPIK is critical for this suppression. These studies suggest that over-expression of SPIK may prevent GzmA-mediated immune-killing, thereby establishing the tolerance of cancer cells to the body's immune surveillance system. Suppression of over-expressed SPIK can restore the susceptibility of these cells to apoptotic death triggered by GzmA. This finding implies that it is possible to overcome tolerance of cancer cells to the body's immune surveillance system and restore the GzmA-mediated immune-killing by suppressing the over-expression of SPIK.     

Modulation of dendritic cell function and immune response by cysteine protease inhibitor from murine nematode parasite Heligmosomoides polygyrus

Modulation and suppression of the immune response of the host by nematode parasites have been reported extensively and the cysteine protease inhibitor (CPI or cystatin) is identified as one of the major immunomodulators. In the present study, we cloned and produced recombinant CPI protein from the murine nematode parasite Heligmosomoides polygyrus (rHp‐CPI) and investigated its immunomodulatory effects on dendritic cell (DC) function and immune responses in mice. Bone‐marrow‐derived CD11c⁺ DC (BMDC) that were exposed to rHp‐CPI during the differentiation stage showed reduced MHC‐II molecule expression compared with BMDC that were generated in normal culture conditions. The BMDC generated in the presence of rHp‐CPI also exhibited reduced expression of CD40, CD86 and MHC‐II molecules and reduced interleukin‐6 and tumour necrosis factor‐α cytokine production when stimulated with Toll‐like receptor ligand CpG. Activation of BMDC generated in normal conditions induced by lipopolysaccharide and CpG was also suppressed by rHp‐CPI, as shown by reduced co‐stimulatory molecule expression and cytokine production. Furthermore, BMDC treated with rHp‐CPI before ovalbumin (OVA) antigen pulsing induced a weaker proliferation response and less interferon‐γ production of OVA‐specific CD4⁺ T cells compared with BMDC without rHp‐CPI pre‐treatment. Adoptive transfer of rHp‐CPI‐treated and OVA‐loaded BMDC to mice induced significantly lower levels of antigen‐specific antibody response than the BMDC loaded with antigen alone. These results demonstrated that the CPI from nematode parasites is able to modulate differentiation and activation stages of BMDC. It also interferes with antigen and MHC‐II molecule processing and Toll‐like receptor signalling pathway, resulting in functionally deficient DC that induce a suboptimum immune response.