Experimentally induced colon cancer metastases in rat liver increase the proliferation rate and capacity for purine catabolism in liver cells

Metastases in rat liver were generated experimentally by intraportal injection of colon cancer cells to investigate the effects of cancerous growth on the metabolism of surrounding liver tissue. Maximum activities (capacity) of glucose-6-phosphate dehydrogenase, phosphogluconate dehydrogenase, lactate dehydrogenase, succinate dehydrogenase, alkaline phosphatase, 5-nucleotidase, xanthine oxidoreductase, purine nucleoside phosphorylase and adenosine triphosphatase have been determined. Two types of metastases were found, a small type surrounded by stroma and a larger type in direct contact with hepatocytes. Both types affected the adjacent tissue in a similar way suggesting that the interactions were not mediated by stroma. High capacity of the degradation pathway of extracellular purines released from dead cells of either tumours or host tissue was found in stroma and sinusoidal cells. Metastases induced both an increase in the number of Kupffer cells and proliferation of hepatocytes. The distribution pattern in the liver lobulus of most enzymes investigated did not change distinctly. However, activity of alkaline phosphatase, succinate dehydrogenase and phosphogluconate dehydrogenase was increased in hepatocytes directly surrounding metastases. These data imply that the overall metabolic zonation in liver lobuli is not dramatically disturbed by the presence of cancer cells despite the fact that various metabolic processes in liver cells are affected.In honour of Prof. Dr. Z. Lojda for his 65th birthday     

Cultured porcine urinary bladder epithelial cells as a screening model for genotoxic effects of aromatic amines: characterisation and application of the cell culture model

Isolated epithelial cells from porcine urinary bladders were maintained in dividing long-term monolayer cultures, and were used as a model system for the urinary bladder in toxicological studies in vitro. To examine the state of differentiation during the culture period, the culture system was characterised morphologically by light and transmission electron microscopy and by immune fluorescence labelling with antibodies against cytokeratins 7,13 and pan. The cultured cells were identified as urothelial epithelium by their polarised structure, and by their expression of several uroepithelial specific morphological features, such as fusiform vesicles, tight junctions and an asymmetric apical cell membrane. Additionally, the cells were labelled with anti-cytokeratin 7,13 and pan antibodies, and negatively with anti-vimentin antibodies. The maintenance of suitable culture conditions was shown by the stable enzyme activities of (gamma-glutamyltranspeptidase, alkaline phosphatase and acid phosphatase over a culture period of 4 weeks. A good viability of the cultured cells under the chosen culture conditions was shown by the presence of low amounts of lactate dehydrogenase (< of = 5%) in the culture medium. The activities of the chosen marker enzymes for cell differentiation (gamma-glutamyltranspeptidase), lysosomes (acid phosphatase) and luminal membranes (alkaline phosphatase) were relatively stable over the observed culture period. Enzyme activities involved in metabolism of xenobiotics were determined, to define the ability for metabolism in cultured cells compared with bladder tissue in situ. Several constitutive phase I and II enzyme activities were found to be stable during the culture period, indicating that the cultured cells should be able to metabolise xenobiotics in a comparable manner to the urothelium in vivo. The cytotoxic effects of xenobiotics were investigated and IC50 values were determined by means of lactate dehydrogenase leakage and inhibition of neutral red uptake. The induction of sister chromatid exchanges was used as a parameter for the genotoxic effects of several xenobiotics. This cell culture system was found to be a very good screening system for the testing of substances that affect the bladder, especially aromatic amines.     

LHPP在人胃腺癌组织中的表达及意义

目的 通过检测LHPP（phospholysine phosphohistidine inorganic pyrophosphate phosphatase）在人胃腺癌组织和胃腺癌细胞系,以及幽门螺杆菌（Helicobacter pylori, H. pylori）感染正常胃上皮细胞系后的表达水平,评估LHPP表达及LHPP与胃腺癌的关系。方法 （1）使用TCGA（The Cancer Genome Atlas）数据库和GEO（Gene Expression Omnibus）数据库中的胃腺癌基因表达数据,分析LHPP在胃腺癌组织和正常胃黏膜组织中的表达差异。（2）选取2017年3月至2018年5月行胃部分切除术或全胃切除术的41例胃腺癌患者,取胃癌组织及匹配的正常胃黏膜上皮组织,应用免疫组化检测LHPP在胃腺癌组织和正常胃黏膜组织中的表达差异。（3）取常规传代培养的第三代正常胃黏膜上皮细胞系GES-1和胃腺癌细胞系AGS,qRT-PCR检测两组LHPP基因表达差异。（4）应用H. pylori P12标准菌株以感染复数（multiplicity of infection,MOI）50感染GES-1细胞系6、12、18和24 h后,Western blot检测各个时间点LHPP和H. pylori毒力蛋白CagA表达量。结果 TCGA数据库中的LHPP表达数据表明LHPP在胃腺癌中表达下调（t=7.064,P<0.0001）,来自GEO数据库中GSE65801表达数据的芯片测序结果得到相同的结论（P=0.001）。免疫组化证实LHPP在低分化胃腺癌中低表达,且表达与分化程度相关（χ2=5.559,P=0.018）,而与性别（χ2=2.304,P=0.129）、年龄（χ2=0.001,P=0.986）、浸润深度（χ2=1.007,P=0.316）、淋巴结转移（χ2=0.693,P=0.405）及肿瘤分期（χ2=0.003,P=0.953）不相关。LHPP在胃腺癌细胞系AGS中表达低于正常胃上皮细胞系GES-1（t=10.38,P=0.005）。H. pylori感染GES-1细胞后LHPP表达随着感染时间延长总体有升高趋势。结论 LHPP在胃腺癌中低表达并与腺癌分化程度相关,有望成为胃腺癌诊断治疗的新靶点。     

Behavior of subcellular marker enzymes during the Hela cell cycle

Six enzymes associated with the activities of the nucleus (thymidine kinase), mitochondria (succinate dehydrogenase), lysosomes (acid phosphatase), peroxisomes (catalase), (cytosol lactate dehydrogenase), and the intermembranal mitochondrial space (alkaline DNase) were assayed at 2 hr intervals over the division cycle of repetitively resynchronized HeLa cells. The results indicated a high degree of reproductibility for cells synchronized by the method of perpetual resynchronization and may be of direct use to those interested in subcellular organellogenesis.     

Potential preventive effect of carvacrol against diethylnitrosamine-induced hepatocellular carcinoma in rats

Antioxidants are one of the key players in tumorigenesis, several natural and synthetic antioxidants were shown to have anticancer effects. The aim of the present study is to divulge the chemopreventive nature of carvacrol during diethylnitrosamine (DEN)-induced liver cancer in male wistar albino rats. Administration of DEN to rats resulted in increased relative liver weight and serum marker enzymes aspartate transaminase (AST), alanine transaminase (ALT), alkaline phosphatase (ALP), lactate dehydrogenase (LDH), and gamma glutamyl transpeptidase (γGT). The levels of lipid peroxides elevated (in both serum and tissue) with subsequent decrease in the final body weight and tissue antioxidants like superoxide dismutase (SOD), catalase (CAT), reduced glutathione (GSH), glutathione peroxidase (GPx), and glutathione reductase (GR). Carvacrol supplementation (15 mg/kg body weight) significantly attenuated these alterations, thereby showing potent anticancer effect in liver cancer. Histological observations and transmission electron microscopy studies were also carried out, which added supports to the chemopreventive action of the carvacrol against DEN-induction during liver cancer progression. These findings suggest that carvacrol prevents lipid peroxidation, hepatic cell damage, and protects the antioxidant system in DEN-induced hepatocellular carcinogenesis.     

Enzyme activities in the plasma of rainbow trout, Salmo gairdneri Richardson; the effects of nutritional status and salinity

The levels of lactate dehydrogenase, hydroxy butyric dehydrogenase, glutamic oxalacetic transaminase, glutamic pyruvic transaminase, glutamate dehydrogenase, alkaline phosphatase and leucine amino peptidase were determined in the plasma of rainbow trout. The protein concentration and the amount of alkaline phosphatase were reduced in starving trout. Fed trout showed reduced lactate dehydrogenase activity. There was a significant correlation between the condition factor, most of the enzyme activities and the protein concentration. At 10 parts per thousand salinity the activities of alkaline phosphatase and leucine amino peptidase were significantly elevated, while lactate dehydrogenase activity was significantly decreased.     

Melatonin regulates cancer migration and stemness and enhances the anti-tumour effect of cisplatin

Melatonin, a lipophilic hormone released from the pineal gland, has oncostatic effects on various types of cancers. However, its cancer treatment potential needs to be improved by deciphering its corresponding mechanisms of action and optimising therapeutic strategy. In the present study, melatonin inhibited gastric cancer cell migration and soft agar colony formation. Magnetic-activated cell sorting was applied to isolate CD133⁺ cancer stem cells. Gene expression analysis showed that melatonin lowered the upregulation of LC3-II expression in CD133⁺ cells compared to CD133⁻ cells. Several long non-coding RNAs and many components in the canonical Wnt signalling pathway were altered in melatonin-treated cells. In addition, knockdown of long non-coding RNA H19 enhanced the expression of pro-apoptotic genes, Bax and Bak, induced by melatonin treatment. Combinatorial treatment with melatonin and cisplatin was investigated to improve the applicability of melatonin as an anticancer therapy. Combinatorial treatment increased the apoptosis rate and induced G0/G1 cell cycle arrest. Melatonin can regulate migration and stemness in gastric cancer cells by modifying many signalling pathways. Combinatorial treatment with melatonin and cisplatin has the potential to improve the therapeutic efficacy of both.     

PCC4azal teratocarcinoma stem cell differentiation in culture: I. Biochemical studies

PCC4azal embryonal carcinoma cells were observed to spontaneously differentiate under defined culture conditions to endoderm-like cells and subsequently to giant cells. This differentiation was examined by determining the specific activities of several enzymes in the stem and endoderm-like cell populations. With differentiation, the level of alkaline and acid phosphatase activities remained unchanged, plasminogen activator specific activity increased fivefold, and lactate dehydrogenase (LDH) specific activity decreased to 40% of its original level. Isozyme analysis revealed a shift of the LDH isozymes toward LDH₁ with the appearance of LDH₂ for the first time in the endoderm-like cells. The surface antigen SSEA-1 was detected by indirect immunofluorescence on virtually all of the stem cells. However, the SSEA-1 antigen was not present on many of the endoderm-like cells, and it was completely undetectable on giant cells as assayed by immunofluorescence. The expression of H-2 antigen was examined in a similar manner using anti-H-2^b antiserum; this antigen was not detected on the stem, endoderm-like, or giant cells. Thus, there are defined biochemical changes that accompany the differentiation of PCC4azal stem cells in culture.     

Changes in the activity of different classes of enzymes in the cerebral cortex of rats in ontogeny and after the transplantation of embryonic nerve tissue

The activity of lactate dehydrogenase (LDH), indophenol oxidase, aspartate aminotransferase (AsAT), alkaline phosphatase, acid phosphatase and aldolase at different stages of rat development was measured. We have also determined changes in the activity of these enzymes resulting from transplantation of embryonic nerve tissue (ENT) into the brain of adult animals. During development from the embryo to the adult animal, LDH and AsAT activities increased, while alkaline phosphatase activity diminished. After ENT transplantation, the most prominent changes were in the alkaline phosphatase activity whereas the activity of LDH, AsAT and acid phosphatase remained unchanged and similar to that in the brain cortex of intact adult animals. Changes in the enzyme activity resulting from ENT transplantation changed in a manner characteristic of the transplant. Local brain damage did not change the activity of the studied enzymes fifty days after surgery.     

Cytochemical study of cells of primary and disseminated ascite Yoshida tumor cells

The different distribution of cytochemically demonstrable enzymes: lactate dehydrogenase (LDH, 1.1.1.27), succinate dehydrogenase (SDH, 1.3.99.1), dihydrofolate reductase (DHFR, 1.5.1.3), acid phosphatase (AcP, 3.1.3.2) and alkaline phosphatase (ALP, 3.1.3.1), has been documented in Yoshida ascites hepatoma cells in vivo or stored at 80 degrees C. The dehydrogenase activities (LDH, SDH, DHFR) show a strong reaction in all samples. An increased level of these enzyme activities has been observed in the malignant cells spreading through the organs of tumor bearing rats. On the contrary, in the same samples, acid and alkaline phosphatase activities are very low. The strong dehydrogenase activities observed in Yoshida ascite cells stress the rapid turnover of tumor cells. Our results indicate that the histochemical method may be a useful tool to detect the scattered tumor cells. Furthermore, the cytochemical methods allow the characterization of the metabolic pathways employed by the primary and disseminated tumor cells.     

Overexpression of ZEB1 associated with metastasis and invasion in patients with gastric carcinoma

The aim of this study was to investigate the expression of ZEB1 in gastric carcinoma, its correlation with the clinicopathology of gastric carcinoma, and the role of ZEB1 in invasion and metastasis in gastric carcinoma. ZEB1 expression was analyzed by immunohistochemistry and Western blot in 45 gastric carcinoma tissue samples that contained the adjacent gastric mucosa. The correlation between ZEB1 expression, the occurrence and development of gastric cancer, and clinical pathology was investigated. ZEB1 expression in the human gastric carcinoma cell line AGS was downregulated by RNA interference, and changes in ZEB1 expression corresponded with changes in the invasive and metastatic ability of AGS cells. Immunohistochemistry revealed that ZEB1 protein expression in gastric carcinoma tissues was significantly higher than in normal gastric mucosa tissues (p < 0.001). A lower degree of differentiation of gastric cancer (p = 0.009), a higher TNM (tumor, node, and metastasis) stage (p = 0.010), and a larger scope of invasion were correlated with higher expression of ZEB1 (p = 0.041, 0.002). However, the expression of ZEB1 in gastric carcinoma tissue was independent of gender, age, and tumor size (p > 0.05). Western blot results also showed that ZEB1 protein expression was significantly higher in gastric carcinoma tissue than in the adjacent normal gastric mucosa tissue (p = 0.008). A lower degree of differentiation of the gastric carcinoma correlated with a higher TNM stage, and a larger scope of invasion correlated with increased ZEB1 expression (p = 0.023). Transfection of ZEB1 siRNA in AGS cells significantly decreased the expression level of ZEB1 protein (p = 0.035). Furthermore, the number of cells that could pass through the Transwell chamber was significantly lower in the transfected group than in the non-transfected control group (p = 0.039), indicating that the suppression of ZEB1 expression could significantly reduce the invasive and metastatic ability of AGS cells (p = 0.005). Concluding, in gastric carcinoma tissue, overexpression of ZEB1 may be related to the occurrence and development as well as invasion and metastasis of gastric carcinoma.     

A light- and electron-microscopic study of enzymes in the embryonic shell-forming tissue of the freshwater snail, Biomphalaria glabrata

Abstract. The mode of formation of the molluscan exoskeleton is still poorly understood, but studies on adult snails indicate that enzymes involved in vertebrate bone formation also participate in mollusc shell formation. The enzymes peroxidase, alkaline phosphatase, and acid phosphatase are expressed in a constant pattern and help to identify the different zones of the adult shell-forming tissue. The present study evaluates whether the expression of these enzymes is also a tool for the identification of the developing zones of the embryonic shell-forming tissue. Thus, we analyzed the temporal and spatial activity of the above-mentioned enzymes and of tartrate-resistant acid phosphatase in the shell forming tissues in Biomphalaria glabrata. Embryos of different age groups and adults were studied; alkaline phosphatase activity was seen in very young embryos in the shell field invagination prior to the secretion of any shell material, while peroxidase activity was present from the start of the periostracum production. Acid phosphatase, found in considerable amounts in yolk granules and albumen cells, appeared in the embryonic shell-forming tissue in relatively few Golgi stacks. Tartrate-resistant phosphatase was not present in embryos, but was found in adults in the same zone of the mantle edge as acid phosphatase. Using the enzymes as cell markers, the differentiation of the embryonic shell-forming tissue to the different zones of the adult mantle edge could clearly be followed.     

Alkaline phosphatase, gamma-glutamyl transferase, lactate dehydrogenase and creatine kinase in bronchial aspirate from neoplastic and normal pulmonary tissue

The activities of 4 enzymes, i.e. alkaline phosphatase, gamma-glutamyl transferase, lactate dehydrogenase and creatine kinase were studied in bronchial aspirates and serums from two groups of subjects, the first one was composed of 14 subjects without active bronchopulmonary pathology and the other of 20 patients with lung cancer. The results showed a statistically significant decrease of the activities of alkaline phosphatase and beta-glutamyl transferase in bronchial aspirate from patients with bronchogenic malignant tumors in relation to normal subjects. This finding could be explained by the 'fetalism' principle, which states that the quantitative pattern of enzymes of immature human tissues resembles those of neoplastic tissues.     

Developmental genetics of teleosts: a biochemical analysis of lake chubsucker ontogeny

Developing embryos of the lake chubsucker, Erimyzon sucetta, were analyzed with regard to both gross morphological changes and specific enzymatic changes from the unfertilized egg stage until some 3 weeks posthatching. Total activities of three enzymes—lactate dehydrogenase, glucose-6-phosphate dehydrogenase, and isocitrate dehydrogenase—were determined throughout the course of development. Each of these different enzymes exhibited a different pattern of change during ontogeny. Electrophoretic analysis of qualitative changes in isozyme patterns was accomplished for these three enzymes and for α-amylase, glucosephosphate isomerase, mannosephosphate isomerase, creatine kinase, esterase, glutamate dehydrogenase, alkaline phosphatase, aspartate aminotransferase, malate dehydrogenase, hexose diphosphatase, phosphoglucomutase, and phosphogluconate dehydrogenase. Many of the enzyme systems investigated exhibited rich patterns of ontogenetic change, while a few remained relatively unchanged throughout the interval studied. Several of the enzymes in particular metabolic pathways exhibited coincident changes suggestive of coordinate control. The appearance of several rather “tissue-specific” isozymes was closely correlated with the morphological and functional differentiation of these particular tissues or organs.     

胃癌组织中HER2、PD-L1的表达与临床病理特征的相关性

目的:检测胃癌组织中人表皮生长因子受体2(HER2)、程序性死亡因子1配体(PD-L1)的表达,并分析其与胃癌患者临床病理特征的相关性。方法:采用分层整群抽样回顾性分析的方法抽取我院2016年1月至2018年12月经手术病理诊断为胃癌的100例原发性胃癌患者,全部患者术后均经病理组织切片免疫组化染色检测其HER2、PD-L1表达,对比不同性别、年龄、肿瘤大小、肿瘤部位、分化程度、病理类型等临床病理特征胃癌患者HER2、PD-L1的表达,分析二者与胃癌患者临床病理特征的相关性。结果:100例胃癌患者中男女比例为2.8:1,年龄60岁占比高,肿瘤大小多超过4 cm,WHO分型以分化不良与分化较好为主,肿瘤部位主要位于胃下2/3,浸润深度多为T4,TNM分期集中在Ⅰ～Ⅲ期,多伴淋巴结转移,几乎无远处转移,多存在脉管侵犯,部分有神经侵犯。100例胃癌患者胃癌组织HER2表达阴性、弱阳性、强阳性检出率分别为42.00%、31.00%、27.00%;PD-L1阴性、阳性检出率分别为57.00%、43.00%。胃癌组织HER2阳性表达、PD-L1阳性表达均高于癌旁正常胃组织,差异有统计学意义(P0.05)。胃癌组织中HER2表达与疾病分化程度呈负相关(r0,P0.05),PD-L1的表达与肿瘤分化程度、浸润深度、与远处转移均呈显著正相关(r0,P0.05)。结论:HER2的阳性表达可能提示胃癌患者较低的恶性程度,PD-L1的高表达可能提示胃癌患者存在远处转移、浸润深度深、恶性程度高。HER2和PD-L1有望成为胃癌患者诊断参考指标及药物干预的靶点。     

The localization of enzymes in the cotyledons of Pisum arvense L. during germination

Summary Enzyme activity is not uniformly distributed through the cotyledon of Pisum arvense. Initially the peripheral region, certain scattered cells of the storage tissue and the procambium show a high level of activity of succinic dehydrogenase, cytochrome oxidase, acid phosphatase and esterase. Activity of acid phosphatase declines sharply after the first day of germination; activity of the other enzymes declines after about three days. In the storage tissue, where activity is lower initially, it declines after about five days and is correlated with the disappearance of the reserves. The pattern of alkaline phosphatase activity is similar except that activity is lower in the procambium but increases in the sieve-elements during differentiation of the phloem. 5-nucleotidase and glucose-6-phosphatase activity is low throughout the cotyledon but it also increases to a significant level in the sieve-elements. Activity of starch synthesizing enzymes is high in the parenchymatous bundle sheath, where they may be involved in the pathway from lipids to soluble carbohydrates.     

The palmitoleate: a natural selective denaturant of enzymes

A study has been carried out in order to explain the enzyme-palmitoleate interaction. The highly purified and crystalline enzymes representative of fundamental metabolic pathways were: alcohol dehydrogenase (ADH), lactate dehydrogenase (LDH), malate dehydrogenase (MDH), isocitrate dehydrogenase (ICDH), glucose-6-phosphate dehydrogenase (G6P-DH), alkaline phosphatase. The enzyme-palmitoleate interaction was studied as a phenomenon time-independent (inhibition) and time-dependent (inactivation). Palmitoleate inhibited remarkably LDH, MDH, ICDH and G6P-DH. A kinetic analysis of the inhibitory action of palmitoleate on LDH and MDH was also carried out. Inactivation studies have shown that ADH and alkaline phosphatase are not sensitive to palmitoleate action, unlike the other enzymes. A comparison was made between the action of palmitoleate and that of a synthetic anionic detergent, sodium dodecyl sulfate (SDS).     

喉癌中SP100 表达及其与临床病理关系

目的：通过对不同喉癌病人癌组织SP100蛋白进行测量,明确SP100 在喉癌的发生发展过程中的作用并探究其与临床病理 的关系。方法：通过对喉癌手术病人切除的癌组织和癌周组织进行固定、脱水、包埋、切片、免疫组织化学染色判断SP100 阳性细 胞个数以及探究其与临床病理学之间的关系。结果：在正常黏膜上皮细胞和分化良好的癌细胞中,以细胞核内染色为主,在低分 化癌细胞中,在细胞质内呈弥漫性分布。SP100 蛋白在癌旁正常黏膜上皮组织中表达的阳性率比喉癌原发灶中高。SP100 蛋白在 96 例喉癌组织中的表达水平与病理分化程度密切相关(P<0.05 ),而与患者性别、年龄、P-TNM 分期、淋巴结转移无相关性 (P>0.05)。结论：喉癌组织中SP100蛋白表达水平、细胞内分布状况在不同分化程度癌细提示在喉癌的不同阶段可能发挥不同的 作用。     

Specific interaction among some enzymes and sodium dodecyl sulfate

The effect of 1-butanesulfonic acid sodium salt and sodium dodecyl sulfate on the activity of highly purified and crystalline enzymes with marked differences in structure and function has been studied. The enzymes were: alcohol dehydrogenase; lactate dehydrogenase; malate dehydrogenase; isocitrate dehydrogenase; glucose-6-phosphate dehydrogenase; lipase; alkaline phosphatase. While 1-butanesulfonic acid sodium salt, at the studied concentrations, resulted generally inactive, sodium dedecyl sulfate showed a selective inhibitory effect, always under the critical micellar concentration. A kinetic analysis of the inhibitory action was also carried out.     

Variation in some enzymes in amniotic fluid and maternal serum during pregnancy

The following 10 enzymes were assayed in 187 amniotic fluid and maternal serum samples at 15-42 weeks of gestation: alkaline phosphatase, heat-stable alkaline phosphatase (only in amniotic fluid), acid phosphatase, alanine aminotransferase, aspartate aminotransferase, alpha-amylase, gamma-glutamyltransferase, creatine kinase, lactate dehydrogenase, and lysozyme. The normal reference ranges are reported for amniotic fluid and maternal serum enzymes, together with the abnormal values accompanying neural tube defects and EPH-gestosis. The determination of gamma-glutamyltransferase, heat-stable alkaline phosphatase and creatine kinase was found to be of appreciable diagnostic significance in clinical practice.