Evaluation of localization and intensity of the reactions of succinic dehydrogenase, NADH tetrazole reductase and lactate dehydrogenase in developing rat testis. I. Physiological conditions

The localization, intensity, and character of the enzymatic reactions of the following enzymes responsible for the oxygenic and nonoxygenic respiration were under examination: succinic dehydrogenase (SDH), NADH tetrazole reductase (NADH-R), and lactate dehydrogenase (LDH). The features of the enzymatic reactions enumerated above were tested in male gonads of the rats on their 1st, 4th, 7th, 15th, 30th, 45th, 60th, 90th d and 1.5 year of post-fetal life. It was estimated on the examined reactions that their metabolic maturity in the gonad was reached on the 45th d of the rat's life.     

短链酰基辅酶A脱氢酶在大鼠心脏发育中的表达及其与心肌肥厚的关系

 目的：研究大鼠心脏发育过程中短链酰基辅酶A脱氢酶(short-chain acyl-CoA dehydrogenase, SCAD)的表达变化规律,并探讨其与高血压大鼠心肌肥厚的关系。方法：观察不同时期Wistar大鼠和不同周龄自发性高血压大鼠心肌组织的SCAD蛋白表达及酶活性变化,检测大鼠的血清和心肌游离脂肪酸含量。结果：与胚胎期19 d Wistar大鼠组比较,出生后1 d、2周、6周及16周龄Wistar大鼠组心肌的SCAD蛋白表达及酶活性增加,血清和心肌游离脂肪酸含量明显减少,二者之间呈负相关,其中,从2周龄Wistar大鼠组开始差异有统计学意义。与周龄匹配的WKY大鼠组比较,2周龄自发性高血压大鼠组收缩压尚未升高,6周龄及16周龄自发性高血压大鼠组收缩压显著增高;各时点自发性高血压大鼠组的左室重量指数均明显增高,提示自发性高血压大鼠在血压升高之前,已经发生了明显的心肌肥厚。与周龄匹配的WKY大鼠组比较,2周、6周及16周龄自发性高血压大鼠组心肌的SCAD蛋白表达及酶活性明显下降,血清和心肌游离脂肪酸含量明显增加,呈显著负相关。结论：(1)SCAD蛋白表达随大鼠心脏的生长发育逐渐上调,可能与心脏对脂肪酸的利用增加密切相关。(2)SCAD的蛋白表达及其酶活性显著下降, 可能是导致自发性高血压大鼠肥厚心肌能量代谢“胚胎型再演”的分子基础。     

Histochemical studies on the early proliferative lesion induced in the rat liver by aflatoxin

Male inbred Fischer rats were fed a diet containing 5 p.p.m. aflatoxin for 1, 3, 4½ and 6 weeks at which times groups were killed for histological and histochemical study. Aflatoxin produced a scattered individual cell necrosis of parenchymal cells by 1 week. At 3 weeks small basophilic proliferative foci were seen which increased in size and abundance to 6 weeks. These foci showed starvation-resistant glycogen, variable depletion of glucose-6-phosphatase, succinic dehydrogenase, aniline hydrogenase, membrane ATPase and acid phosphatase. At 6 weeks the foci showed the presence of gamma glutamyl transpeptidase and glucose-6-phosphate dehydrogenase. The basophilic foci were not preceded by other focal histological and histochemical change. The basophilic proliferative lesions are observed when an irreversible change has been induced in the liver. The role of such lesions in the histogenesis of hepatocellular carcinoma is discussed.     

Structural-metabolic changes in histaminergic neurons of the rat hypothalamus in conditions of loss of bile

The aim of the present work was to evaluate structural and metabolic changes in histaminergic neurons in hypothalamic nucleus E2 in rats in conditions of complete external drainage of bile. Studies were performed on male Wistar rats (n = 45). Controls consisted of animals subjected to sham surgery with preservation of physiological bile flow throughout the experiment. Quantitative histological and histochemical methods were used. Serial frontal cryostat sections cut from the posterior hypothalamus were used for detection of the activity of the following enzymes: monoamine oxidase B, succinate dehydrogenase, NADH dehydrogenase, NADPH dehydrogenase, glucose-6-phosphate dehydrogenase, lactate dehydrogenase, and acid phosphatase. Morphological studies of histaminergic neurons were performed on preparations stained with thionine. These studies showed that complete external drainage of bile led to transient size reductions and rounding of cell perikarya. Metabolic changes were seen within a day of bile loss and subsequently progressed. All energy metabolic pathways were suppressed and acid phosphatase activity was increased on day 5. __________ Translated from Morfologiya, Vol. 132, No. 6, pp. 22–25, November–December, 2007.     

A technique for demonstrating of dehydrogenase activities in Epon-embedded, semithin sections of the nervous tissue.

Experiments were performed aiming at establishing optimal conditions for demonstrating the activity of dehydrogenases in Epon-embedded, semithin sections. Small fragments of the nervous tissue were incubated 1, 4, 10 and 24 h 37, 20 and 4 degrees C in the presence of NBT or TC NBT. The incubated fragments were embedded in Epon and semithin sections prepared thereof. The activities of succinate dehydrogenase and of alpha-glycerol dehydrogenase was evaluated. The experiments have shown that semithin, Epon-embedded sections may be used for demonstration and cytological localization of respiratory enzymes. Various experimental conditions were tried. Optimal results obtained when unfixed tissue fragments were incubated for 24 h at 4 degrees C.     

The pyridoindole antioxidant stobadine attenuates histochemical changes in kidney of streptozotocin-induced diabetic rats

The aim of the present study was to investigate the effects of dietary supplementation with the pyridoindole antioxidant stobadine on histochemical parameters in kidney of streptozotocin-induced diabetic rats. Diabetic male Wistar rats were fed a standard diet or a diet supplemented with stobadine (0.05% w/w) for 24 weeks. The diabetic state was characterized by significantly elevated plasma levels of glucose and glycated hemoglobin, severe reduction of total body weight and relatively enlarged kidneys. Kidney alkaline phosphatase activity was not changed by diabetes. Activity of 5'-nucleotidase, K(+)-dependent p-nitrophenylphosphatase, ATPase and mitochondrial succinic dehydrogenase were markedly decreased in kidneys of diabetic rats. In contrast, activity of beta-hydroxybutyrate dehydrogenase was moderately increased in kidney of diabetic rats as compared to controls. Long-term treatment of diabetic animals with stobadine attenuated histochemical changes in kidney tissue.     

Exocrine cell mitochondria of the rat pancreas after lead intoxication.

The aim of the study was to evaluate alterations in exocrine cell mitochondria of the rat pancreas after lead acetate intoxication. The experiment used 45 rats divided into 2 experimental groups receiving lead acetate to drink, of lead concentration 50 and 500 mg/dm3 (ppm), and a control group given tap water. The animals from the experimental group were decapitated after 2, 4, 6 and 8 weeks, 5 rats from the control group after 8 weeks of the experiment. Rats from experimental groups decapitated after 8 weeks had lead administration stopped after six weeks and then, for two weeks tap water was given. Pancreatic sections were examined with biochemical methods for the activity of cytochrome oxidase and succinic dehydrogenase. Ultrastructural and morphometric examinations were also performed. It was demonstrated that: a) exocrine cell mitochondria are particularly predisposed to lead effect, b) intoxication of rats with lower lead doses (50 ppm) causes reversible adaptative or compensatory changes in these organelles, c) intoxication of rats with higher lead doses (500 ppm) induces irreversible ultrastructural alterations in numerous mitochondria, including damage to inner and to outer mitochondrial membranes, d) structural changes in the mitochondria in the course of lead intoxication are the morphological expression of the impairment of metabolic processes, associated with the inhibited activity of the respiratory enzymes: succinic dehydrogenase and cytochrome oxidase.     

Histophotometry--the method of choice in quantifying dehydrogenase histochemistry

Succinate, malate, and lactate dehydrogenase were demonstrated histochemically and measured histophotometrically in the heart and skeletal muscle (m. extensor digitorum longus and m. soleus) of rats at different ages. To prove the value of histophotometry, the enzymes of the tissues were estimated biochemically. The gel film technique cannot sufficiently prevent the diffusion of the soluble enzymes (malate-, lactate dehydrogenase) out of the tissue sections. Because of the different mobility, various isoenzymes, histophotometry cannot give reliable results. But, as far as membrane-bound dehydrogenases (succinate dehydrogenase) are concerned, histophotometry is the method of choice for basic measurements as in routine practical work, especially with tissues where the enzyme activities are heterogeneously distributed, e.g. in different types of muscle fibres in skeletal muscles.     

Age-related changes in liver and adipose tissue pyruvate dehydrogenase of genetically obese mice

Changes of the pyruvate dehydrogenase complex in liver and epididymal fat pad were examined longitudinally in obese mice (C57BL/6J-ob/ob) and their lean controls as a function of age. Total pyruvate dehydrogenase in liver was expressed on several reference bases because of differences in hepatic cellularity and protein content between obese mice and their age-matched lean controls. When total hepatic pyruvate dehydrogenase was expressed on a protein basis, the enzyme activity was elevated in obese mice older than 28 weeks in age when compared to lean controls of a similar age. However, when expressed on a DNA basis, total pyruvate dehydrogenase activity in livers of obese mice up to 10 weeks in age was increased when compared to the age-matched lean control. The proportion of hepatic pyruvate dehydrogenase in the active form was also augmented significantly in obese mice from 5 to 28 weeks of age. In 18-week-old obese mice, the proportion of total pyruvate dehydrogenase in the active form of adipose tissue was significantly higher than that of the lean controls. When expressed on a DNA basis, total pyruvate dehydrogenase in the fat pad was also increased in obese mice up to 10 weeks in age when compared to age-matched controls. Total pyruvate dehydrogenase activity in the epididymal fat pad was higher in obese mice than the lean controls in animals as old as 32 weeks in age when the enzyme activity was expressed per 100 g body weight. The increase in the active form and total activity of pyruvate dehydrogenase in both liver and epididymal fat pad during the dynamic early phase of obesity would augment the capacity for acetyl-coenzyme A formation necessary in the support of an accelerated lipogenesis and fat deposition.     

Accumulation of cadmium and its effects on liver and kidney functions in rats given diet containing cadmium-polluted radish bulb

The aim of this study was to examine the accumulation of cadmium (Cd) incorporated in radish bulb and its effects on liver and kidney functions in male rats. Control animals were given diet containing ordinary radish bulb for 4, 8 and 12 weeks, while contaminated animals were given diet containing Cd-polluted radish bulb (1.1 microg Cd/g of diet) for the same periods as in controls. At each time point, rats were killed and plasma was collected, and the liver and the kidneys were removed. Results indicated that body weight gain of contaminated rats was identical to that of control rats. Cd concentration in the liver and the kidney increased significantly and gradually from the 4th to the 12th week of treatment. Plasma alanine aminotrasfase (ALT) and lactate dehydrogenase (LDH) activities increased significantly after 8 and 12 weeks of treatment, while plasma alkaline phosphatase (ALP) activity was increased significantly only after 12 weeks. Plasma urea concentration was comparable in the two groups during the experimental period, while plasma creatinine concentration increased significantly after 12 weeks of treatment.     

Immunofluorescent antibody response to lactic dehydrogenase virus in different strains of mice.

The development of antibody against lactic dehydrogenase virus in five strains of mice (NZB x NZWF1, BALB/c, C.B-17, ICR and C.B-17 scid or SCID mice) was examined by indirect immunofluorescence (IIF) of infected liver sections. IIF antibody appeared 1 to 3 weeks and rose progressively 2 to 4 weeks after infection in four strains of mice (NZB x NZWF1, BALB/c, C.B-17 and ICR mice). SCID mice did not develop antibody. These results suggest that IIF may be applicable for detecting LDV infection in many other ordinary strains of mice.     

Bone-specific alkaline phosphatase protein, total alkaline phosphatase activity and lactate dehydrogenase in sera of patients with sickle cell disease

Serum bone-specific alkaline phosphatase protein (bAP) was evaluated as indicator of bone turnover by immunoradiometric assay (IRMA) in twenty patients with sickle cell disease and in twenty healthy control subjects. Serum bAP was also compared with serum total alkaline phosphatase activity and serum lactate dehydrogenase in the same group. The concentrations of serum bAP and serum lactate dehydrogenase were significantly higher in the study group than in the control group (p < 0.05, p < 0.01, respectively). The serum total alkaline phosphatase activity showed no significant difference with the control healthy subjects. There was no correlation between serum bAP and total alkaline phosphatase or lactate dehydrogenase levels in the patient group. In conclusion, serum bAP protein measured by IRMA can be considered a sensitive marker of bone turnover and could be especially useful as valuable non-invasive biochemical marker for identifying sickle cell patients with skeletal complications.     

Metabolic changes in rat brain histaminergic neurons during subhepatic cholestasis

The aim of the present work was to assess metabolic changes in histaminergic neurons in the rat brain during subhepatic cholestasis. Studies were performed on male Wistar rats using quantitative histochemical methods. The results showed that in cholestasis, histaminergic neurons in the rat hypothalamus developed significant changes in succinate dehydrogenase, lactate dehydrogenase, and glucose-6-phosphate dehydrogenase activity, in NADH and NADPH, and in acid phosphatase and monoamine oxidase B. These changes depended on the duration of cholestasis and had a dynamic, wave-like nature. The changes were apparent after five days of cholestasis, reached a maximum at 10–20 days, decreased at 45 days, and completely disappeared at 90 days. __________ Translated from Morfologiya, Vol. 132, No. 4, pp. 27–30, July–August, 2007.     

Metabolic myopathy produced by acute inhibition of glyceraldehyde-3-phosphate dehydrogenase with ortho-iodosobenzoic acid

A previously developed model of exercise-induced muscle contracture using iodoacetate to inhibit glyceraldehyde-3-phosphate dehydrogenase in rat hindlimb muscles produced selective type II myofiber damage. Utilizing a modification of the same model system, rats were given intra-aortic ortho-iodosobenzoic acid (700 nmol/kg body weight), which cleaves tryptophanyl peptides from glyceraldehyde-3-phosphate dehydrogenase. Within 2-4 h, spontaneous electrically-silent contracture developed in the injected musculature resulting in a plantar-flexed position of the hindlimb. After 24 h, the extensor digitorum longus and tibialis anterior muscles appeared grossly swollen (edematous) and discolored. Microscopically, the extensor digitorum longus (composed predominantly of type II myofibers) contained many randomly scattered, damaged myofibers, reduced glycogen content, absent glyceraldehyde-3-phosphate dehydrogenase activity, interstitial edema and focal collections of mononuclear phagocytes. Damaged fibers showed degenerative changes and contained stainable intracellular calcium. On modified trichrome-stained sections, an outer red staining rim of material was identifiable in many fibers. The fibers of the soleus muscle (composed predominantly of type I myofibers) were not damaged, indicating a preferential ortho-iodosobenzoic acid effect on type II myofibers.     

Effects of ischaemia on enzyme-activities in the soleus muscle of the rat

1. The changes with the time of the activities of some energy-supplying enzymes and of the hydrolytic enzyme, acid phosphatase, were studied over 2 weeks of complete ischaemia, produced in the rat soleus muscle by section of the abdominal aorta and terminal devascularization, leaving nerve and tendon intact. 2. Activities of glycolytic enzymes, oxidative enzymes, hexokinase and acid phosphatase are affected in a different manner. Activities of the glycolytic enzymes, lactate dehydrogenase, triosephosphate dehydrogenase and glycerolphosphate dehydrogenase, are lowest on the 1st day and increase thereafter. The first two reach the control values again on the 4th and 14th day, respectively, while glycerolphosphate dehydrogenase reaches about 50% of the control value on the 14th day. The maximum decrease in activity of the oxidative enzymes, citrate synthase, beta-hydroxyacyl-CoA-dehydrogenase and malate dehydrogenase occurs later (4th day); thereafter their activity returns slowly to control values, but does not reach them even on the 14th day. Hexokinase activity is slightly decreased on the 1st day; then it increased and reached on the 7th day twice the control value. Thus on the 1st day the activity of the enzymes of aerobic metabolism prevail, and on the 4th day those of anaerobic carbohydrate (glucose) metabolism; the recovery of enzyme activity of aerobic oxidation occurs later. 3. Acid phosphatase activity increased from the 2nd day onwards, reaching up to 3 times the control value on the 4th day and still twice that value on the 14th day. This agrees well with the histochemical picture of acid phosphatase. 4. Histochemical changes of alkaline phosphatase activity reveal destruction of capillary endothelial cells during the first few days after operation and their later proliferation from the periphery, correlating with the loss and recovery of oxidative enzyme activity.     

The evaluation of liver function after partial hepatectomy in the rat: serum changes.

In serial studies of hepatic function in rats after 70% partial hepatectomy, quantitative changes were found in several of the serum components used clinically to assess liver status. The activities of the following enzymes were found to increase: gamma-glutamyl transpeptidase and lactic dehydrogenase were maximal 6 h postoperatively, while glutamic oxaloacetic transaminase and alkaline phosphatase reached peak values at 24 and 48 h respectively. Albumin levels were found to be relatively constant during the study; however, total protein concentration was lowest 6--12 h postoperatively, paralleling a decrease in globulin concentration. Bilirubin levels were elevated to 4x normal within 12 h after surgery. After partial hepatectomy calcium and phosphorus concentrations were significantly decreased at 24 and 12 h respectively. With the exception of alkaline phosphatase, the activities of all serum components measured returned to normal levels by 1 week after surgery; the alkaline phosphatase concentration continued to be elevated 2 weeks postoperatively.     

The evaluation of liver function after partial hepatectomy in the rat: serum changes

In serial studies of hepatic function in rats after 70% partial hepatectomy, quantitative changes were found in several of the serum components used clinically to assess liver status. The activities of the following enzymes were found to increase: gamma-glutamyl transpeptidase and lactic dehydrogenase were maximal 6 h postoperatively, while glutamic oxaloacetic transaminase and alkaline phosphatase reached peak values at 24 and 48 h respectively. Albumin levels were found to be relatively constant during the study; however, total protein concentration was lowest 6--12 h postoperatively, paralleling a decrease in globulin concentration. Bilirubin levels were elevated to 4x normal within 12 h after surgery. After partial hepatectomy calcium and phosphorus concentrations were significantly decreased at 24 and 12 h respectively. With the exception of alkaline phosphatase, the activities of all serum components measured returned to normal levels by 1 week after surgery; the alkaline phosphatase concentration continued to be elevated 2 weeks postoperatively.     

Chronic hypoxia delays myocardial lactate dehydrogenase maturation in young rats

The effect of exposure to hypobaric hypoxia for 4 weeks (oxygen pressure = 106 hPa), equivalent to 5500 m in altitude) on myocardial total lactate dehydrogenase (tLDH) activity and isoform (H and M) composition was comparatively studied in growing (4.5 weeks old) and in adult (4.5 months old) male rats. The consequences of the hypoxia-induced anorexia were checked in growing rats using a pair-fed group. Exposure to hypoxia induced a significant decrease in the H/tLDH ratio in the left (LV) and right ventricle (RV) of growing and adult rats. In adult rats this alteration was mainly a consequence of the significant increase in the specific activity of the M isomer, which resulted in an increase in the overall LDH activity. In contrast, in the LV of young rats exposed to hypoxia, the specific activity of the M isomer was similar to that of normoxic animals while the H isomer activity was significantly lower than in normoxic rats, and the overall LDH activity remained unchanged. These effects were specifically due to hypoxia per se since no significant alterations were observed in pair-fed animals. In the hypertrophied RV, the alteration of H and M isomers following hypoxia was similar to that observed in adults (i.e. no change in H and an increase in M isoform). We conclude that the well-known hypoxia-induced decrease in the H/tLDH ratio is governed by different age-dependent mechanisms. In adult rats, hypoxia may induce in both ventricles a stimulating effect on M isomer expression. In the LV of growing rats this stress could inhibit the H isomer maturation without any effect on the M isomer. In the RV of growing rats this effect could have been counteracted by the growth effect of the hypertrophying process.     

Islet graft-induced changes of beta-hydroxybutyrate dehydrogenase and glucose-6-phosphatase activity in liver cells of diabetic recipient rats.

It is known that the liver is a favourable site for implantation of pancreatic islets since the grafted islets remain metabolically intact and provide long-term normoglycemia in diabetic animals. However, the long-term effects exerted by the grafted tissue on the host organ are not well defined. We therefore investigated by light and electron microscopy the effects of syngeneic islets on the host organ after intraportal transplantation into the liver of streptozotocin (STZ)-induced diabetic LEW.1W rats. In addition, tissue sections of graft-bearing liver were stained by enzyme histochemical methods for beta-hydroxybutyrate dehydrogenase (HBDH) and glucose-6-phosphatase (G6Pase). At 12 weeks after transplantation, the changes seen in the hepatocytes surrounding the grafted islets were hyperproliferation and accumulation of glycogen. Hepatocytes adjacent to the implanted islets displayed increased HBDH activity, whereas G6Pase activity was variable, either decreased or increased. Increased HBDH activity was also observed in the periportal region and in liver cells extending to the central veins. The results demonstrate that intraportal islet grafts, in addition to normalizing glucose homeostasis, exert remarkable effects on the liver parenchyma of experimentally diabetic recipient rats.     

不同剂量地塞米松对大鼠骨质量的作用

观察不同剂量地塞米松(Dex)对大鼠骨质量的影响并探讨其作用机制。31只3月龄SPF级雌性SD大鼠随机分为4组:对照组(saline 7只)、Dex-L组(1 mg/kg 8只)、Dex-M组(2.5 mg/kg 8只)、Dex-H组(5 mg/kg 8只);尾静脉注射Dex,2次/周,8周后处死。取胫骨上段制作不脱钙骨切片进行骨形态计量学分析,取左侧尺骨用于测定钙、磷和羟脯氨酸含量,股骨进行骨密度(BMD)和生物力学检测。结果表明:与对照组相比,Dex各组体重均显著下降;胫骨荧光周长百分率、矿化沉积率、骨形成率等显著降低,骨小梁面积百分数,骨小梁数目显著增加,骨小梁分离度显著减少;Dex-L BMD和Dex-M羟脯氨酸含量明显下降,Dex各组力学参数显著降低。提示:Dex显著抑制骨形成,降低骨转换,随着剂量增加,骨小梁面积百分数增加,BMD和骨生物力学性能下降,同时骨内有机质合成降低,非矿化骨形成增加,骨质量降低。以1 mg/kg的Dex对骨的质量影响最明显,该剂量略增加骨小梁面积百分数,但骨形成、骨生物力学性能、骨内有机质含量明显下降。不同剂量的Dex对骨质量的影响不同,剂量与骨质量之间无比例依赖关系。