Expression of Angiotensinogen in the Uterus induced by Coagulating Gland Renin in Mice

Coagulating gland (CG) renin is one of the components of the local renin-angiotensin system (RAS), which plays important roles in the maintenance of homeostasis in several tissues. Although the existence of local renin has been reported in many tissues, its function is not yet well understood. In the present study, the relationship between CG renin and uterine angiotensinogen was investigated by immunohistochemical and in situ hybridization techniques. In mating experiments with male and female C57BL/6 mice, renin was demonstrated immunohistochemically on the epithelial cells of the uterus on the first day after coitus; however, it was not detected on the second and third days after coitus. Neither immunoreactive cells nor messenger signals for renin were demonstrated on the epithelial cells of the uterus throughout the experiments. On the first day after mating, it was noted that positive signals for angiotensinogen mRNA were expressed on the epithelial cells of the uterus in situ hybridization, but not on the second and third days after coitus. These results suggest the possibility that CG renin is transferred to the uterine epithelium at mating and temporarily activates the expression of angiotensinogen in the uterus. Angiotensin II produced by angiotensinogen may act as an important mediator for vascularization of the first step of pregnancy.     

Comparative study of renin-containing cells. Histological approaches

The juxtaglomerular apparatus is known to be the functional unit of renin control. In the present review, the author will describe the comparative characteristics of renin-containing (RC) cells as well as extrarenal distribution, paying special attention to developmental and topographical approaches. The characteristic locality of RC cells suggests that the secretion of renin is performed at a site beside the adventitia or via the glomerular capillaries. Ontogenetical and phylogenetical investigations of RC cells have provided interesting findings on their morphogenesis. Analysis of the endocrine kidney after unilateral obstruction of the ureter provides some findings about the origin of RC cells and the processing of renin granules. Observation of developing adrenal renin suggests that there is important involvement of angiotensin II produced by renin synthesis in the morphogenesis of the adrenal gland in the fetal stage. Coagulating gland (CG) renin is characterized by testosterone-regulated and exocrine mechanisms. Recently, all or some of the components of the renin-angiotensin system (RAS) have been reported to be synthesized and secreted outside of classical organs or tissues. In the future, the real function of local RAS will be clarified by using gene targeting in mice.     

Formation of monoethylglycinexylidide (MEGX) in clinically healthy dogs

Liver function tests help to investigate actual liver function. In dogs, only a few tests are available. We evaluated the formation of monoethylglycinexylidide (MEGX) in clinically healthy dogs to assess the usefulness of this liver function test in dogs. Twenty-five healthy dogs were used in this study. The MEGX test was done according to human protocols. The results of our study showed that dogs synthesize MEGX after the administration of lidocaine. There was no age dependence of this test in dogs and no significant difference between measurements obtained at 15 and 30 min after administration of lidocaine. Female dogs had significantly (P < 0.05) higher concentrations of MEGX 15 min after administration. The reference interval for dogs after 15 min is 34 to 79 μg/L and after 30 min 39 to 89 μg/L. In conclusion, the MEGX test may be an additional liver function test in dogs.     

Immunity to Toxocara vitulorum repeated infections in a rabbit model.

Eleven rabbits were infected with 10 embryonated eggs of Toxocara vitulorum per g body weight on Days 0, 35 and 72. Embryonated eggs and larvae were enumerated in feces on Days 1-3 after each infection. Two rabbits were killed and larvae were enumerated in small intestine, liver, lungs, skeletal muscles, heart, kidney, brain, eye, uterus, and mammary glands on Days 5, 15, 30, 65 and 101. Serum was obtained on Days 0, 5, 15, 30, 42, 50, 65, 78, 86 and 101 to perform enzyme-linked immunosorbent assays (ELISAs) and Western blots against an extract of embryonated eggs. Between 4 and 10% of the administered parasites, almost all embryonated eggs, were found in the feces after the first or second infection, but 32% (27% of them larvae) after the third. Yields of tissue parasites were 4.1% of the administered dose on Day 5, 2% on Day 15, and 0.8% on Day 30 of the first infection, 0.1% on Day 30 of the second infection, and 0.06% on Day 30 of the third. Larvae were found only in liver, lungs and muscle, including heart. Larva content declined steadily in liver and lungs from Day 5 to 30 of the first infection, was absent in the liver at Day 30 of the second, and in both organs at Day 30 of the third. Muscle larva content increased from Day 15 to 30 of the first infection, and persisted throughout the third infection. Production of IgM antibody was minimal, IgG and the sum of IgMGA antibodies increased slightly or moderately after the first and second infections, but dramatically after the third. Western blots revealed the first antigens (12) by Day 15 of the first infection. Their total number increased with time and number of infections, but some antigens disappeared, whereas new antigens appeared in the course of the observations. Four antigens (32,500-41,000 mol.wt.) may be related to protection. Comparison of the Western blot patterns of two rabbits showed differences in the antigens, recognizable for each rabbit.     

Characterization of key genes of the renin–angiotensin system in mature feline adipocytes and during in vitro adipogenesis

Obesity is a growing health problem in humans as well as companion animals. In the development and progression of obesity‐associated diseases, the members of the renin–angiotensin system (RAS) are proposed to be involved. Particularly, the prevalence of type 2 diabetes mellitus in cats has increased enormously which is often been linked to obesity as well as to RAS. So far, reports about the expression of a local RAS in cat adipocytes are missing. Therefore, we investigated the mRNA expression of various RAS genes as well as the adipocyte marker genes adiponectin, leptin and PPAR‐γ in feline adipocytes using quantitative PCR. To characterize the gene expression during adipogenesis, feline pre‐adipocytes were differentiated into adipocytes in a primary cell culture and the expression of RAS key genes measured. All major RAS components were expressed in feline cells, but obvious differences in the expression between pre‐adipocytes and the various differentiation stages were found. Interestingly, the two enzymes ACE and ACE2 showed an opposite expression course. In addition to the in vitro experiments, mature adipocytes were isolated from subcutaneous and visceral adipose tissue. Significant differences between both fat depots were found for ACE as well as AT1 receptor with greater expression in subcutaneous than in visceral adipocytes. Visceral adipocytes had significantly higher adiponectin and PPAR‐γ mRNA level compared to the subcutaneous fat cells. Concerning the nutritional status, a significant lower expression of ACE2 was measured in subcutaneous adipocytes of overweight cats. In summary, the results show the existence of a potentially functional local RAS in feline adipose tissue which is differentially regulated during adipogenesis and dependent on the fat tissue depot and nutritional status. These findings are relevant for understanding the development of obesity‐associated diseases in cats such as diabetes mellitus.     

Time-related morphological changes of porcine ovarian granulosa cells incubated with urea-EDTA solution.

Morphological profiles of porcine granulosa cells incubated with 10 mM Tris-HCl containing 1 M urea and 5 mM EDTA (urea-EDTA solution) were investigated. The percentages of granulosa cells incorporating the dye, trypan blue on incubation with urea-EDTA solution did not change during the initial 30 min. Thereafter, granulosa cells gradually took up the dye throughout the incubation. The amount of protein released from granulosa cells increased dramatically during initial 15 min of incubation, but decreased during the following 15-30 min of incubation. Thereafter, the amount of proteins released from granulosa cells increased gradually again. The releasing profile of 51Cr-bounded substances in granulosa cells increased markedly during the initial 15 min of incubation, and decreased during the next 15 min of incubation. Subsequently the amount of 51Cr released was enhanced. The min of incubation. Subsequently the amount of 51Cr released was enhanced. The plasma membranes of granulosa cells remained intact at 30 min of incubation, although chromatin clusters of granulosa cells disappeared. Thereafter, a number of cells showed signs of degeneration, including broken plasma membrane and cytolysis. The present study revealed that urea-EDTA solution is useful in extracting materials from porcine granulosa cells. The majority of the materials extracted from granulosa cells during the initial 30 min of incubation with urea-EDTA solution is considered to be from the cell surfaces and/or intercellular matrix.     

外源胰岛素和能量限饲对鸡PPP1R3C表达的效应

旨在检测PPP1R3C在爱拔益加肉鸡不同组织中的表达情况，探究外源胰岛素和能量限饲对鸡体胰岛素敏感组织中PPP1R3C表达的影响。试验一，取不同发育阶段的雌性肉鸡（E14、E19、D7和D21，n=10）组织样，通过qRT-PCR技术检测PPP1R3C在不同时期胸肌中的表达；试验二，腹腔注射胰岛素（INS）或PBS，取两组注射后不同时间点（0、15、120和240 min，n=5）的D24雄性肉鸡组织样，检测PPP1R3C在肉鸡不同组织中的表达，探究外源胰岛素处理对肉鸡胰岛素敏感组织中PPP1R3C表达的影响；试验三，取D18雌性肉鸡，一组饲喂常规日粮（n=20），另一组饲喂限制30%能量的日粮（n=20），饲喂至48天屠宰取样，探究30%能量限饲对肉鸡PPP1R3C表达的影响；试验四，D7雌性肉鸡随机分为3组，对照组、15%能量限饲组和15%蛋白限饲组（n=10），分别饲养至D21屠宰取样，探究能量限饲是否具有剂量依赖性。结果表明：1）PPP1R3C在胸肌组织中高表达，其次是心和腿肌组织（P<0.05）。2）PPP1R3C表达量呈现了明显随鸡发育而升高的趋势。3） INS注射显著下调了胸肌中PPP1R3C的表达，在注射后120 min时PPP1R3C的表达显著低于0和15 min （P<0.05）；PBS注射后胸肌中PPP1R3C的表达没有显著差异，在注射后120和240 min时，INS组PPP1R3C表达显著低于PBS组（P<0.05）。INS注射也降低了肝中PPP1R3C的表达，在INS处理15 min时PPP1R3C的表达已显著低于0 min （P<0.05）；PBS注射后肝中PPP1R3C表达处于动态平衡，在注射后120 min时，INS组PPP1R3C表达显著低于PBS组（P<0.05）。胰岛素注射后在腹脂中出现了与胸肌和肝相反的结果，在胰岛素注射后15 min时PPP1R3C的表达显著高于0、120、和240 min （P<0.05），而0、120、和240 min之间没有显著差异；PBS注射后腹脂中PPP1R3C表达没有显著变化，在注射后15 min时，INS组PPP1R3C表达量显著高于PBS组（P<0.05）。4）30%能量限饲导致PPP1R3C在胸肌和肝中的表达均显著下调（P<0.05）。5）15%能量限饲和15%蛋白限饲不能显著影响胸肌中PPP1R3C的表达。以上研究表明，PPP1R3C在胸肌中表达最高，并随着个体发育表达上升，且外源胰岛素对PPP1R3C的表达效应具有明显的组织特异性。30%能量限饲可产生类似于外源胰岛素的效应，且能量限饲存在一定的剂量依赖性，本研究结果为进一步揭示鸡PPP1R3C功能奠定了基础。     

The effects of microwave-irradiated fixation for postmortem changes of the kidney

In the present study, we evaluated the advantages of microwave-irradiated fixation for postmortem autolysis of the kidney. Mouse kidneys, sampled at 0, 1, 3, 5, 10, 15, 20 and 25 hr after death, were fixed with 10% neutral formalin by microwave irradiation (MWI; 20 sec/500 W) and by conventional immersion. They were then examined with light and electron microscopy, morphometrics and immunohistochemicals. Light microscopic and morphometric observations showed that structural preservation effect of MWI was limited to the proximal convoluted tubules at 25 hr. Contrary, mild ultrastructural damage by MWI was found in the glomeruli at 0 and 15 hr. Immunohistochemistry for renin and alpha-smooth muscle actin showed no apparent differences between MWI and the immersion.     

Biology and pathophysiology of Toxocara vitulorum infections in a rabbit model.

Ten female New Zealand rabbits were infected via stomach intubation with eggs of Toxocara vitulorum at a dosage of 10 embryonated eggs per gram of body weight on Days 0, 35 and 72. Ten or 4% of the administered parasites passed in the feces during the 3 days following the first or second infection, but 32% after the third infection. Many larvae were passed in the third infection, but not in the first or second. Tissue parasite yields were 4.1% on Day 5, 2% on Day 15, 0.8% on Day 30, 0.1% on Day 65 and 0.06% on Day 101. Five hundred and ninety-three larvae were recovered from liver, 243 from lungs and 0 from muscles on Day 5; 282 from liver, 138 from lungs and 21 from muscles on Day 15; 151 from liver, 21 from lungs and 50 from muscles on Day 30; 0 from liver, 26 from lungs and 15 from muscles on Day 65; 0 from liver, 0 from lungs and 9 from muscles on Day 101. No larvae were found in other tissues. The size of the muscle larvae at 30, 65 and 101 days indicated that the parasites did not develop beyond the infective stage and suggested that they were probably hypobiotic organisms. Erythrocytes, packed cell volume and monocytes decreased, but eosinophils and basophils increased, after each infection. Serum enzyme levels indicated that liver damage occured only after the first infection, but muscle injury occurred after each infection and was increasingly more precocious after each infection.     

Biology and pathophysiology of Toxocara vitulorum infections in a rabbit model

Ten female New Zealand rabbits were infected via stomach intubation with eggs of Toxocara vitulorum at a dosage of 10 embryonated eggs per gram of body weight on Days 0, 35 and 72. Ten or 4% of the administered parasites passed in the feces during the 3 days following the first or second infection, but 32% after the third infection. Many larvae were passed in the third infection, but not in the first or second. Tissue parasite yields were 4.1% on Day 5, 2% on Day 15, 0.8% on Day 30, 0.1% on Day 65 and 0.06% on Day 101. Five hundred and ninety-three larvae were recovered from liver, 243 from lungs and 0 from muscles on Day 5; 282 from liver, 138 from lungs and 21 from muscles on Day 15; 151 from liver, 21 from lungs and 50 from muscles on Day 30; 0 from liver, 26 from lungs and 15 from muscles on Day 65; 0 from liver, 0 from lungs and 9 from muscles on Day 101. No larvae were found in other tissues. The size of the muscle larvae at 30, 65 and 101 days indicated that the parasites did not develop beyond the infective stage and suggested that they were probably hypobiotic organisms. Erythrocytes, packed cell volume and monocytes decreased, but eosinophils and basophils increased, after each infection. Serum enzyme levels indicated that liver damage occured only after the first infection, but muscle injury occurred after each infection and was increasingly more precocious after each infection.     

Comparative oxidative metabolic profiles of clomipramine in cats, rats and dogs: preliminary results from an in vitro study

The objectives of this in vitro study were to describe cytochrome-dependent metabolism of clomipramine in canine and feline microsomes, compare metabolic profiles between cats, rats and dogs, and investigate a potential gender-related difference in metabolic activity between male and female cats. Pooled liver microsomes were incubated with clomipramine, where species and gender-specific reactions were initiated by the addition of a nicotinamide adenine dinucleotide phosphate regenerating system and quenched with methanol at 0, 5, 15, 30, 45 and 60 min, and 0, 30, 60, 90, 120, 180, 240 and 360 min respectively. Liquid chromatography tandem mass spectrometry was used to measure clomipramine and its metabolites. Preliminary results showed that cat microsomes biotransformed clomipramine slower and less efficiently than rat and dog microsomes. Moreover, gender differences in metabolic profiles suggested that male cat microsomes may be less efficient demethylators and hydroxylators than female cat microsomes. As gender metabolic differences may carry clinical significance for this antidepressant, further studies are warranted.     

Cellular localization of Visudyne® as a function of time after local injection in an in vivo model of squamous cell carcinoma: an investigation into tumor cell death

Objective To evaluate the effects of time on cellular localization of Visudyne^® after local injection. Animals Twenty athymic nude mice. Procedures A squamous cell carcinoma (SCC) cell line (A‐431) was injected into right and left dorsolumbar subcutaneous tissue of each mouse, representing treatment (T) and control (C) tumors. In experiment 1 (Exp 1; n = 10) and 2 (Exp 2; n = 10), the T tumors received a local injection of Visudyne^® (0.1 mg/cm³), and C tumors received an equal dose of 5% dextrose in water (D5W). Mice were randomly subdivided into two groups (A and B; n = 5 per group). Mice in Exp 1A and B were sacrificed 1 and 30 min after local injection, respectively. Experiment 1A and B tumors were evaluated by fluorescence microscopy to determine drug localization. Experiment 2A and B tumors were exposed to LED illumination 1 and 30 min after injection, respectively, and evaluated by transmission electron microscopy (TEM) to determine ultrastructural tumor cell damage. Results Fluorescence was detected within the cytoplasm of T tumors in both Exp 1A and B. Significance was detected in fluorescence intensity between T1 min vs. T30 min (P = 0.03) and between T1 min and C1 min tumors (P = 0.01), respectively. Tumors in Exp 2A and B demonstrated evidence of apoptotic cell death. Conclusions Fluorescence microscopy demonstrated higher Visudyne^® concentration within SCC cytoplasm of 1 min compared with 30‐min tumors. Transmission electron microscopy results revealed that tumors treated by photodynamic therapy (PDT) within 30 min of local injection undergo cellular apoptosis.     

No significant differences were observed in the amounts of DNA strand breaks produced by copper between male and female liver cells of long-evans cinnamon (LEC) strain rats.

The amounts of DNA single strand breaks that are oxidative damage produced by copper were examined by comet assay in the liver cells of an inbred strain of Long-Evans Cinnamon (LEC) rats that spontaneously develops fulminant hepatitis. At 4 weeks of age, copper contents in the liver of LEC rats were approximately 30-fold higher than those of WKAH rats that are control rats used in the present study. Copper accumulated in the liver of LEC rats in an age-dependent manner and no significant differences were observed between copper contents in the livers of males and females at each week of age from 4 to 15 weeks. No significant amounts of DNA strand breaks were found in the liver cells of both male and female WKAH rats from 4 to 15 weeks of age. DNA strand breaks were produced in the substantial population of LEC rat liver cells at 10 weeks of age and induced in an age-dependent manner from 10 to 15 weeks of age. The amounts of DNA strand breaks produced by copper accumulation in the liver cells of female LEC rats are not more abundant than those in the cells of male rats, although it has been reported that hepatitis in female rats is more serious than that in male rats.     

热处理对牛初乳理化性质和功能性蛋白的影响

科学控制牛初乳的加工工艺参数对初乳产品的开发与加工具有重要意义。本试验旨在探究热处理条件对初乳理化性质和功能性蛋白的影响,本研究设置7 种不同的热处理条件（55 ℃/30 min、55 ℃/60 min、60 ℃/30 min、60 ℃/60 min、63 ℃/30 min、63 ℃/60 min和72 ℃/15 s）,探究热处理后初乳理化指标、微生物菌落数和免疫球蛋白IgG以及乳铁蛋白的变化规律。结果表明,63 ℃条件下处理30 min和60 min,以及在72 ℃条件下处理15 s,出现明显的蛋白凝块;72 ℃处理15 s后显著降低初乳密度、酸度、乳蛋白、非脂固形物（SNF）、总固形物（TS）、柠檬酸、酪蛋白和尿素等理化指标;本研究所有热处理条件均显著降低菌落总数,有效控制大肠菌群和金黄色葡萄球菌数;经本研究设置的热处理后初乳中IgG水平显著降低,乳铁蛋白含量不受影响。综上考虑,建议初乳热加工温度控制在63 ℃以下,并根据生产目的针对性控制热加工参数。本研究可为牛初乳加工工艺参数控制提供详实科学数据支持。     

Renal lesions in spontaneous insulin-dependent diabetes mellitus in the nonobese diabetic mouse: acute phase of diabetes

The nonobese diabetic mouse is a model of spontaneous insulin-dependent diabetes mellitus. The present study made longitudinal observations of renal lesions in the acute-progressive phase of diabetic mice 0, 10, 20, 30, and 40 days after onset of diabetes without insulin therapy. Plasma creatinine and blood urea nitrogen concentrations gradually increased after onset of diabetes. Kidney weight increased and plateaued at day 20. Under electron microscopy the glomeruli demonstrated only mild changes on day 40. In the proximal tubules proliferating cell nuclear antigen-positive nuclei and nuclear divisions were increased on days 10 and 20. On day 40 of diabetes, increased periodic acid-Schiff-positive granules, confirmed as lysosomal dense bodies, increased neuronal nitric oxide synthase (nNOS) positive reaction, and decreased periodic acid-Schiff staining in the brush border were observed in the proximal straight tubules. In the juxtaglomerular apparatus stratified macula densa were decreased with time in diabetes compared with the findings on day 0, and this macula densa positively reacted with nNOS. No changes in renin levels were observed. In addition, apoptotic cells were not detected. In conclusion, this research represents the first thorough characterization of acute changes in nonobese diabetic mouse kidneys. The results demonstrated renal hypertrophy and slight glomerular injury in early stages and structural alteration of the proximal straight tubules at later stages during the acute phase of diabetes. Furthermore, increased nNOS may represent one of the pathogenic factors of diabetic nephropathy.     

Weaning causes a prolonged but transient change in immune gene expression in the intestine of piglets

Controlling gut inflammation is important in managing gut disorders in the piglet after weaning. Establishing patterns of inflammation markers in the time subsequent to weaning is important for future research to determine whether interventions are effective in controlling gut inflammation. The objective of this study was to evaluate the intestinal inflammatory response during the postweaning period in piglets. A 45-d study included 108 piglets (weaned at 22 d, body weight 5.53 ± 1.19 kg), distributed in 12 pens with nine pigs per pen. Histomorphometry, gene expression of pro- and anti-inflammatory cytokines, and the quantity of immunoglobulin (Ig) A producing cells were measured in jejunum, ileum, and colon on days 0, 15, 30, and 45 postweaning. Cytokine gene expression in peripheral blood mononuclear cells and Ig quantities were analyzed in blood from piglets on days 0, 15, 30, and 45 postweaning. Histomorphometrical results showed a lower villus length directly after weaning. Results demonstrated a postweaning intestinal inflammation response for at least 15 d postweaning by upregulation of IgA producing cells and IFN-γ, IL-1α, IL-8, IL-10, IL-12α, and TGF-β in jejunum, ileum, and colon. IgM and IgA were upregulated at day 30 postweaning. IgG was downregulated at day 15 postweaning. The results indicate that weaning in piglets is associated with a prolonged and transient response in gene expression of pro- and anti-inflammatory cytokines and IgA producing cells in the intestine.     

Pathophysiologic response of the juxtaglomerular apparatus to dietary sodium restriction in the dog.

Pathophysiologic changes in the juxtaglomerular apparatus (JGA) of the dog were induced by 10 days of dietary sodium restriction (less than 1 mEq of Na+/day). Plasma renin activity increased 12-fold and plasma aldosterone values increased 60-fold, whereas urinary sodium excretion decreased precipitously. Urinary potassium excretion remained within normal values throughout the period of sodium restriction. The JGA cell counts, determined by light microscopy, were significantly (P = less than 0.05) increased after 2 days of sodium restriction and remained increased through day 10. Adrenal gland weights and the cross-sectional width of the zona glomerulosa were not altered. Ultrastructurally, JGA cells showed progressive hypertrophy and hyperplasia. The Golgi apparatus became more prominent. The endoplasmic reticulum increased, as did the number of ribosomes. Cytoplasmic secretory granules increased in number and size from day 2 through day 6. On days 8 and 10, fewer and smaller secretory granules were encountered, even though plasma renin and aldosterone values continued to increase. In the dog maintained in a balanced sodium state, little renin is stored in cytoplasmic granules of the juxtaglomerular cells. Short-term stimulation results in increased plasma renin values and increased production and storage of renin in JGA cells. Continued stimulation results in depletion of cytoplasmic stores, although plasma renin content continues to rise, suggesting that renin is produced and secreted directly during more prolonged stimulation.     

游泳应激对小鼠血液和肝脏脂质过氧化水平的影响

为探讨游泳应激对小鼠血液和肝脏脂质过氧化水平的影响,试验选取50只昆明小鼠进行游泳试验,于游泳前和游泳10、20、30、50min后分别随机选取10只小鼠眼眶采血致死,采集血液和肝脏,测定样品中MDA含量、SOD和GSH-Px活性。结果表明,游泳应激使小鼠血清MDA含量持续升高并显著高于游泳前对照组(P<0.05);SOD活性先显著下降(P<0.05),之后缓慢升高,游泳50min后仍显著低于对照组(P<0.05);10min游泳应激使小鼠血清GSH-Px活性显著上升(P<0.05),之后缓慢下降,50min后与对照组无显著差异。游泳应激使小鼠肝脏MDA含量显著升高(P<0.05),游泳50min后肝脏SOD活性显著升高(P<0.05),GSH-Px活性在游泳20、30、50min后均显著下降(P<0.05)。结果提示游泳应激能显著影响小鼠血液和肝脏组织脂质过氧化水平。