Sexual Dimorphism in Development of Kidney Damage in Aging Fischer-344 Rats

BackgroundAging kidneys exhibit slowly developing injury and women are usually protected compared with men, in association with maintained renal nitric oxide.ObjectivesOur purpose was to test 2 hypotheses: (1) that aging intact Fischer-344 (F344) female rats exhibit less glomerular damage than similarly aged males, and (2) that loss of female ovarian hormones would lead to greater structural injury and dysregulation of the nitric oxide synthase (NOS) system in aging F344 rat kidneys.MethodsWe compared renal injury in F344 rats in intact, ovariectomized, and ovariectomized with estrogen replaced young (6 month) and old (24 month) female rats with young and old intact male rats and measured renal protein abundance of NOS isoforms and oxidative stress.ResultsThere was no difference in age-dependent glomerular damage between young or old intact male and female F344 rats, and neither ovariectomy nor estrogen replacement affected renal injury; however, tubulointerstitial injury was greater in old males than in old females. These data suggest that ovarian hormones do not influence these aspects of kidney aging in F344 rats and that the greater tubulointerstitial injury is caused by male sex. Old males had greater kidney cortex NOS3 abundance than females, and NOS1 abundance (alpha and beta isoforms) was increased in old males compared with both young males and old females. NOS abundance was preserved with age in intact females, ovariectomy did not reduce NOS1 or NOS3 protein abundance, and estrogen replacement did not uniformly elevate NOS proteins, suggesting that estrogens are not primary regulators of renal NOS abundance in this strain. Nicotinamide adenine dinucleotide phosphate oxidase-dependent superoxide production and nitrotyrosine immunoreactivity were increased in aging male rat kidneys compared with females, which could compromise renal nitric oxide production and/or bioavailability.ConclusionsThe kidney damage expressed in aging F344 rats is fairly mild and is not related to loss of renal cortex NOS3 or NOS1 alpha.     

Subregional Expression of Hippocampal Glutamatergic and GABAergic Genes in F344 Rats with Social Isolation after Weaning

Many studies have shown that postweaning social isolation (pwSI) alters various behavioral phenotypes, including hippocampus-dependent tasks. Here, we report the comprehensive analysis of the expression of glutamatergic and GABAergic neurotransmission-related genes in the distinct hippocampal subregions of pwSI rats. Male F344 rats (age, 4 wk) experienced either pwSI or group housing (controls). At 7 wk of age, the hippocampus of each rat was removed and laser-microdissected into the CA1 and CA3 layers of pyramidal cells and the granule cell layer of the dentate gyrus. Subsequently, the expression of glutamatergic- and GABAergic-related genes was analyzed by quantitative RT-PCR. In the CA1 and CA3 pyramidal cell layers, 18 of 24 glutamate receptor subunit genes were at least 1.5-fold increased in expression after pwSI. In particular, the expression of several N-methyl-D-aspartate and kainate receptors (for example, Grin2a in CA1, Grik4 in CA3) was significantly increased after pwSI. In contrast, pwSI tended to decrease the expression of GABA_A receptor subunit genes, and Gabra1, Gabra2, Gabra4, Gabra5, Gabrb2, Gabrg1, and Gabrg2 were all significantly decreased in expression compared with the levels in the group-housed rats. These results indicate a subregion-specific increase of glutamate receptors and reduction of GABA_A receptors, suggesting that the hippocampal circuits of pwSI rats may be in more excitable states than those of group-housed rats.Abbreviations: AMPA, α-amino-2,3-dihydro-5-methyl-3-oxo-4-isoxazolepropanoic acid, LMD, laser microdissection, NMDA;N-methyl-D-aspartic acid, pwSI, postweaning social isolation, DG;dentate gyrusBehavioral phenotypes differ among strains of laboratory animals, including mice and rats. The acquisition of strain-specific behavioral traits is governed not only by the genetic background but also by the postnatal rearing environment. The level of maternal care is a representative environmental factor. For example, inbred Fischer 344 pups raised by Wistar dams showed Wistar-like behavioral traits in adulthood in socially interactive situations.³⁵ Furthermore, the effect of environmental factors is not limited to neonatal periods. In animals that form a society among conspecifics, such as mice and rats, postweaning social environments have a marked effect on behavioral traits. Many studies have reported that postweaning social isolation (pwSI) alters various strain-specific behavioral phenotypes, including aggressiveness,^15,32,34 novelty preference,²² locomotor activity,²⁹ anxiety-like behavior,²⁰ and learning and memory.^13,15,20,31 In other words, pwSI means the deprivation of several social interactions among conspecifics. The expression of social contact begins fundamentally in the postweaning periods, its frequency increases to a peak at 4 to 5 wk of age, and it declines thereafter until sexual maturity.^2,17,30 Therefore, rats were isolated from their conspecifics to deprive them of social contact during the current study.In the brain, the hippocampus is necessary for the acquisition of episodic and spatial memory,²⁸ but the influences of pwSI on hippocampal functions remain largely unclear. The hippocampus has a lamellar organization of neurons, and the intrinsic neuronal circuit of the hippocampus, termed the trisynaptic circuit, consists of 3 topographically and morphologically distinct neuronal layers: the pyramidal cell layer in subfields CA1 and CA3 and the granule cell layer in the dentate gyrus (DG). Sensory information is carried first to the DG by a perforant pathway that originates in the entorhinal cortex. DG granule cells project to the apical dendrites of the CA3 pyramidal cells through mossy fibers. In turn, CA3 pyramidal cells project to the CA1 layer through Schaffer collaterals.^1,18 Glutamate is a key excitatory neurotransmitter in the hippocampus and plays a central role in the activation of the trisynaptic circuit, whereas the inhibitory neurotransmitter GABA modulates the activated circuit. This excitatory–inhibitory balance is critical for the appropriate functioning of the hippocampal circuit. Here, we comprehensively investigated the effect of pwSI on the expression of glutamatergic and GABAergic neurotransmission-related genes in the 3 hippocampal subregions of inbred F344 rats.     

Strain Differences in SA Gene Expression in Brain and Kidney of Normotensive and Hypertensive Rats

SUMMARY 1. In situ hybridization done using a ³⁵S-cRNA probe was carried out to obtain information on the expressions of the SA gene in brains and kidneys of the spontaneously hypertensive rat (SHR) strain obtained from the Izumo colony (/Izm) and from Charles River Laboratories (/Crj).2. In the brain, SA mRNA expression was most abundantly observed in epithelial cells of the choroid plexus. High to moderate levels was present on neurons of the CA1–CA4 pyramidal cell layer and the dentate gyrus of the hippocampus and the cerebellar Purkinje cell layer. The solitary tract nucleus and the dorsal motor nucleus of the vagus expressed the SA gene at very low levels. An increase in the expression was noted in the choroid plexus of WKY/Crj; there was no difference, however, in expression levels of other brain areas between WKY/Izm, SHR/Izm, and SHRSP/Izm, and between WKY/Crj and SHR/Crj.3. In the kidney, expression signals of SA mRNA were observed in renal medullary rays and focal cortex of WKY/Izm, SHR/Izm, SHRSP/Izm, and SHR/Crj, whereas mRNA expression in the WKY/Crj kidney was observed in medullary rays and outer strips of the outer medulla. Microscopically, hybridization signals were predominant in the proximal tubules.4. Expression densities decreased only in the kidney of WKY/Crj in 4-and 8-week-old rats, but not in the WKY/Izm kidney, compared with findings in SHR and SHRSP kidneys. These observations are in good agreement with data from Northern blot analysis.5. The SA gene expressions in the brain and the kidney seem not to relate to states of elevated blood pressure, but rather to strain differences. Abundant expressions in the brain and the kidney may mean that the SA gene plays a role in the water–electrolyte transport system. It is noteworthy that there are neuronal expressions of the SA gene in hippocampal pyramidal cells and cerebellar Purkinje cells.     

青花菜雄性不育相关基因BoDHAR的克隆与表达分析

以一个与甘蓝显性核不育相关的差异表达片段的序列为信息探针,通过在NCBI与TAIR网站数据库中进行同源EST序列搜索,经人工拼接、RT-PCR、PCR克隆与序列分析,获得了青花菜脱氢抗坏血酸还原酶DHARdehydroascorbatereductase基因的cDNA与DNA全长序列,命名为BoDHAR。并利用双链接头介导PCR的染色体步行技术(genomewalking)克隆了其上游644bp的5′端序列。所获的BoDHAR基因全长1486bp,存在两个内含子,DNA编码区序列633bp,编码210个氨基酸;序列分析表明BoDHAR与同源基因AT1G19570.1cDNA序列有82.3%的一致性,推导的氨基酸序列有79.6%的一致性;编码的水溶性蛋白存在多个磷酸化位点;5′端上游区存在明显的转录调控序列。半定量RT-PCR结果表明BoDHAR在可育系花蕾中的表达量明显高于不育系花蕾,在花药中的表达明显高于其它部位。     

F344大鼠常见非增殖性病变病理学观察

目的建立F344大鼠自发性非增殖性病变背景数据,为药物安全性评价提供基础。方法 SPF级F344大鼠336只,雌雄各半,8周龄,本中心屏障环境下饲养,随机分为4个组,分别于6、12、18和24个月4个时间点处死动物32、64、64、176只,常规病理学取材制片后显微镜检查,观察统计四个时间点动物自发性病变的病变类型和发病率。结果主要的非增殖性病变包括乳头肌纤维化、肺细支气管旁慢性炎症、肾脏病变包括肾小管扩张、蛋白管型、肾小管萎缩和钙沉积等,睾丸精子生成障碍、卵巢生长卵泡数量减少等生殖器官组织的退行性改变等。结论在本中心现有条件下,随动物生存期的延长,F344大鼠自发性非增殖性疾病病变程度和发病率均有所增加。本实验丰富了F344大鼠自发性病变的背景资料,为药物安全性评价提供基础。     

亚硒酸钠对糖尿病肾病大鼠肾脏Nephrin表达的影响与意义

探讨亚硒酸钠对糖尿病肾病大鼠肾脏Nephrin表达的影响及二者间的关系,从而研究亚硒酸钠和Nephrin在糖尿病肾病中的作用机制.通过链脲佐菌素法及给予高脂饮食诱导模拟大鼠糖尿病肾病模型,实验设空白对照组、糖尿病肾病对照组、亚硒酸钠干预组,亚硒酸钠干预组每日给予亚硒酸钠溶液灌胃,其它组给予等量生理盐水灌胃.灌胃10周后处死大鼠,取血、尿标本测相关生化指标.取肾脏组织戊二醛固定制作切片电镜下观察超微结构改变,取肾脏组织多聚甲醛固定制石蜡切片光镜下观察病理改变和免疫组化定位蛋白表达.取肾脏组织RT-PCR检测Nephrin的mRNA表达、Western Blotting检测nephrin的蛋白表达,分析各组数据的统计差异.结果发现亚硒酸钠干预组大鼠基本状况和生化指标较糖尿病肾病对照组明显改善,光镜和电镜下观察病理改变和超微结构病变较糖尿病肾病对照组明显减轻.免疫组化nephrin蛋白表达着色糖尿病肾病对照组较空白对照组减少,亚硒酸钠干预组较糖尿病肾病对照组着色明显增多.Nephrin mRNA和蛋白表达糖尿病肾病对照组较空白对照组明显降低,而亚硒酸钠干预组较糖尿病肾病对照组升高,但低于空白对照组,差异均有统计学意义（P〈0．05）．亚硒酸钠明显促进肾脏Nephrin表达,改善了糖尿病肾病,表明亚硒酸钠和Nephrin在防治和延缓糖尿病肾病的发生发展中可能起重要作用．     

Dietary Chemoprevention of PhIP Induced Carcinogenesis in Male Fischer 344 Rats with Tomato and Broccoli

The heterocyclic amine, 2-amino-1-methyl-6-phenylimidazo[4,5-B]pyridine (PhIP), found in meats cooked at high temperatures, has been implicated in epidemiological and rodent studies for causing breast, prostate, and colorectal cancers. A previous animal study using a xenograft model has shown that whole tomato and broccoli, when eaten in combination, exhibit a marked effect on tumor reduction compared to when eaten alone. Our aim was to determine if PhIP-induced carcinogenesis can be prevented by dietary consumption of whole tomato + broccoli powders. Male Fischer 344 rats (n = 45) were randomized into the following treatment groups: control (AIN93G diet), PhIP (200 ppm in AIN93G diet for the first 20 weeks of the study), or tomato + broccoli + PhIP (mixed in AIN93G diet at 10% each and fed with PhIP for 20 weeks, and then without PhIP for 32 weeks). Study animals were monitored for 52 weeks and were euthanized as necessary based on a set of criteria for health status and tumor burden. Although there appeared to be some hepatic and intestinal toxicity due to the combination of PhIP and tomato + broccoli, these rodents had improved survival and reduced incidence and/or severity of PhIP-induced neoplastic lesions compared to the PhIP-alone treated group. Rats eating tomato + broccoli exhibited a marked decrease in the number and size of cribiform prostatic intraepitheilial neoplasia/carcinoma in situ (cribiform PIN/CIS) lesions and in the incidence of invasive intestinal adenocarcinomas and skin carcinomas. Although the apparent toxic effects of combined PhIP and tomato + broccoli need additional study, the results of this study support the hypothesis that a diet rich in tomato and broccoli can reduce or prevent dietary carcinogen-induced cancers.     

Exercise Is More Effective at Altering Gut Microbial Composition and Producing Stable Changes in Lean Mass in Juvenile versus Adult Male F344 Rats

The mammalian intestine harbors a complex microbial ecosystem that influences many aspects of host physiology. Exposure to specific microbes early in development affects host metabolism, immune function, and behavior across the lifespan. Just as the physiology of the developing organism undergoes a period of plasticity, the developing microbial ecosystem is characterized by instability and may also be more sensitive to change. Early life thus presents a window of opportunity for manipulations that produce adaptive changes in microbial composition. Recent insights have revealed that increasing physical activity can increase the abundance of beneficial microbial species. We therefore investigated whether six weeks of wheel running initiated in the juvenile period (postnatal day 24) would produce more robust and stable changes in microbial communities versus exercise initiated in adulthood (postnatal day 70) in male F344 rats. 16S rRNA gene sequencing was used to characterize the microbial composition of juvenile versus adult runners and their sedentary counterparts across multiple time points during exercise and following exercise cessation. Alpha diversity measures revealed that the microbial communities of young runners were less even and diverse, a community structure that reflects volatility and malleability. Juvenile onset exercise altered several phyla and, notably, increased Bacteroidetes and decreased Firmicutes, a configuration associated with leanness. At the genus level of taxonomy, exercise altered more genera in juveniles than in the adults and produced patterns associated with adaptive metabolic consequences. Given the potential of these changes to contribute to a lean phenotype, we examined body composition in juvenile versus adult runners. Interestingly, exercise produced persistent increases in lean body mass in juvenile but not adult runners. Taken together, these results indicate that the impact of exercise on gut microbiota composition as well as body composition may depend on the developmental stage during which exercise is initiated.     

小麦中雄性不育同源序列的分离、鉴定及表达分析

利用拟南芥中已克隆的雄性核不育基因MS2和水稻中假定雄性不育蛋白的保守区域,设计一对简并引物,并在太谷核不育小麦可育株及不育株花药中进行扩增,得到了一条134bp的片段。以该片段为基础,通过电子延伸得到一个长为1604bp的序列,该序列编码的氨基酸包含一段由200个氨基酸组成的雄性不育保守区。RT-PCR结果表明,该雄性不育同源序列只在小麦可育花药中表达,而在小麦败育花药、叶片和根中不表达,说明该雄性不育同源序列为花药发育特异基因。     

Spontaneous calcification in F344/Slc and F344/JCL rats

F344/Slc and F344/JCL rats 2 years of age were histologically examined for the incidence and distribution of calcification. In the male rats of both strains, calcification was observed in the testis, lung, brain, kidney, heart, aorta, cornea, prostate and seminal vesicle respectively. In the female F344/JCL rats, calcification appeared in the kidney, lung, cornea, brain, stomach, ovary and heart. Among these of both sexes, the lung was one of the most affected organs for calcification. On the other hand, calcification in the kidney was more severe and frequent in the females than in the males, suggesting that the sex may be one of enhancing factors for calcification.     

SHR与WKY大鼠肾脏组织基因表达差异分析

为了观察SHR与WKY大鼠肾脏组织的基因表达的差异,用T11G、AP6引物对12周龄的SHR与WKY大鼠肾脏mRNA进行了差异显示,得到了一条在SHR大鼠肾脏组织中高表达的差异显示条带,进一步分析证明这一差异显示条带含有170 bp,与GenBank 收录的已知的核苷酸序列的同源性低于80%,可能为未知基因的cDNA片段,并定位于SHR大鼠的肾小管上皮细胞内.     

Expression of Tropomyosin-Encoding Gene in the Kidney Depends on Functioning of Vasopressin Gene

Brattleboro diabetes insipidus mutant rats and normal WAG rats were subjected to water loading or thirst during 3 days. It was found that tropomyosin-encoding gene expression has a tissue-specific pattern in the kidney. Northern blot and western blot analysis had shown that the main expression of Tpm3(3) takes place in the renal medulla, and its intensity differs in normal and mutant rats. The differences between mutants with an ineffective vasopressin synthesizing system and the rats having an intact vasopressin gene were more distinct under long-lasting dehydration. The ratio between renal medullary tropomyosin of Brattleboro and WAG lines of rats was 39.76 +/- 0.90 versus 18.29 +/- 0.86 under water loading, and 46.12 +/- 2.14 versus 13.83 +/- 0.66 in thirst.