| The genus Nothobranchius is a kind of tropical freshwater fish which has short lifespan,fast growth and reproduction.It has become a new model animal for studying aging.In recent years,Nothobranchius has drawn more and more attention in immunology research.The thymus and kidney are important immune and hematopoietic organs of fish.On account of its small size,taking samples of thymus and kidney is difficult.And there are few reports about thymus and kidney of Nothobranchius in morphology so far.So it is necessary to understand the ontogenic development of the thymus and kidney.Studies of histological features on thymus and kidney in Nothobranchius can provide a histomorphological reference for related pathological and toxicological experiments.In the present study,we used the annual fish N.guentheri as experimental animal to explore their histological structure and age-related changes of thymus and kidney.In the process of aging,due to decline of immune function,aging cells accumulate in tissues[1],which may promote fibrosis and organ dysfunction through releasing factors such as IL-6,IL-8 and GRO[2,3].Resveratrol,an activator of SIRT1,has many pharmacological effects such as anti-inflammatory,anti-aging,antioxidant.Previous studies in our laboratory have shown that resveratrol significantly prolongs the life span of N.guentheri[4].Therefore,we have explored the effects of resveratrol feeding on kidney aging in the annual fish.ⅠThe development of thymus of N.guentheriFirstly,the age-related changes of thymus structure were observed in N.guentheri according to the results of HE staining.At least 5 fish were observed at each age stage.During the early development of the fish,the alymphoid thymic primordium was observed at 2 day after hatching（dah）.At 4 dah,the thymus showed a homogeneous distribution of lymphocytes and its shape was obvious.It rapidly increased in size and filled with lymphocytes.We observed a small aggregate of thymic epithelial cells at the inner of the organ.The thymus at 3-month-old fish could be divided into the cortex and the medulla.The extracortical epithelial cells were flat.The cortex was full of smaller lymphocytes,and the quantity of the lymphocytes in the medulla was less than that in cortical area.Secondly,the thymus of 4-month-old fish showed aging according to the results of?-galactosidase staining and the number of lymphocytes decreased.The thymus parenchyma showed an increase in the number of epithelium cysts at 6-month-old fish.Plasmocyte and lymphatic capillary were observed in the thymus.Furthermore,Masson staining showed increased connective tissue in thymus of9-month-old fish,and the thymus atrophied,vacuolization was severe,and macrophages and capillary vessel were observed.At 12-month-old fish,thymus was further atrophied,but not completely disappeared.ⅡThe development of kidney and the effects of resveratrol on kidney aging of N.guentheriThe age-related changes in kidney structure of N.guentheri were observed according to the results of HE staining.At least 5 fish were observed at each age stage.The kidney primordium was observed at 1 dah.The glomerulus was first visible at 4 dah.Glomerulus and renal tubules were distributed uniformly at 1-month-old fish.A large number of lymphoid hematopoietic cells appeared in the kidney at 2-,3-month-old fish.The kidney showed glomerular cystic enlargement in 6-month-old fish,and with many changes in 12-month-old fish,such as glomerular atrophy,tubular atrophy,and increased center of melanin macrophages.We used tissue staining,immunohistochemistry,and q PCR to detect kidney senescence and the effects of resveratrol on kidney with aging.The fish grew to sexual maturity at 4-month-old and they were randomly divided into a control group and resveratrol feeding group.The control group was fed with standard food and the treatment group was fed with resveratrol-supplemented food（25μg/fish/day）.The fish were killed as rapidly as possible when they were 6-,9-,12-month-old（Totally 90 fish,45 fish for controls and another 45fish were fed with resveratrol）and the experimental results as following:1.We found that?-galactosidase activity in the kidney showed a rising trend,and glomerular and renal interstitial fibrosis increased with aging,while resveratrol reduced the activity of?-galactosidase and improved renal fibrosis in the fish.2.We found that protein level of SIRT1 gradually decreased and gene level of NF-?B gradually increased with age of the fish,while resveratrol increased SIRT1 expression and decreased NF-?B expression significantly.Previous studies have shown that SIRT1 can inhibit transcription of NF-?B by deacetylating its subunit the Rel A/p65[5],and NF-?B regulates the release of inflammatory factors.Next,we detected the changes of inflammatory factors in the kidney of the fish.We found that resveratrol significantly decreased the level of pro-inflammatory factor IL-8 in kidney of 6-month-old fish,and increased anti-inflammatory cytokine IL-10expression in kidney of 9-month-old fish.In conclusion,the thymus and kidney of N.guentheri showed histological and morphological changes with age.During renal aging,resveratrol activated SIRT1,inhibited the transcriptional activity of NF-?B,and regulated the release of inflammatory factors.Finally,resveratrol decreased SA-?-gal activity with aging process,and improved renal fibrosis of the annual fish. |
