Age-related Expression Of AQP-3, 4 And Na⁺, K⁺-ATPase α₁ In Rat Kidneys

Objective: This study was aimed to investigate age-related expression of renal morphology and Aquaporin 4(AQP3), Aquaporin 4(AQP4) and Na+, K+-ATPase α1 in Sprague Dawley rats under physiological conditions, which could provide data for further physiological function study of AQP-3, 4 and Na+, K+-ATPase α1 at different ages of rats.Methods: The experiment utilized kidneys from pregnant 19 days(-2d) and postnatal 1d, 7d, 14 d, 21 d, 28 d, 35 d, 60 d, 180 d, 420 d, 540 d, and 850d-old Sprague Dawley(SD) rats. Total RNA was isolated from left kidneys and subjected to real-time quantitative polymerase chain(RT-PCR) analysis to test AQP3, AQP4, Na+, K+-ATPase α1 expression at gene levels. The 180 d, 420 d, 540 d, and 850d-old right kidneys were fixed for 24 hours in 4% paraformaldehyde solution. Hematoxylin-Eosin(HE) staining was observed for renal morphological changes at 180 d, 420 d, 540 d, and 850d-old rats. AQP3 and AQP4 protein expression at 420 d, 540 d, and 850 d kidneys were measured and analyzed by immunohistochemical(IHC) staining.Results:(1) The results of HE staining showed that kidney structure was obvious at postnatal 180 days. With the increase of time, glomerulus volume shrinked, accompanying with leaved glomerular, thickening of the glomerular basement membrane, lumen fusion, increased tubular contents and thinner wall thickness. Cell nucleus from180 to 850 days showed that granular degeneration increased gradually in renal tubular.(2) Results of IHC: The expression levels of AQP3 and AQP4 in renal cortex and medulla were higher at 420 d, lower at 540 d, and again higher at 850 d. Comparison of the expression amounts at 850 days with the expression amounts at 540 days showed increases(P<0.05). The expression of AQP3 and AQP4 in renal medulla was more than the cortex. The expression of AQP3 at 420 d was higher than that at 850 d, and the expression of AQP4 at 420 d was higher than that at 850 d.(3) Results of RT-PCR: AQP3 m RNA, AQP4 m RNA, and Na+, K+-ATPase α1 m RNA expression levels from pregnancy 19 days to 14 days after birth were low and were subsequently striking augment until 850 days of age. AQP3 m RNA, AQP4 m RNA expressions reached the peak at 35 days and then decreased. The comparison of expression quantities of AQP3 m RNA at 850 days with expression quantities of AQP3 m RNA at 1 days, 7 days, 14 days, 21 days, 180 days and 420 days after birth showed no statistical difference(P>0.05), and the comparison of expression quantities of AQP3 m RNA at 850 days with expression quantities of AQP3 m RNA at-2 days, 28 days, 35 days and 540 days after birth showed differences(P<0.05). Comparison of expression quantities of AQP4 m RNA at 850 days with expression quantities of AQP4 m RNA at 1 days, 7 days, 14 days, 21 days, 60 days, 180 days, 420 days and 540 days after birth showed no statistical differences(P>0.05), and comparison of expression quantities of AQP4 m RNA at 850 days with expression quantities of AQP4 m RNA at-2 days, 28 days and 35 days after birth showed statistical difference(P<0.05). Na+, K+-ATPase α1 m RNA expression quantities was in a state of high expression after postnatal 21 days, with a peak of expression at postnatal 180 days, and then reduced. Comparison of expression quantities of Na+, K+-ATPase α1 m RNA at 850 days with expression quantities of Na+, K+-ATPase α1 m RNA at 28 days, 35 days and 60 days after birth showed no statistical differences(P>0.05). Comparison of expression quantities of Na+, K+-ATPase α1 m RNA at 850 days with expression quantities of Na+, K+-ATPase α1 m RNA at 1 days, 7 days, 14 days, 21 days, 60 days, 180 days, 420 days and 540 days after birth showed statistical differences(P<0.05).Conclusion:(1) With the increase of age, glomerulus and renal tubules in rats show structural changes. Glomerular volume, glomerular cyst cavity and renal tubular cavity become small, wall thickness of renal tubule become thinner, glomerular basement membrane become thickened, and tubular contents and lumen fusion are increase in rats.(2) Renal cortex and medulla expression trends of AQP4 and AQP3 in rats in different periods are consistent and they express mainly in the medulla.(3) AQP3 m RNA, AQP4 m RNA and Na+, K+-ATPase α1 m RNA in rats are in the low expression levels before weaning. The amount of expression increase significantly after weaning and then decrease after the expression peak.(4) In different periods, the renal structure and expression of AQP3, AQP4, Na+, K+-ATPase α1 in rat kidneys vary in accordance with the general rules of the occurrence, development and senescence of life.