Effects of Dryk1A,ASF and alternative splicing of CaMKⅡδ on myocardial hypertrophy in renovascular hypertensive rats

AIM: To investigate the role of dual-specificity tyrosine phosporylation-regulated kinase 1A (Dyrk1A)-alternative splicing factor (ASF)-calcium/calmodulin-dependent protein kinase Ⅱδ (CaMK Ⅱδ) pathway in the progression of myocardial hypertrophy in renovascular hypertensive rats. METHODS: The renovascular hypertension was induced by two-kidney one-clip (2K1C) method. The changes of blood pressure and myocardial hypertrophy were measured. The techniques of RT-PCR and Western blotting were used to detect CaMKⅡδ alternative splicing and the protein expression of Dyrk1A and ASF, respectively. RESULTS: Eight weeks after operation, systolic blood pressure (SBP) and diastolic blood pressure (DBP) in 2K1C rats increased (P<0.05). The increases in left ventricular weight (LVW), the ratio of LVW to body weight (BW) and the area of myocardial cells indicated that the hypertensive rats developed significant cardiac hypertrophy. The protein expression of Dyrk1A and mRNA expression of CaMKⅡδA and δB were significantly increased, while the protein expression of ASF and mRNA expression of CaMKⅡδC were decreased compared with sham-operated control rats (P<0.05). Treatment with Dryk1A inhibitor epigallocatechin gallate (EGCG) or harmine effectively attenuated cardiac hypertrophy and reversed the changes in the protein expression of Dyrk1A, ASF and alternative splicing of CaMKⅡδ (all P<0.05). CONCLUSION: Dyrk1A-ASF-CaMKⅡδ pathway plays a role in the development of myocardial hypertrophy in renovascular hypertensive rats.     

Change of synapsins in hippocampal formation during spacial learning and memory in rats

AIM and METHODS: The immunohistochemical method was used to study the on synapsin express in hippocampal formation during spatial learning and memory in rats.RESULTS: (1) In control group there are diffuse brown granules without any obvious granular product in hippocampal formation and in model group there are synapsin products in the hippocampal formation ,especially in the subregion CA3 , CA4 and the dentate gyrus after water maze trained for one week. After two weeks water maze trained the hippocampal formation appear the same distribution of the product granules with darker staining. (2) The light dendity of synapsin products between the model and the control groups showed significant differences, that in the model group was highter than that in the control group (P＜0.05). The light dendity of synapsin products increased with the training time from the first week to the second week in the model group. And there is no increase with the training time from the second week to the third week . The light dendity is higher in the CA3, CA4 subregion and the dentate gyrus than the CA1 and CA2 subregion (P＜0.05).CONCLUSIONS: (1) As mentioned above, it is believed that the new synapes are formed in the hippocampal formation of the rat by spacial learning and memory training. (2) The CA3, CA4 subregion and the dentate gyrus are correlated with the spacial learning and memory.     

Effect of 2, 3-butanedione monoxime (BDM) on myocardial protection in cold cardioplegia

AIM: The effects of BDM on isolated rat heart in cold cardioplegia were studied. METHODS: Rat heart were subjected to cold cardioplegia at 4℃ for 8, 18 and 24 h.Then each heart was perfused (90 cm H₂O) in Langendorff model at 37℃ for 40 min. In the high K⁺ group(n=24) the hearts were preserved in St.Thomas cardioplegic solution, in BDM group(n=24) hearts were preserved in K-H solution with BDM 30 mmoL/L. RESULTS: After 18 h, heart rate and the coronary flow in BDM group were significantly higher than in high K⁺ group(P＜0.05). Activity of Na⁺-K⁺-ATPase in cell membrane and in mitochondrial membrane in the BDM group was significantly higher than high K⁺ group(P＜0.01). After 24 h, all hearts in high K⁺ group were dead, but were alive in BDM group. CONCLUSION: Under given experiment conditions, BDM did enhance the tolerance to cold ischemia significantly. The results showed that BDM may become a useful agent for prolong the storage period of heart in cold cardioglegia.     

p38MAPK expression and apoptosis in the rat unilateral ureteral obstruction model

AIM: To investigate the expression of p38 mitogen-activated protein kinase (p38MAPK) in the kidney after unilateral ureteral obstruction (UUO) in rats and the functional role of it on apoptosis and fibrosis.METHODS: Eighteen Wistar rats underwent UUO were killed at 3, 7, 14 days. Additional 7 rats were sham operated. Histological changes were observed by HE and Masson staining. Immunohistochemistry study was performed on renal tissue for proliferating cell nuclear antigen (PCNA). Apoptotic cells were determined by terminal deoxynucleotidyl transferase- mediated dUTP nick end labeling (TUNEL) and the electrophoresis analysis of genomic DNA. Western blotting of cysteinyl aspartate specific proteinase-3 (caspase-3), p38MAPK and p-p38MAPK were measured.RESULTS: UUO induced a significant increase in renal tubular and interstitial cell apoptosis, immunohistochemistry of PCNA and Western blotting of caspase-3, p-p38MAPK as well as severe morphology changes. However, there was no significant difference between UUO and the control in Western blotting of p38MAPK.CONCLUSION: An in vivo model of renal fibrosis after UUO demonstrates that activated or phosphorylated p38MAPK plays a role in apoptosis of renal tubulointerstitial cells.     

Roles of four liver cells in regulation of PA-plasmin system during liver fibrogenesis in rats

AIM: To investigate the roles of hepatocytes, hepatic stellate cells (HSCs), Kupffer cells (KCs) and endothelial cells (ECs) in the regulation of PA-plasmin system during liver fibrogenesis in rats. METHODS: Experimental liver fibrosis was induced in rats by injection of carbon tetrachloride (CCl₄) twice a week for 12 weeks. Four kinds of liver cells were separated from the normal and fibrotic livers of the rats. The expression levels of urokinase plasminogen activator (uPA), uPA receptor (uPAR) and PA inhibitor-1 (PAI-1) in liver cells were determined by Northern and Western blotting. RESULTS: The expression of PAI-1 and uPAR was markedly increased in HSCs during liver fibrosis in rats as compared to those in the ECs. CONCLUSION: HSCs and ECs may play very important roles in the regulation of PA-plasmin system during liver fibrogenesis in rats. The activated HSCs are main cells to secrete PAI-1 and uPAR.     

Effect of hypothalamic arcuate nucleus damaged by monosodium glutamate on bone histomorphometry parameters in rat

AIM: To investigate the effect of hypothalamic arcuate nucleus(ARC) damaged by monosodium glutamate (MSG) on bone histomorphometry parameters in rats. METHODS: (1) Newborn SD rats of experimental group were hypodermically injected 10% MSG (4 g/kg BW) on the postnatal lst, 3rd, 5th, 7th, 9th day. Meanwhile, the newborn rats of the control group were given equal volume of normal saline and the rats after the postnatal 70th, 72th, 74th, 76th and 78th day were hypodermically injected 10% MSG (4 g/kg BW) as the MSG toxicity control group. After survivial for 160 days, all rats were killed. (2) Morphological methods were used to estimate the ARC neurons and the bone histomorphometry parameters. (3) Radioimmunoassay was used to measure the levels of the serum growth hormoen(GH), estradiol(E2) and testosterone(T). RESULTS: The number of the hypothalamic arcuate nucleus neurons, the levels of serum E2, T, GH and the %Tb·Ar, Tb·Th, Tb·N of proximal tibial metaphysis(PTM) significantly decreased, while Tb·Sp of proximal tibial metaphysis(PTM) significantly increased, and the osteoporotic alterations appeared obviously. All these changes did not appear in the rats of NS group and MSG toxicity control group. CONCLUSION: (1) The changes of the bone in MSG guoup rats are not the effect of the MSG toxicity on the bone directly. (2) The hypothalamic arcuate nucleus participates in the regulation of the bone metabolism. (3) ARC regulates bone metabolism via altering the functions of the hypothalamus-GH-IGF-1 axis and the hypothalamus-pituitary-gonal axis.     

Cell proliferation and nestin expression in cortex and SEZ of MCAO rats

AIM:To explore the effect of brain ischemia injury on cell proliferation and nestin expression in cortex and subependymal zone (SEZ).METHODS:Using a local brain ischemia model(MCAO), BrdU positive cells of cortex and subependymal zone (SEZ), also nestin positive cells, were observed by immunohistochemistry.RESULTS:BrdU and nestin positive cells in SEZ of MCAO rats were obviously increased. In cortex, only nestin positive cells were observed.CONCLUSION:Neural stem cells in SEZ and cortex were activated after brain ischemia, it may be related with neural recovery after brain ischemia injury.     

Effects of matrix metalloproteinases on ventricular remodelingfollowing myocardial ischemia in the rat

AIM:To examine the relationship between the activity of matrix metallproteinases(MMPs) and ventricular remodeling following myocardial ischemia in the rat.METHODS:The model of myocardial ischemia(MI) in the rat was established by isoprenaline(ISP). The activity of MMPs was measured by zymography and collagen concentration was assessed by the method of chloramine T, so did I/III collagen ratio by immunohistochemical staining. The microstructure of myocardium was also observed by electron microscope.RESULTS:The activity of MMP-2 in myocardial ischemia group (group M) increased by 5.8 folds at 1 st week(P＜0.01), 2.3 folds at 2 nd week(P＜0.01) and 1.7 flods at 4 th week(P＜0.05) compared with control group (group C) and MMP-9's activity in group M increased by 4.9 folds (P＜0.01), 1.9 flods(P＜0.01) and 1.4 folds(P＜0.05), respectively. Collagen amount and I/III collagen ratio in group M increased compared with that in group C at 2 nd week and 4 th week. It showed that cardiac myocytes in group M were necrosed and collagen grew abundantly in interstitium under electron microscope.CONCLUSION:The activity of MMPs in the myocardial interstitium increased following myocardial ischemia, then collagen amount and I/III collagen ratio increased, which may be the major causes of ventricular remodeling.