Objective To clarify the association between non-dipping circadian blood pressure(BP) rhythm and left ventricular hypertrophy(LVH) in chronic kidney disease (CKD) patients. Methods A total of 257 CKD patients of stage 1 to 5 were enrolled in the study. The parameters of BP and circadian rhythm were measured by ambulatory BP monitoring (ABPM) and the cardiac structure was examined by echocardiography. The association between circadian BP rhythm and echocardiographic parameters was studied. Results The prevalence of abnormal circadian BP rhythm (non-dipping rhythm) was quite high (75.4%) in CKD patients and increased with the deterioration of renal function. Even if in the normal BP group, the prevalence of non-dipping rhythm was 71.3%. The change of cardiac structure such as LVH in non-dipping patients was more obvious than the dipping patients. The left ventricular mass index(LVMI) was positively correlated with BP, non-dipping rhythm. Multiple regression analysis showed that 24 h-SBP (β=0.417, P<0.01), triglyceride (TG) (β=-0.132, P=0.007), Hb (β=-0.394, P=0.016) and gender(β=0.158, P=0.039) were independent risk factors of LVMI. Conclusions The prevalence of non- dipping rhythm is quite high in CKD patients and increases with the deterioration of renal function. The change of cardiac structure such as LVH is obvious in CKD patients, especially in non-dipping group. The non-dipping rhythm is related with LVMI.

Objective To elucidate the prevalence of vitamin D insufficiency and deficiency in chronic kidney diseases (CKD) patients and provide the evidence for treatment of these patients. Methods Clinical data of 358 inpatients with CKD from stage 1 to stage 5 were analyzed retrospectively. Level of 25（OH）D3 in these inpatients, as well as the levels of intact parathyroid hormone (iPTH), hemoglobin (Hb), serum creatinine (Scr), urea nitrogen (BUN), carbon dioxide combining power (CO2CP), albumin (Alb), serum calcium (Ca) and blood serum (P) were examined. Correlation between 25（OH）D3 and parameters was analyzed. Results The mean level of 25（OH）D3 in these CKD patients was (18.58±11.7) µg/L, which was significantly lower than that of normal reference (P<0.01). The 25（OH）D3 levels of CKD patients from stage 1 to stage 5 were (25.84±9.71) µg/L, (20.76±6.99) µg/L, (20.40±17.02) µg/L, (19.49±11.29) µg/L, and (14.16±7.98) µg/L respectively. The prevalence of vitamin D deficiency was 39.66%, and from CKD stage 1 to stage 5 was 5.00%, 17.50%, 37.21%, 42.37% and 57.14%. The prevalence of vitamin D insufficiency was 44.97%, and from CKD stage 1 to stage 5 was 72.50%, 47.50%, 45.35%, 33.90% and 40.60%. The prevalence of decreased vitamin D level was 84.63%, and from CKD stage 1 to stage 5 was 77.50%, 65.00%, 82.56%, 76.27% and 97.74%. Single factor correlation analysis showed 25(OH)D3 was correlated with Hb, Alb, Scr, eGRF and iPTH. Regression analysis indicated that 25(OH)D3 was negatively correlated with iPTH and Scr, and positively correlated with Alb. According to K/DOQI, percentage of CKD patients from stage 3 to stage 5 who were consistent with vitamin D treatment was 87.20%, 83.05% and 26.31% based on 25(OH)3 and iPTH levels, but such percentage was 16.28%, 35.59% and 26.31% based on iPTH level only. Conclusions The prevalence of vitamin D insufficiency and deficiency in patients with CKD is quite high. Alb, iPTH and Scr are key factors influencing vitamin D level. Vitamin D level should be measured among CKD patients in order to carry out corresponding treatment.

Objective To investigate the cell types of renal allograft intimal arteritis in posttransplantation patients and its relationship with peritubular capillaries（PTCs）C4d deposition. Methods Twenty allograft kidney transplant recipients from Jun 2006 to Jun 2008 were enrolled in the retrospective study. Twenty-one biopsy specimens with acute vascular rejection were immunostained for macrophages, T cells and C4d. In each biopsy specimen, arterial intimal macrophages and T cells were counted, and mean number of macrophages per artery and T cells per artery were determined. The recipients were divided into C4d+ and C4d- group according to whether C4d deposition in the PTCs. Results In the intimal arteritis of biopsies diagnosed as acute vascular rejection, the infiltrating cells were predominantly macrophages including C4d+ and C4d- group. T cells were the minority. The mean macrophage number per artery cross-section was significantly higher than the mean T cell number per artery cross-section in the C4d+ group (12.45±9.86 vs 3.91±3.03, P=0.007) and in the C4d- group (3.47±1.89 vs 1.45±1.37, P=0.006). Forther more, the mean number of macrophage per artery cross-section in the C4d+ group was significantly higher than that in the C4d- group (P=0.007). Conclusions In the intimal arteritis of biopsies diagnosed as acute vascular rejection, the infiltrating cells are predominantly macrophages which has relationship with peritubular capillary C4d deposition. The mean number of macrophage per artery cross-section in the C4d+ biopsies is significantly higher than that in the C4d- biopses.

Objective To investigate the inhibitory effect and associated mechanism of rapamycin on proliferation and extracellular matrix（ECM） secretion in myofibroblasts stimulated by connective tissue growth factor（CTGF）. Methods Primary cultivated myofibroblasts were divided into 6 groups: control, CTGF(100 μg/L), rapamycin 20 μg/L+CTGF 100 μg/L, rapamycin 40 μg/L+CTGF 100 μg/L, rapamycin 20 μg/L, and rapamycin 40 μg/L alone. 5’-bromodeoxyuridine (BrdU) incorporation assay was used to detect the myofibroblast proliferation. Western blot was used to analysis the secretory FN protein in the supernatant medium of cultured myofibroblasts and the ERK1/2 phosphorylation in myofibroblasts. Results CTGF (100 μg/L) incubation significantly increased the number of Brdu positive myofibroblasts（P<0.01） and the level of FN protein secretory（P<0.05） in cell supernatant medium compared with control group, respectively. The number of Brdu positive myofibroblasts markedly decreased by 62% and 70% （P <0.05） in rapamycin 20 μg/L+CTGF 100 μg/L and rapamycin 40 μg/L+CTGF 100 μg/L groups, respectively. The FN protein levels in supernatant were decreased by 15% and 44% compared with CTGF 100 μg/L group, respectively; but the difference of FN protein levels was significant only in rapamycin 40 μg/L group（P<0.05）. CTGF could activate ERK1/2 at 10 minutes; but as myofibroblasts were pretreated with rapamycin 40 μg/L for 30 min, it abolished CTGF-induced ERK1/2 phosphoralation. PD98059, the specific inhibitor of ERK1/2, could block the effect of CTGF-induced proliferation (7%±5% vs 85%±7%, P<0.01) and FN secretion （1.0±0.1 vs 1.6±0.3, P<0.05）. Conclusions Rapamycin partially suppresses the proliferation and ECM secretion of myofibroblasts induced by CTGF. Its effect may be through inhibiting CTGF-induced activation of ERK1/2 signaling pathway.

Objective To determine the biological characteristics of human embryonic MSC (hMSCs) and their potential ability of differentiation, and to investigate the survival and distribution of hMSCs after transplantation into the kidney of newborn mice. Methods hMSCs were derived from 4-7 week-old embryos, then primary culture was done. The biological characteristics of hMSCs were detected by immunohistochemical methods and flow cytometry. Their differentiation potential was determined by coculture with conditioning medium. The survival and distribution of PKH-26-stained hMSCs in mice were observed by laser scanning confocal microscope. Results Flow cytometry and immunochemistry staining revealed that the expression of CD29, CD44, CD90, SH-2, OCT-4 was positive significantly, and CD34, CD45 was negative. The cells could be induced to differentiate to osteocytes and adipocytes under special conditions. After transplantation for 1 month, PKH-26-stained hMSCs still existed in the kidney of mice and co-localized in tubular epithelium by confocal microscope. Conclusion hMSCs derived from the early human embryo have the ability of proliferation and differentiation with low immunity, and may be involved in the development of renal tubule in newborn mice.

Objective To investigate the effect of rosiglitazone (RGTZ) on anti-oxidation in aging rat kidney. Methods Twenty-four-month-old male Sprague-Dawley rats were randomly divided into three groups (n=10): control group (CON), rosiglitazone group (RGTZ) and caloric restriction group (CR). The CON rats were allowed ad libitum access to feed and tap water. The RGTZ rats received intragastric administration of RGTZ (4 mg&#8226;kg－1&#8226;d－1), and the CR rats were provided with a vitamin and mineral fortified version of the same diet at a level of 40% less food (by weight) than the CON rats. After 12 weeks all the animals were sacrificed by decapitation, and both the body weight and the percentage of kidney and heart in each group were measured. Western blot was performed to analyze the expression of PPARγ protein. The content of MDA and the activity of SOD and GSH-PX in kidney tissue were detected. Besides, frozen sections of kidney tissue were stained for senescence-associated-β galactosidase （SA-β-Gal）. Results The body weight of CR rats decreased obviously, in contrast, which did not change in CON and RGTZ group. Percentage of kidney and heart to body weight was normal in CR or RGTZ group after intervention. Western blot result showed that PPARγ protein expression in rat kidney was significantly higher in RGTZ and CR group as compared to CON group（P＜0.05）. Compared with RGTZ and CR rats, obviously lower activities of SOD and GSH-Px were noted in CON rats, however, the content of MDA was higher in CON rats. Additionally, the positive staining area of β-Gal in CR and RGTZ group was significantly smaller than that in CON rats（P＜0.05, P＜0.01）. Conclusion RGTZ can defer the kidney aging in senescence SD rat, and the mechanism may be related to amelioration of oxidative damage and enhancement of antioxidation.

Objective To investigate the effects of high glucose and insulin on the expression of glucose transporter 4 (GLUT4), Cbl-associated protein (CAP) and cytoskeleton protein F-actin of glomerular mesangial cells (GMCs), in order to explore the function of GLUT4, Cbl-associated protein and F-actin in the pathogenesis and development of diabetic nephropathy (DN). Methods Cultured 1097 rat glomerular mesangial cells were divided into 8 groups: control, 10-9 mol/L insulin, 10－8 mol/L insulin, 10－6 mol/L insulin, high glucose (30 mmol/L), mannitol (25 mmol/L mannitol＋5 mmol/L glucose), high glucose plus 10－6 mol/L insulin, high glucose plus 10－9 mol/L insulin. Expression of CAP mRNA and GLUT4 was measured by RT-PCR and immunohistochemistry method. F-actin was stained by rhodamine-pholloidin and the fluorescent intensity was calculated by image analysis system. Results The expression of GLUT4 mRNA and protein, CAP mRNA was found in normal glomerular mesangial cells（control）, and there was no significant difference in 10-9 mol/L insulin group. The expression of GLUT4 mRNA (P<0.05) and protein (P<0.01), CAP mRNA (P<0.01) level was decreased in high glucose group compared with that of control group, but there was no significant difference in mannitol group. The expression of GLUT4 and CAP mRNA up-regulated with the increase of concentration of insulin. The expressions of GLUT4 mRNA in 10-8 mol/L insulin and 10－6 mol/L insulin groups were 2.06-fold and 2.66-fold of 10－9 mol/L insulin group, of GLUT4 protein were 1.93-fold and 2.83-fold of control, and of CAP mRNA were 1.91-fold and 2.15-fold of control, respectively. The expressions of GLUT4 mRNA, GLUT4 protein, CAP mRNA in high glucose plus insulin group were 2.15-fold, 2.08-fold, 2.14-fold of high glucose group respectively. High glucose decreased the fluorescent intensity of F-actin to 44.5% (P<0.01). 10-8 mol/L insulin and 10－6 mol/L insulin groups increased to 1.224-fold (P<0.05), 1.296-fold (P<0.01) in a concentration-dependent manner. The spearman correlation coefficient between GLUT4 and F-actin was 0.929 (P=0.001), between GLUT4 mRNA and CAP mRNA was 0.905 (P=0.002). Conclusions (1) A certain expression of GLUT4 mRNA and protein, CAP mRNA from GMC is found in normal glomerular mesangial cells. (2) High glucose can inhibit the expression of GLUT4 and CAP mRNA significantly, and facilitate the depolymerization of F-actin. (3) Insulin can reverse down-regulation of GLUT4 and CAP mRNA caused by high glucose. (4) GLUT4, CAP and F-actin are important factors in the development of DN.

Objective To investigate the expression of vascular endothelial growth factor(VEGF), angiopoietin and their receptors (VEGFR and Tie2) in aging mice kidney and the possible roles in aging mice. Methods Mice were divided as follows: 4-month old group (n=6), 9-month old group (n=6), 12-month old group (n=6) and 20-month old group (n=6). Paraffin sections of the mice kidneys were stained by PAS. The density of glomerular microvascular was determined by renal perfusion with fluorescent dyes. The level of VEGF, VEGFR2(Flk-1), Ang-1, Ang-2, Tie2 mRNA expression and protein abundance in kidney was determined by real-time PCR, immunochemistry, immunofluorescence and Western blot. Results Compared with other three groups, in the 20-month old group, the glomerulosclerosis index (GSI) increased remarkbly (2.48±0.79 vs 0.53±0.19, 0.69±0.18, 1.50±0.70, P<0.05); the fluorescence intensity in glomeruli decreased (P<0.05). Immunohistochemistry demonstrated that the TGF-β1 level in the aging kidneys showed an increase trend in the glomerular tubulointerstitium, and especially in the glomeruli. Real-time PCR results revealed that compared with 4-month old group mice, the mRNA expression of VEGF, Flk-1, Ang-1, Ang-2, Tie2 of the other three groups decreased，the gene levels of VEGF, Flk-1, and Ang-2 fell about 90%, 50% and 80% (all P>0.05), and the gene levels of Ang-1 and Tie2 fell about 75% and 40% in 20-month-old group(all P<0.05). Western blot domonstrated that the protein abundace of VEGF, Flk-1, Ang-1, Ang-2, Tie2 also declined with aging, the protein level of VEGF, Flk-1, Ang-1, Ang-2 and Tie2 dropped by about 35%, 50%, 15%, 13% and 21% respectively in 20-month-old group as compared to 4-month-old group (all P<0.05). Expression of above 5 factors and glomerular fluorescence intensity were negatively correlated with Scr (P<0.05). Conclusions The mRNA expression and protein abundance of VEGF, Flk-1, Ang-1, Ang-2, Tie2 in mice kidneys decreases with aging. Angiogenesis regulatory factors may play important roles in aging progression of the mice kidney.

Objective To investigate the effects of the serum from chronic renal failure(CRF) rats on endoplasmic reticulum stress(ERS) and activation of activating transcription factor 4(ATF4) of rat arterial smooth muscle cells(ASMCs) cultured in vitro and explore the possible mechanism. Methods The rat model of CRF was established by 5/6 nephrectomy. ASMCs were incubated in the media with 10% or 20% serum of CRF rats cultured in vitro. ATF4 nuclear translocation were analyzed by immunofluorescence. GRP78(glucose regulated protein 78), p-PERK (pancreatic ER kinase) and ATF4 proteins in response to the CRF serum in ASMCs were determined by Western blot. The ATF4-DNA binding activity was analyzed by electrophoretic mobility shift assay(EMSA). Results The CRF serum could significantly promote the expression of GRP78 of ASMCs in dose-dependent manner. Under the stimulation of 20% CRF serum, nuclear translocation of ATF4 in ASMCs was found. Compared with control serum group, the expression of ATF4 and p-PERK of CRF serum group was increased significantly (P<0.01). The CRF serum could increase the ATF4-DNA binding activity. Conclusion The CRF rat serum can effectively induce ERS of ASMCs and activate the pathway of PERK-ATF4.

Objective To investigate the effect and possible mechanism of bone marrow stem cell mobilized by stem cell factor(SCF) with granulocyte colony-stimulating factor（G-CSF）on renal peritubular capillary, fibrosis and renal function in unilateral ureteral obstruction (UUO) rats. Methods One hundred and twenty eight healthy male Wistar rats were randomly divided into four groups: Sham group, SCF-G group, UUO group and UUO+SCF-G group. Eight rats of each group were randomly selected and killed on the 5th, 14th, 21st and 28th day. Serum creatinine, CD34 positive cells and factor Ⅷ positive cells in renal interstitium, histopathologic lesion scores of interstitial fibrosis and interstitial pathology in kidney were measured. The mRNA expression of vascular endothelial growth factor (VEGF) and thrombospondin-1(TSP-1) in the renal cortex was detected. Results (1) The renal interstitial fibrosis and the loss of peritubular capillary were observed in UUO group after two weeks. (2) The number of bone marrow stem cells homing to renal interstitium in UUO+SCF-G group was significantly higher than that in UUO and Sham groups (P<0.05). (3) The loss of peritubular capillary in UUO+SCF-G group appeared later than that in UUO group (P<0.05). (4) The interstitial fibrosis and tubule injury was milder in UUO+SCF-G group than that in UUO group (P<0.05). (5) The decrease of VEGF mRNA expression of renal cortex in UUO+SCF-G group was seen later than that in UUO group. VEGF mRNA expression in UUO+SCF-G group was higher than that in UUO group. (6) The increase of TSP-1 mRNA expression of renal cortex in UUO+SCF-G group was seen later than that in UUO group. TSP-1 mRNA expression in UUO+SCF-G group was lower than that in UUO group (P<0.05). (7) In UUO and UUO+SCF-G groups, peritubular capillary index was negatively correlated with serum creatinine, interstitial fibrosis and interstitial lesion scores. VEGF mRNA expression of renal cortex was positively correlated with peritubular capillary index, and TSP-1 mRNA expression of renal cortex was positively correlated with peritubular capillary index. Conclusions （1）The loss of peritubular capillary is found in UUO group, and is correlated with interstitial fibrosis and interstitial lesion. (2) Application of SCF with G-CSF can effectively motivate stem cells to injured renal tissue, contribute to decrease the loss of peritubular capillary, lessen interstitial fibrosis and interstitial lesion, and ameliorate renal function. (3) Application of SCF with G-CSF can up-regulate VEGF mRNA expression and down-regulate TSP-1 mRNA expression, which may contribute to promote the repair of endothelial cells and protect peritubular capillary.