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Objective:To explore the predictive values of tissue inhibitor of metalloproteinase-2 (TIMP-2) and insulin-like growth factor binding protein 7 (IGFBP7) in donor sera and lavage fluid on delayed graft function (DGF) in donation after circulatory death (DCD) kidney transplant recipients.Methods:A total of 33 eligible kidney donors and 33 corresponding recipients were recruited. Preoperative serum and renal perfusion fluid samples of donors were collected to determine the levels of TIMP-2 and IGFBP7. Patients were grouped according to whether DGF occurred after kidney transplantation and measured indicators analyzed. Independent sample t test was utilized for comparing the groups with normal distribution measurement data. And χ2 test was employed for comparing the groups with normal distribution counting data and Mann-Whitney test for comparing the groups with non-normal distribution measurement data. Receiver operating characteristic (ROC) curve and area under curve (AUC) were used for evaluating the diagnostic efficacy of indicators. Results:In donor-DGF group, lavage fluid TIMP-2, product of lavage fluid TIMP-2 and IGFBP7 (TIMP-2×IGFBP7), serum IGFBP7 and product of serum TIMP-2 and IGFBP7 (TIMP-2×IGFBP7) were higher than those in donor-non-DGF group ( P<0.05). The AUC of TIMP-2, TIMP-2×IGFBP7, serum IGFBP7 and serum TIMP-2×IGFBP7 in the diagnosis of DGF were 0.753 (95%CI 0.546~0.959), 0.747 (95%CI 0.510~0.984), 0.824 (95%CI 0.615~1.000) and 0.852 (95%CI 0.660~1.000) respectively. Conclusions:Donor serum IGFBP7, donor serum TIMP-2×IGFBP7, lavage fluid TIMP-2 and lavage fluid TIMP-2×IGFBP7 may be used for predicting the occurrence of early DGF after kidney transplantation. Among them, serum TIMP-2×IGFBP7 has the highest diagnostic efficiency and may be an excellent predictor of DGF occurrence.
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Objective@#To investigate the effects and underlying mechanisms of aspirin on endoplasmic reticulum stress in podocytes induced by hyperlipemia.@*Methods@#Cultured podocytes were divided into four groups: control group, aspirin (100 μg/ml) group, oxidized low density lipoprotein (ox-LDL, 100 μg/ml) group, aspirin+ox-LDL group. The expression of protein kinase R-1ike endoplasmic reticulum kinase (PERK), eukaryotic translation initiation factor 2α (eIF2α), activating transcription factor-4 (ATF4) and CAAT/enhancer binding protein homologous protein (CHOP) at 6 h, 12 h, 24 h, 48 h were evaluated by real-time PCR. The related proteins of p-PERK and p-eIF2α at 24 h and ATF4 at 12 h were evaluated by Western blotting, respectively.@*Results@#The expressions of PERK, eIF2α peaked at 24 h, while ATF4 and CHOP peaked at 12 h in ox-LDL group and aspirin+ox-LDL group. Compared with control group, the expressions of PERK, eIF2α, ATF4 and CHOP were significantly higher in ox-LDL group at each times (all P<0.05). Compared with ox-LDL group, the expressions of the above indicators were significantly lower in aspirin+ox-LDL group at each times (all P<0.05). At 24 h, compared with control group, the expressions of p-PERK and p-eIF2α were significantly higher in ox-LDL group (both P<0.05). Compared with ox-LDL group, the expressions of p-PERK and p-eIF2α were significantly lower in aspirin+ox-LDL group (both P<0.05). At 12 h, the expression of ATF4 protein in each group was similar to that of mRNA. There were no significant difference in the expressions of all indicators between aspirin group and control group.@*Conclusions@#Hyperlipidemia may cause endoplasmic reticulum stress in podocytes by inducing phosphorylation of PERK and eIF2α, activating ATF4 transcription and inducing high expression of CHOP. Aspirin may partially block the PERK pathway, which may have protective effects for podocytes.
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Objective:To investigate the effects and underlying mechanisms of aspirin on endoplasmic reticulum stress in podocytes induced by hyperlipemia.Methods:Cultured podocytes were divided into four groups: control group, aspirin (100 μg/ml) group, oxidized low density lipoprotein (ox-LDL, 100 μg/ml) group, aspirin+ox-LDL group. The expression of protein kinase R-1ike endoplasmic reticulum kinase (PERK), eukaryotic translation initiation factor 2α (eIF2α), activating transcription factor-4 (ATF4) and CAAT/enhancer binding protein homologous protein (CHOP) at 6 h, 12 h, 24 h, 48 h were evaluated by real-time PCR. The related proteins of p-PERK and p-eIF2α at 24 h and ATF4 at 12 h were evaluated by Western blotting, respectively.Results:The expressions of PERK, eIF2α peaked at 24 h, while ATF4 and CHOP peaked at 12 h in ox-LDL group and aspirin+ox-LDL group. Compared with control group, the expressions of PERK, eIF2α, ATF4 and CHOP were significantly higher in ox-LDL group at each times (all P<0.05). Compared with ox-LDL group, the expressions of the above indicators were significantly lower in aspirin+ox-LDL group at each times (all P<0.05). At 24 h, compared with control group, the expressions of p-PERK and p-eIF2α were significantly higher in ox-LDL group (both P<0.05). Compared with ox-LDL group, the expressions of p-PERK and p-eIF2α were significantly lower in aspirin+ox-LDL group (both P<0.05). At 12 h, the expression of ATF4 protein in each group was similar to that of mRNA. There were no significant difference in the expressions of all indicators between aspirin group and control group. Conclusions:Hyperlipidemia may cause endoplasmic reticulum stress in podocytes by inducing phosphorylation of PERK and eIF2α, activating ATF4 transcription and inducing high expression of CHOP. Aspirin may partially block the PERK pathway, which may have protective effects for podocytes.
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Objective To investigate the effect of curcumin on autophagy and inflammatory response in mice subject to renal ischemia reperfusion.Method Forty male C57BL/6 mice were randomly divided into four groups (n =10 each):sham-operated (SO) group (Abdominal incision was made to expose the kidneys,bilateral renal pedicle dissociated and the abdomen sewed),ischemia reperfusion (IR) group,curcumin (CM) group (given CM 10 mg/kg) and 3-methyladenine (3-MA) group (given 3-MA 15 mg/kg).Six and 24 h after reperfusion,renal function was tested by determining the serum creatinine (Scr) and blood urea nitrogen (BUN) levels,and the morphological changes in the kidney tissue were observed.The expression of LC3,Beclin 1,Rab7 and LAMP2 in kidney tissue was detected by fluorescence quantitative RT-PCR and Western blotting.Fluorescence quantitative reverse transcription PCR and enzyme-linked immunosorbent assay were used to examine the expression of IL-6,IL-10,IL-17 and TNF-α in kidney tissue and serum.Result As compared with IR group and 3-MA group,the Scr and BUN levels in the CM group were significantly decreased (P＜0.01),and the renal morphological changes were improved significantly (P＜0.01).The mRNA and protein expression of LC3,Beclin 1,Rab7 and LAMP2 was significantly increased in kidney tissue,and the expression of IL-6,IL-17 and TNF-α was reduced,while IL-10 was increased (P＜ 0.01) in the CM group as compared with IR group and 3-MA group.Conclusion Curcumin possesses a protective effect against renal ischemia reperfusion injury in mice,which is probably mediated by promoting autophagy and subsequently inhibiting inflammatory response.
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Objective To establish a culture protocol for Th17 cells in vitro and evaluate the effect of curcumin on the differentiation and functions of Th17 cells and explore the related mechanisms. Methods Splenic CD4+CD25- T cells of C57BL/6 mice were isolated and purified with magnetic bead methods, and were co-cultured with plate-bound anti-CD3 and anti-CD28 antibodies and with transforming growth factor (TGF)-β, interleukin (IL)-6, IL-23, anti-interferon-γ antibody and anti-IL-2 antibody for Th17-polarization. The cultured Th17 cells were allocated into four groups: the control group, in which T cells were cultured on the basis of the above protocol; the low-concentration curcumin (CM-L, 5 μmol/L) and high-concentration (CM-H, 25 μmol/L) curcumin groups, and sirolimus (SRL, 100 ng/ml) group. The proportion of Th17 cells was detected with flow cytometry, and the mRNA expression of retinoic acid-related orphan receptor (ROR)γt, IL-17A, IL-21 was examined with real-time quantitative polymerase chain reaction. Finally, the protein expression of RORγt and the phosphorylation levels of signal transducer and activator of transcription (STAT) 3 were measured using western blotting analysis. Results The proportion of Th17 cells in the freshly isolated CD4+CD25- T cells was (3.1 ±0.4)%, whereas the proportion of the cells cultured with the protocol could get [(54.1±3.4)%, P0.05). Moreover, the mRNA levels of RORγt, IL-17A and IL-21, the protein expression of RORγt and the phosphorylation levels of STAT 3 in CM-L, CM-H and SRL groups were also lower than those in the control group (IL-17A mRNA: CM-L 0.81±0.05, CM-H 0.61±0.05, SRL 0.58±0.05, Control 1.01 ±0.11, t=4.81, 8.52, 8.89; IL-21 mRNA: CM-L 0.73±0.06, CM-H 0.49±0.03,SRL 0.59±0.03, Control 1.12±0.11, t=5.98, 9.22, 7.95, P<0.01). The mRNA level of IL-21 in the CM-H group was lower than that in the SRL group (P<0.05). Conclusion In vitro, curcumin can inhibit the differentiaton of splenic CD4+CD25- T cells into Th17 cells in the setting of Th 17-polarization and inhibit the expression of IL-17A and IL-21 mRNA, which is associated with the inhibitive effect of curcumin on RORγt expression and STAT 3 phosphorylation.
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Objective To study the pathologic pattern and clinical feature of type 2 diabetes mellitus complicated with chronic kidney diseases (CKD). Methods Clinicopathological features of 155 type 2 diabetic patients complicated with CKD were collected and analyzed retrospectively. The patients were divided into four groups: typical diabetic glomerulopathy (DG),atypical diabetes-related renal disease (ADRD), non-diabetic renal diseases (NDRD) and DG complicated with NDRD. Results Renal biopsies revealed DG accounted for 18.7% of the patients, ADRD accounted for 12.9%, NDRD accounted for 60.0%, and DG complicated with NDRD accounted for 8.4%. In DG group, duration of type 2 diabetes was longer;the level of fast blood glucose, systolic blood pressure, mean arterial pressure and prevalence of diabetic retinopathy (DR) were higher;proteinuria was heavier and evaluated glomerular filtration rate (eGFR) was lower. In ADRD group, body mass index and prevalence of obesity were higher;dyslipidemia was more severe. Gross hematuria and acute renal insufficiency could be only found in NDRD group.Without DR, duration of diabetes under 5 years, gross hematuria, acute renal insufficiency,evidences of autoimmune diseases and proteinuria≥3.5 g/24 h but eGFR ≥60 ml/min were specific valuable predictors for NDRD. Conclusions Renal injuries in type 2 diabetic patients are structural heterogeneous, in which NDRD is more common and is different from ADRD and DG.Renal biopsy should be considered when type 2 diabetic patients complicated with CKD present at least one characteristic as follows: duration of diabetes under 5 years, without DR, history of gross hematuria, acute decrease of renal function, evidences of autoimmune diseases and proteinuria ≥ 3.5 g/24 h but eGFR ≥ 60 ml/min.
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Objective To find out a more rational pathological classification criteria for renal injury in patients with type 2 diabetes mellitus. Methods The renal clinicopathological features of forty-nine type 2 diabetic patients with maeroalbuminuria were collected and were compared retrospectively. The patients without diabetic renal disease were excluded. According to the pathological features, the patients were divided into two groups: typical diabetic glomerulopathy (DG) and atypical diabetes-related renal disease (ADRD). Results The renal biopsy revealed DG accounted for 59.2% of the patients, while the remaining 40.8% presented atypical renal injury defined as ADRD. In DG group, volume fraction of mesangium per glomerulus, glomerular basement membrane width, atrophic tubules index, intersititium injury index and prevalence of hyalinization of renal arteriole were higher; podocyte density per glomerulus was lower; duration of type 2 diabetes was longer; the level of fast blood glucose, systolic blood pressure, mean arterial pressure, proteinuria and prevalence of diabetic retinopathy (DR) were higher; glomerular filtration rate (GFR) was lower. In ADRD group, body mass index and prevalence of obesity were higher; dyslipidemia was more severe. GFR was negatively correlated with glomerular global sclerosis rate in both DG and ADRD group. Proteinuria was positively correlated with volume fraction of mesangium per glomerulus in DG. No correlation between proteinuria and pathological features was found in ADRD. DR (94.8%) and duration of type 2 diabetes over five years (90.7%) had high negative predictive value for DG. Conclusions Renal injuries in type 2 diabetes patients are heterogeneous. ADRD is an atypical renal injury in type 2 diabetes patients whieh is different from DG. DR and duration of diabetes are more helpful in predicting DG separating from ADRD.