BACKGROUND: Renal tubulointerstitial fibrosis is mainly featured as the accumulation of extracellular matrix (ECM) in renal interstitium. The tubular epithelial-myofibroblast transdifferentiation (TEMT) is important to the pathogenesy of renal tubulointerstitial fibrosis. OBJECTIVE: To examine the effects of sodium ferulate (SF) on TEMT, and ECM main components such as collagen Ⅰ, collagen Ⅲ and fibronectin, in rat renal tubular epithelial cellsinduced by transforming growth factor-beta 1 (TGF- β1)- DESIGN: Randomized and controlled experimental study based on cells. SETTING: Department of Kidney in West China Hospital of Sichuan University. MATERIALS: Rat renal tubular epithelial cells (NRK-52E) originated from American Type Culture Collection (ATCC), were offered by the laboratory of Department of Nephrology in Australian Monash Medical Center. Cell strain used in this study was cultured at the 36th passage. SF white crystal with water solubility and more than 98.0% purify, was from Chengdu Hengda Pharmaceutical Co., Ltd. Different concentrations of SF (125,250, 500μreel/L) were designed in this study. Rabbit anti-rat α-smooth muscle actin (α -SMA) was produced by Wuhan Boster Company. Enzyme-linked immunosorbent assay (ELISA) kit was the produced of Shanghai Senxiong Science and Technology Co.,Ltd. Human recombinant TGF- β1 was produced by R&D Company. DNA Engine OpticonTM real-time fluorescence quantitative polymerase chain reaction apparatus was the product of MJ Research Company. METHODS: Rat renal tubular epithelial cells (NRK-52E) cultured in vitro were divided into five groups. Control group was added with serum-contained DMEM; TGF-β1-induced group was added with TGF-β1 at final concentration of 5 ng/L; SF at different concentrations groups were added with 125, 250, 500 μ mol/L SF and TGF- β1 at final concentration of 5 ng/L,respectively. MAIN OUTCOME MEASURES: The contrast phase microscope, real-time fluorescence quantitative polymerase chain reaction and ELISA method were used to detect TEMT of NRK52E cells induced by TGF-β1 and levels of collagen Ⅰ, collagen Ⅲ and fibronectin in the supernatant. RESULTS: Morphology of NRK52E cells: Compared with control group, TGF-β1 could induce the transdifferentiation of NRK52E cells, showing fibroblast-like in morphology after 3 days, which were previously the typical road stone-like epithelial cells. In three different concentration SF groups, the morphologic transformation stimulated by TGF-β1 could be partly ameliorated in a dose-dependent manner. Expression of α-SMA mRNA: Compared with control group, 5 ng/L TGF- β1 enhanced expression of α-SMA at 6 hours, and reached a peak at 72 hours; SF depressed the expression in a dose-dependent manner at 72 hours (P < 0.05). Changes of ECM: After induced by 5 ng/L TGF- β1 for 72 hours, the levels of collagen Ⅰ, collagen Ⅲ and fibronectin in the supernatant increased significantly (P < 0.05), whereas SF decreased these levels in a dose-dependent manner (P < 0.05). CONCLUSION: TGF- β1 induces the TEMT, and promotes the secretion of collagen Ⅰ, collagen Ⅲ and fibronectin. SF can inhibit TGF- β1-induced TEMT In a dose-dependent manner.

BACKGROUND: Connective tissue growth factor (CTGF) is a kind of factor that can mediate downstream action of transforming growth factor-β 1 (TGF- β 1). The upregulation of connective tissue growth factor expression plays an important role in pathological changes of renal interstitial fibrosis.OBJECTIVE: To explore the effect of Sodium Ferulate on the expression of CTGF mRNA and protein in rats with unilateral ureteral obstruction (UUO) and pathological changes of renal interstitial fibrosis, and to compare with Losartan.DESIGN: Randomized and controlled animal trial.SETTING: Department of Nephrology, West China Hospital of Sichuan University, and College of Public Health, Sichuan University.MATERIALS: Twenty-four healthy adult male SD rats were selected from the Experimental Animal Center of Sichuan University. Sodium Ferulate was provided by Sichuan Hengda Pharmacy Co, Ltd (No. 050302); rabbit anti-rat CTGF by Santa Cruz; Western blotting by BioRAD, USA; DNA Engine OpticonTM real-time fluorescent quantitation PCR device by MJ Research, USA.METHODS: The experiment was performed at Research Laboratory of Clinical Medicine (grade BSL-1), College of Public Health, Sichuan University from May to December 2006. Twenty-four healthy rats were randomly divided into 4 groups (n=6): UUO model group, Sodium Ferulate group, Losartan group, and sham-operation group. According to the previous protocol, UUO models were established in UUO model group, Sodium Ferulate group, and Losartan group, and the other rats were subjected to sham operation. From the first day after UUO, Sodium Ferulate group was intragastrically (i.g.)administrated with 150 mg/kg/d Sodium Ferulate; Losartan group was administrated ig. with 20 mg/kg/d. Losartan; UUO and sham operation groups were administrated i.g. with matching normal saline. All rats were executed 14 days after UUO to harvest partial renal tissues. All experimental procedure was accorded with animal ethical standards.MAIN OUTCOME MEASURES: The mRNA and protein expressions of CTGF were quantified by real-time PCR and Western blot. The pathological changes of renal interstitial tissues were observed by hematoxylin/eosin (HE) and Masson staining.RESULTS: Twenty-four rats were included in final analysis. Fourteen days after UUO, CTGF mRNA and protein expressions in UUO model group were significantly increased compared with sham operation group, but the expressions in Sodium Ferulate group were significantly lower than model group (P < 0.05). Compared with Losartan treated group, there was no significant difference (P > 0.05). HE and Masson staining showed inflammatory cell infiltration and tubular and interstitial changes as well as collagen deposition in renal interstitial tissues on day 14 after UUO. Sodium Ferulate obviously improved the renal pathological changes in UUO rats (P < 0.05), and the effect was similar to Losartan (P > 0.05).CONCLUSION: Sodium Ferulate inhibits UUO-induced renal interstitial fibrosis. This action, similar to the effect of Losartan, might be due to downregulation of CTGF expression.

Objective To compare health related quality of life (HRQOL) between modified and traditional cutaneous ureterostomy, and explore the reasons for these differences, in order to provide the basis of HRQOL for the choice of cutaneous ureterostomy. Methods A total of 53 patients underwent cutaneous ureterostomy were selected, and the patients were divided into traditional cutaneous ureterostomy group (traditional group, 21 cases) and modified cutaneous ureterostomy group (modified group, 32 cases) according to the surgery method. The patients were evaluated by functional assessment of cancer therapy-bladder (FACT-BL), and the HRQOL was compared between 2 groups. Results There were no statistical differences in HRQOL score at 1, 3, 6 and 9 months after surgery between 2 groups (P>0.05). The HRQOL score at 12 months after surgery was significantly higher in modified group than that in traditional group:(141.5 ± 10.4) scores vs. (123.1 ± 5.2) scores, and there was statistical difference (P<0.01). There were no statistical differences in the scores of physiology status, society/family status, emotional state and functional assessment of cancer therapy-general (FACT-G) at 12 months after surgery between 2 group (P>0.05). But the scores of functional status and bladder cancer special scale (BSS), total score of FACT-BL in modified group were significantly higher than those in traditional group:(26.0 ± 2.5) scores vs. (23.8 ± 3.5) scores, (46.7 ± 6.2) scores vs. (34.8 ± 5.5) scores, (143.9 ± 15.7) scores vs. (117.5 ± 8.1) scores, and there were statistical differences (P<0.01). Conclusions The HRQOL at 12 months after surgery in modified cutaneous ureterostomy is better than that in traditional cutaneous ureterostomy. Therefore, if the patient's physical condition permits, priority should be given to modified cutaneous ureterostomy to reduce the complications and improve the quality of life.

ObjectiveTo study the amplification and expression of HER-2 gene and protein in breast cancer and to investigate the relationship between HER-2 gene and Ki-67 、P53 、ER、PR、lymph node metastasisandTNMstaging.MethodsFluorescenceinsituhybridization(FISH) and immunohistochemistry(IHC) method were used to detect the amplification and expression of HER-2 and the expression of Ki-67、P53、ER、PR in 150 cases of breast cancer. Results49 out of 150 cases were amplificated,positive rate of HER-2 gene was 32.68％.In the 32 patients whose expression of HER-2 protein was ( + + + ),HER-2 gene was expressed in 28 cases.In The 46 patients whose expression of HER-2 protein was ( + + ),HER-2 was expressed in 16 cases.In the 26 patients whose expression of HER-2 protein was ( + ),HER-2 was expressed in 3 cases.In the 47 cases that the expression of HER-2 protein was negative,there were 45 cases that the expression HER-2 gene was negative.The expression of HER-2 was correlated with ER,PR expression and TNM staging,but not significantly related with age,Ki-67,P53 and lymph node metastasis.ConclusionsFISH and IHC correlate well with each other in the determination of HER-2 gene amplication and protein expression in cases of invasive breast cancer.

Objective To investigate the significance of tumor markers CEA and CA19-9 in predicting the clinicopathologic characteristics and prognosis of primary duodenal carcinoma.Methods A retrospective analysis of 110 cases with primary duodenal carcinoma treated in our hospital from January 1999 to December 2016 was conducted.ROC analysis,univariate and multivariate analysiswere performed to investigate the relationship between CEA,CA19-9 and the clinicopathologic characteristics of primary duodenal carcinoma.Kaplain-Meier method was used to analyze the relationship between CEA and CA19-9 and the prognosis of primary duodenal carcinoma.Results CEA level was of value for predicting the depth of infiltration,lymphatic involvement,metastasis and TNM stage.The receiver operating characteristic was 0.629,0.672,0.749,0.692 respectively.Univariate analysis showed serum CA19-9 lever was related to the depth of infiltration and serum CEA lever were related to tumor differentiation,lymphatic invasion,metastasis and TNM stage.Logistic analysis showed that CEA value was only associated with metastasis (OR:9.853,P ＜ 0.01).Patients with elevated serum CEA level had a significant worse prognosis than patients with normal serum CEA level (P ＜ 0.05).Conclusion Serum CEA level was closely associated with the clinicopathologic characteristics and prognosis of primary duodenal carcinoma.

Objective: Taking human A549 cells as the research object, to construct the paraquat-induced pulmonary fibrosis model in vitro, and to explore the role of TSP-1 (Thrombospondin-1, TSP-1) and its receptor CD47 in PQ-induced pulmonary fibrosis. Methods: Human A549 cells were cultured in vitro, divided into normal control group, PQ group, Anti-TSP1 group (PQ with neutralizing anti-TSP1 antibody at a final concentration of 10 μg/ml) . A549 cells were stimulated with different concentrations (50, 100, 200, 400, 600, 800, 1 000 μmol/L) for different time (12, 24, 48 h) , and then CCK8 method was used to detect the cell viability to screen out the concentration and time of half cell viability. The subsequent test will be performed at this concentration point.The morphology of the cells was observed under inverted microscope. The expression levels of Fibronectin (FN) and type I collagen were determined by Enzyme Linked Immunosorbent Assay (ELISA) . Immunocytochemistry (ICC) and Immunofluorescence (IF) were used to observe the expression of TSP-1 and CD47 protein and the co-expression.The mRNA expression of TSP-1 and CD47 was detected by Real Time PCR (RT-PCR) . The protien expression of TSP-1 and CD47 was detected by Western Blot (WB) . The levels of Reactive Oxygen Species (ROS) were measured by flow cytometry. Results: Before neutralizing anti-TSP1 antibody intervention: (1) When the time of PQ was constant, the cell viability decreased with the increase of PQ concentration. (2) The cells in the control group were closely connected, cobble-like, arranged neatly; with the increase of PQ concentration, the cell gap of PQ group gradually increased, spindle shape or long spindle shape. (3) With the increase of PQ concentration, the relative expression of FN and I collagen in PQ group was gradually increased compared with the control group in a concentration-dependent manner, and 200 μmol/L is the most obvious. (4) Compared with the control group, the mRNA level and the protein expression of TSP-1 and CD47 in PQ group was significantly increased, and 200 μmol/L is the most obvious, and Immunofluorescence showed they were co-expression in cytoplasm. (5) Compared with the normal group, the level of ROS in A549 cells was significantly increased at 24 h after PQ stimulation. (6) Compared with PQ group, the cell viability of Anti-TSP1 group was significantly increased, and the morphology changed to normal cell morphology, and the mRNA level and the protein expression of TSP-1 and CD47 decreased, and the overexpression of ROS was inhibited, and the relative expression of FN and I collagen decreased. Conclusion PQ stimulation induced morphological changes of A549 cells, increased expression of TSP-1, CD47, FN and type I collagen, and increased production of ROS.Neutralizing anti-TSP1 antibodies against TSP-1 can partially improve the above lesions. TSP-1-CD47 may be associated with oxidative stress-mediated PQ-induced pulmonary fibrosis.

This study sought to assess the effect of Ginsenoside Rg1 on streptozocin-induced diabetic nephropathy in rats and to unveil the underlying mechanism. Diabetic nephropathy (DN) was induced by intraperitoneal injection of streptozocin (STZ). Eight weeks after drug administration, the rats from each group were sacrificed. Serum creatine (Scr) and 24 hours urine protein, cross reaction protein (CRP) and tumor necrosis factor-alpha (TNF-alpha) were measured at the end of the study. The histological changes of renal interstitial tissues were observed by periodic acid-Schiff staining (PAS). Immunohistochemical method was used to examine the expression levels of ectodermal dysplasia (ED-1). The mRNA of transforming growth factor-beta1 (TGF-beta1) was measured by real-time PCR (RT-PCR), and the protein expression of TGF-beta1 was surveyed by Enzyme-Linked Immunosorbent Assay (ELISA). The renal pathological changes in DN rats given ginsenoside Rg1 treatment were ameliorated, and the expression levels of 24 h urine protein, serum creatinine, CRP, TNF-alpha, ED-1 and TGF-beta1 were significantly lower than those in the diabetic nephropathy group (P < 0.05). So, we reach a conclusion that, in the experiment, Ginsenoside Rg1 obviously reduced TGF-beta1 expression and the already-mentioned inflammatory reaction factors in the renal tissues and improved the renal pathological changes in DN rats.
Animals ; Diabetes Mellitus, Experimental ; complications ; Diabetic Nephropathies ; drug therapy ; Drugs, Chinese Herbal ; therapeutic use ; Ginsenosides ; therapeutic use ; Male ; Phytotherapy ; RNA, Messenger ; genetics ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Transforming Growth Factor beta1 ; genetics ; metabolism

OBJECTIVETo study the effect of Astragalus mongholicus (AM) on the expression of transforming growth factor-beta1 (TGF-beta1) in SD rats with unilateral ureteral occlusion (UUO) and to elucidate the mechanisms underlying the renoprotective effects of AM.

METHODFifty-four Sprague-Dawley rats were randomly divided into 4 groups: sham-operation group, the UUO group and AM treatment group. After administration of AM (10 g kg(-1) d(-1)) for 3, 7 and 14 days, the dynamic histological changes of renal interstitial tissues were observed and renal damage including tubular impairment and interstitial fibrosis were quantified on HE and Masson stained tissue sections. The expression of TGF-beta1 and alpha-smooth muscle actin (alpha-SMA) was measured by immunohistochemistry staining sections. The mRNA of TGF-beta1 and alpha-SMA were reverse transcribed and quantified by real-time PCR. The expression of TGF-beta1 protein were assessed by Western blot.

RESULTRenal damage was exacerbated and the expression of alpha-SMA and TGF-beta1 were all significantly increased in UUO group compared with those of sham-operation group (P<0.05) at each time point. Tubular impairment and interstitial fibrosis were alleviated, and up-regulations of expressions of TGF-beta1 and alpha-SMA were significantly suppressed by AM treatment (P<0.05).

CONCLUSIONAM can ameliorate renal interstitial fibrosis induced by UUO in vivo. The mechanisms of its antifibrotic effects might be related with the down-regulation of TGF-beta1 expression and suppression of tubular epithelial myofibroblast transdifferentiation in the progress of renal interstitial fibrosis.

Actins ; genetics ; Animals ; Astragalus Plant ; chemistry ; Drugs, Chinese Herbal ; pharmacology ; Gene Expression Regulation ; drug effects ; Kidney Tubules ; drug effects ; metabolism ; Male ; RNA, Messenger ; genetics ; metabolism ; Rats ; Transforming Growth Factor beta1 ; genetics ; Ureteral Obstruction ; metabolism ; pathology ; prevention & control

Objective: To establish a rat model of paraquat (PQ) -induced pulmonary fibrosis and observe the changes in thrombospondin-1 (TSP-1) and its receptor CD47 in lung tissue, and to investigate their roles in the pathogenesis of PQ-induced pulmonary fibrosis. Methods: Fifty-four clean adult male Sprague-Dawley rats were randomly divided into normal control group (n=6) and 2 h, 12 h, 1 d, 3 d, 7 d, and 14 d PQ poisoning groups (n=8 per group). A rat model of PQ poisoning was established by a single gavage of 20 wt.% PQ solution (50 mg/kg). Flow cytometry was used to determine the concentration of reactive oxygen species (ROS) in blood and lung tissue. Enzyme-linked immunosorbent assay was used to determine the concentrations of hydroxyl radicals, malondialdehyde, and hydroxyproline in lung tissue. HE staining and Masson staining were used to observe the pathological damage of lung tissue after PQ poisoning. The expression of TSP-1 and CD47 in lung tissue was measured by Immunohistochemistry. Results: Compared with the normal control group, the 2 h to 7 d PQ poisoning groups showed significant increases in ROS fluorescence intensity in red blood cells and lung tissues and the concentrations of malondialdehyde and hydroxyl radicals in lung tissue (P<0.05) , and the 14 d PQ poisoning group had a significant increase in the concentration of hydroxyproline in lung tissue (P<0.05). HE staining showed that the 2 h to 7 d PQ poisoning groups had significantly higher semiquantitative pathological scores of pulmonary alveolitis than the normal control group (P<0.05). The Masson staining showed that the 7 d and 14 d PQ poisoning groups had significantly higher semiquantitative pathological scores of pulmonary fibrosis than the normal control group (P<0.05). Compared with the normal control group, all PQ poisoning groups (except the 12 h group) had significantly increased expression of TSP-1 in lung tissue (P<0.05) , and all PQ poisoning groups (except the 1 d group) had significantly increased expression of CD47 in lung tissue (P<0.05). Within 2 h after PQ poisoning, the expression of TSP-1 and CD47 was positively correlated with the concentrations of ROS, hydroxyl radicals, and malondialdehyde and the degree of pulmonary alveolitis (P<0.01) ; at 1 d after PQ poisoning, the expression of TSP-1 and CD47 was positively correlated with the concentration of hydroxyproline in lung tissue (P<0.01) . Conclusion The expression of TSP-1 and CD47 is closely related to oxidative stress and subsequent pulmonary fibrosis, and they may be involved in the development and progression of pulmonary alveolitis and subsequent pulmonary fibrosis in rats with PQ poisoning.

Objective To investigate the clinical efficacy and prognostic factors for combined caudate lobectomy radical resection plus broad lymph node dissection in patients of hilar cholangiocarcinoma.Methods The clinical data and follow-up results of patients with hilar cholangiocarcinoma surgically treated from Feb 2008 to Feb 2017 were retrospectively analyzed.Result The R0 resection rate[72.2％ (13/18) vs 43.9％ (18/41)],operation time [(433 ± 136) min vs (302 ± 122) min],intraoperative blood loss [(1 789 ± 1 091) ml vs (776 ± 755) ml] and postoperative complication rate [66.7％ (12/18) vs 36.6％ (15/41)]were significantly higher in the hepatic lobe combined with caudate lobe resection group than that without caudate lobe resection group (P ＜ 0.05).The median survival time of patients with enlarged lymph node dissection was longer than that of patients with regional lymph node dissection (33 months vs 13 months,P ＜0.05).Univariate and multivariate analysis showed that the preoperative serum CA199 level ＞ 1 000 U/ml,the degree of microscopic margin and tumor TNM stage were significantly correlated with the prognosis of the patients (P ＜ 0.05).Conclusion Combined with caudate lobe resection can improve R0 resection rate.Targeted lymph node dissection helps prolong survival.The degree of microscopic margin,preoperative CA199 and TNM staging are independent risk factors for the prognosis of patients with hilar cholangiocarcinoma.