INTRODUCTION

Renal fibrosis is the major pathological basis of chronic renal failure(CRF),and also a clinical feature of the development and progression of chronic kidney disease(CKD),which is closely related to the degree of renal inflammation and renal function damage.1Renal fibrosis is mainly manifested by glomerular sclerosis,tubular atrophy and disappearance,and excessive accumulation of extracellular matrix,which is caused by continuous synthesis and degradation reduction.In our previous study,we found that the levels of serum leptin and various inflammatory factors were significantly increased in CRF patients,suggesting that leptin and inflammatory factors may be closely related to the process of renal fibrosis.2JAK/STAT pathway is a major signal transduction pathway mediated by leptin.3JAK/STAT signaling is activated in the rat model of unilateral ureteral obstruction(UUO),which accelerates the progression of renal fibrosis.4Furthermore,the nuclear transcription factor kappa B(NF-kB)signaling pathway,which is the downstream regulated by the JAK/STAT pathway,is the main signal transduction of the inflammatory reaction directly involved in the pathological process of renal fibrosis.5Therefore,it is reasonable to infer that leptin can activate JAK/STAT signaling pathway,leading to the activation of NF-kB signaling pathway which induces the production of inflammatory factors,and ultimately resulting in renal tubular epithelial-to-mesenchymal transition and renal interstitial fibrosis.Inhibition of leptin-mediated JAK/STAT activity is therefore a potential strategy to treat renal fibrosis.Previous research has confirmed that Qingshen granules(QSG),which has the effects of clearing heat and dampness and promoting blood circulation,can alleviate the inflammatory response in the kidney by reducing the levels of serum leptin,anti-renal fibrosis,and improving the kidney function.However,the mechanisms for the signal transduction pathway of leptin and the regulation of the inflammatory response in renal fibrosis are not yet clear.In this study,a rat model of UUO was established to explore the role of leptin-mediated JAK/STAT signaling pathway in NF-kB pathway activation and inflammatory effects,and the role in renal tubule transdifferentiation and renal interstitial fibrosis.Additionally,we discuss the mechanism of QSG in regulating the leptin signaling pathway.

MATERIALS AND METHODS

Experimental animals

概而言之，传统知识服务商力图将学术圈形成循环，在科研人员做科研、写论文/专著、发论文/专著、出版论文/专著、论文影响力评价、获取论文等全流程进行介入和服务，对即将进入学术圈的高校学生产品使用习惯等进行培养，获取用户忠诚度。

该厂家使用的牧羊公司全套膨化生产线，其配套调质器为新型组合调质器，调质时间为240～360 s，测试产品为乌龟料。在调质膨化工段前后取5组样，并测试样品的糊化度进行比较。这5组样品分别为：调质前（样品1），调质20 s左右（样品2），调质170 s左右（样品3），调质278 s（样品4），膨化颗粒成品（样品5）。使用三层组合调质器，可以使物料的淀粉糊化度达到85%以上，成品的淀粉糊化度达到96%以上。新型五轴组合调质器对调质时间的延长，对于膨化沉性料的品质提高起到了决定性作用。

Drugs and reagents

QSG,10 g/bag,was provided by the First Affiliated Hospital of Anhui Medical Center(Hefei,China,batch No.20141023).Valsartan capsules,80 mg/capsule,were obtained from Beijing Novartis pharmaceutical factory(Beijing,China,batch No.H20040217).Serum creatinine(batch No.20151102,Nanjing Institute of biological Engineering,Nanjing,China),Leptin(batch No.20121215A,Shanghai Biotechnology Co.,Ltd.,Shanghai,China),P-JAK2(batch No.CA36131,Bioworld,Nanjing,China),p-STAT3(batch No.561201,Bioworld,Nanjing,China),MCP-1(batch No.131120w,Bioworld,Nanjing,China),NF-kBp65(batch No.GR194772-1,Abcam,Cambridge,England),FN(batch No.AE011910,Bioss,Beijing,China),and Col-Ⅳ(batch No.AD091929,Bioss,Beijing,China)were used in this study.

Instruments

SPSS Version 17.0(SPSS Inc.,Chicago,IL,USA)was used for data analyses.Continuous variables were expressed as mean ±standard deviation(xˉ±s).All the samples were tested to ascertain if they followed a normal distribution.Data comparison among the groups was performed using analysis of variance and homogeneity of variance tests.Comparison between groups was carried out using the independent samplest-test.P＜0.05 was considered to be statistically significant.

UUO modeling and drug administration

The total kidney RNA was extracted,and was dissolved by adding 30 μL RNase-free ddH2O.After that,500 ng of total RNA was reverse transcribed in 10 μL of reaction system[5× prime script buffer 2 μL,Prime Script RT Enzyme MixⅠ 0.5 μL,Oligo dT Primer 0.5 μL,random 6-mers 0.5 μL,and diethypyrocarbonate(DEPC)water 6.5 μL].cDNA was then reverse transcribed(37℃15 min,70℃60 min).PCR instrument amplification:the reaction system 20 μL(cDNA 2 μL,2× Premix ExTaqII 10 μL,10 μmol/L PCR Forward/Reverse Primer 0.8 μL,50× Roxanne Brown Reference DyeII 0.4 μL,dH2O 6 μL).PCR amplification program:95℃denaturation for 30 s after the start cycle;95℃5 s,60℃34 s,40 cycles.Ct values were recorded using the relative quantitative analysis,mRNA relative expression=2ΔCt× 100%,whereΔCt=Ct(target gene)－ Ct(internal reference β-actin).Primer sequences are shown in Table 1.

Detection of creatinine and leptin in serum using enzyme-linked immunosorbent assay(ELISA)

3 Ibars M,Ardid-Ruiz A,Suárez M,Muguerza B,Bladé C,Aragonès G.Proanthocyanidins potentiate hypothalamic leptin/STAT3 signalling and Pomc gene expression in rats with diet-induced obesity.Int J Obes(Lond)2016;18(10):1-8.

Detection of leptin receptor(OB-R),p-JAK2,p- STAT3,nuclear factors-κBp6(NF-kBp65),and monocytechemotatic protein-1(MCP-1)proteins by Western blotting

The total protein in the kidney tissues was extracted with RIPA buffer.The proteins were analyzed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis(SDS-PAGE).The proteins were then transferred to nitrocellulose membranes,and blocked with 5%skim milk powder for 2 h.The blots were incubated with 1∶500 diluted OB-R,p-JAK2,p-STAT3,NF-kBp65 and MCP-1 monoclonal antibodies for 2 h.After that,the membranes were washed in PBS three times,for 10 min each time,and then incubated with 1∶1000 sheep-horseradish peroxidase anti mouse IgG for 1 h.Finally,the blots were washed in PBS three times,for 10 min each time,and electrochemiluminescence(ECL)reagent was used for developing the stained bands,and the blots scanned.The stained bands were analyzed with BANDSCAN software to calculate the densitometry grey value.

Detection of JAK2,STAT3,E-cadherin,α-SMA mRNA in kidney tissues by real-time quantitative polymerase chain reaction(PCR)

After acclimatization for 1 week in cages,the rats were randomly divided into the sham-operated group(normal group,n=10)and the UUO group(renal fibrosis group,n=50).Then,the renal fibrosis rats were randomly divided into five groups(the model group,n=10;QSG high-dose group,QSG medium-dose group,QSG low-dose group,n=10 respectively;and the valsartan group,n=10).The renal fibrosis rats were anaesthetized with 10%hydrate of chlorine aldehyde(0.3 mL/100 g,i.p.),and then the left ureter was exposed and ligated with 3-0 silk sutures.The ureter was mobilized but not ligated in the sham-operated rats.Administration of QSG began on the 7th d after the surgery.According to the literature,6the normal group and model group were given 0.9%physiological saline(1 mL/100 g per day).The QSG groups were given 4.0,8.0 and 16.0 g·kg－1·d－1in the low-dose,medium-dose and high-dose groups respectively,and the dosage of valsartan group was 20 mg·kg－1·d－1.The entire intervention course was 4 weeks.

Immunohistochemistry of typeⅣcollagen(Col-Ⅳ)and fibronectin(FN)in kidney tissues

EnVision IHC technique was used to examine the expressions of Col-Ⅳand FN in kidney tissues.Paraffin-embedded tissues were dewaxed,subjected to antigen retrieval,blocking of endogenous peroxidase,and then stained with anti-FN and anti-COL-Ⅳantibodies(1∶100 each).PBS was used in place of antibodies in the negative control samples.The tissues were stained with DAB reagent and counterstained with hematoxylin.Finally,the tissues were dehydrated by 1%HCL differentiation for 5 s,tap water washing,gradient alcohol dehydration,xylene transparent,and then dried with neutral resin under dry observation.Image-Pro plus analysis software was used for the results analysis,with the average light density of positive expression(indicated by yellow color)detected.

Table 1 Primer sequences of detection index

Notes:JAK:janus-activated kinase;STAT:signal transducers and activator of transcriptions;α-SMA:alphasmooth muscle actin;E-cadherin:calcium-dependent adhesion.

Tm(℃)77 79 82 79 75 84 Index Leptin JAK2 STAT3 α-SMA E-cadherin β-actin Upstream primer 5'-ACACCCTTAGAGGGGGCTAA-3'5'-AGGGAACCTCCACATCTCCT-3'5'-TGATGCGCTCTTATGTGAGG-3'5'-TTCAATGTCCCTGCCATGTA-3'5'-GGAGCATGTGAAGAACAGCA-3'5'-CCCATCTATGAGGGTTACGC-3'Downstream primer 5'-GGGCTCACTGTCTCCAATGT-3'5'-TTCAGCTTGCCCAAGAGAAT-3'5'-GGCGGACAGAACATAGGTGT-3'5'-CATCTCCAGAGTCCAGCACA-3'5'-CAGGAGAAGAGTCCCTGTGC-3'5'-TTTAATGTCACGCACGATTTC-3'Length(bp)91 119 104 94 91 150

Histological examination of kidney tissues by hematoxylin and eosin stain(HE)and Periodic Acid-Schiff stain(PAS)

The standard protocols were used for HE staining.For PAS staining,the samples were dewaxed,rinsed,and incubated with 1%periodate oxidation for 10 min after rinsing;Schiff reagent was added and the samples incubated in the dark for 20 min,followed by 5-min standard hematoxylin staining,and 1-min rinsing.Then,samples were dehydrated,mounted,and imaged under a microscope.

不求朋友成群，但求知己一人；不求财富无数，但求够花够用；不求万人怜惜，但求一人懂得；不求房子多大，但求一家温暖；不求车子豪华，但求一生平安；不求衣着华丽，但求穿着合体；不求才高八斗，但求思想丰富；不求地位显赫，但求工作顺心。

Statistical analysis

Semi-automatic biochemical analyzer(AT-648)was purchased from Shanghai Bomaijie Biotechnology Co.,Ltd.,(Shanghai,China).The electrophoresis apparatus(EPS-301)wasfrom Amersham (Piscataway,NJ,USA).Gel image analyzer was from Bio-Rad Corporation(Gel Doc XR,Hercules,CA,USA).PCR instrument was provided by Biometra Company Biometra GmbH(T-Gradient,Goettingen,Germany).Pathological section machine(LEICA-213)and paraffin-embedding station(LEICA-115)were from Leica Inc.(Frankfurt,Germany).

目前出版的SPSS教材不下数十种,多数教材在给出案例后,直接跳入菜单或命令的分析,穿插介绍每个选项的中文含义,最后解释软件的输出结果,仅花很少的篇幅介绍统计理论[3]。这种处理方式有其可取之处，如方便学生理清思路、方便上机模仿练习等，但轻视统计理论教学的缺点也是显而易见的：学生对知识的掌握一知半解，经常出现明显的模型误用。例如，在因变量不服从正态分布的情况下，却采用多元回归方法进行建模；明显的单边检验问题，给出的却是双边情形的P-值。

RESULTS

BUN,Scr and serum Leptin

Compared with the normal group,the BUN,Scr and serum leptin levels in the UUO model group were significantly higher(P＜0.01).Treatment with QSG and valsartan significantly lowered BUN,Scr,and serum leptin as compared to the model group(P＜0.05 orP＜0.01),with the lowest effect seen in the QSG medium-dose group(P＜0.05)(Table 2).

OB-R,p-JAK2,p-STAT3,NF-kBp65,and MCP-1 proteins

Compared with the normal group,the expressions of MCP-1,p-JAK2,p-STAT3,NF-kBp65 and OB-R in renal tissues were significantly higher in the UUO model group(P＜0.01).As compared to the model control,the expressions of these proteins were lower in all the QSG groups and the valsartan group(P＜0.01),with the lowest expression seen in the QSG medium-dose group(P＞0.05)(Table 3,Figure 1).

JAK2,STAT3,E-cadherin,andα-SMA mRNA

Compared with the normal group,the mRNA expressions of leptin,JAK2,STAT3,and α-SMA in the UUO model group were significantly increased(P＜0.01)(Table 4).Following treatment,these expressions were significantly lower in all the QSG groups and the valsartan group than the model group(P＜0.01),with the lowest expression seen in the QSG medium-dose group(P＞0.05).Compared with the normal group the expression of E-cadherin protein in renal tissues in the UUO model group were significantly decreased(P＜0.01).E-cadherin expression in all the QSG groups and the valsartan group were higher than the model group(P＜0.01),with the highest effect seen in the QSG medium-dose group(P＜0.05).

Col-Ⅳand FN in kidney tissues

IHC analysis revealed that there were minimal expression levels of FN and Col-Ⅳin the renal tubular epithelial cells and interstitial cells in the normal group.The expressions of FN and Col-Ⅳwere significantly increased in the UUO model group(P＜0.05),which were abrogated with QSG and valsartan treatment(P＜0.05).The lowest expression was found in the QSG medium-dose group(P＜0.05)(Figures 2,3).

Pathomorphology

The glomerular and tubular morphology in the normal group showed clear structures,with normal sizes and shapes.There were no apparent edema,fibrosis,and inflammatory cell infiltration in the renal interstitial cells.In the UUO model group,there was a change in the structure of renal tubules,with visible tubular epithelial cell atrophy,vacuole degeneration,tubular dilation,interstitial edema,inflammatory cell infiltration,and fibroblast proliferation(Figures 4,5).Nevertheless,this damage was alleviated following the QSG and valsartan treatment,with the least damage found in the QSG medium-dose group.

坚持以人为本，人水和谐。坚持突出重点，兼顾一般；统一规划，统筹兼顾；分期实施，注重实效；与城市景观相结合；政府负责，分级管理。

DISCUSSION

Leptin is taken known as the obese gene.Some studies have found that 81%of leptin in the blood circulation is excreted by the kidneys in rats.7In recent years,it has been demonstrated that leptin,as a mitogenic growth factor,has the immune and inflammatory regulation capacity.8Leptin is likely involved in the proliferation,hypertrophy,and fibrosis of the tissues.Experimental studies9,10showed that leptin can cause the hyperplasia of myocardial fibroblasts and the hypertrophy of vascular smooth muscle and cardiac muscle cells.Leptin levels are closely related to the degree of liver fibrosis.11Moreover,the rise of leptin level may be an important factor in the process of renal fibrosis development.Leptin receptors exist in kidney tissues(specifically in glomerular endothelial cells,mesangial cells,and renal interstitial fibroblasts),which are activated upon leptin ligation,and thus participate in renal fibrosis.Studies have found that the degree of fibrosis in rat model of renal fibrosis is associated with the leptin level.12

It is suggested that leptin may stimulate the glomerular endothelial cell proliferation and the expression of mesangial cells.Due to the obesity gene mutation,ob/ob rats cannot produce leptin;although these rats are prone to diabetes and obesity,they rarely show kidney damage.When these rats were injected with recombinant leptin,significant proteinuria was seen,and the expressions of TGF-β1 and collagen type Ⅳ in renaltissues were increased,suggesting that leptin may promote the occurrence of renal fibrosis.13

Table 2 Comparison of BUN,Scr and Leptin levels in each group(ˉ±s)

Notes:the normal group and model group were given 0.9%physiological saline(1 mL/100 g per day).The QSG groups were given 4.0,8.0 and 16.0 g·kg－1·d－1in the low-dose,medium-dose and high-dose groups respectively,and the dosage of valsartan group was 20 mg·kg－1·d－1.The entire intervention course was 4 weeks.QSG:Qingshen granules;BUN:urea nitrogen;Scr:serum creatinine.Compared with the normal group,aP＜0.01;compared with the model group,bP＜0.05,cP＜0.01;compared with the valsartan group,dP＞0.05.

Group Normal Model QSG high-dose(16.0 g·kg－1·d－1)QSG medium-dose(8.0 g·kg－1·d－1)QSG low-dose(4.0 g·kg－1·d－1)Valsartann10 10 10 10 10 10 BUN(mmol/L)4.77±0.27 9.74±0.83a7.27±0.66b6.37±0.70cd8.07±0.34b6.30±0.29bScr(μmol/L)40.70±12.54 114.93±18.22a88.59±9.03b77.30±16.19cd95.77±13.59b79.70±27.76bLeptin(ng/mL)2.15±0.34 5.79±0.82a4.56±0.40b3.98±0.21cd4.93±0.67b4.00±0.22b

Table 3 Relative expression quantity of OB-R,p-JAK2,p-STAT3,NF-kBp65,and MCP-1 proteins in each group(ˉ±s)

Notes:the normal group and model group were given 0.9%physiological saline(1 mL/100 g per day).The QSG groups were given 4.0,8.0 and 16.0 g·kg－1·d－1in the low-dose,medium-dose and high-dose groups respectively,and the dosage of valsartan group was 20 mg·kg－1·d－1.The entire intervention course was 4 weeks.QSG:Qingshen granules;OB-R:leptin receptor;NF-kBp65:nuclear factors-κBp65;MCP-1:monocytechemotatic protein-1;JAK:janus-activated kinase;STAT:signal transducers and activator of transcriptions.Compared with the normal group,aP＜0.01;compared with the model group,bP＜0.01;compared with the valsartan group,cP＞0.05.

Group Normal Model QSG high-dose(16.0 g·kg－1·d－1)QSG medium-dose(8.0 g·kg－1·d－1)QSG low-dose(4.0 g·kg－1·d－1)Valsartann10 10 10 10 10 10 OB-R 0.374±0.015 1.189±0.074a0.782±0.042b0.653±0.038bc0.983±0.056b0.741±0.036bp-JAK2 0.609±0.082 1.270±0.019a1.010±0.074b0.788±0.056bc1.018±0.081b0.860±0.0.40bp-STAT3 0.608±0.022 1.470±0.058a1.057±0.047b0.754±0.054bc1.077±0.076b0.835±0.058bNF-kBp65 0.318±0.094 1.067±0.025a0.549±0.019b0.415±0.034bc0.564±0.032b0.453±0.027bMCP-1 0.259±0.048 1.211±0.048a0.607±0.037b0.366±0.014bc0.832±0.032b0.637±0.048b

Figure 1 Expressions of OB-R,p-JAK2,p-STAT3,NF-kBp65,and MCP-1 proteins in each group

1:normal group;2:model group;3:QSG high-dose(16.0 g·kg－1·d－1);4:QSG medium-dose(8.0 g·kg－1·d－1);5:QSG low-dose(4.0 g·kg－1·d－1);6:valsartan group.The normal group and model group were given 0.9%physiological saline(1 mL/100 g per day).The QSG groups were given 4.0,8.0 and 16.0 g·kg－1·d－1in the low-dose,medium-dose and high-dose groups respectively,and the dosage of valsartan group was 20 mg·kg－1·d－1.The entire intervention course was 4 weeks.QSG:Qingshen granules;OB-R:leptin receptor;NF-kBp65:nuclear factors-κBp65;MCP-1:monocytechemotatic protein-1;JAK:janus-activated kinase;STAT:signal transducers and activator of transcriptions.

OB-R is a single transmembrane molecule,which is composed of 3 domains:extracellular,transmembrane and intracellular domains.OB-R falls into the class I cytokine receptor family,of which the subtypes are A,B,C,D,E and F.14Leptin binds OB-R to exert its biological effect.A study found that the level of OB-R in the renal tissues of UUO rats was positively correlated with the degree of renal fibrosis.Leptin was shown to combine with OB-R in renal tubular epithelial cells to promote interstitial fibrosis.In addition,leptin-OB-R ligation can activate JAK2,15then promote the renal interstitial fibroblasts proliferation induced by connective tissue growth factor,which is involved in the process of renal fibrosis.In this study,we found that the serum levels of leptin and OB-R were significantly higher in UUO model rats than in the normal group,indicating that leptin and OB-R were involved in the pathological changes of renal fibrosis.After a 4-week course of treatment,the serum leptin and OB-R levels of the rats in all the groups were lower than the UUO model group,suggesting that the renal fibrosis progression was delayed.

JAK/STAT signaling pathway is closely related to the occurrence and development of inflammation,which is the main pathway for leptin signal transduction.JAK kinase family consists of JAK1,JAK2,JAK3 and TYK2,all of which belong to soluble and non-receptor tyrosine protein kinases.16STAT is a downstream substrate of the JAK,and its family members include STAT1,2,3,4,5a,5b and 6.STAT3 and its upstream factor AK2 are more closely related to renal interstitial fibrosis.5JAK2 is widely expressed in various tissuesand cells,and its expression was increased in high glucose-cultured fibroblasts and mesangial cells.17STAT3 plays an important role in the regulation of inflammatory response,and its phosphorylation can lead to the occurrence of inflammation.In the present study,it has been confirmed that the expressions of serum leptin,JAK2,STAT3 and p-STAT3 in the kidney tissues of UUO model group were increased,and the expression of p-JAK2 in the kidney could be explained by the activation of the leptin-mediated JAK/STAT signaling pathway in the renal tissues.

Table 4 mRNA expressions of JAK2,STAT3,E-cadherin,and α-SMA in each group(ˉ±s)

Notes:the normal group and model group were given 0.9%physiological saline(1 mL/100 g per day).The QSG groups were given 4.0,8.0 and 16.0 g·kg－1·d－1in the low-dose,medium-dose and high-dose groups respectively,and the dosage of valsartan group was 20 mg·kg－1·d－1.The entire intervention course was 4 weeks.QSG:Qingshen granules;JAK:janus-activated kinase;STAT:signal transducers and activator of transcriptions;α-SMA:alphasmooth muscle actin;E-cadherin:calcium-dependent adhesion.Compared with the normal group,aP＜0.01;compared with the model group,bP＜0.01,cP＜0.05;compared with the valsartan group,dP＞0.05.

α-SMA 1.00±0.11 4.28±0.42a2.56±0.64b1.52±0.20bd3.19±0.31b1.77±0.13bGroup Normal Model QSG high-dose(16.0 g·kg－1·d－1)QSG medium-dose(8.0 g·kg－1·d－1)QSG low-dose(4.0 g·kg－1·d－1)Valsartann10 10 10 10 10 10 Leptin 1.00±0.12 15.40±1.39a6.27±1.33b5.18±0.97bd7.34±1.43b6.00±0.25bJAK2 1.00±0.04 8.25±0.73a4.76±1.08b3.54±0.81bd6.39±0.65c4.02±1.17bSTAT3 1.00±0.08 2.74±0.23a1.84±0.08b1.26±0.09bd2.51±0.14c1.34±0.24bE-cadherin 1.00±0.11 0.31±0.07a0.59±0.06b0.68±0.15bd0.48±0.05c0.61±0.15b

Table 5 Expressions of FN and Col-Ⅳ in renal tissues(AO ×10-2,(xˉ±s)

Notes:the normal group and model group were given 0.9%physiological saline(1 mL/100 g per day).The QSG groups were given 4.0,8.0 and 16.0 g·kg－1·d－1in the low-dose,medium-dose and high-dose groups respectively,and the dosage of valsartan group was 20 mg·kg－1·d－1.The entire intervention course was 4 weeks.QSG:Qingshen granules;FN:fibronectin;Col-Ⅳ:typeⅣ collagen;AO:average optical.Compared with the normal group,aP＜0.01;compared with the model group,bP＜0.01,cP＜0.05;compared with the valsartan group,dP＜0.05.

nGroup Normal Model QSG high-dose(16.0 g·kg－1·d－1)QSG medium-dose(8.0 g·kg－1·d－1)QSG low-dose(4.0 g·kg－1·d－1)Valsartan 10 10 10 10 10 10 FN 18.1±3.0 25.6±3.4a22.4±2.0b20.4±2.3bd22.3±3.18b22.3±3.2bCol-Ⅳ18.0±3.3 27.4±4.1a24.6±5.6c21.7±3.7bd24.8±3.9c24.4±3.4c

Figure 2 Immunohistochemical staining of fibronectin in rat tissue in each group(×400)

A:normal group;B:model group;C:QSG high-dose group;D:QSG medium-dose group;E:QSG low-dose group;F:valsartan group.The normal group and model group were given 0.9%physiological saline(1 mL/100 g per day).The QSG groups were given 4.0,8.0 and 16.0 g·kg－1·d－1in the low-dose,medium-dose and high-dose groups respectively,and the dosage of valsartan group was 20 mg·kg－1·d－1.The entire intervention course was 4 weeks.QSG:Qingshen granules.

Figure 3 Immunohistochemical staining of typeⅣcollagen in rat tissue in each group(×400)

A:normal group;B:model group;C:QSG high-dose group;D:QSG medium-dose group;E:QSG low-dose group;F:valsartan group.The normal group and model group were given 0.9%physiological saline(1 mL/100 g per day).The QSG groups were given 4.0,8.0 and 16.0 g·kg－1·d－1in the low-dose,medium-dose and high-dose groups respectively,and the dosage of valsartan group was 20 mg·kg－1·d－1.The entire intervention course was 4 weeks.QSG:Qingshen granules.

Figure 4 Hematoxylin and eosin staining of rat tissue in each group(×400)

A:normal group;B:model group;C:QSG high-dose group;D:QSG medium-dose group;E:QSG low-dose group;F:valsartan group.The normal group and model group were given 0.9%physiological saline(1 mL/100 g per day).The QSG groups were given 4.0,8.0 and 16.0 g·kg－1·d－1in the low-dose,medium-dose and high-dose groups respectively,and the dosage of valsartan group was 20 mg·kg－1·d－1.The entire intervention course was 4 weeks.QSG:Qingshen granules.

Figure 5 Periodic Acid-Schiff staining of rat tissue in each group(×400)

A:normal group;B:model group;C:QSG high-dose group;D:QSG medium-dose group;E:QSG low-dose group;F:valsartan group.The normal group and model group were given 0.9%physiological saline(1 mL/100 g per day).The QSG groups were given 4.0,8.0 and 16.0 g·kg－1·d－1in the low-dose,medium-dose and high-dose groups respectively,and the dosage of valsartan group was 20 mg·kg－1·d－1.The entire intervention course was 4 weeks.QSG:Qingshen granules.

13 Wolf G,Hamann A,Han DC,et al.Leptin stimulates proliferation and TGF-beta expression in renal glomemlar endothelial cells：potential role in glomemlosclerosis.Kidney Int l999;56(3):860-872.

REFERENCES

会计准则的经济后果，是指各种财富的转移是既得利益在不同利益集团之间的重新分割，而这种“社会性后果”的表现是会计报告对企业、政府、投资人和债权人决策行为的影响，这种影响使会计信息具备了财富分配效应和决策效应，起到了资源调配作用。

1 Lovisa S,Zeisberg M,Kalluri R.Partial epithelial-to-mesenchymal transition and other new mechanisms of kidney fibrosis.Trends Endocrinol Metab 2016;27(10):681-695.

2 Wang YP,Li WJ,Cao EZ,et al.Effects of Qingshen decoction on serum leptin and interleukin-6 in patients with chronic renal failure and acute exacerbation of damp heat syndrome.Zhong Guo Zhong Xi Jie He Ji Jiu Za Zhi 2005;12(2):71-75.

因为平均输出电压为20 V,平均输入电压为30 V,则PWM占空比D为0.67,最小输入电压Uimin为24 V,设电感电流的最大变换值即纹波电流ΔILmax为满载输入电流的20%,代入式(1)和式(2),得电感H为476 μH。假设变换器输出纹波电压为10 mV,计算滤波电容C

The detection was performed in accordance with the manufacture's instructions described of the ELISA kit.The optical density(OD)was read at 450 nm using a microtiter plate reader within15-mincolor development.

4 Koike K,Ueda S,Yamagishi S,et al.Protective role of JAK/STAT signaling against renal fibrosis in mice with unilateral ureteral obstruction.Clin Immunol 2014;150(1):78-87.

目前，我国人民群众的环保诉求日益提高，生态环境在群众幸福指数中的地位不断凸显，改善环境质量不仅仅是简单的环保问题，更是严肃的政治问题、重要的经济问题和重大的民生问题。汽车维修行业作为“汽车医院”，对超标排放车辆维修治理具有先天优势，具有重要作用，可以主动承担社会责任，可以大有作为。实施I/M制度可使排放超标车辆得到真正治理，进而有效降低汽车尾气排放，达到改善区域空气质量的目的，这是利国利民的大事，是行业勇于承担社会责任的重要体现，也有助于提升整个行业在经济社会生活中的地位和形象。

在婚姻中，不要试图改造或改变伴侣。人无完人，每个人身上都存在优点，也存在缺点。当初，你之所以选择和对方结婚，肯定是因为对方身上有你欣赏的地方，不妨学会欣赏对方的优点，忽略对方的缺点。一个善解人意的妻子或丈夫，应该尊重对方的个性，不要把自己的意志强加给对方，要给对方保留一定的自由空间，允许对方有自己的社交圈子。

5 Chuang PY,He JC.JAK/STAT signaling in renal diseases.Kidney Int 2010;78(3):231-234.

6 Chen Q.Research methodology of pharmacology of Traditional Chinese Medicine.3 rd ed.Beijing:People's Medical Publishing House,2011:412-413.

参考译文:Konka Group has been granted“National Advanced Quality and Benefit Enterprise”and“National Customer Satisfied Enterprise”for several years．

7 Yaylali A,Ergin K,Ceçen S.Effect of resveratrol on leptin and sirtuin 2 expression in the kidneys in streptozotocin-induced diabetic rats.Anal Quant Cytopathol Histpathol 2015;37(4):243-251.

8 Lourenço EV,Liu A,Matarese G,La Cava A.Leptin promotes systemic lupus erythematosus by increasing autoantibody production and inhibiting immune regulation.Proc Natl Acad Sci USA 2016;113(38):10637-10642.

这件事上报后，上级克扣了丁主任和甲洛洛两个月的工资。这个惩罚很轻，甲洛洛知道自己罩在丁主任的光里。惩罚归惩罚，更重要的是名声，那可不能就此不了了之啊，小偷这名号，甲洛洛是无论如何都消受不起的，如果落这一恶名，那他大半辈子人生所树起来的光辉形象就会一下全毁了。甲洛洛想到这些，一阵阵寒意从后背袭来，在心口一层层摺叠，他不由走出屋子，坐在门口的木桩上，长长地吐气。

9 Shin HJ,Oh J,Kang SM.Leptin induces hypertrophyviap38 mitogen-activated protein kinase in rat vascular smooth muscle cells.Biochem Biophys Res Commun 2005;329(1):18-24.

10 Rajapurohitam V,Can XT,Kirshenbaum LA.The obesity-associated peptide leptin induces hypertrophy in neonatal rat ventricular myocytes.Circ Res 2003;93:277-279.

11 Rachakonda V,Borhani AA,Dunn MA,Andrzejewski M,Martin K,Behari J.Serum leptin is a biomarker of malnutrition in decompensated cirrhosis.PLoS One 2016;11(9):159-175.

12 Kümpers P,Gueler F,Rong S,Mengel M,Tossidou I,Peters I.Leptin is a coactivator of TGF-beta in unilateral ureteral obstructive kidney disease.Am J Physiol Renal Physiol 2007;293(4):F1355-F1362.

Sixty male Sprague-Dawley rats[(200±20)g]were purchased from Beijing Weitong Lihua Experimental Animal Technology Limited Company.All the animals were housed in Xin'an Medical Laboratory of Anhui University of Traditional Chinese Medicine with specific pathogen-free conditions,and had free access to water and standard rat feed.

Leptin can stimulate the expression of NF-kB in rat airway smooth muscle cells,promote the proliferation of airway smooth muscle cells,and the expression of NF-kB is related to the concentration of leptin.On the contrary,Leptin can promote the proliferation of vascular smooth muscle cells by activating the NF-kB signaling pathway.18It has been shown that there is crosstalk between JAK/STAT and NF-kB signaling pathways:NF-kB is a downstream factor of JAK/STAT signaling pathway,and is regulated by JAK/STAT signaling pathway.A study determined that following myocardial ischemia reperfusion injury in rats,NF-kB expression was increased by inhibiting the activation of JAK/STAT signaling pathway.Moreover,inhibiting the expression of myocardial NF-kB reduced the incidence of acute inflammation.19JAK/STAT signaling pathway mediates the pathological changes of renal interstitial fibrosis,which needs to be further activated by regulat-ing gene transcription and downstream signal transduction mechanism in the nucleus,such asviaNF-kB signaling.In the present study,serum leptin,JAK2 STAT3,p-JAK2,and p-STAT3 expression were decreased in all QSG groups when compared with the UUO model group.In the QSG groups,the expressions of MCP-1,FN,Col-Ⅳ, α-SMA were lower groups while E-cadherin was higher than the model group.Three doses of QSG were used,and the highest therapeutic effect was seen in the medium-dose.In all,these results confirm that QSG has an anti-renal fibrosis effect,whose mechanism may beviainhibition of the activity of leptin-mediated JAK/STAT signaling pathway,reduction of the activity of NF-kB and its inflammatory effect,and transdifferentiation in renal tubular epithelial cells.Combined,these effects of QSG delay renal fibrosis and protect renal function in rats following UUO injury.

14 Schaab M,Kratzsch J.The soluble leptin receptor.Best Pract Res Clin Endocrinol Metab 2015;29(5):661-670.

15 Mullen M,Gonzalez-Perez RR.Leptin-induced JAK/STAt signaling and cancer growth.Vaccines(Basel)2016;4(3):E26.

16 Villarino AV,Kanno Y,Ferdinand JR,O'Shea JJ.Mechanisms of Jak/STAT signaling in immunity and disease.J Immunol 2015;194(1):21-27.

17 Mao T,Chen H,Hong L,Li J.Pigment epithelium-derived factor inhibits high glucose-induced JAK/STAT signalling pathway activation in human glomerular mesangial cells.Saudi Med J 2013;34(8):793-800.

18 Huang F,Xiong X,Wang H,You S,Zeng H.Leptin-induced vascular smooth muscle cell proliferation via regulating cell cycle,activating ERK1/2 and NF-kappaB.Acta Biochim Biophys Sin(Shanghai)2010;42(5):325-331.

19 Lou L,Liu Y,Zhou J,et al.Chlorogenic acid and luteolin synergistically inhibit the proliferation ofinterleukin-1β-induced fibroblast-like synoviocytes through regulating the activation of NF-κB and JAK/STAT-signaling pathways.Immunopharmacol Immunotoxicol 2015;37(6):499-507.