Objective

2

To investigate the potential mechanism of

Yiqi Yangyin Huoxue Formula

（益气养阴活血方， YYHF） in the treatment of membranous nephropathy （MN） based on the NOX4-mediated stromal cell-derived factor-1α（SDF-1α）/CXC chemokine receptor 4 （CXCR4） signaling pathway.

Methods

2

A total of 42 male SD rats were randomly divided into blank group （n=6） and modeling group （n=36）. The modeling rats were given a single injection of sheep antirat Fx1A （6 ml/kg） serum into the tail vein to establish MN model. Thirty-six rats with successful modeling were further divided into model group （normal sali

ne， 10 ml/kg）， benazepril group （10 mg·kg

-1

·d

-1

）， the medium-dose YYHF group （11.0 g·kg

-1

·d

-1

） and high-dose YYHF group （22.0 g·kg

-1

·d

-1

）， with 9 rats in each group. All groups received daily intragastric administration for 6 consecutive weeks. During the experimental period， the general condition of rats in each group was observed. After gavage administration finished， a fully automated biochemical analyzer was used to measure the 24-hour urine total protein （24 h-UTP） and serum biochemical parameters， including albumin （ALB）， aspartate aminotransferase （AST）， blood urea nitrogen （BUN）， serum creatinine （SCr）， total cholesterol （TC）， and triglyceride （TG）. Hematoxylin-eosin （HE） staining and periodic acid-silver metheramine （PASM） staining were used to observe the histopathological changes of renal tissues in each group of rats. The levels of superoxide dismutase （SOD） and malondialdehyde （MDA） in renal tissues were detected by ELISA. The protein expression levels of NOX4， SDF-1α， CXCR4， Nrf2， nuclear factor kappa B subunit p65 （NF-κB p65） and podocyte slit diaphragm protein nephrin in rat renal tissues were detected by Western Blotting. The mRNA expressions of

NOX4

，

SDF

-

1α

，

CXCR4

，

Nrf2

，

NF

-

κB p65

and Nephrin in renal tissues were detected by real-time fluorescence quantitative polymerase chain reaction （RT-PCR）.

Results

2

Compared to the blank group， rats in the model group exhibited typical features of MN， including lethargy， reduced activity， dull and coarse fur， marked edema， and massive proteinuria. Compared to the model group， these abnormal conditions were improved to varying degrees in all treatment groups. The high-dose YYHF group showed a level of improvement comparable to that of the benazepril group， with overall recovery superior to that of the medium-dose YYHF group. Compared to the blank

group， the model group showed increased levels of 24 h-UTP， BUN， SCr， TC and TG， decreased level of ALB， increased levels of MDA and protein and mRNA expressions of

NOX4

，

SDF

-

1α

，

CXCR4

and

NF

-

κB p65

in renal tissue， along with decreased levels of SOD and protein and mRNA expressions of

Nrf2

，

Nephrin

（

P

＜0.01）， accompanied by severe renal pathological damage. Compared to the model group， the benazepril and the medium- and high-dose YYHF groups showed significant improvements in the above indicators （

P

＜0.01）， along with alleviation of renal pathological injury. The 24 h-UTP level in the high-dose YYHF group was lower than that in the medium-dose group （

P

＜0.01）， while no statistically significant difference was observed among the other treatment groups （

P

＞0.01）.

Conclusion

2

YYHF may alleviate oxidative stress and inflammatory responses by down-regulating the expression of NOX4 and inhibiting the SDF-1α/CXCR4-Nrf2 signaling pathway， thereby improving podocyte damage and protecting renal function.