Objective

2

To observe the effects of acupuncture on podocyte autophagy and long non-coding RNA SOX2 overlapping transcript (LncRNA SOX2OT)/mammalian target of rapamycin C1 (mTORC1)/Unc-51-like kinase 1 (ULK1) pathway in rats with diabetic kidney disease (DKD)

and to explore the mechanism by which acupuncture reduces urinary protein.

Methods

2

A total of 40 SPF-grade male Sprague-Dawley rats were randomly divided into a control group (

n

=10) and a modeling group (

n

=30). The DKD model was established by feeding a high-fat

high-sugar diet combined with intraperitoneal injection of streptozotocin (STZ) in the modeling group. Twenty rats with successful DKD model were randomly divided into a model group (

n

=10) and an acupuncture group (

n

=10). The acupuncture group received "spleen and stomach-regulating" acupuncture at bilateral "Zusanli" (ST36)

"Fenglong" (ST40)

"Yinlingquan" (SP9)

and "Zhongwan" (CV12)

30 min per session

once daily

five times per week

for four weeks. The general condition

fasting blood glucose (FBG)

2-hour postprandial glucose (2hPG)

serum creatinine (SCr)

blood urea nitrogen (BUN)

24-hour urinary protein quantification

and urine albumin-to-creatinine ratio (UACR) were compared before and after the intervention. After intervention

urinary podocyte injury marker SPON2 was measured by ELISA. Podocyte autophagosomes and glomerular basement membrane ultrastructure in renal tissue were observed via transmission electron microscopy. Podocyte apoptosis was assessed by TUNEL staining. The protein expression of microtubule-associated protein 1 light chain 3Ⅱ (LC3-Ⅱ)

mTORC1

ULK1

Beclin-1

and p62 in renal tissue was detected by Western blot. LncRNA SOX2OT expression in renal tissue was measured by real-time PCR.

Results

2

After the intervention

compared with the control group

the model group exhibited increased food and water intake

increased urine output

weight loss

and loose stools; compared with the model group

the food and water intake

urine volume

and loose stools were improved in the acupuncture group. Compared with the control group

FBG

2hPG

SCr

BUN

24-hour urinary protein quantification

UACR

and urinary SPON2 were all higher in the model group (

P

<

0.01); compared with the model group

the FBG

2hPG

SCr

BUN

24-hour urinary protein quantification

UACR

and urinary SPON2 were all lower in the acupuncture group (

P

<

0.01). Compared with the control group

the model group showed reduced podocyte autophagosomes and thickened glomerular basement membrane; compared with the model group

the acupuncture group had increased podocyte autophagosomes and less thickened basement membrane. Compared with the control group

the podocyte apoptosis index (AI) was higher in the model group (

P

<

0.01); compared with the model group

the AI was lower in the acupuncture group (

P

<

0.01). Compared with the control group

the expression of ULK1

Beclin-1

and LC3-Ⅱ proteins was lower

and the expression of mTORC1 and p62 proteins was higher in the model group (

P

<

0.01). Compared with the model group

the expression of ULK1

Beclin-1

and LC3-Ⅱ proteins was higher

and the expression of mTORC1 and p62 proteins was lower in the acupuncture group (

P

<

0.01). Compared with the control group

the LncRNA SOX2OT expression was lower in the model group (

P

<

0.01). Compared with the model group

LncRNA SOX2OT expression was higher in the acupuncture group (

P

<

0.01).

Conclusion

2

The "spleen and stomach-regulating" acupuncture method could improve renal function in DKD rats

reduce blood glucose and urinary protein excretion

alleviate podocyte injury

and enhance podocyte autophagy. The mechanism may be related to modulation of the renal LncRNA SOX2OT/mTORC1/ULK1 pathway.