Effect of nicotinamide adenine dinucleotide phosphate oxidase 4 blocker on sciatic nerve apoptosis in diabetic neuropathy rats

doi:10.3877/cma.j.issn.2095-655X.2020.02.015

Abstract

Abstract:

Objective

To investigate the nicotinamide adenine dinucleotide phosphate oxidase 4 (Nox4) expression in sciatic nerve of diabetic peripheral neuropathy (DPN) rats, and the effect of VAS2870, a Nox4 blocker, on the apoptosis of the sciatic nerve of DPN rats.

Methods

8 weeks old male SD rats, with an average weight of (203.5±6.4)g, were developed diabetes after one time intraperitoneal injection of STZ. The other rats were intraperitoneally injected with equal volume citric acid solution as the control group (n=10). DPN model (n=20) was established in diabetic rats after 8 weeks, and DPN rats were divided into two groups by random number table method: VAS2870 group (n=10, VAS2870 1 mg/kg was injected into tail vein once a day for 7 days) and DPN group (n=10, 0.9% sterile NaCl solution was injected into tail vein once a day for 7 days). The motor nerve conduction velocity (MNCV) and sensory nerve conduction velocity (SNCV) of sciatic nerve were detected by PowerLab data recording and analysis system. The expressions of Nox4, caspase-3, B-cell lymphoma factor-2(Bcl-2) and Bcl-2 associated X protein(Bax) mRNA of sciatic nerve were detected by fluorescence real time quantitative PCR reaction, and the expressions of Nox4, caspase-3, Bcl-2 and Bax protein of sciatic nerve were detected by western blot assay.

Results

The MNCV [(41.21±7.36)m/s] and SNCV [(30.39±7.66)m/s] in DPN group were lower than those in VAS2870 group [(49.14±8.67)m/s, (38.95±6.24)m/s], VAS2870 could alleviate the decrease of MNCV and SNCV of sciatic nerve(q=3.58, 4.53, P<0.05). Compared with the expression of mRNA (5.79±0.53, 6.90±0.79, 7.39±0.67) and protein (0.59±0.13, 0.45±0.14, 0.64±0.14) in control group, the expressions of Nox4 (4.72±0.42), caspase-3 (5.02±0.43), Bax (5.63±0.82) mRNA (q=7.02, 6.40, 7.65, all P<0.05) and protein (0.86±0.10, 0.78±0.17, 0.95±0.13, q=7.83, 9.15, 6.87, all P<0.05) in DPN rats were up-regulated, while the expressions of Bcl-2 mRNA (6.81±0.60, q=6.31, P<0.05) and protein (0.44±0.12, q=6.20, P<0.05)were down-regulated. Compared with DPN group, the expression of Nox4, caspase-3, Bax mRNA (5.31±0.44, 5.71±0.60, 6.41±0.95)(q=3.66, 2.98, 4.02, 3.38, all P<0.05) and protein (0.72±0.13, 0.60±0.18, 0.79±0.15) (q=4.24, 5.78, 3.83, all P<0.05) in VAS2870 group were down-regulated, Bcl-2 mRNA (6.29±0.47) and protein (0.56±0.12) were up-regulated (q=3.38, 2.81, all P<0.05), VAS2870 partially reversed the expression changes of Nox4, caspase-3, Bax, Bcl-2 mRNA and protein.

Conclusions

The expression of Nox4 is increased in the sciatic nerve of DPN rats, and the expression of caspase-3, Bax and Bcl-2 is regulated by Nox4 to promote the apoptosis of the sciatic nerve. Nox4 blocker can restore the conduction velocity of the sciatic nerve in DPN rats by reducing the apoptosis of sciatic nerve.

Key words: Diabetic neuropathies, Sciatic nerve, Nicotinamide adenine dinucleotide phosphate oxidase 4, Apoptosis

Ting Yu, Bailiu Ya, Hongju Cheng, Qing Xin, Fei Xu, Lei Li. Effect of nicotinamide adenine dinucleotide phosphate oxidase 4 blocker on sciatic nerve apoptosis in diabetic neuropathy rats[J]. Chinese Journal of Diagnostics(Electronic Edition), 2020, 08(02): 135-140.

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Figures/Tables 5

References 21

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基因	引物序列
烟酰胺腺嘌呤二核苷酸磷酸氧化酶4	5′-CCAGAATGAGGATCCCAGAA-3′(F)
烟酰胺腺嘌呤二核苷酸磷酸氧化酶4	5′-AGCAGCAGCAGCATGTAGAA-3′(R)
半胱氨酸天冬酶-3	5′-ATGCTTACTCTACCGCACCCG-3′(F)
?	5′-GGTTAACACGAGTGAGGATGTGC-3′(R)
B-细胞淋巴瘤因子-2	5′-GTGAACTGGGGGAGGATTGT-3′(F)
B-细胞淋巴瘤因子-2	5′-GCATCCCAGCCTCCGTTA-3′(R)
B-细胞淋巴瘤因子-2相关X蛋白	5′-CCCGAGAGGTCTTCTTCCG-3′(F)
B-细胞淋巴瘤因子-2相关X蛋白	5′-GAAGTCCAGTGTCCAGCCCA-3′(R)
β-肌动蛋白	5′-CACCCGCGAGTACAACCTTC-3′(F)
?	5′-CCCATACCCACCATCACACC-3'(R)

基因	引物序列
烟酰胺腺嘌呤二核苷酸磷酸氧化酶4	5′-CCAGAATGAGGATCCCAGAA-3′(F)
烟酰胺腺嘌呤二核苷酸磷酸氧化酶4	5′-AGCAGCAGCAGCATGTAGAA-3′(R)
半胱氨酸天冬酶-3	5′-ATGCTTACTCTACCGCACCCG-3′(F)
?	5′-GGTTAACACGAGTGAGGATGTGC-3′(R)
B-细胞淋巴瘤因子-2	5′-GTGAACTGGGGGAGGATTGT-3′(F)
B-细胞淋巴瘤因子-2	5′-GCATCCCAGCCTCCGTTA-3′(R)
B-细胞淋巴瘤因子-2相关X蛋白	5′-CCCGAGAGGTCTTCTTCCG-3′(F)
B-细胞淋巴瘤因子-2相关X蛋白	5′-GAAGTCCAGTGTCCAGCCCA-3′(R)
β-肌动蛋白	5′-CACCCGCGAGTACAACCTTC-3′(F)
?	5′-CCCATACCCACCATCACACC-3'(R)

组别	鼠数(只)	体重(g)	空腹血糖(mmol/L)
VAS2870组	10	437.69±28.17^ab	12.15±1.09^ab
DPN组	10	425.34±30.45^a	11.54±1.22^a
对照组	10	576.58±24.27	4.23±0.64
F值	?	78.54	182.35
P值	?	<0.01	<0.01

组别	鼠数(只)	体重(g)	空腹血糖(mmol/L)
VAS2870组	10	437.69±28.17^ab	12.15±1.09^ab
DPN组	10	425.34±30.45^a	11.54±1.22^a
对照组	10	576.58±24.27	4.23±0.64
F值	?	78.54	182.35
P值	?	<0.01	<0.01

组别	鼠数(只)	MNCV	SNCV
VAS2870组	10	49.14±8.67^b	38.95±6.24^b
DPN组	10	41.21±7.36^a	30.39±7.66^a
对照组	10	57.32±8.15	46.58±6.97
F值	?	10.82	13.77
P值	?	<0.01	<0.01

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