ALKBH5通过TRAF1/NF⁃κB通路减轻脓毒症心肌损伤的机制

doi:10.3969/j.issn.1006-5725.2024.17.005

Abstract

Abstract:

Objective To investigate the molecular mechanism of ALKBH5 reducing sepsis-induced myocardial dysfunction （SIMD）. Methods The expression levels of ALKBH5 and TRAF1 in the blood of 50 SIMD patients and 50 healthy individuals were detected using reverse transcription fluorescence quantitative polymerase chain reaction （RT-qPCR）， and the correlation between their expression levels was analyzed by person analysis； In vitro experiments， H9C2 myocardial cells were divided into 7 groups according to over expression of TARF1 and knockdown ALKBH5. The molecular mechanism of ALKBH5 targeting TRAF1 to regulate lipopolysaccharide （LPS）induced myocardial cell damage was studied through experiments such as CCK8， ELISA， and Western blot； In the in vivo experiment of rats， LPS induced rats were divided into 6 groups according to over expression of TARF1 and knockdown ALKBH5. Experimental methods such as colorimetry， ELISA， Western blot， HE staining， and immunohistochemistry were used to study the mechanism of ALKBH5 targeting TRAF1 through NF- κB pathway in reducing myocardial cell damage. Results The expression levels of ALKBH5 and TRAF 1 were downregulated in SIMD， and the Pearson analysis showed a positive correlation between them（P < 0.001）； In vitro experiments showed that overexpression of TRAF1 promotes cell proliferation， inhibits the expression of inflammatory factors and proteins involved in the NF- κB pathway， and knockdown ALKBH5 obtain the opposite resulst； In vivo experiments in rats showed that knockdown ALKBH5 promotes injury in cardiomyocytes， expression of inflammatory factors and NF- κB-related pathway proteins， and nuclear translocation of NF-κB p65 protein， but the overexpression of TRAF 1 yielded the opposite results. Conclusion ALKBH5 increases the stability of TRAF1 by reducing its methylation， thereby inhibiting NF- κB pathway， thereby reducing SIMD.

Key words: sepsis -induced myocardial dysfunction, ALKBH5, TRAF1, NF-κB

CLC Number:

R364.5

Min LIU,Xiyun CHEN,Jianlei LÜ,Jie. FENG. ALKBH5 reduce septic-induced myocardial dysfunction by regulating the TRAF1/NF-κB pathway[J]. The Journal of Practical Medicine, 2024, 40(17): 2381-2389.

Figures/Tables 11

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References 25

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时间	Control	LPS	LPS+sh-NC	LPS+sh-ALKBH5
0 h	1.01 ± 0.03	1.03 ± 0.14	0.98 ± 0.15	0.98 ± 0.18
24 h	2.03 ± 0.32	1.42 ± 0.28	1.35 ± 0.35	1.12 ± 0.29
48 h	3.01 ± 0.24	2.15 ± 0.25	2.23 ± 0.27	1.58 ± 0.32
72 h	4.12 ± 0.42	3.19 ± 0.31	3.35 ± 0.26	2.24 ± 0.35
t值	3.086^a		4.410^b
P值	0.037^a		0.012^b

组别	TNF-α	IL-6	IL-8
Control	5.67 ± 1.05	2.56 ± 1.28	2.88 ± 1.35
LPS	20.03 ± 2.09	15.95 ± 1.32	16.05 ± 2.32
t值	10.634	12.613	8.500
P值	< 0.001	< 0.001	0.001
LPS+sh-NC	19.83 ± 2.85	15.23 ± 1.45	16.55 ± 2.23
LPS+sh-ALKBH5	28.45 ± 2.07	25.98 ± 3.56	26.92 ± 3.34
t值	4.239	4.844	4.472
P值	0.013	0.008	0.011

组别	Control	LPS	LPS + pcDNA	LPS + pcDNA-TRAF1	LPS + pcDNA-TRAF1 + sh-NC	LPS + pcDNA-TRAF1 + sh-ALKBH5	LPS + pcDNA-TRAF1 + NF-κΒ activator 1
0 h	1.01 ± 0.02	1.03 ± 0.05	0.99 ± 0.11	0.98 ± 0.12	1.03 ± 0.05	0.95 ± 0.15	1.03 ± 0.05
24 h	2.05 ± 0.28	1.45 ± 0.16	1.38 ± 0.27	1.83 ± 0.34	1.85 ± 0.28	1.39 ± 0.32	1.42 ± 0.24
48 h	3.04 ± 0.22	2.14 ± 0.28	2.25 ± 0.24	2.96 ± 0.23	2.84 ± 0.16	2.22 ± 0.24	2.24 ± 0.21
72 h	4.15 ± 0.32	3.15 ± 0.21	3.46 ± 0.27	4.86 ± 0.25	4.89 ± 0.34	3.23 ± 0.23	3.35 ± 0.14
t值	4.525^a		6.600^b		8.311^e		0.772^d
P值	0.011^a		0.003^b		0.001^c		> 0.05^d

分组	TNF-α	IL-6	IL-8
Control	5.58 ± 1.08	2.66 ± 0.84	3.45 ± 1.32
LPS	20.15 ± 2.59	16.35 ± 2.28	16.25 ± 2.22
t值	8.993	9.759	6.305
P值	< 0.001	< 0.001	0.003
LPS + pcDNA	20.83 ± 1.67	16.23 ± 1.98	16.45 ± 1.23
LPS + pcDNA-TRAF1	15.46 ± 2.57	11.98 ± 1.56	11.86 ± 1.52
t值	3.035	2.920	4.066
P值	0.038	0.043	0.015
LPS + pcDNA-TRAF1 + sh-NC	15.35 ± 1.35	11.34 ± 1.54	11.45 ± 2.24
LPS + pcDNA-TRAF1 + sh-ALKBH5	20.42 ± 1.43	16.43 ± 1.34	16.82 ± 1.83
t值	2.962	3.748	3.611
P值	0.041	0.020	0.023
LPS + pcDNA-TRAF1 + NF-κΒ activator 1	21.45 ± 2.67	16.34 ± 2.21	17.02 ± 2.56
t值	0.589	0.048	0.110
P值^a	> 0.05	> 0.05	> 0.05

组别	TNF-α	IL-6	IL-8	cTnI
Control	5.58 ± 1.08	2.66 ± 0.84	3.45 ± 1.32	20.54 ± 3.45
LPS	20.15 ± 2.59	16.35 ± 2.28	16.25 ± 2.25	65.43 ± 6.65
t值	8.993	9.766	8.499	18.877
P值	0.001	0.001	0.001	< 0.001
LPS+pcDNA	20.83 ± 2.67	16.23 ± 2.98	16.45 ± 3.23	64.53 ± 5.65
LPS+pcDNA-TRAF1	10.46 ± 1.57	8.98 ± 1.47	8.92 ± 1.52	22.56 ± 3.21
t值	5.799	3.779	3.653	11.187
P值	0.004	0.020	0.022	< 0.001
LPS+pcDNA-TRAF1+sh-NC	9.35 ± 1.35	8.34 ± 1.54	8.45 ± 1.24	23.24 ± 2.84
LPS+pcDNA-TRAF1+sh-ALKBH5	20.42 ± 2.43	16.23 ± 2.34	16.82 ± 2.45	62.45 ± 7.23
t值	6.898	4.878	5.280	8.743
P值	0.002	0.008	0.006	0.001

ALKBH5 reduce septic-induced myocardial dysfunction by regulating the TRAF1/NF-κB pathway

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