生脉散对热应激大鼠肺脏AMPK-mTOR通路及自噬的影响

doi:10.11843/j.issn.0366-6964.2025.10.046

摘要/Abstract

摘要：

旨在探究热应激发生和生脉散治疗热应激肺损伤的作用及其机制，为生脉散治疗热应激肺损伤提供理论依据。选择48只SD大鼠根据热应激恢复时间建立热应激模型。将60只SD雄性大鼠随机分为6组(n=10)，各试验组大鼠每天热应激前2 h灌胃给药，对照(Control)组、热应激(Heat stress, HS)组、生脉散(Sheng Mai San, SMS)组按5.04、2.52、1.26 g·kg^-1 3个剂量给药，N-乙酰半胱氨酸(NAC)按150 mg·kg^-1给药为阳性药物组。第7天热应激结束后采样。利用HE、Masson染色、Western blot、实时荧光定量PCR等技术评估SMS对大鼠HS模型的干预作用及其潜在机制。结果显示，在热应激后6~12 h引起明显肺损伤，以6 h肺损伤最为严重(P<0.05)。实时荧光定量PCR、Western blot和生化试验结果显示，SMS能够有效降低相关炎症因子TNF-α、IL-1β、IL-10、IL6和HSP70的表达量，缓解ATP的异常累积(P<0.05)。SMS能够显著降低p-mTOR的表达量和p62堆积，激活LKB1和AMPK磷酸化，增加Beclin1和LC3表达量(P<0.05)。热应激后6 h对大鼠肺脏组织造成的损伤最严重，且SMS可通过激活AMPK-mTOR通路，促进细胞自噬，清除损伤肺细胞，从而达到治疗HS致肺损伤的效果。

关键词: 热应激, 生脉散, 肺损伤, AMPK-mTOR通路, 自噬

Abstract:

This study aims to investigate the occurrence of heat stress and the therapeutic effects and mechanisms of Sheng Mai San (SMS) in treating heat stress-induced lung injury, providing theoretical support for the application of SMS in this context. Forty-eight SD rats were selected to establish heat stress models based on the recovery time of heat stress. Another 60 SD male rats were randomly divided into 6 groups (n=10 per group). Each group was given intragastradized administration 2 hours before heat stress every day, and the Control group and heat stress group (HS) were given normal saline. SMS group was administered at SMS-H (5.04 g·kg^-1), SMS-M (2.52 g·kg^-1) and SMS-L (1.26 g·kg^-1) doses, and N-acetylcysteine (NAC) (150 mg·kg^-1) was set up as a positive control group. Sampling was performed after the end of heat stress on day 7. Biological techniques including HE staining, Masson staining, Western blot, and real-time quantitative PCR were employed to evaluate the effects of SMS on heat stress-induced lung injury and its potential mechanisms. Heat stress caused significant lung injury between 6-12 h post-exposure, with the most severe damage observed at 6 h post-stress (P<0.05). Real-time quantitative PCR, Western blot, and biochemical analyses revealed that SMS significantly reduced the expression levels of inflammatory cytokines TNF-α, IL-1β, IL-10, IL-6, and HSP70, alleviating the abnormal accumulation of ATP (P<0.05). Moreover, SMS notably decreased p-mTOR expression and p62 accumulation, activated LKB1 and AMPK phosphorylation, and increased Beclin1 and LC3 expression levels (P<0.05). Heat stress-induced lung injury was most severe at 6 h post-exposure. SMS demonstrated therapeutic effects by activating the AMPK-mTOR pathway, promoting autophagy, and clearing damaged lung cells, thereby mitigating heat stress-induced lung injury.

Key words: heat stress, Sheng Mai San, lung injury, AMPK-mTOR pathway, autophagy

中图分类号:

S853.74

肖薇, 董嘉琪, 张晓松, 周轲, 魏彦明. 生脉散对热应激大鼠肺脏AMPK-mTOR通路及自噬的影响[J]. 畜牧兽医学报, 2025, 56(10): 5277-5288.

XIAO Wei, DONG Jiaqi, ZHANG Xiaosong, ZHOU Ke, WEI Yanming. The Effects of Sheng Mai San on the AMPK-mTOR Pathway and Autophagy in the Lungs of Rats under Heat Stress[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(10): 5277-5288.

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引物名称 Primer name	上游引物(5′→3′) Forward sequence	下游引物(5′→3′) Reverse sequence
热休克蛋白70 HSP70	TCGAACAGTTCAAGATCAAACG	CCGGAACTTCTTGTAGTAGTCA
白细胞介素1β IL-1β	AACTGTGAAATAGCAGCTTTCG	CTGTGAGATTTGAAGCTGGATG
白细胞介素6 IL-6	CACTGCCTTCCCTACTTC	CATCATCGCTGTTCATAC
白细胞介素10 IL-10	GTTGCCTGCTCTTACTGG	ATGCTCCTTGATTTCTGG
肿瘤坏死因子α TNF-α	CATGGATCTCAAAGACAACCAA	CTCCTGGTATGAAATGGCAAAT
肝激酶B1 LKB1	GGAGGACGAGGACTTGTTT	CTCTGTCCATTCTGACCCAC
腺苷酸活化蛋白激酶 AMPK	AGGGTTGAAGAGGTGGAAGC	GGCAATAGAACGGTTGAGATACT
哺乳动物雷帕霉素靶蛋白 mTOR	GAGATACGCCGTCATTCC	GCATCAGAGTCAGGTGGTC
自噬效应蛋白 Beclin1	CGCTGTTTGGAGATGTT	TGAAATTATTGATCGTGCC
泛素结合蛋白 P62	AGGCTTTCAGGCGCACTACC	TGGCCATTGTCAGTTCCTCATC
微管相关蛋白轻链3 ILC3	ATAGAGCGATACAAGGGTGAGAAG	AGAAGGCTTGGTTAGCATTGAG
甘油醛-3-磷酸脱氢酶 GAPDH	ACGGGAAACCCATCACCATC	CTCGTGGTTCACACCCATCA

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