Effect of asperuloside on pyroptosis of lung tissue in septic rats by regulating NLRP3/Caspase-1/GSDMD signaling pathway

doi:10.11958/20221430

Abstract

Abstract:

Objective To investigate the influence of asperuloside on pyroptosis of lung tissue in septic rats by regulating NOD-like receptor protein 3 (NLRP3)/cysteine-containing aspartate-specific proteases 3 (Caspase-1)/gasdermin D (GSDMD) signaling pathway. Methods Fifty SD rats were injected intraperitoneally with 5 mg/kg lipopolysaccharide (LPS) to establish the septic lung injury model. Rats were randomly grouped into the model (M) group, the low-dose asperuloside (AL) group (17.5 mg/kg), the medium-dose asperuloside (AM) group (35 mg/kg), the asperuloside high-dose (AH) group (70 mg/kg) and the high-dose asperuloside (70 mg/kg) + nigericin (NLRP3 activator, 1 mg/kg) group (AH+N group), 10 rats in each group. Another 10 SD rats were intraperitoneally injected with the same dose of normal saline and used as the control (C) group. After rats in each group were intervened with terpenoside and nigericin, pulmonary function indexes {minute ventilation (MV), inspiratory resistance (Ri)} and arterial oxygen partial pressure [p(O₂)] were detected in each group. HE staining was applied to detect the pathological morphology of lung tissue of rats in each group. Wright-Giemsa staining was applied to classify and count numbers of neutrophils, macrophages and lymphocytes in bronchoalveolar lavage fluid (BALF) of rats in each group. Enzyme-linked immunosorbent assay (ELISA) was used to detect serum levels of inflammatory cytokines interleukin (IL)-6, IL-8 and IL-1β in each group of rats. Immunofluorescence staining was used to detect the pyroptosis of lung tissue in each group. Expression levels of NLRP3 and GSDMD in lung tissue were compared. Western blot assay was used to detect expression levels of NLRP3/Caspase-1/GSDMD signaling pathway-related proteins in lung tissue of rats in each group. Results Compared with the C group, MV and p(O₂) were obviously decreased in the M group, and Ri, counts of neutrophils and macrophages, lymphocyte in BALF, serum levels of IL-6, IL-8, IL-1β, relative fluorescence intensity of NLRP3 and GSDMD in lung tissue, and protein expressions of NLRP3, Caspase-1 and GSDMD were obviously increased (P<0.05). Compared with the M group, MV and p(O₂) were increased in the AL group, the AM group and the AH group. Ri, counts of neutrophils and macrophages, lymphocyte in BALF, serum levels of IL-6, IL-8, IL-1β, relative fluorescence intensity of NLRP3 and GSDMD in lung tissue, and protein expressions of NLRP3, Caspase-1 and GSDMD were all decreased in a dose-dependent manner (P<0.05). Compared with the AH group, MV and p(O₂) decreased in the AH+N group, Ri, counts of neutrophils and macrophages, lymphocyte in BALF, serum levels of IL-6, IL-8, IL-1β, relative fluorescence intensity of NLRP3 and GSDMD in lung tissue, and protein expressions of NLRP3, Caspase-1 and GSDMD increased (P<0.05). Conclusion Asperuloside can inhibit lung tissue inflammation in sepsis rats by down-regulating NLRP3/Caspase-1/GSDMD pathway, attenuate lung tissue inflammatory damage and lung tissue pyroptosis, and restore lung function.

Key words: sepsis, lung injury, pyroptosis, asperuloside, NLRP3, Caspase-1, GSDMD

CLC Number:

R563.9

Figures/Tables 8

References 19

[1]	EVANS L, RHODES A, ALHAZZANI W, et al. Surviving sepsis campaign:international guidelines for management of sepsis and septic shock 2021[J]. Intensive Care Med, 2021, 47(11):1181-1247. doi:10.1007/s00134-021-06506-y.
[2]	MA J, XU L Y, SUN Q H, et al. Inhibition of miR-1298-5p attenuates sepsis lung injury by targeting SOCS6[J]. Mol Cell Biochem, 2021, 476(10):3745-3756. doi:10.1007/s11010-021-04170-w.
[3]	JIAO Y, ZHANG T, ZHANG C, et al. Exosomal miR-30d-5p of neutrophils induces M1 macrophage polarization and primes macrophage pyroptosis in sepsis-related acute lung injury[J]. Crit Care, 2021, 25(1):356-370. doi:10.1186/s13054-021-03775-3.
[4]	ZHANG Z T, ZHANG D Y, XIE K, et al. Luteolin activates Tregs to promote IL-10 expression and alleviating caspase-11-dependent pyroptosis in sepsis-induced lung injury[J]. Int Immunopharmacol, 2021, 99(5):107914-107922. doi:10.1016/j.intimp.2021.107914.
[5]	YAO F, JIN Z, ZHENG Z, et al. HDAC11 promotes both NLRP3/caspase-1/GSDMD and caspase-3/GSDME pathways causing pyroptosis via ERG in vascular endothelial cells[J]. Cell Death Discov, 2022, 8(1):112-122. doi:10.1038/s41420-022-00906-9.
[6]	LIU B, WANG Z, HE R, et al. Buformin alleviates sepsis-induced acute lung injury via inhibiting NLRP3-mediated pyroptosis through an AMPK-dependent pathway[J]. Clin Sci (Lond), 2022, 136(4):273-289. doi:10.1042/CS20211156.
[7]	洪庆, 徐曼丽, 汤建. 车叶草苷的研究进展[J]. 中国野生植物资源, 2018, 37(4):43-45,69.
	HONG Q, XU M L, TANG J. Advances of study on the iridoid asperuloside[J]. Chin Wild Plant Res, 2018, 37(4):43-45,69. doi:10.3969/j.issn.1006-9690.2018.04.009.
[8]	屠万倩, 李宁, 张留记, 等. 不同加工方式生产的杜仲叶中8种化学成分的含量及抗氧化活性研究[J]. 中国新药杂志, 2020, 29(16):1863-1867.
	TU W Q, LI N, ZHANG L J, et al. Comparison of contents and antioxidant activity of eight components in Eucommia ulmoides leaves processed by different methods[J]. Chin J New Drugs, 2020, 29(16):1863-1867. doi:10.3969/j.issn.1003-3734.2020.16.013.
[9]	王海燕, 高令心, 莫美娜, 等. 补肾益肺法联合家庭氧疗治疗慢性阻塞性肺疾病稳定期40例[J]. 中医研究, 2019, 32(3):26-29.
	WANG H Y, GAO L X, MO M N, et al. Treatment of 40 patients with chronic obstructive pulmonary disease by Tonifying kidney and nourishing lung combined with home oxygen therapy[J]. Trad Chin Med Res, 2019, 32(3):26-29. doi:10.3969/j.issn.1001-6910.2019.03.11.
[10]	陈剑明, 彭美哲, 陈奕杉, 等. 参附黄制剂联合亚低温对脓毒症大鼠下丘脑AMPK-POMC相关靶点的影响[J]. 中华中医药杂志, 2022, 37(5):2475-2480.
	CHEN J M, PENG M Z, CHEN Y S, et al. Effects of Shenfuhuang Preparation combined with subhypothermia on AMPK-POMC-related targets in the hypothalamic brain of septic rats[J]. Chin J Tradit Chin Med Pharm, 2022, 37(5):2475-2480.
[11]	初巍巍, 袁菲阳, 丁国臣, 等. 车叶草苷对小鼠Lewis肺癌的抑瘤、抗炎作用及其机制[J]. 武警医学, 2020, 31(5):401-404.
	CHU W W, YUAN F Y, DING G C, et al. Anti-tumor and anti-inflammatory effect of asperuloside on mice with Lewis lung cancer[J]. Med J Chin PAPF, 2020, 31(5):401-404. doi:10.3969/j.issn.1004-3594.2020.05.009.
[12]	SHAN W, LIAO X, TANG Y, et al. Dexmedetomidine alleviates inflammation in neuropathic pain by suppressing NLRP3 via Nrf2 activation[J]. Exp Ther Med, 2021, 22(4):1046-1054. doi:10.3892/etm.2021.10479.
[13]	LV X, ZHANG X Y, ZHANG Q, et al. lncRNA NEAT1 aggravates sepsis-induced lung injury by regulating the miR-27a/PTEN axis[J]. Lab Invest, 2021, 101(10):1371-1381. doi:10.1038/s41374-021-00620-7.
[14]	ACKERMAN M H, AHRENS T, KELLY J, et al. Sepsis[J]. Crit Care Nurs Clin North Am, 2021, 33(4):407-418. doi:10.1016/j.cnc.2021.08.003.
[15]	ZHUO Y, YANG L, LI D, et al. Syringaresinol resisted sepsis-induced acute lung injury by suppressing pyroptosis via the oestrogen receptor-β signalling pathway[J]. Inflammation, 2022, 45(2):824-837. doi:10.1007/s10753-021-01587-9.
[16]	DING H, YANG J, CHEN L, et al. Memantine alleviates acute lung injury via inhibiting macrophage pyroptosis[J]. Shock, 2021, 56(6):1040-1048. doi:10.1097/SHK.0000000000001790.
[17]	WANG L X, REN C, YAO R Q, et al. Sestrin2 protects against lethal sepsis by suppressing the pyroptosis of dendritic cells[J]. Cell Mol Life Sci, 2021, 78(24):8209-8227. doi:10.1007/s00018-021-03970-z.
[18]	LI S, SUN Y, SONG M, et al. NLRP3/caspase-1/GSDMD-mediated pyroptosis exerts a crucial role in astrocyte pathological injury in mouse model of depression[J]. JCI Insight, 2021, 6(23):e146852-e146865. doi:10.1172/jci.insight.146852.
[19]	WANG Y C, LIU Q X, ZHENG Q, et al. Dihydromyricetin alleviates sepsis-induced acute lung injury through inhibiting NLRP3 inflammasome-dependent pyroptosis in mice model[J]. Inflammation, 2019, 42(4):1301-1310. doi:10.1007/s10753-019-00990-7.

组别	MV（mL）	Ri（kPa^-1·L^-1·s^-1）	p（O₂）（mmHg）
C组	7.69±0.86	24.97±4.65	98.82±10.61
M组	2.71±0.42^a	59.68±6.80^a	54.75±5.15^a
AL组	3.91±0.48^b	50.98±6.12^b	65.12±6.24^b
AM组	4.80±0.51^bc	42.24±5.16^bc	75.82±7.43^bc
AH组	7.13±0.73^bcd	26.43±4.52^bcd	95.93±9.52^bcd
AH+N组	2.93±0.38^e	55.32±6.03^e	58.12±5.36^e
F	129.083^＊＊	69.525^＊＊	61.205^＊＊

组别	MV（mL）	Ri（kPa^-1·L^-1·s^-1）	p（O₂）（mmHg）
C组	7.69±0.86	24.97±4.65	98.82±10.61
M组	2.71±0.42^a	59.68±6.80^a	54.75±5.15^a
AL组	3.91±0.48^b	50.98±6.12^b	65.12±6.24^b
AM组	4.80±0.51^bc	42.24±5.16^bc	75.82±7.43^bc
AH组	7.13±0.73^bcd	26.43±4.52^bcd	95.93±9.52^bcd
AH+N组	2.93±0.38^e	55.32±6.03^e	58.12±5.36^e
F	129.083^＊＊	69.525^＊＊	61.205^＊＊

组别	中性粒细胞	巨噬细胞	淋巴细胞
C组	2.68±0.65	22.76±2.43	5.09±0.67
M组	13.10±1.12^a	54.20±6.51^a	81.94±17.62^a
AL组	10.68±0.95^b	46.59±5.27^b	64.75±13.41^b
AM组	8.19±0.70^bc	39.84±4.25^bc	45.01±9.46^bc
AH组	3.15±0.57^bcd	25.90±2.56^bcd	6.13±0.81^bcd
AH+N组	12.38±1.31^e	53.32±6.03^e	80.61±15.73^e
F	242.906^＊＊	80.336^＊＊	88.312^＊＊

组别	中性粒细胞	巨噬细胞	淋巴细胞
C组	2.68±0.65	22.76±2.43	5.09±0.67
M组	13.10±1.12^a	54.20±6.51^a	81.94±17.62^a
AL组	10.68±0.95^b	46.59±5.27^b	64.75±13.41^b
AM组	8.19±0.70^bc	39.84±4.25^bc	45.01±9.46^bc
AH组	3.15±0.57^bcd	25.90±2.56^bcd	6.13±0.81^bcd
AH+N组	12.38±1.31^e	53.32±6.03^e	80.61±15.73^e
F	242.906^＊＊	80.336^＊＊	88.312^＊＊

组别	TNF-α	IL-8	IL-1β
C组	27.35±5.02	204.94±32.25	16.87±3.03
M组	183.46±21.53^a	968.40±92.30^a	242.64±28.12^a
AL组	136.59±17.84^b	761.25±77.62^b	193.45±20.51^b
AM组	107.86±13.46^bc	589.32±54.92^bc	134.45±12.51^bc
AH组	31.12±6.32^bcd	229.86±30.74^bcd	21.13±3.54^bcd
AH+N组	164.63±20.14^e	960.11±90.58^e	230.36±25.92^e
F	185.239^＊＊	251.860^＊＊	296.171^＊＊