产气荚膜梭菌Beta1毒素通过P2X7-Ca2+轴诱导巨噬细胞焦亡和铁死亡

doi:10.12122/j.issn.1673-4254.2025.10.09

摘要/Abstract

摘要：

目的探索P2X7受体调控钙稳态失调介导的产气荚膜梭菌Beta1毒素毒理机制，为Beta1毒素致病机制的研究提供新思路。方法将20只10日龄BALB/c乳鼠随机分为对照组、rCPB1组、PD151746组、PD151746＋rCPB1组，5只/组。对照组灌胃PBS，rCPB1组灌胃rCPB1，PD151746组灌胃抑制剂PD151746，PD151746＋rCPB1组先灌胃PD151746，2 h后再灌胃rCPB1。采用抗体芯片技术检测空肠的炎性因子的表达，在体内水平揭示钙稳态失调在Beta1毒素导致机体炎性损伤中的调控作用；进一步使用si-RNA-P2X7并经rCPB1处理THP-1细胞后，分别检测细胞存活率，Ca²⁺、ROS和ATP水平，以及细胞焦亡和铁死亡的标志性检测指标，在体外水平揭示P2X7受体调控钙稳态失调介导的Beta1毒素的毒理机制。结果 rCPB1灌胃乳鼠后，空肠组织中有多种炎性细胞因子表达升高（P<0.05），而用PD151746治疗后，其表达水平下降（P<0.05）。成功建立P2X7受体沉默细胞模型；P2X7受体沉默后，细胞存活率升高（P<0.05）、Ca²⁺、活性氧、三磷酸腺苷水平以及细胞焦亡和铁死亡的标志性检测指标均下降（P<0.01）。结论 P2X7受体介导的Beta1毒素功能性孔形成，能够进一步导致钙稳态失调，从而触发活性氧过度积累，进而诱导细胞焦亡和铁死亡共同发生。

关键词: 产气荚膜梭菌Beta1毒素, P2X7受体, 钙稳态失调, 细胞焦亡, 铁死亡

Abstract:

Objective To explore the toxic mechanism of Clostridium perfringens Beta1 toxin mediated by P2X7 receptor-induced calcium dyshomeostasis. Methods Ten-day-old BALB/c mice were randomly divided into control group, recombinant Beta1 toxin (rCPB1) group, PD151746 group, and PD151746+rCPB1 group, and all the treatment agents were administered by gavage. The changes in expressions of inflammatory factors in the jejunum of the mice were detected using antibody chip technology to explore the regulatory role of calcium dyshomeostasis in Beta1 toxin-induced inflammatory injury level. In the cell experiment, THP-1 cells were transfected with a si-RNA targeting P2X7 receptor and treated with rCPB1, and the changes in cell survival rate, levels of Ca²⁺, ROS and ATP, and expressions of pyroptosis and ferroptosis markers were determined. Results Oral administration of rCPB1 significantly increased the levels of inflammatory cytokines in the jejunal tissue of the neonatal mice, but their levels were significantly decreased after treatment with PD151746. In THP-1 cells, rCPB1 treatment significantly decreased cell survival and increased the levels of Ca²⁺, ROS, ATP and the expressions of pyroptosis and ferroptosis markers, and these changes were obviously attenuated by P2X7 receptor knockdown. Conclusion P2X7 receptor-mediated functional pore formation by Beta1 toxin can further lead to calcium dyshomeostasis, thereby triggering excessive accumulation of ROS to subsequently induce the co-occurrence of pyroptosis and ferroptosis.

Key words: Clostridium perfringens Beta1 toxin, P2X7 receptor, calcium dyshomeostasis, pyroptosis, ferroptosis

张思雨, 冉林武, 曾瑾, 王玉炯. 产气荚膜梭菌Beta1毒素通过P2X7-Ca²⁺轴诱导巨噬细胞焦亡和铁死亡[J]. 南方医科大学学报, 2025, 45(10): 2126-2134.

Siyu ZHANG, Linwu RAN, Jin ZENG, Yujiong WANG. Clostridium perfringens Beta1 toxin induces macrophage pyroptosis and ferroptosis through the purinergic receptor P2X7-Ca²⁺ axis[J]. Journal of Southern Medical University, 2025, 45(10): 2126-2134.

图/表 11

参考文献 30

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siRNA	Sequence
siRNA-P2X7-1	F: GGAUAGCAGAGGUGAAAGATT
siRNA-P2X7-1	R: UCUUUCACCUCUGCUAUCCTT
siRNA-P2X7-2	F: AGAUCUACUGGGACUGCAATT
siRNA-P2X7-2	R: UUGCAGUCCCAGUAGAUCUTT
siRNA-P2X7-3	F: GGAUGGACCCGCAGAGCAATT
siRNA-P2X7-3	R: UUGCUCUGCGGGUCCAUCCTT

siRNA	Sequence
siRNA-P2X7-1	F: GGAUAGCAGAGGUGAAAGATT
siRNA-P2X7-1	R: UCUUUCACCUCUGCUAUCCTT
siRNA-P2X7-2	F: AGAUCUACUGGGACUGCAATT
siRNA-P2X7-2	R: UUGCAGUCCCAGUAGAUCUTT
siRNA-P2X7-3	F: GGAUGGACCCGCAGAGCAATT
siRNA-P2X7-3	R: UUGCUCUGCGGGUCCAUCCTT

Protein ID	AveExp.R	AveExp.NC	logFC	P	Foldchange	Regulation	Threshold
IL-1β	11.038471	9.952830	1.085640	0.014842	2.122318	Up	True
IL-4	10.367004	8.840474	1.526530	0.012641	2.880921	Up	True
IL-17	11.149417	9.917431	1.231985	0.011102	2.348900	Up	True
IL-21	12.077335	11.492716	0.584619	0.016853	1.499643	Up	True
IL-23	11.225443	9.751727	1.473716	0.020574	2.777363	Up	True
TNF-α	11.054078	7.873779	3.180298	0.024354	9.064948	Up	True
IL-10	11.594167	10.449359	1.144807	0.032358	2.211166	Up	True
IL-2	11.624989	8.212568	3.412420	0.039846	10.647333	Up	True
IL-9	10.103333	7.685385	2.417947	0.053630	5.344102	Up	False
IL-5	10.910604	9.888081	1.022522	0.072574	2.031467	Up	False
IL-15	8.952885	6.671663	2.281222	0.076890	4.860896	Up	False
IL-12p70	10.626989	9.659774	0.967215	0.093436	1.955062	Up	False
IL-13	8.279287	5.954063	2.325224	0.190672	5.011438	Up	False
GM-CSF	11.105001	10.092616	1.012384	0.353971	2.017243	Up	False
IL-6	10.714897	9.905783	0.809113	0.430292	1.752134	Up	False
IFN-γ	11.600352	11.329201	0.271151	0.441625	1.206770	Up	False
IL-3	10.335866	9.441554	0.894311	0.495537	1.858723	Up	False
G-CSF	6.169542	4.829909	1.339633	0.575135	2.530870	Up	False
IL-7	4.832278	4.495829	0.336449	0.857724	1.262645	Up	False
IL-1α	10.786292	10.839428	-0.053135	0.954795	0.963839	Down	False

Protein ID	AveExp.R	AveExp.NC	logFC	P	Foldchange	Regulation	Threshold
IL-1β	11.038471	9.952830	1.085640	0.014842	2.122318	Up	True
IL-4	10.367004	8.840474	1.526530	0.012641	2.880921	Up	True
IL-17	11.149417	9.917431	1.231985	0.011102	2.348900	Up	True
IL-21	12.077335	11.492716	0.584619	0.016853	1.499643	Up	True
IL-23	11.225443	9.751727	1.473716	0.020574	2.777363	Up	True
TNF-α	11.054078	7.873779	3.180298	0.024354	9.064948	Up	True
IL-10	11.594167	10.449359	1.144807	0.032358	2.211166	Up	True
IL-2	11.624989	8.212568	3.412420	0.039846	10.647333	Up	True
IL-9	10.103333	7.685385	2.417947	0.053630	5.344102	Up	False
IL-5	10.910604	9.888081	1.022522	0.072574	2.031467	Up	False
IL-15	8.952885	6.671663	2.281222	0.076890	4.860896	Up	False
IL-12p70	10.626989	9.659774	0.967215	0.093436	1.955062	Up	False
IL-13	8.279287	5.954063	2.325224	0.190672	5.011438	Up	False
GM-CSF	11.105001	10.092616	1.012384	0.353971	2.017243	Up	False
IL-6	10.714897	9.905783	0.809113	0.430292	1.752134	Up	False
IFN-γ	11.600352	11.329201	0.271151	0.441625	1.206770	Up	False
IL-3	10.335866	9.441554	0.894311	0.495537	1.858723	Up	False
G-CSF	6.169542	4.829909	1.339633	0.575135	2.530870	Up	False
IL-7	4.832278	4.495829	0.336449	0.857724	1.262645	Up	False
IL-1α	10.786292	10.839428	-0.053135	0.954795	0.963839	Down	False

Protein ID	AveExp.PR	AveExp.R	logFC	adj.P.Val	Foldchange	Regulation	Threshold
IL-1β	9.017981	11.038472	-2.020491	0.000628	0.246474	Down	True
IL-4	7.738409	10.367005	-2.628596	0.000628	0.161701	Down	True
IL-10	9.194662	11.594168	-2.399506	0.000628	0.189529	Down	True
IFN-γ	10.175170	11.600353	-1.425182	0.001782	0.372372	Down	True
GM-CSF	6.999299	11.105001	-4.105702	0.002697	0.058085	Down	True
IL-3	5.430662	10.335866	-4.905204	0.002697	0.033372	Down	True
IL-23	9.085735	11.225443	-2.139708	0.003959	0.226926	Down	True
IL-5	9.121355	10.910604	-1.789249	0.007069	0.289323	Down	True
IL-17	9.684801	11.149417	-1.464616	0.007069	0.362332	Down	True
IL-2	7.235229	11.624989	-4.389760	0.019791	0.047704	Down	True
IL-12p70	9.127082	10.626990	-1.499907	0.021880	0.353576	Down	True
IL-21	11.530132	12.077336	-0.547204	0.040580	0.684345	Down	True
IL-7	0.713847	4.832278	-4.118431	0.054878	0.057574	Down	False
IL-6	8.623386	10.714897	-2.091512	0.069235	0.234635	Down	False
IL-9	8.054050	10.103334	-2.049284	0.131383	0.241604	Down	False
TNF-α	8.907458	11.054079	-2.146621	0.148291	0.225841	Down	False
IL-13	5.552009	8.279288	-2.727278	0.149572	0.151011	Down	False
G-CSF	3.761456	6.169543	-2.408087	0.337903	0.188406	Down	False
IL-15	7.700203	8.952886	-1.252682	0.337903	0.419667	Down	False
IL-1-α	10.168991	10.786292	-0.617301	0.511964	0.651889	Down	False

产气荚膜梭菌Beta1毒素通过P2X7-Ca²⁺轴诱导巨噬细胞焦亡和铁死亡

Clostridium perfringens Beta1 toxin induces macrophage pyroptosis and ferroptosis through the purinergic receptor P2X7-Ca²⁺ axis

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