前列地尔联合血必净对脓毒症大鼠心肌线粒体的影响

Effects of alprostadil combined with Xuebijing on myocardial mitochondria in septic rats

摘要

目的 分析脓毒症对大鼠心肌组织及心肌线粒体的损伤情况，并探讨前列地尔联合血必净对脓毒症大鼠心肌损伤及线粒体结构的影响。方法 选取90只SPF级健康成年雄性SD大鼠，随机分为假手术组（Sham组）、脓毒症组（CLP组）、前列地尔组（Alp组）、血必净组（XBJ组）和前列地尔联合血必净（Alp+XBJ）组，每组18只。除Sham组仅行开腹游离盲肠外，其余4组均通过盲肠结扎穿孔法建立脓毒症模型。造模后3 h用药组大鼠分别给予前列地尔（10 μg/kg）、血必净（10 mL/kg）及前列地尔联合血必净腹腔注射干预，而脓毒症组则给予等量生理盐水腹腔注射干预。每组取10只大鼠观察7 d生存情况，其余大鼠于术后24 h处死，检测血清肌酸激酶同工酶（CK-MB）及肌钙蛋白Ⅰ（cTn Ⅰ）水平，并通过苏木精-伊红（HE）染色及透射电镜观察心肌组织病理学及线粒体超微结构变化，随后通过透射电镜下图像进一步对线粒体横截面积及体密度进行定量分析。结果 各组大鼠7 d生存率比较差异有统计学意义（log-rank检验： χ² = 33.587， P ＜ 0.001），CLP组7 d生存率低于Alp+XBJ组（ P = 0.003），XBJ组、Alp组和Alp+XBJ组之间大鼠7 d生存情况差异无统计学意义（ P值分别为0.464、0.330、0.074）。与假手术组相比，脓毒症组血清CK-MB和cTn Ⅰ水平升高（ P均 ＜ 0.001），心肌组织结构紊乱并伴炎性细胞浸润；电镜下可见线粒体肿胀、嵴断裂及空泡样变性，线粒体横截面积和体密度增加（ P均 ＜ 0.001）。在脓毒症模型下，双因素方差分析显示，前列地尔与血必净在降低CK-MB和cTn Ⅰ水平方面存在交互效应（ F = 10.98， P = 0.003； F = 9.63， P = 0.004），估算边际均值图显示，联合用药具有协同增效作用。超微结构方面，两药均可改善线粒体横截面积和体密度（ P均 ＜ 0.001），且联合用药对线粒体横截面积的改善作用优于各单药治疗组（均 P<0.05）。结论 脓毒症可导致大鼠心肌损伤及线粒体结构破坏，前列地尔与血必净联合应用可增强心肌保护效果，并在部分线粒体结构指标上表现出协同作用。

Abstract

Objective To analyze the injury of sepsis to myocardial tissue and myocardial mitochondria in rats, and to investigate the effects of alprostadil combined with Xuebijing on myocardial injury and mitochondrial structure in septic rats. Methods Ninety SPF healthy adult male SD rats were randomly divided into the sham operation group (Sham group), sepsis group (CLP group), alprostadil group (Alp group), Xuebijing group (XBJ group), and alprostadil combined with Xuebijing group (Alp + XBJ group), with 18 rats in each group. Except for the Sham group, in which only laparotomy and cecal mobilization were performed, sepsis models were established in the other four groups by cecal ligation and puncture. At 3 h after model establishment, rats in the treatment groups were given intraperitoneal injections of alprostadil (10 μg/kg), Xuebijing (10 mL/kg), or alprostadil combined with Xuebijing, respectively, while rats in the sepsis group were given an equal volume of normal saline by intraperitoneal injection. Ten rats from each group were selected to observe 7-day survival, and the remaining rats were euthanized 24 h after surgery. Serum levels of creatine kinase-MB (CK-MB) and cardiac troponin Ⅰ (cTn Ⅰ) were measured. Histopathological changes in myocardial tissue and mitochondrial ultrastructural changes were observed by HE staining and transmission electron microscopy, respectively. The mitochondrial cross-sectional area and volume density were further quantitatively analyzed using images obtained under transmission electron microscopy. Results There was a statistically significant difference in the 7-day survival rate among the groups (log-rank: χ ² = 33.587, P ＜ 0.001). The 7-day survival rate in the CLP group was lower than that in the Alp + XBJ group ( P = 0.003). No statistically significant differences in 7-day survival were observed among the XBJ, Alp, and Alp + XBJ groups ( P = 0.464, 0.330, and 0.074, respectively). Compared with the Sham group, the sepsis group showed increased serum CK-MB and cTn Ⅰ levels (both P ＜ 0.001), disordered myocardial tissue structure, and inflammatory cell infiltration. Under electron microscopy, mitochondrial swelling, cristae disruption, and vacuolar degeneration were observed, with increases in mitochondrial cross-sectional area and volume density (both P ＜ 0.001). Under the sepsis model, two-way analysis of variance showed that an interaction effect was also observed between alprostadil and Xuebijing in reducing CK-MB and cTn Ⅰ levels ( F = 10.98, P = 0.003; F = 9.63, P = 0.004), the estimated marginal means plot showed that the combination therapy had a synergistic effect. In terms of ultrastructure, both drugs improved mitochondrial cross-sectional area and volume density (both P ＜ 0.001), and the combination therapy showed superior improvement in mitochondrial cross-sectional area compared to each monotherapy group (both P < 0.05). Conclusions Sepsis can lead to myocardial injury and mitochondrial structural damage in rats. The combined application of alprostadil and Xuebijing can enhance the cardioprotective effect and shows synergistic effects on some mitochondrial structural indicators.

关键词

脓毒症 / 心肌损伤 / 心肌线粒体 / 血必净 / 前列地尔

Key words

Sepsis / Myocardial injury / Myocardial mitochondria / Xuebijing / Alprostadil

参考文献

[1] Srzić I, Nesek Adam V, Tunjić Pejak D . Sepsis definition: what’s new in the treatment guidelines[J]. Acta Clin Croat, 2022, 61(Suppl 1): 67-72. DOI: 10.20471/acc.2022.61.s1.11.

[2] Coloretti I, Tosi M, Biagioni E, et al. Management of sepsis in the first 24 hours: bundles of care and individualized approach[J]. Semin Respir Crit Care Med, 2024, 45(4): 503-509. DOI: 10.1055/s-0044-1789185.

[3] Chiu C, Legrand M . Epidemiology of sepsis and septic shock[J]. Curr Opin Anaesthesiol, 2021, 34(2): 71-76. DOI: 10.1097/aco.0000000000000958.

[4] Zhang X, Zhang Y, Yuan S, et al. The potential immunological mechanisms of sepsis[J]. Front Immunol, 2024, 15: 1434688. DOI: 10.3389/fimmu.2024.1434688.

[5] 梁旭, 李国旗, 库尔班江·吐尔逊, 等 . 脓毒症患者系统免疫炎症指数、D-二聚体水平与SOFA评分的变化及其对预后的评估价值[J]. 暨南大学学报(自然科学与医学版), 2024, 45(6): 622-630. DOI: 10.11778/j.jdxb.20240013.

[6] Liang X, Li G Q, Tu'erxun K, et al. Changes in systemic immune inflammatory index, D-dimer level and SOFA score in patients with sepsis and their prognostic value assessment[J]. J Jinan Univ (Sci Med Ed), 2024, 45(6): 622-630. DOI: 10.11778/j.jdxb.20240013.

[7] Liu D, Huang S Y, Sun J H, et al. Sepsis-induced immunosuppression: mechanisms, diagnosis and current treatment options[J]. Mil Med Res, 2022, 9(1): 56. DOI: 10.1186/s40779-022-00422-y.

[8] Gao X, Cai S, Li X, et al. Sepsis-induced immunosuppression: mechanisms, biomarkers and immunotherapy[J]. Front Immunol, 2025, 16: 1577105. DOI: 10.3389/fimmu.2025.1577105.

[9] Oczkowski S, Alshamsi F, Belley-Cote E, et al. Surviving Sepsis Campaign Guidelines 2021: highlights for the practicing clinician[J]. Pol Arch Intern Med, 2022, 132(7/8): 16290. DOI: 10.20452/pamw.16290.

[10] Srdić T, Đurašević S, Lakić I, et al. From molecular mechanisms to clinical therapy: understanding sepsis-induced multiple organ dysfunction[J]. Int J Mol Sci, 2024, 25(14): 7770. DOI: 10.3390/ijms25147770.

[11] Wang J, Luo C, Luo M, et al. Targets and mechanisms of xuebijing in the treatment of acute kidney injury associated with sepsis: a network pharmacology-based study[J]. CAD, 2024, 20(6): 752-763. DOI: 10.2174/1573409919666230519121138.

[12] Wang M, Cai X F, Zhang S M, et al. Alprostadil alleviates liver injury in septic rats via TLR4/NF-κB pathway [J]. Eur Rev Med Pharmacol Sci, 2021, 25(3): 1592-1599. DOI: 10.26355/eurrev_202102_24869.

[13] Liao X, Rello J . Efficacy of xuebijing injection for sepsis (EXIT-SEP): lost in translation[J]. Anaesth Crit Care Pain Med, 2023, 42(4): 101257. DOI: 10.1016/j.accpm.2023.101257.

[14] Singer M, Deutschman C S, Seymour C W, et al. The third international consensus definitions for sepsis and septic shock (sepsis-3)[J]. JAMA, 2016, 315(8): 801. DOI: 10.1001/jama.2016.0287.

[15] Wang S, He S, Hu X, et al. Nrf2 mediated signaling axis in sepsis-induced cardiomyopathy: potential pharmacological receptor[J]. Inflamm Res, 2025, 74(1): 76. DOI: 10.1007/s00011-025-02037-0.

[16] 黄颖, 唐立丽, 关于琳, 等 . 脓毒症心肌损伤发病机制及治疗研究进展[J]. 实用医学杂志, 2023, 39(14): 1848-1852. DOI: 10.3969/j.issn.1006-5725.2023.14.021.

[17] Huang Y, Tang L L, Guan Y L, et al. Research progress on the pathogenesis and treatment of sepsis-induced myocardial injury[J]. J Pract Med, 2023, 39(14): 1848-1852. DOI: 10.3969/j.issn.1006-5725.2023.14.021.

[18] Manetti A C, Maiese A, Paolo M D, et al. microRNAs and sepsis-induced cardiac dysfunction: a systematic review[J]. Int J Mol Sci, 2020, 22(1): 321. DOI: 10.3390/ijms22010321.

[19] 于紫英, 刘峰涛, 曾勇智. 线粒体功能障碍与脓毒症心肌病的研究进展[J]. 中国心血管杂志, 2024, 29(6): 593-596. DOI: 10.3969/j.issn.1007-5410.2024.06.017.

[20] Yu Z Y, Liu F T, Zeng Y Z. Research progress on mitochondrial dysfunction and septic cardiomyopathy[J]. Chin J Cardiovasc Med, 2024, 29(6): 593-596. DOI: 10.3969/j.issn.1007-5410.2024.06.017.

[21] Yang H, Zhang Z . Sepsis-induced myocardial dysfunction: the role of mitochondrial dysfunction[J]. Inflamm Res, 2021, 70(4): 379-387. DOI: 10.1007/s00011-021-01447-0.

[22] Gao M, Xu C, Su Z, et al. Gstk1 confers cardioprotection in sepsis by regulating mitochondrial function and inhibiting cGAS/STING-dependent inflammation and pyroptosis[J]. Int Immunopharmacol, 2026, 172: 116225. DOI: 10.1016/j.intimp.2026.116225.

[23] Zou R, Tao J, Qiu J, et al. DNA-PKcs promotes sepsis-induced multiple organ failure by triggering mitochondrial dysfunction[J]. J Adv Res, 2022, 41: 39-48. DOI: 10.1016/j.jare.2022.01.014.

[24] Zheng D, Cao T, Zhang L L, et al. Targeted inhibition of calpain in mitochondria alleviates oxidative stress-induced myocardial injury[J]. Acta Pharmacol Sin, 2021, 42(6): 909-920. DOI: 10.1038/s41401-020-00526-y.

[25] Tang W, Guo R, Hu C, et al. BMAL1 alleviates myocardial damage in sepsis by activating SIRT1 signaling and promoting mitochondrial autophagy[J]. Int Immunopharmacol, 2024, 133: 112111. DOI: 10.1016/j.intimp.2024.112111.

[26] Kuroshima T, Kawaguchi S, Okada M . Current perspectives of mitochondria in sepsis-induced cardiomyopathy[J]. Int J Mol Sci, 2024, 25(9): 4710. DOI: 10.3390/ijms25094710.

[27] 陈秋华, 庄燕, 蒋华, 等 . 基于网络药理学和分子对接技术的血必净注射液治疗脓毒症的机制研究[J]. 徐州医科大学学报, 2024, 44(6): 415-421. DOI: 10.3969/j.issn.2096-3882.2024.06.005.

[28] Chen Q H, Zhuang Y, Jiang H, et al. Mechanism of Xuebijing Injection in the treatment of sepsis based on network pharmacology and molecular docking[J]. Acta Acad Med Xuzhou, 2024, 44(6): 415-421. DOI: 10.3969/j.issn.2096-3882.2024.06.005.

[29] Li C, Wang P, Li M, et al. The current evidence for the treatment of sepsis with Xuebijing injection: bioactive constituents, findings of clinical studies and potential mechanisms[J]. J Ethnopharmacol, 2021, 265: 113301. DOI: 10.1016/j.jep.2020.113301.

[30] Yu T, Dong D, Guan J, et al. Alprostadil attenuates LPS-induced cardiomyocyte injury by inhibiting the Wnt5a/JNK/NF-κB pathway[J]. Herz, 2020, 45(Suppl 1): 130-138. DOI: 10.1007/s00059-019-4837-0.

[31] 郑坚, 吴云龙, 王梅, 等 . 左西孟旦联合中药制剂治疗严重脓毒症心肌损伤患者的临床效果[J]. 中华医院感染学杂志, 2019, 29(1): 44-47, 51. DOI: 10.11816/cn.ni.2018-174135.

[32] Zheng J, Wu Y L, Wang M, et al. Clinical effect of levosimendan combined with traditional Chinese medicine on treatment of severe sepsis patients with myocardial injury[J]. Chin J Nosocomiology, 2019, 29(1): 44-47, 51. DOI: 10.11816/cn.ni.2018-174135.

[33] 刘芳宇, 金悦婷, 杜元, 等 . 不同中药注射液联合常规治疗对脓毒症心肌损伤的网状Meta分析[J]. 中国中医药信息杂志, 2023, 30(5): 10-18. DOI: 10.19879/j.cnki.1005-5304.202209609.

[34] Liu F Y, Jin Y T, Du Y, et al. Different TCM injections combined with conventional therapy for septic myocardial injury: a network meta-analysis[J]. Chin J Inf Tradit Chin Med, 2023, 30(5): 10-18. DOI: 10.19879/j.cnki.1005-5304.202209609.

[35] Verra C, Paulmann M K, Wegener J, et al. Spleen tyrosine kinase: a novel pharmacological target for sepsis-induced cardiac dysfunction and multi-organ failure[J]. Front Immunol, 2024, 15: 1447901. DOI: 10.3389/fimmu.2024.1447901.

[36] Park Y J, Lee M J, Bae J, et al. Effects of glucocorticoid therapy on sepsis depend both on the dose of steroids and on the severity and phase of the animal sepsis model[J]. Life (Basel), 2022, 12(3): 421. DOI: 10.3390/life12030421.

[37] Jin N, Zhang M, Zhou L, et al. Mitochondria transplantation alleviates cardiomyocytes apoptosis through inhibiting AMPKα-mTOR mediated excessive autophagy[J]. FASEB J, 2024, 38(10): e23655. DOI: 10.1096/fj.202400375R.

[38] Yu Q, Liu Z, Yu J, et al. Cordycepin alleviates sepsis-induced cardiomyopathy by attenuating mitochondrial oxidative stress and apoptosis through modulation of the PI3K/Akt/mTOR pathway[J]. Cell Signal, 2026, 139: 112354. DOI: 10.1016/j.cellsig.2026.112354.

[39] Song R, Huang C, Ma Y, et al. Cytoskeletal remodeling promotes tunneling nanotube formation and drives cardiac resident cell mitochondrial transfer in sepsis[J]. Sci Adv, 2026, 12(11): eadz3266. DOI: 10.1126/sciadv.adz3266.

[40] Chen W, Zhao Z, Geng Z, et al. Advances in mitochondria-nucleus crosstalk in septic cardiomyopathy[J]. Cell Biol Toxicol, 2025, 41(1): 136. DOI: 10.1007/s10565-025-10090-y.

[41] Yuan L, Tang Y, Yin L, et al. Nucleolin alleviates endotoxemia-induced myocardial dysfunction via inhibiting Drp1-mediated mitochondrial fission [J]. Tissue Cell, 2025, 96: 102964. DOI: 10.1016/j.tice.2025.102964.

[42] Shi J, Chen Y, Zhi H, et al. Levosimendan protects from sepsis-inducing cardiac dysfunction by suppressing inflammation, oxidative stress and regulating cardiac mitophagy via the PINK-1-Parkin pathway in mice [J]. Ann Transl Med, 2022, 10(4): 212. DOI: 10.21037/atm-22-483.

[43] Liu P, Xu T, Luo Y, et al. SIRT3 attenuates sepsis-induced EndMT and cardiac remodeling by facilitating mitophagy process via PINK1/Parkin signaling [J]. Int Immunopharmacol, 2025, 164: 115377. DOI: 10.1016/j.intimp.2025.115377.

[44] Pesta D. Mitochondrial density in skeletal and cardiac muscle[J]. Mitochondrion, 2024, 75: 101838. DOI: 10.1016/j.mito.2023.101838.

[45] Kang X F, Lu X L, Bi C F, et al. Xuebijing injection protects sepsis induced myocardial injury by mediating TLR4/NF-κB/IKKα and JAK2/STAT3 signaling pathways[J]. Aging, 2023, 15(16): 8501-8517. DOI: 10.18632/aging.204990.

基金资助

湖北省中医药管理局中医药科研项目(ZY2023F088)

中华慈善总会“英才科研基金”临床公益项目(ZKL-KY-2024-037)

湖北医药学院襄阳市第一人民医院科技创新项目(XYY2025SD21)