The correlation analysis between the NLRP3 inflammasome and inflammatory reaction in venous ulcer of the lower extremity_CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

Authors：

WANG Tao ¹ , FAN Longhua ^1,2 , CHEN Bin ^1,2 ,  LIU Jianjun ¹ , LI Xu ¹ , TAO Yue ¹ , WANG Xiaojun ¹ , LI Limeng ¹

1. Department of Vascular Surgery, Qingpu Branch of Zhongshan Hospital, Fudan University, Shanghai 201700, P. R. China;
2. Department of Vascular Surgery, Zhongshan Hospital, Fudan University, Shanghai 200032, P. R. China;

Corresponding author：

LIU Jianjun, Email: liu1921@sina.com

Keywords：

nucleotide binding oligomerization domain like receptor protein 3 inflammasome; venous ulcer of lower extremity; inflammatory reactory

DOI：

10.7507/1007-9424.202101029

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Objective To investigate the relationship between the nucleotide binding oligomerization domain like receptor protein 3 (NLRP3) inflammasome and inflammatory reaction of venous ulcer of lower extremity.Methods Twenty-four patients with active venous ulcer of lower extremity (active ulcer group), 24 patients with non exudative venous ulcer of lower extremity as positive control (non-active ulcer group), and 24 patients with traumatic wound as negative control (traumatic-wound group) were enrolled. The clinical data of the three groups were compared, the tissue samples around the wound were harvested, and the expressions of NLRP3 protein were detected by immunohistochemistry among the three groups. Enzyme linked immunosorbent assay (ELISA) was used to detect the IL-1β and IL-18 protein levels, RT-PCR was used to detect the mRNA expressions of apoptosis associated speck like protein containing CARD (ASC), caspase-1, c-Jun N-terminal kinase (JNK), p38, nuclear factor (NF)-κB p65 and NF-κB inhibitor alpha (NF-κB IkBα), and Western blotting was performed to evaluate the level of NLRP3 inflammasome in wound tissues.Results The inflammatory response in the non-active ulcer group and trauma-wound group were milder than that in the active ulcer group. The levels of IL-1β and IL-18 proteins in the active ulcer group were higher than those in the non-active ulcer group and the traumatic-wound group [IL-1β: (146.621±11.597) ng/L vs. (80.967±14.213) ng/L vs. (84.962±19.484) ng/L, F=136.200, P<0.001; IL-18: (119.814±12.788) ng/L vs. (72.899±17.220) ng/L vs. (48.131±10.407) ng/L, F=167.910, P<0.001]. The results of RT-PCR showed that the mRNA expressions of ASC [(0.030±0.012) ng/L vs. (0.021±0.005) ng/L vs. (0.016±0.004) ng/L, F=18.106, P<0.001], caspase-1 [(0.054±0.012) ng/L vs. (0.013±0.009) ng/L vs. (0.018±0.006) ng/L, F=130.372, P<0.001], NF-κB p65 [(0.093±0.015) ng/L vs. (0.038±0.013) ng/L vs. (0.043±0.014) ng/L, F=110.950, P<0.001], NF-κB IkB-α [(0.085±0.015) ng/L vs. (0.078±0.015) ng/L vs. (0.041±0.016) ng/L, F=53.070, P<0.001], and JNK [(0.075±0.018) ng/L vs. (0.042±0.013) ng/L vs. (0.039±0.014) ng/L, F=41.271, P<0.001] in the wound tissues of the active ulcer group were higher than those in the non-active ulcer group and the traumatic-wound group. And the mRNA expression of p38 in the wound tissues of the active ulcer group was lower than that in the non-active ulcer group [(0.050±0.008) ng/L vs. (0.064±0.014) ng/L, P<0.05]. The result of Western blotting showed that the relative expression level of NLRP3 protein in the wound tissues of the active ulcer group was higher than that in the trauma-wound group and non-active ulcer group (0.767±0.272 vs. 0.605±0.212 vs. 0.556±0.183, F=4.804, P=0.012).Conclusion NLRP3 inflammasome is closely related to the wound in venous ulcer of lower extremity and provides a new target to the therapy of venous ulcer of lower extremity.

Citation： WANG Tao, FAN Longhua, CHEN Bin, LIU Jianjun, LI Xu, TAO Yue, WANG Xiaojun, LI Limeng. The correlation analysis between the NLRP3 inflammasome and inflammatory reaction in venous ulcer of the lower extremity. CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY, 2021, 28(10): 1287-1293. doi: 10.7507/1007-9424.202101029 Copy

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3.	Crawford JM, Lal BK, Durán WN, et al. Pathophysiology of venous ulceration. J Vasc Surg Venous Lymphat Disord, 2017, 5(4): 596-605.
4.	曾彦彰, 杨轶, 赵纪春. 大隐静脉结扎剥脱联合泡沫硬化剂与单纯泡沫硬化剂治疗下肢静脉性溃疡的疗效比较. 中国普外基础与临床杂志, 2018, 25(3): 312-316.
5.	刘国全, 王雁南, 刘天娇. 下肢静脉性溃疡差异表达基因挖掘及目标基因筛选. 中国普外基础与临床杂志, 2021, 28(4): 445-451.
6.	Coleridge Smith PD, Thomas P, Scurr JH, et al. Causes of venous ulceration: a new hypothesis. Br Med J (Clin Res Ed), 1988, 296(6638): 1726-1727.
7.	Signorelli SS, Malaponte MG, Di Pino L, et al. Venous stasis causes release of interleukin 1beta (IL-1beta), interleukin 6 (IL-6) and tumor necrosis factor alpha (TNFalpha) by monocyte-macrophage. Clin Hemorheol Microcirc, 2000, 22(4): 311-316.
8.	潘晓燕, 龚小花, 沈飞霞, 等. 高糖环境对HBZY-1大鼠肾小球系膜细胞IL-18、IL-1β、IL-6、TNF-α表达的影响. 中国中西医结合肾病杂志, 2012, 13(6): 519-521.
9.	Zhu X, Zhang HW, Chen HN, et al. Perivascular adipose tissue dysfunction aggravates adventitial remodeling in obese mini pigs via NLRP3 inflammasome/IL-1 signaling pathway. Acta Pharmacol Sin, 2019, 40(1): 46-54.
10.	Starobova H, Nadar EI, Vetter I. The NLRP3 inflammasome: Role and therapeutic potential in pain treatment. Front Physiol, 2020, 11: 1016.
11.	任改艳, 黄剑林. NLRP3炎症小体与炎症性肠病的研究进展. 中国免疫学杂志, 2019, 35(13): 1662-1665.
12.	童晓鹏, 张群辉. 巨噬细胞炎症反应的内外控制. 湖北民族学院学报(医学版), 2015, 32(3): 70-73.
13.	Liu YC, Margolis DJ, Isseroff RR. Does inflammation have a role in the pathogenesis of venous ulcers? A critical review of the evidence. J Invest Dermatol, 2011, 131(4): 818-827.
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15.	Boniakowski AE, Kimball AS, Jacobs BN, et al. Macrophage-mediated inflammation in normal and diabetic wound healing. J Immunol, 2017, 199(1): 17-24.
16.	Liu W, Yu M, Xie D, et al. Melatonin-stimulated MSC-derived exosomes improve diabetic wound healing through regulating macrophage M1 and M2 polarization by targeting the PTEN/AKT pathway. Stem Cell Res Ther, 2020, 11(1): 259.
17.	Cavassan NRV, Camargo CC, de Pontes LG, et al. Correlation between chronic venous ulcer exudate proteins and clinical profile: A cross-sectional study. J Proteomics, 2019, 192: 280-290.
18.	Mirza R, Koh TJ. Dysregulation of monocyte/macrophage phenotype in wounds of diabetic mice. Cytokine, 2011, 56(2): 256-264.
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20.	Fang R, Uchiyama R, Sakai S, et al. ASC and NLRP3 maintain innate immune homeostasis in the airway through an inflammasome-independent mechanism. Mucosal Immunol, 2019, 12(5): 1092-1103.
21.	Wang Z, Meng S, Cao L, et al. Critical role of NLRP3-caspase-1 pathway in age-dependent isoflurane-induced microglial inflammatory response and cognitive impairment. J Neuroinflammation, 2018, 15(1): 109.
22.	Li Z, Guo J, Bi L. Role of the NLRP3 inflammasome in autoimmune diseases. Biomed Pharmacother, 2020, 130: 110542.
23.	Wang F, Huang L, Peng ZZ, et al. Losartan inhibits LPS+ATP-induced IL-1beta secretion from mouse primary macrophages by suppressing NALP3 inflammasome. Pharmazie, 2014, 69(9): 680-684.
24.	Mulero MC, Huxford T, Ghosh G. NF-κB, IκB, and IKK: Integral components of immune system signaling. Adv Exp Med Biol, 2019, 1172: 207-226.
25.	He R, Li Y, Han C, et al. L-Fucose ameliorates DSS-induced acute colitis via inhibiting macrophage M1 polarization and inhibiting NLRP3 inflammasome and NF-kB activation. Int Immunopharmacol, 2019, 73: 379-388.

1. Lim CS, Baruah M, Bahia SS. Diagnosis and management of venous leg ulcers. BMJ, 2018, 362: k3115.
2. Polak MW, Siudut J, Plens K, et al. Prothrombotic clot properties can predict venous ulcers in patients following deep vein thrombosis: a cohort study. J Thromb Thrombolysis, 2019, 48(4): 603-609.
3. Crawford JM, Lal BK, Durán WN, et al. Pathophysiology of venous ulceration. J Vasc Surg Venous Lymphat Disord, 2017, 5(4): 596-605.
4. 曾彦彰, 杨轶, 赵纪春. 大隐静脉结扎剥脱联合泡沫硬化剂与单纯泡沫硬化剂治疗下肢静脉性溃疡的疗效比较. 中国普外基础与临床杂志, 2018, 25(3): 312-316.
5. 刘国全, 王雁南, 刘天娇. 下肢静脉性溃疡差异表达基因挖掘及目标基因筛选. 中国普外基础与临床杂志, 2021, 28(4): 445-451.
6. Coleridge Smith PD, Thomas P, Scurr JH, et al. Causes of venous ulceration: a new hypothesis. Br Med J (Clin Res Ed), 1988, 296(6638): 1726-1727.
7. Signorelli SS, Malaponte MG, Di Pino L, et al. Venous stasis causes release of interleukin 1beta (IL-1beta), interleukin 6 (IL-6) and tumor necrosis factor alpha (TNFalpha) by monocyte-macrophage. Clin Hemorheol Microcirc, 2000, 22(4): 311-316.
8. 潘晓燕, 龚小花, 沈飞霞, 等. 高糖环境对HBZY-1大鼠肾小球系膜细胞IL-18、IL-1β、IL-6、TNF-α表达的影响. 中国中西医结合肾病杂志, 2012, 13(6): 519-521.
9. Zhu X, Zhang HW, Chen HN, et al. Perivascular adipose tissue dysfunction aggravates adventitial remodeling in obese mini pigs via NLRP3 inflammasome/IL-1 signaling pathway. Acta Pharmacol Sin, 2019, 40(1): 46-54.
10. Starobova H, Nadar EI, Vetter I. The NLRP3 inflammasome: Role and therapeutic potential in pain treatment. Front Physiol, 2020, 11: 1016.
11. 任改艳, 黄剑林. NLRP3炎症小体与炎症性肠病的研究进展. 中国免疫学杂志, 2019, 35(13): 1662-1665.
12. 童晓鹏, 张群辉. 巨噬细胞炎症反应的内外控制. 湖北民族学院学报(医学版), 2015, 32(3): 70-73.
13. Liu YC, Margolis DJ, Isseroff RR. Does inflammation have a role in the pathogenesis of venous ulcers? A critical review of the evidence. J Invest Dermatol, 2011, 131(4): 818-827.
14. Liu D, Zhang L, Li T, et al. Negative-pressure wound therapy enhances local inflammatory responses in acute infected soft-tissue wound. Cell Biochem Biophys, 2014, 70(1): 539-547.
15. Boniakowski AE, Kimball AS, Jacobs BN, et al. Macrophage-mediated inflammation in normal and diabetic wound healing. J Immunol, 2017, 199(1): 17-24.
16. Liu W, Yu M, Xie D, et al. Melatonin-stimulated MSC-derived exosomes improve diabetic wound healing through regulating macrophage M1 and M2 polarization by targeting the PTEN/AKT pathway. Stem Cell Res Ther, 2020, 11(1): 259.
17. Cavassan NRV, Camargo CC, de Pontes LG, et al. Correlation between chronic venous ulcer exudate proteins and clinical profile: A cross-sectional study. J Proteomics, 2019, 192: 280-290.
18. Mirza R, Koh TJ. Dysregulation of monocyte/macrophage phenotype in wounds of diabetic mice. Cytokine, 2011, 56(2): 256-264.
19. Kroeze KL, Vink L, de Boer EM, et al. Simple wound exudate collection method identifies bioactive cytokines and chemokines in (arterio) venous ulcers. Wound Repair Regen, 2012, 20(3): 294-303.
20. Fang R, Uchiyama R, Sakai S, et al. ASC and NLRP3 maintain innate immune homeostasis in the airway through an inflammasome-independent mechanism. Mucosal Immunol, 2019, 12(5): 1092-1103.
21. Wang Z, Meng S, Cao L, et al. Critical role of NLRP3-caspase-1 pathway in age-dependent isoflurane-induced microglial inflammatory response and cognitive impairment. J Neuroinflammation, 2018, 15(1): 109.
22. Li Z, Guo J, Bi L. Role of the NLRP3 inflammasome in autoimmune diseases. Biomed Pharmacother, 2020, 130: 110542.
23. Wang F, Huang L, Peng ZZ, et al. Losartan inhibits LPS+ATP-induced IL-1beta secretion from mouse primary macrophages by suppressing NALP3 inflammasome. Pharmazie, 2014, 69(9): 680-684.
24. Mulero MC, Huxford T, Ghosh G. NF-κB, IκB, and IKK: Integral components of immune system signaling. Adv Exp Med Biol, 2019, 1172: 207-226.
25. He R, Li Y, Han C, et al. L-Fucose ameliorates DSS-induced acute colitis via inhibiting macrophage M1 polarization and inhibiting NLRP3 inflammasome and NF-kB activation. Int Immunopharmacol, 2019, 73: 379-388.

CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

The correlation analysis between the NLRP3 inflammasome and inflammatory reaction in venous ulcer of the lower extremity

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