Effect and mechanism of sADAM33 on the proliferation and lumen formation of airway vascular endothelial cells_Chinese Journal of Respiratory and Critical Care Medicine

Authors：

HU Xiaojuan , MANGUNNU Shi Nabek , HU Xin , YU Minjie ,  YAN Fang

The first ward of Respiratory and respiratory Critical Care Center, the First Affiliated Hospital of Xinjiang Medical University, Urumuqi Xinjiang 810000, P.R.China;

Corresponding author：

YAN Fang, Email: 58078817@qq.com

Keywords：

Asthma; vascular smooth muscle cells; vascular endothelial cell; adisintegrin and metalloproteinase-33; airway vascular remodeling

DOI：

10.7507/1671-6205.202404057

Video：

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Objective To investigate the effect of ADAM33 gene silencing in VSMCs on the proliferation and lumen formation of airway vascular endothelial cells (VECs) in a co-culture system and the possible regulatory mechanism. Methods The Human aortic smooth muscle cells (HASMCs) and human pulmonary microvascular endothelial cells (HPMECs) were used to construct a cell co-culture system. ADAM33 gene expression was silenced by lentivirus transfection technique, and the subjects were divided into endothelial cell blank group, co-culture group, co-culture +shRNA negative control group, and co-culture + ADAM33-SHRNA group. The expressions of sADAM33, VEGFA,VEGER2, ang-1 and ang-2 in co-culture system were detected by ELISA. The proliferation and lumen formation of HPMECs were observed by CCK-8 and Transwell experiments. The protein expression of Tie2, PI3K, Akt, and mTOR key molecules in Tie2/PI3K/Akt/mTOR signaling pathway and the phosphorylation levels of AKT and mTOR were detected by Western-blotting method. Results ① Compared with the co-culture group (0.851±0.036) and the co-culture + shRNA negative control group (0.828±0.047), the OD value of the co-culture + ADAM33shRNA group (0.699±0.038) was significantly decreased (P<0.05). ② Compared with the co-culture group (159.169±15.740) and the co-culture +shRNA negative control group (157.357±21.612), the tube length of the co-culture +ADAM33shRNA group (120.812±2.791) was also significantly decreased (P<0.05). ③ After ADAM33 gene expression of HASMCs was silted in co-culture system, the expression levels of VEGFA, VEGFR2, ang-1 and ang-2 were significantly decreased (P<0.05), while the expression levels of Tie2, PI3K, P-Akt and P-mtor were decreased (P<0.05). Conclusions Silencing the expression of the ADAM33 gene could reduce the release of sADAM33 from the membrane of the airway VSMCs, regulate the proliferation and lumen formation of airway VECs by reducing the expression of VEGF/VEGFR and inhibiting the activities of the Tie2/PI3K/Akt/mTOR signaling pathways,and then participate in airway vascular remodeling in asthma.

Citation： HU Xiaojuan, MANGUNNU Shi Nabek, HU Xin, YU Minjie, YAN Fang. Effect and mechanism of sADAM33 on the proliferation and lumen formation of airway vascular endothelial cells. Chinese Journal of Respiratory and Critical Care Medicine, 2024, 23(8): 569-574. doi: 10.7507/1671-6205.202404057 Copy

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15.	Janulaityte I, Januskevicius A, Rimkunas A, et al. Asthmatic Eosinophils Alter the Gene Expression of Extracellular Matrix Proteins in Airway Smooth Muscle Cells and Pulmonary Fibroblasts. International Journal of Molecular Sciences, Int J Mol Sci, 2022, 23(8): 4086.
16.	Duan Y, Long J, Chen J, et al. Overexpression of soluble ADAM33 promotes a hypercontractile phenotype of the airway smooth muscle cell in rat. Exp Cell Res, 2016, 15;349(1): 109-118.
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18.	Davies ER, Kelly JF, Howarth PH, et al. Soluble ADAM33 initiates airway remodeling to promote susceptibility for allergic asthma in early life. JCI Insight, 2016, 1(11): e87632.
19.	Puxeddu I, Pang YY, Harvey A, et al. The soluble form of a disintegrin and metalloprotease 33 promotes angiogenesis: implications for airway remodeling in asthma. J Allergy Clin Immunol, 2008, 121(6): 1400-6, 1406. e1-4.
20.	Tripathi P, Awasthi S, Husain N, et al. Increased expression of ADAM33 protein in asthmatic patients as compared to non-asthmatic controls. Indian J Med Res, 2013, 137: 507-514.
21.	Tripathi P, Awasthi S, Gao P. ADAM metallopeptidase domain 33 (ADAM33): a promising target for asthma. Mediators Inflamm, 2014, 2014: 572025.
22.	Pan L, Liu C, Kong Y, Piao Z, Cheng B. Phentolamine inhibits angiogenesis in vitro: Suppression of proliferation migration and differentiation of human endothelial cells. Clin Hemorheol Microcirc, 2017, 65(1): 31-41.
23.	Karaman S, Leppänen VM, Alitalo K, et al. Vascular endothelial growth factor signaling in development and disease. Development, 2018, 145(14): dev151019.
24.	Tsai JL, Lee YM, Pan CY, Lee AY. The Novel VEGF121-VEGF165 Fusion Attenuates Angiogenesis and Drug Resistance via Targeting VEGFR2-HIF-1α-VEGF165/Lon Signaling Through PI3K-AKT-mTOR Pathway. Curr Cancer Drug Targets. 2016, 16(3): 275-286.
25.	王师英, 孙博云, 林江等. VEGFR2在血管新生与重塑中的调控作用. 中国老年学杂志, 2017, 37(19): 4924-4927.
26.	Bakakos P, Patentalakis G, Papi A. Vascular Biomarkers in Asthma and COPD. Curr Top Med Chem. 2016;16(14): 1599-1609.
27.	Yao S, Su C, Wu SH, et al. Aliskiren Improved the Endothelial Repair Capacity of Endothelial Progenitor Cells from Patients with Hypertension via the Tie2/PI3k/Akt/eNOS Signalling Pathway. Cardiol Res Pract. 2020, 2020: 6534512.
28.	Piotr Kobialka1 and Mariona Graupera. Revisiting PI3-kinase signalling in angiogenesis. Vasc Biol. 2019, 1(1): H125-H134.
29.	雷佳慧, 祝贺, 肖亚丽, 等. AMPK在支气管哮喘中的作用及其研究进展. 中国呼吸与危重监护杂志, 2022, 21(12): 893-898.

1. Guida G, Riccio AM. Immune induction of airway remodeling. Semin Immunol, 2019, 46: 101346.
2. Barbato A, Turato G, Baraldo S, et al. Epithelial damage and angiogenesis in the airways of children with asthma. Am Respir Crit Care Med 2006, 174(9): 975-981.
3. Holgate ST, Yang Y, Haitchi HM, et al. The genetics of asthma: ADAM33 as an example of a susceptibility gene. Proc Am Thorac Soc, 2006, 3(5): 440-443.
4. Banno A, Reddy AT, Lakshmi SP, et al. Bidirectional interaction of airway epithelial remodeling and inflammation in asthma. Clin Sci(Lond), 2020, 134(9): 1063-1079.
5. Dreymueller D, Pruessmeyer J, Groth E, et al. The Role of ADAM-mediated Shedding in Vascular Biology. Eur J Cell Biol, 2012, 91(6-7): 472-485.
6. Camoretti-Mercado B, Lockey RF. Airway smooth muscle pathophysiology in asthma. J Allergy Clin Immunol, 2021, 147(6): 1983-1995.
7. Dreymueller D, Uhlig S, Ludwig A. ADAM-family met alloproteinases in lung inflammation: potential therapeutic targets. Am J Physiol Lung Cell Mol Physiol, 2015, 308(4): L325-343.
8. 闫芳. ADAM33在哮喘气道血管重塑中的作用和分子机制. 新疆医科大学, 2023.
9. Breikaa RM, Denman K, Ueyama Y, et al. Loss of Jagged1 in mature endothelial cells causes vascular dysfunction with alterations in smooth muscle phenotypes. Vascul Pharmacol. 2022, 145: 107087.
10. Duan Y , Long J , Lin F , et al. Soluble ADAM33 determines mechanical behaviors of airway smooth muscle cells, 医用生物力学, 2013, 28(S1): 107-108.
11. D Amato M, Cecchi L, Annesi-Maesano I,et al. News on Climate Change, Air Pollution, and Allergic Triggers of Asthma. J Investig Allergol Clin Immunol, 2018, 28(2): 91-97.
12. Pinto LA, Stein RT, Kabesch M. Impact of genetics in childhood asthma. J Pediatr (Rio J), 2008, 84(4 Suppl): S68-S75.
13. Fehrenbach H, Wagner C, Wegmann M. Airway remodeling in asthma: what really matters. Cell Tissue Res, 2017, 367(3): 551-569.
14. Deng R, Zhao F, Zhong X. T1 polymorphism in a disintegrin and metalloproteinase 33 (ADAM33) gene may contribute to the risk of childhood asthma in Asians. Inflamm Res, 2017, 66(5): 413-424.
15. Janulaityte I, Januskevicius A, Rimkunas A, et al. Asthmatic Eosinophils Alter the Gene Expression of Extracellular Matrix Proteins in Airway Smooth Muscle Cells and Pulmonary Fibroblasts. International Journal of Molecular Sciences, Int J Mol Sci, 2022, 23(8): 4086.
16. Duan Y, Long J, Chen J, et al. Overexpression of soluble ADAM33 promotes a hypercontractile phenotype of the airway smooth muscle cell in rat. Exp Cell Res, 2016, 15;349(1): 109-118.
17. Yan F, Hao Y, Gong X, et al. Silencing a disintegrin and metalloproteinase-33 attenuates the proliferation of vascular smooth muscle cells via PI3K/AKT pathway: Implications in the pathogenesis of airway vascular remodeling. Mol Med Rep, 2021, 24(1): 502.
18. Davies ER, Kelly JF, Howarth PH, et al. Soluble ADAM33 initiates airway remodeling to promote susceptibility for allergic asthma in early life. JCI Insight, 2016, 1(11): e87632.
19. Puxeddu I, Pang YY, Harvey A, et al. The soluble form of a disintegrin and metalloprotease 33 promotes angiogenesis: implications for airway remodeling in asthma. J Allergy Clin Immunol, 2008, 121(6): 1400-6, 1406. e1-4.
20. Tripathi P, Awasthi S, Husain N, et al. Increased expression of ADAM33 protein in asthmatic patients as compared to non-asthmatic controls. Indian J Med Res, 2013, 137: 507-514.
21. Tripathi P, Awasthi S, Gao P. ADAM metallopeptidase domain 33 (ADAM33): a promising target for asthma. Mediators Inflamm, 2014, 2014: 572025.
22. Pan L, Liu C, Kong Y, Piao Z, Cheng B. Phentolamine inhibits angiogenesis in vitro: Suppression of proliferation migration and differentiation of human endothelial cells. Clin Hemorheol Microcirc, 2017, 65(1): 31-41.
23. Karaman S, Leppänen VM, Alitalo K, et al. Vascular endothelial growth factor signaling in development and disease. Development, 2018, 145(14): dev151019.
24. Tsai JL, Lee YM, Pan CY, Lee AY. The Novel VEGF121-VEGF165 Fusion Attenuates Angiogenesis and Drug Resistance via Targeting VEGFR2-HIF-1α-VEGF165/Lon Signaling Through PI3K-AKT-mTOR Pathway. Curr Cancer Drug Targets. 2016, 16(3): 275-286.
25. 王师英, 孙博云, 林江等. VEGFR2在血管新生与重塑中的调控作用. 中国老年学杂志, 2017, 37(19): 4924-4927.
26. Bakakos P, Patentalakis G, Papi A. Vascular Biomarkers in Asthma and COPD. Curr Top Med Chem. 2016;16(14): 1599-1609.
27. Yao S, Su C, Wu SH, et al. Aliskiren Improved the Endothelial Repair Capacity of Endothelial Progenitor Cells from Patients with Hypertension via the Tie2/PI3k/Akt/eNOS Signalling Pathway. Cardiol Res Pract. 2020, 2020: 6534512.
28. Piotr Kobialka1 and Mariona Graupera. Revisiting PI3-kinase signalling in angiogenesis. Vasc Biol. 2019, 1(1): H125-H134.
29. 雷佳慧, 祝贺, 肖亚丽, 等. AMPK在支气管哮喘中的作用及其研究进展. 中国呼吸与危重监护杂志, 2022, 21(12): 893-898.

Chinese Journal of Respiratory and Critical Care Medicine

Effect and mechanism of sADAM33 on the proliferation and lumen formation of airway vascular endothelial cells

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