Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Role of thrombospondin-1 in high-salt–induced mesenteric artery endothelial impairment in rats

Acta Pharmacologica Sinica volume 45, pages 545–557 (2024)Cite this article

Abstract

The matrix glycoprotein thrombospondin-1 (THBS1) modulates nitric oxide (NO) signaling in endothelial cells. A high-salt diet induces deficiencies of NO production and bioavailability, thereby leading to endothelial dysfunction. In this study we investigated the changes of THBS1 expression and its pathological role in the dysfunction of mesenteric artery endothelial cells (MAECs) induced by a high-salt diet. Wild-type rats, and wild-type and Thbs1−/− mice were fed chow containing 8% w/w NaCl for 4 weeks. We showed that a high salt diet significantly increased THBS1 expression and secretion in plasma and MAECs, and damaged endothelium-dependent vasodilation of mesenteric resistance arteries in wild-type animals, but not in Thbs1−/− mice. In rat MAECs, we demonstrated that a high salt environment (10–40 mM) dose-dependently increased THBS1 expression accompanied by suppressed endothelial nitric oxide synthase (eNOS) and phospho-eNOS S1177 production as well as NO release. Blockade of transforming growth factor-β1 (TGF-β1) activity by a TGF-β1 inhibitor SB 431542 reversed THBS1 up-regulation, rescued the eNOS decrease, enhanced phospho-eNOS S1177 expression, and inhibited Smad4 translocation to the nucleus. By conducting dual-luciferase reporter experiments in HEK293T cells, we demonstrated that Smad4, a transcription promoter, upregulated Thbs1 transcription. We conclude that THBS1 contributes to endothelial dysfunction in a high-salt environment and may be a potential target for treatment of high-salt-induced endothelium dysfunction.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: THBS1 is highly expressed in mesenteric artery endothelial cells collected from rats exposed to a high-salt diet.
Fig. 2: Effects of a high-salt diet and THBS1 on endothelium-dependent relaxation in the rat mesenteric resistant artery (MRA).
Fig. 3: Effects of a high-salt diet on endothelium-dependent relaxation in the mesenteric resistant artery (MRA) of wild-type and Thbs1−/− mice.
Fig. 4: Effects of high-salt environment on THBS1, p-eNOS, and eNOS expression and THBS1 secretion in rat mesenteric artery endothelial cells (MAECs).
Fig. 5: Effects of THBS1 on eNOS expression and NO production in rat mesenteric artery endothelial cells (MAECs).
Fig. 6: Effects of THBS1 knockdown on p-eNOS and eNOS expression in rat mesenteric artery endothelial cells cultured in a high-salt medium.
Fig. 7: Effects of TGF-β1 inhibition on p-eNOS, eNOS, THBS1, and downstream protein expression in rat mesenteric artery endothelial cells (MAECs).
Fig. 8: Effects of high-salt medium with or without SB 431542 on Smad4 translocation and effects of Smad4 siRNA on Smad4, THBS1, p-eNOS, and eNOS expression in rat mesenteric artery endothelial cells (MAECs).

Similar content being viewed by others

References

  1. Pilic L, Pedlar CR, Mavrommatis Y. Salt-sensitive hypertension: mechanisms and effects of dietary and other lifestyle factors. Nutr Rev. 2016;74:645–58.

    Article  PubMed  Google Scholar 

  2. Rust P, Ekmekcioglu C. Impact of salt intake on the pathogenesis and treatment of hypertension. Adv Exp Med Biol. 2017;956:61–84.

    Article  PubMed  Google Scholar 

  3. Feng W, Dell’Italia LJ, Sanders PW. Novel paradigms of salt and hypertension. J Am Soc Nephrol. 2017;28:1362–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Kozina N, Mihaljevic Z, Loncar MB, Mihalj M, Misir M, Radmilovic MD, et al. Impact of high salt diet on cerebral vascular function and stroke in Tff3-/-/C57BL/6N knockout and WT (C57BL/6N) control mice. Int J Mol Sci. 2019;20:1–25.

    Article  CAS  Google Scholar 

  5. Cosic A, Jukic I, Stupin A, Mihalj M, Mihaljevic Z, Novak S, et al. Attenuated flow-induced dilatation of middle cerebral arteries is related to increased vascular oxidative stress in rats on a short-term high salt diet. J Physiol. 2016;594:4917–31.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Raffai G, Durand MJ, Lombard JH. Acute and chronic angiotensin-(1-7) restores vasodilation and reduces oxidative stress in mesenteric arteries of salt-fed rats. Am J Physiol Heart Circ Physiol. 2011;301:H1341–1352.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Rogers NM, Sharifi-Sanjani M, Yao M, Ghimire K, Bienes-Martinez R, Mutchler SM, et al. TSP1-CD47 signaling is upregulated in clinical pulmonary hypertension and contributes to pulmonary arterial vasculopathy and dysfunction. Cardiovasc Res. 2017;113:15–29.

    Article  CAS  PubMed  Google Scholar 

  8. Shimoda LA, Kuebler WM. ‘Hypoxio-spondin’: thrombospondin and its emerging role in pulmonary hypertension. Cardiovasc Res. 2016;109:1–3.

    Article  CAS  PubMed  Google Scholar 

  9. Buda V, Andor M, Cristescu C, Tomescu MC, Muntean DM, Baibata DE, et al. Thrombospondin-1 serum levels in hypertensive patients with endothelial dysfunction after one year of treatment with perindopril. Drug Des Devel Ther. 2019;13:3515–26.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Matsuo Y, Tanaka M, Yamakage H, Sasaki Y, Muranaka K, Hata H, et al. Thrombospondin 1 as a novel biological marker of obesity and metabolic syndrome. Metabolism. 2015;64:1490–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Rogers NM, Ghimire K, Calzada MJ, Isenberg JS. Matricellular protein thrombospondin-1 in pulmonary hypertension: multiple pathways to disease. Cardiovasc Res. 2017;113:858–68.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Motegi K, Harada K, Ohe G, Jones SJ, Ellis IR, Crouch DH, et al. Differential involvement of TGF-beta1 in mediating the motogenic effects of TSP-1 on endothelial cells, fibroblasts and oral tumour cells. Exp Cell Res. 2008;314:2323–33.

    Article  CAS  PubMed  Google Scholar 

  13. Yafai Y, Eichler W, Iandiev I, Unterlauft JD, Jochmann C, Wiedemann P, et al. Thrombospondin-1 is produced by retinal glial cells and inhibits the growth of vascular endothelial cells. Ophthalmic Res. 2014;52:81–88.

    Article  CAS  PubMed  Google Scholar 

  14. Zak S, Treven J, Nash N, Gutierrez LS. Lack of thrombospondin-1 increases angiogenesis in a model of chronic inflammatory bowel disease. Int J Colorectal Dis. 2008;23:297–304.

    Article  PubMed  Google Scholar 

  15. Venkatraman L, Tucker-Kellogg L. The CD47-binding peptide of thrombospondin-1 induces defenestration of liver sinusoidal endothelial cells. Liver Int. 2013;33:1386–97.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Bauer EM, Qin Y, Miller TW, Bandle RW, Csanyi G, Pagano PJ, et al. Thrombospondin-1 supports blood pressure by limiting eNOS activation and endothelial-dependent vasorelaxation. Cardiovasc Res. 2010;88:471–81.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Kim J, Lee KS, Kim JH, Lee DK, Park M, Choi S, et al. Aspirin prevents TNF-α-induced endothelial cell dysfunction by regulating the NF-κB-dependent miR-155/eNOS pathway: Role of a miR-155/eNOS axis in preeclampsia. Free Radic Biol Med. 2017;104:185–98.

    Article  CAS  PubMed  Google Scholar 

  18. Sessa WC. eNOS at a glance. J Cell Sci. 2004;117:2427–9.

    Article  CAS  PubMed  Google Scholar 

  19. Feletou M, Kohler R, Vanhoutte PM. Nitric oxide: orchestrator of endothelium-dependent responses. Ann Med. 2012;44:694–716.

    Article  CAS  PubMed  Google Scholar 

  20. Qian J, Fulton D. Post-translational regulation of endothelial nitric oxide synthase in vascular endothelium. Front Physiol. 2013;4:1–11.

    Article  CAS  Google Scholar 

  21. Chen Z, S DSO, Zimnicka AM, Jiang Y, Sharma T, Chen S, et al. Reciprocal regulation of eNOS and caveolin-1 functions in endothelial cells. Mol Biol Cell. 2018;29:1190–202.

    Article  PubMed  PubMed Central  Google Scholar 

  22. Qin Y, Dong T, Jiang W, Ding W, Zhan T, Du J, et al. iTRAQ-based proteomics reveals serum protein changes in hypertensive rats induced by a high-salt diet. EXCLI J. 2020;19:1496–511.

    PubMed  PubMed Central  Google Scholar 

  23. Jiang W, Ye L, Yang Y, Wang P, Pan W, Du J, et al. TRPP2 associates with STIM1 to regulate cerebral vasoconstriction and enhance high salt intake-induced hypertensive cerebrovascular spasm. Hypertens Res. 2019;42:1894–904.

    Article  CAS  PubMed  Google Scholar 

  24. Guo J, Zhao R, Zhou M, Li J, Yao X, Du J, et al. TRPP2 and STIM1 form a microdomain to regulate store-operated Ca2+ entry and blood vessel tone. Cell Commun Signal. 2020;18:1–16.

    Article  Google Scholar 

  25. Shen B, Cheng KT, Leung YK, Kwok YC, Kwan HY, Wong CO, et al. Epinephrine-induced Ca2+ influx in vascular endothelial cells is mediated by CNGA2 channels. J Mol Cell Cardiol. 2008;45:437–45.

    Article  CAS  PubMed  Google Scholar 

  26. Suh SH, Vennekens R, Manolopoulos VG, Freichel M, Schweig U, Prenen J, et al. Characterisation of explanted endothelial cells from mouse aorta: electrophysiology and Ca2+ signalling. Pflug Arch. 1999;438:612–20.

    CAS  Google Scholar 

  27. Xin C, Ren S, Eberhardt W, Pfeilschifter J, Huwiler A. The immunomodulator FTY720 and its phosphorylated derivative activate the Smad signalling cascade and upregulate connective tissue growth factor and collagen type IV expression in renal mesangial cells. Br J Pharmacol. 2006;147:164–74.

    Article  CAS  PubMed  Google Scholar 

  28. Li W, Yong J, Xu Y, Wang Y, Zhang Y, Ren H, et al. Glutathione depletion and dual-model oxygen balance disruption for photodynamic therapy enhancement. Colloids Surf B Biointerf. 2019;183:1–10.

    Article  Google Scholar 

  29. Lee WK, Choi JK, Cha SH. Co-localization and interaction of human organic anion transporter 4 with caveolin-1 in primary cultured human placental trophoblasts. Exp Mol Med. 2008;40:505–13.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Tang EH, Feletou M, Huang Y, Man RY, Vanhoutte PM. Acetylcholine and sodium nitroprusside cause long-term inhibition of EDCF-mediated contractions. Am J Physiol Heart Circ Physiol. 2005;289:H2434–2440.

    Article  CAS  PubMed  Google Scholar 

  31. Castiglione RC, Barros C, Boa BCS, De Souza M, Bouskela E. Effects of selenium in the microcirculation of fructose-fed hamsters. J Vasc Res. 2018;55:203–9.

    Article  CAS  PubMed  Google Scholar 

  32. Daubon T, Leon C, Clarke K, Andrique L, Salabert L, Darbo E, et al. Deciphering the complex role of thrombospondin-1 in glioblastoma development. Nat Commun. 2019;10:1–15.

    Article  CAS  Google Scholar 

  33. Dennler S, Itoh S, Vivien D, ten Dijke P, Huet S, Gauthier JM. Direct binding of Smad3 and Smad4 to critical TGF β-inducible elements in the promoter of human plasminogen activator inhibitor-type 1 gene. EMBO J. 1998;17:3091–100.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Takekawa M, Tatebayashi K, Itoh F, Adachi M, Imai K, Saito H. Smad-dependent GADD45β expression mediates delayed activation of p38 MAP kinase by TGF-β. EMBO J. 2002;21:6473–82.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. von Toerne C, Huth C, de Las Heras Gala T, Kronenberg F, Herder C, Koenig W, et al. MASP1, THBS1, GPLD1 and ApoA-IV are novel biomarkers associated with prediabetes: the KORA F4 study. Diabetologia. 2016;59:1882–92.

    Article  Google Scholar 

  36. Kong P, Gonzalez-Quesada C, Li N, Cavalera M, Lee DW, Frangogiannis NG. Thrombospondin-1 regulates adiposity and metabolic dysfunction in diet-induced obesity enhancing adipose inflammation and stimulating adipocyte proliferation. Am J Physiol Endocrinol Metab. 2013;305:E439–450.

    Article  PubMed  PubMed Central  Google Scholar 

  37. Olerud J, Mokhtari D, Johansson M, Christoffersson G, Lawler J, Welsh N, et al. Thrombospondin-1: an islet endothelial cell signal of importance for β-cell function. Diabetes. 2011;60:1946–54.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Kong P, Cavalera M, Frangogiannis NG. The role of thrombospondin (TSP)-1 in obesity and diabetes. Adipocyte. 2014;3:81–84.

    Article  CAS  PubMed  Google Scholar 

  39. Nurkiewicz TR, Boegehold MA. High salt intake reduces endothelium-dependent dilation of mouse arterioles via superoxide anion generated from nitric oxide synthase. Am J Physiol Regul Integr Comp Physiol. 2007;292:R1550–1556.

    Article  CAS  PubMed  Google Scholar 

  40. Maruhashi T, Kihara Y, Higashi Y. Assessment of endothelium-independent vasodilation: from methodology to clinical perspectives. J Hypertens. 2018;36:1460–7.

    Article  CAS  PubMed  Google Scholar 

  41. Yamamoto E, Kataoka K, Shintaku H, Yamashita T, Tokutomi Y, Dong YF, et al. Novel mechanism and role of angiotensin II induced vascular endothelial injury in hypertensive diastolic heart failure. Arterioscler Thromb Vasc Biol. 2007;27:2569–75.

    Article  CAS  PubMed  Google Scholar 

  42. Adejare A, Oloyo A, Anigbogu C, Jaja S. L-arginine supplementation increased only endothelium-dependent relaxation in Sprague-Dawley rats fed a high-salt diet by enhancing abdominal aorta endothelial nitric oxide synthase gene expression. Clin Med Insights Cardiol. 2020;14:1–9.

    Article  Google Scholar 

  43. Bazzazi H, Zhang Y, Jafarnejad M, Isenberg JS, Annex BH, Popel AS. Computer simulation of TSP1 inhibition of VEGF-Akt-eNOS: an angiogenesis triple threat. Front Physiol. 2018;9:1–14.

    Article  Google Scholar 

  44. Schwarte-Waldhoff I, Volpert OV, Bouck NP, Sipos B, Hahn SA, Klein-Scory S, et al. Smad4/DPC4-mediated tumor suppression through suppression of angiogenesis. Proc Natl Acad Sci USA. 2000;97:9624–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We would like to express our gratitude to Yun-xia Lu, Ph.D (and the technicians), in the Comprehensive Experiment Center of Basic Medical Sciences, Anhui Medical University, for the support of these facilities, and the Center for Scientific Research of Anhui Medical University for valuable assistance with this study. The study was supported by the National Natural Science Foundation of China Regional Innovation and Development Joint Fund (grant No. U22A20272), National Natural Science Foundation of China (grant No. 82204381), Natural Science Foundation of Anhui Province (grant No. 2108085MH260 and 2208085QH277), Anhui Province Key Research and Development Project (grant No. 201904a07020032), and Applied Medicine Research Project of Hefei Municipal Health Commission (grant No. 2019-172-2-15).

Author information

Author notes

  1. These authors contributed equally: Fang-fang Xu, Fan Zheng, Ye Chen, Yang Wang

Authors and Affiliations

  1. School of Basic Medical Sciences, Anhui Medical University, Hefei, 230032, China

    Fang-fang Xu, Fan Zheng, Shao-bo Ma, Weng Ding, Le-sha Zhang, Ji-zheng Guo & Bing Shen

  2. Department of Pharmacy, The First Affiliated Hospital of USTC, Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei, 230001, China

    Fang-fang Xu

  3. Department of Pathology, The First Affiliated Hospital of Anhui Medical University, Hefei, 230032, China

    Ye Chen

  4. Department of Otolaryngology-Head and Neck Surgery, Lu’an People’s Hospital, Lu’an Affiliated Hospital of Anhui Medical University, Lu’an, 237000, China

    Yang Wang

  5. School of Pharmaceutical Science, Kunming Medical University, Kunming, 650500, China

    Chang-Bo Zheng

Authors
  1. Fang-fang Xu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Fan Zheng
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ye Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Yang Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Shao-bo Ma
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Weng Ding
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Le-sha Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Ji-zheng Guo
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Chang-Bo Zheng
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Bing Shen
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

FFX, FZ, CBZ and BS conceived and designed the study. FFX, FZ, YC and YW conducted major experiments and analyzed the data. SBM, WD, YC, LSZ and JZG performed primary cell culture, transfection and confocal experiments. BS drafted the manuscript. BS, YW and CBZ supervised the whole study and wrote the manuscript. All authors edited the manuscript and approved the final manuscript.

Corresponding authors

Correspondence to Chang-Bo Zheng or Bing Shen.

Ethics declarations

Competing interests

The authors declare no competing interests.

Supplementary information

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Xu, Ff., Zheng, F., Chen, Y. et al. Role of thrombospondin-1 in high-salt–induced mesenteric artery endothelial impairment in rats. Acta Pharmacol Sin 45, 545–557 (2024). https://doi.org/10.1038/s41401-023-01181-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41401-023-01181-9

Keywords

Search

Advanced search

Quick links