Article | Published:

Preconditioned adipose-derived stem cells ameliorate cardiac fibrosis by regulating macrophage polarization in infarcted rat hearts through the PI3K/STAT3 pathway

Laboratory Investigation (2019) | Download Citation

Abstract

Stem cells can modify macrophage phenotypes; however, the mechanisms remain unclear. We investigated whether n-butylidenephthalide (BP) primed adipose-derived stem cells (ADSCs) attenuated cardiac fibrosis via regulating macrophage phenotype by a PI3K/STAT3-dependent pathway in postinfarcted rats. Male Wistar rats after coronary ligation were allocated to receive either intramyocardial injection of vehicle, ADSCs (1 × 106 cells), BP-preconditioned ADSCs, (BP + lithium)-preconditioned ADSCs, (BP + LY294002)-preconditioned ADSCs, and (BP + S3I-201)-preconditioned ADSCs. ADSCs were primed for 16 h before implantation. BP-pretreated ADSCs increased the cell viability compared with naive ADSCs in the in vitro experiments. Infarct sizes were similar among the infarcted groups at the acute and chronic stages of infarction. At day 3 after infarction, post-infarction was associated with increased M1 macrophage infiltration, which was inhibited by administering naive ADSCs. Compared with naive ADSCs, BP-preconditioned ADSCs provided a significant increase of Akt and STAT3 phosphorylation, STAT3 activity, STAT3 nuclear translocation, myocardial IL-10 levels, and the percentage of M2 macrophage infiltration. The effects of BP on M2 polarization were reversed by LY294002 or S3I-201. Furthermore, the phosphorylation of both Akt and STAT3 was abolished by LY294002, whereas Akt phosphorylation was not affected following the inhibition of STAT3. The addition of lithium did not have additional effects compared with BP alone. After 4 weeks of implantation, ADSCs remained in the myocardium, and reduced fibrosis and improved cardiac function. BP-preconditioned ADSCs provided superior cardioprotection, greater ADSC engraftment, and antifibrotic effects compared with naive ADSCs. These results suggest that BP-pretreated ADSCs polarize macrophages into M2 cells more efficiently than naive ADSCs via the PI3K/STAT3 pathway.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

References

  1. 1.

    Matsushita K, Dzau VJ. Mesenchymal stem cells in obesity: insights for translational applications. Lab Invest. 2017;97:1158–66.

  2. 2.

    Gordon S. Alternative activation of macrophages. Nat Rev Immunol. 2003;3:23–35.

  3. 3.

    Mantovani A, Sica A, Sozzani S, Allavena P, Vecchi A, Locati M. The chemokine system in diverse forms of macrophage activation and polarization. Trends Immunol. 2004;25:677–86.

  4. 4.

    Panizzi P, Swirski FK, Figueiredo JL, Waterman P, Sosnovik DE, Aikawa E, et al. Impaired infarct healing in atherosclerotic mice with Ly-6C(hi) monocytosis. J Am Coll Cardiol. 2010;55:1629–38.

  5. 5.

    Hart JR, Liao L, Yates JR 3rd, Vogt PK. Essential role of Stat3 in PI3K-induced oncogenic transformation. Proc Natl Acad Sci USA. 2011;108:13247–52.

  6. 6.

    Kral JB, Kuttke M, Schrottmaier WC, Birnecker B, Warszawska J, Wernig C, et al. Sustained PI3K activation exacerbates BLM-induced lung fibrosis via activation of pro-inflammatory and pro-fibrotic pathways. Sci Rep. 2016;6:23034.

  7. 7.

    Bai X, Yan Y, Song YH, Seidensticker M, Rabinovich B, Metzele R, et al. Both cultured and freshly isolated adipose tissue-derived stem cells enhance cardiac function after acute myocardial infarction. Eur Heart J. 2010;31:489–501.

  8. 8.

    Abumaree MH, Al Jumah MA, Kalionis B, Jawdat D, Al Khaldi A, Abomaray FM, et al. Human placental mesenchymal stem cells (pMSCs) play a role as immune suppressive cells by shifting macrophage differentiation from inflammatory M1 to anti-inflammatory M2 macrophages. Stem Cell Rev. 2013;9:620–41.

  9. 9.

    Maggini J, Mirkin G, Bognanni I, Holmberg J, Piazzón IM, Nepomnaschy I, et al. Mouse bone marrow-derived mesenchymal stromal cells turn activated macrophages into a regulatory-like profile. PLoS ONE. 2010;5:e9252.

  10. 10.

    Dayan V, Yannarelli G, Billia F, Filomeno P, Wang XH, Davies JE, et al. Mesenchymal stromal cells mediate a switch to alternatively activated monocytes/macrophages after acute myocardial infarction. Basic Res Cardiol. 2011;106:1299–310.

  11. 11.

    Ben-Mordechai T, Holbova R, Landa-Rouben N, Harel-Adar T, Feinberg MS, Abd Elrahman I, et al. Macrophage subpopulations are essential for infarct repair with and without stem cell therapy. J Am Coll Cardiol. 2013;62:1890–901.

  12. 12.

    Yi LZ, Liang YZ, Wu H, Yuan DL. The analysis of Radix Angelicae sinensis (Danggui). J Chromatogr A. 2009;1216:1991–2001.

  13. 13.

    Ying L, Si-Wang W, Hong-Hai T, Wei C. Simultaneous quantification of six main active constituents in Chinese Angelica by high-performance liquid chromatography with photodiode array detector. Pharmacogn Mag. 2013;9:114–9.

  14. 14.

    Xiang J, Pan J, Chen F, Zheng L, Chen Y, Zhang S, et al. L-3-n-butylphthalide improves cognitive impairment of APP/PS1 mice by BDNF/TrkB/PI3K/AKT pathway. Int J Clin Exp Med. 2014;7:1706–13.

  15. 15.

    Lee TM, Harn HJ, Chiou TW, Chuang MH, Chen CH, Lin PC, et al. Targeting the pathway of GSK-3β/nerve growth factor to attenuate post-infarction arrhythmias by preconditioned adipose-derived stem cells. J Mol Cell Cardiol. 2017;104:17–30.

  16. 16.

    Lee TM, Lai PY, Chang NC. Effect of N-acetylcysteine on sympathetic hyperinnervation in post-infarcted rat hearts. Cardiovasc Res. 2010;85:137–46.

  17. 17.

    Dong H, Zhang X, Dai X, Lu S, Gui B, Jin W, et al. Lithium ameliorates lipopolysaccharide-induced microglial activation via inhibition of toll-like receptor 4 expression by activating the PI3K/Akt/FoxO1 pathway. J Neuroinflamm. 2014;11:140.

  18. 18.

    Harn HJ, Lin SZ, Lin PC, Liu CY, Liu PY, Chang LF, et al. Local interstitial delivery of z-butylidenephthalide by polymer wafers against malignant human gliomas. Neuro Oncol. 2011;13:635–48.

  19. 19.

    Nemoto T, Kanai T, Yanagita T, Satoh S, Maruta T, Yoshikawa N, et al. Regulation of Akt mRNA and protein levels by glycogen synthase kinase-3beta in adrenal chromaffin cells: effects of LiCl and SB216763. Eur J Pharmacol. 2008;586:82–9.

  20. 20.

    Abdul-Ghani R, Serra V, Györffy B, Jürchott K, Solf A, Dietel M, et al. The PI3K inhibitor LY294002 blocks drug export from resistant colon carcinoma cells overexpressing MRP1. Oncogene. 2006;25:1743–52.

  21. 21.

    Pang M, Ma L, Gong R, Tolbert E, Mao H, Ponnusamy M, et al. A novel STAT3 inhibitor, S3I-201, attenuates renal interstitial fibroblast activation and interstitial fibrosis in obstructive nephropathy. Kidney Int. 2010;78:257–68.

  22. 22.

    Bélichard P, Savard P, Cardinal R, Nadeau R, Gosselin H, Paradis P, et al. Markedly different effects on ventricular remodeling result in a decrease in inducibility of ventricular arrhythmias. J Am Coll Cardiol. 1994;23:505–13.

  23. 23.

    Stegemann H, Stalder K. Determination of hydroxyproline. Clin Chim Acta. 1967;18:267–73.

  24. 24.

    Vasandan AB, Jahnavi S, Shashank C, Prasad P, Kumar A, Prasanna SJ. Human mesenchymal stem cells program macrophage plasticity by altering their metabolic status via a PGE2-dependent mechanism. Sci Rep. 2016;6:38308.

  25. 25.

    Wang G, Shi Y, Jiang X, Leak RK, Hu X, Wu Y, et al. HDAC inhibition prevents white matter injury by modulating microglia/macrophage polarization through the GSK3β/PTEN/Akt axis. Proc Natl Acad Sci USA. 2015;112:2853–8.

  26. 26.

    Enomoto D, Obana M, Miyawaki A, Maeda M, Nakayama H, Fujio Y. Cardiac-specific ablation of the STAT3 gene in the subacute phase of myocardial infarction exacerbated cardiac remodeling. Am J Physiol Heart Circ Physiol. 2015;309:H1471–80.

  27. 27.

    Khan JA, Cao M, Kang BY, Liu Y, Mehta JL, Hermonat PL. AAV/hSTAT3-gene delivery lowers aortic inflammatory cell infiltration in LDLR KO mice on high cholesterol. Atherosclerosis. 2010;213:59–66.

  28. 28.

    Chen YL, Jian MH, Lin CC, Kang JC, Chen SP, Lin PC, et al. The induction of orphan nuclear receptor Nur77 expression by n-butylenephthalide as pharmaceuticals on hepatocellular carcinoma cell therapy. Mol Pharmacol. 2008;74:1046–58.

  29. 29.

    Hossini AM, Quast AS, Plötz M, Grauel K, Exner T, Küchler J, et al. PI3K/AKT signaling pathway is essential for survival of induced pluripotent stem cells. PLoS One. 2016;11:e0154770.

  30. 30.

    Shi J, Li J, Guan H, Cai W, Bai X, Fang X, et al. Anti-fibrotic actions of interleukin-10 against hypertrophic scarring by activation of PI3K/AKT and STAT3 signaling pathways in scar-forming fibroblasts. PLoS One. 2014;9:e98228.

  31. 31.

    Lee TM, Lin SZ, Chang NC. Nicorandil regulates the macrophage skewing and ameliorates myofibroblasts by inhibition of RhoA/Rho-kinase signalling in infarcted rats. J Cell Mol Med. 2018;22:1056–69.

  32. 32.

    Liechty KW, Kim HB, Adzick NS, Crombleholme TM. Fetal wound repair results in scar formation in interleukin-10-deficient mice in a syngeneic murine model of scarless fetal wound repair. J Pediatr Surg. 2000;35:866–72.

  33. 33.

    Gordon A, Kozin ED, Keswani SG, Vaikunth SS, Katz AB, Zoltick PW, et al. Permissive environment in postnatal wounds induced by adenoviral-mediated overexpression of the anti-inflammatory cytokine interleukin-10 prevents scar formation. Wound Repair Regen. 2008;16:70–9.

  34. 34.

    Li L, Zhang S, Zhang Y, Yu B, Xu Y, Guan Z. Paracrine action mediates the antifibrotic effect of transplanted mesenchymal stem cells in a rat model of global heart failure. Mol Biol Rep. 2009;36:725–31.

  35. 35.

    Yang J, Dai C, Liu Y. A novel mechanism by which hepatocyte growth factor b locks tubular epithelial to mesenchymal transition. J Am Soc Nephrol. 2005;16:68–78.

  36. 36.

    Poli G. Pathogenesis of liver fibrosis: role of oxidative stress. Mol Asp Med. 2000;21:49–98.

  37. 37.

    Wink DA, Vodovotz Y, Grisham MB, DeGraff W, Cook JC, Pacelli R, et al. Antioxidant effects of nitric oxide. Methods Enzymol. 1999;301:413–24.

  38. 38.

    Powell DW, Mifflin RC, Valentich JD, Crowe SE, Saada JI, West AB. Myofibroblasts. I. Paracrine cells important in health and disease. Am J Physiol. 1999;277:C1–9.

  39. 39.

    Jousset F, Maguy A, Rohr S, Kucera JP. Myofibroblasts electrotonically coupled to cardiomyocytes alter conduction: insights at the cellular level from a detailed in silico tissue structure model. Front Physiol. 2016;7:496.

  40. 40.

    Mias C, Coatrieux C, Denis C, Genet G, Seguelas MH, Laplace N, et al. Cardiac fibroblasts regulate sympathetic nerve sprouting and neurocardiac synapse stability. PLoS One. 2013;8:e79068.

  41. 41.

    Gyöngyösi M, Winkler J, Ramos I, Do QT, Firat H, McDonald K, et al. Myocardial fibrosis: biomedical research from bench to bedside. Eur J Heart Fail. 2017;19:177–91.

  42. 42.

    Lim GB. Heart failure: macrophages promote cardiac fibrosis and diastolic dysfunction. Nat Rev Cardiol. 2018;15:196–7.

  43. 43.

    Vallania F, Schiavone D, Dewilde S, Pupo E, Garbay S, Calogero R, et al. Genome-wide discovery of functional transcription factor binding sites by comparative genomics: the case of Stat3. Proc Natl Acad Sci USA. 2009;106:5117–22.

  44. 44.

    Sun Y, Iyer M, McEachin R, Zhao M, Wu YM, Cao X, et al. Genome-wide STAT3 binding analysis after histone deacetylase inhibition reveals novel target genes in dendritic cells. J Innate Immun. 2017;9:126–44.

  45. 45.

    Hutchins AP, Poulain S, Miranda-Saavedra D. Genome-wide analysis of STAT3 binding in vivo predicts effectors of the anti-inflammatory response in macrophages. Blood. 2012;119:e110–9.

  46. 46.

    Fu W, Hu W, Shi L, Mundra JJ, Xiao G, Dustin ML, et al. Foxo4- and Stat3-dependent IL-10 production by progranulin in regulatory T cells restrains inflammatory arthritis. FASEB J. 2017;31:1354–67.

Download references

Funding

This work was supported by the grants of An-Nan Hospital (ANHRF107-04) and Ministry of Science and Technology (MOST-106-2314-B-039-049), Taiwan.

Author information

Affiliations

  1. Cardiovascular Institute, An Nan Hospital, China Medical University, Tainan, Taiwan

    • Tsung-Ming Lee
  2. Department of Medicine, China Medical University, Taichung, Taiwan

    • Tsung-Ming Lee
  3. Bioinnovation Center, Tzu Chi Foundation, Hualien, Taiwan

    • Horng-Jyh Harn
    •  & Shinn-Zong Lin
  4. Department of Pathology, Buddhist Tzu Chi General Hospital, Tzu Chi University, Hualien, Taiwan

    • Horng-Jyh Harn
  5. Department of Life Science and Graduate Institute of Biotechnology, National Dong Hwa University, Hualien, Taiwan

    • Tzyy-Wen Chiou
  6. Department of Technology Management, Chung Hua University, Hsinchu, Taiwan

    • Ming-Hsi Chuang
  7. Gwo Xi Stem Cell Applied Technology, Hsinchu, Taiwan

    • Ming-Hsi Chuang
    • , Chun-Hung Chen
    •  & Po-Cheng Lin
  8. Genomics Research Center, Academia Sinica, Taipei, Taiwan

    • Chi-Hsuan Chuang
  9. Department of Neurosurgery, Buddhist Tzu Chi General Hospital, Tzu Chi University, Hualien, Taiwan

    • Shinn-Zong Lin

Authors

  1. Search for Tsung-Ming Lee in:

  2. Search for Horng-Jyh Harn in:

  3. Search for Tzyy-Wen Chiou in:

  4. Search for Ming-Hsi Chuang in:

  5. Search for Chun-Hung Chen in:

  6. Search for Chi-Hsuan Chuang in:

  7. Search for Po-Cheng Lin in:

  8. Search for Shinn-Zong Lin in:

Conflict of interest

The authors declare that they have no conflict of interest.

Ethics approval

All rats had unrestricted access to food/water in accordance with the China Medical University Committee on Animal Care (Permit Number: 2016-070). The investigation conforms to the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH Publication No. 85-23, revised 1996).

Corresponding author

Correspondence to Shinn-Zong Lin.

Supplementary information

About this article

Publication history

Received

Revised

Accepted

Published

DOI

https://doi.org/10.1038/s41374-018-0181-x