Cell and Tissue Research

, Volume 376, Issue 2, pp 257–271 | Cite as

Early injection of human adipose tissue-derived mesenchymal stem cell after inflammation ameliorates dextran sulfate sodium-induced colitis in mice through the induction of M2 macrophages and regulatory T cells

  • Yuzo Kawata
  • Atsunori TsuchiyaEmail author
  • Satoshi Seino
  • Yusuke Watanabe
  • Yuichi Kojima
  • Shunzo Ikarashi
  • Kentaro Tominaga
  • Junji Yokoyama
  • Satoshi Yamagiwa
  • Shuji TeraiEmail author
Regular Article


Inflammatory bowel diseases (IBDs) are sometimes refractory to current therapy or associated with severe adverse events during immunosuppressive therapy; thus, new therapies are urgently needed. Recently, mesenchymal stem cells (MSCs) have attracted attention based on their multitude of functions including anti-inflammatory effects. However, proper timing of MSC therapy and the mechanisms underlying the therapeutic effects of MSCs on colitis are not fully elucidated. Human adipose tissue-derived mesenchymal stem cells (hAdMSCs; 1 × 106) were administrated via the tail vein on day 3 (early) or 11 (delayed) using a 7-day dextran sulfate sodium (DSS)-induced mouse model of colitis. The effects were evaluated based on colon length, disease activity index (DAI) and histological score. Cytokine-encoding mRNA levels T cells and macrophages were evaluated by real-time PCR and flow cytometry. Regarding the timing of administration, early (day 3) injection significantly ameliorated DSS-induced colitis in terms of both DAI and histological score, compared to those parameters with delayed (day 11) injection. With early cell injection, the tissue mRNA levels of anti-inflammatory cytokine genes (Il10, Tgfb) increased, whereas those of inflammatory cytokine genes (Il6, Tnfa and Il17a) decreased significantly. Regarding the associated mechanism, hAdMSCs suppressed T cell proliferation and activation in vitro, increased the number of regulatory T cells in vivo and changed the polarity of macrophages (into the anti-inflammatory M2 phenotype) in vitro. Timing of injection is critical for the effective therapeutic effects of hAdMSCs. Furthermore, part of the associated mechanism includes T cell activation and expansion and altered macrophage polarization.


Mesenchymal stem cells Regulatory T cells Macrophages Adipose tissue Dextran sulfate sodium 



The authors thank Takao Tsuchida (Department of Gastroenterology and Hepatology, Graduate School of Medical and Dental Science, Niigata University) for his cooperation.

Financial support

This work was supported by InterStem Co., Ltd.

Compliance with ethical standards

Conflicts of interest

ST received research funding from InterStem Co., Ltd.

Supplementary material

441_2018_2981_Fig9_ESM.png (184 kb)
Supplemental Figure 1

Evaluation of the therapeutic effect of human adipose tissue-derived mesenchymal stem cells (hAdMSCs) in a mouse model of dextran sulfate sodium-induced colitis, after intra-peritoneal administration on day 3 after initiation. (a and b) Prevention of colon length shortening and improved disease activity index (DAI) scores were observed in the group intra-peritoneally administered hAdMSCs on day 3 and sacrificed on day 21 (N = 18 mice; control, 8 mice, injected, 10 mice). (c) Histological analysis. Significant improvement in histological sores was not observed in this group. (d) Analysis of mRNA expression changes in the middle colon in this group, compared to levels in the untreated group (*P < 0.05). N.S., not significant. (PNG 183 kb)

441_2018_2981_MOESM1_ESM.tif (299 kb)
High Resolution Image (TIF 299 kb)
441_2018_2981_Fig10_ESM.png (177 kb)
Supplemental Figure 2

Evaluation of the therapeutic effect of human adipose tissue-derived mesenchymal stem cells (hAdMSCs) in a mouse model of dextran sulfate sodium-induced colitis, after administration using the tail vein on day 7 after initiation. (a and b) Prevention of colon length shortening was observed; however, improved disease activity index (DAI) scores were not observed in the group administered hAdMSCs using the tail vein on day 7 and sacrificed on day 21 (N = 13 mice; control, 6 mice, injected, 7 mice). (c) Histological analysis. Significant improvement in histological scores was not observed in this group. (d) Analysis of mRNA expression changes in the middle colon in this group, compared to levels in the untreated group (*P < 0.05). N.S., not significant. (PNG 176 kb)

441_2018_2981_MOESM2_ESM.tif (281 kb)
High Resolution Image (TIF 281 kb)
441_2018_2981_Fig11_ESM.png (323 kb)
Supplemental Figure 3

Effect of human adipose tissue-derived mesenchymal stem cells (hAdMSCs) on macrophages. Evaluation of CD206 (M2) macrophages by flow cytometry (a-c). The frequency of CD206-positive cells/F4/80-positive cells significantly increased in the hAdMSC-injected group compared to that in the control group (*P < 0.05). (PNG 322 kb)

441_2018_2981_MOESM3_ESM.tif (447 kb)
High Resolution Image (TIF 446 kb)
441_2018_2981_MOESM4_ESM.doc (94 kb)
Supplemental Table 1 List of primers used for real-time PCR (DOC 94 kb)
441_2018_2981_MOESM5_ESM.doc (77 kb)
Supplemental Table 2 List of antibodies used for immunostaining (DOC 77 kb)
441_2018_2981_MOESM6_ESM.docx (13 kb)
Supplemental Table 3 List of antibodies used for flow cytometry (DOCX 12 kb)


  1. Asakura K, Nishiwaki Y, Inoue N, Hibi T, Watanabe M, Takebayashi T (2009) Prevalence of ulcerative colitis and Crohn’s disease in Japan. J Gastroenterol 44:659–665CrossRefGoogle Scholar
  2. Chen X, Gan Y, Li W, Su J, Zhang Y, Huang Y, Roberts AI, Han Y, Li J, Wang Y et al (2014) The interaction between mesenchymal stem cells and steroids during inflammation. Cell Death Dis 5:e1009CrossRefGoogle Scholar
  3. Cho YB, Lee WY, Park KJ, Kim M, Yoo HW, Yu CS (2013) Autologous adipose tissue-derived stem cells for the treatment of Crohn’s fistula: a phase I clinical study. Cell Transplant 22:279–285CrossRefGoogle Scholar
  4. Cooper HS, Murthy SN, Shah RS, Sedergran DJ (1993) Clinicopathologic study of dextran sulfate sodium experimental murine colitis. Lab Invest 69:238–249Google Scholar
  5. Dave M, Hayashi Y, Gajdos GB, Smyrk TC, Svingen PA, Kvasha SM, Lorincz A, Dong H, Faubion WA Jr, Ordog T (2015) Stem cells for murine interstitial cells of cajal suppress cellular immunity and colitis via prostaglandin E2 secretion. Gastroenterology 148:978–990CrossRefGoogle Scholar
  6. de la Portilla F, Alba F, Garcia-Olmo D, Herrerias JM, Gonzalez FX, Galindo A (2013) Expanded allogeneic adipose-derived stem cells (eASCs) for the treatment of complex perianal fistula in Crohn’s disease: results from a multicenter phase I/IIa clinical trial. Int J Color Dis 28:313–323CrossRefGoogle Scholar
  7. Duijvestein M, Vos AC, Roelofs H, Wildenberg ME, Wendrich BB, Verspaget HW, Kooy-Winkelaar EM, Koning F, Zwaginga JJ, Fidder HH et al (2010) Autologous bone marrow-derived mesenchymal stromal cell treatment for refractory luminal Crohn’s disease: results of a phase I study. Gut 59:1662–1669CrossRefGoogle Scholar
  8. Forbes GM, Sturm MJ, Leong RW, Sparrow MP, Segarajasingam D, Cummins AG, Phillips M, Herrmann RP (2014) A phase 2 study of allogeneic mesenchymal stromal cells for luminal Crohn's disease refractory to biologic therapy. Clin Gastroenterol Hepatol 12:64–71CrossRefGoogle Scholar
  9. Gonzalez MA, Gonzalez-Rey E, Rico L, Buscher D, Delgado M (2009) Adipose-derived mesenchymal stem cells alleviate experimental colitis by inhibiting inflammatory and autoimmune responses. Gastroenterology 136:978–989CrossRefGoogle Scholar
  10. Gonzalez-Rey E, Anderson P, Gonzalez MA, Rico L, Buscher D, Delgado M (2009) Human adult stem cells derived from adipose tissue protect against experimental colitis and sepsis. Gut 58:929–939CrossRefGoogle Scholar
  11. Hu J, Zhao G, Zhang L, Qiao C, Di A, Gao H, Xu H (2016) Safety and therapeutic effect of mesenchymal stem cell infusion on moderate to severe ulcerative colitis. Exp Ther Med 12:2983–2989CrossRefGoogle Scholar
  12. Kappelman MD, Moore KR, Allen JK, Cook SF (2013) Recent trends in the prevalence of Crohn's disease and ulcerative colitis in a commercially insured US population. Dig Dis Sci 58:519–525CrossRefGoogle Scholar
  13. Katsuda T, Kosaka N, Takeshita F, Ochiya T (2013) The therapeutic potential of mesenchymal stem cell-derived extracellular vesicles. Proteomics 13:1637–1653CrossRefGoogle Scholar
  14. Kim HS, Shin TH, Lee BC, Yu KR, Seo Y, Lee S, Seo MS, Hong IS, Choi SW, Seo KW et al (2013) Human umbilical cord blood mesenchymal stem cells reduce colitis in mice by activating NOD2 signaling to COX2. Gastroenterology 145(1392–1403):e1391–e1398Google Scholar
  15. Lee WY, Park KJ, Cho YB, Yoon SN, Song KH, Kim DS, Jung SH, Kim M, Yoo HW, Kim I et al (2013) Autologous adipose tissue-derived stem cells treatment demonstrated favorable and sustainable therapeutic effect for Crohn’s fistula. Stem Cells 31:2575–2581CrossRefGoogle Scholar
  16. Mittal M, Tiruppathi C, Nepal S, Zhao YY, Grzych D, Soni D, Prockop DJ, Malik AB (2016) TNFalpha-stimulated gene-6 (TSG6) activates macrophage phenotype transition to prevent inflammatory lung injury. Proc Natl Acad Sci U S A 113:E8151–E8158CrossRefGoogle Scholar
  17. Molendijk I, Bonsing BA, Roelofs H, Peeters KC, Wasser MN, Dijkstra G, van der Woude CJ, Duijvestein M, Veenendaal RA, Zwaginga JJ et al (2015) Allogeneic bone marrow-derived mesenchymal stromal cells promote healing of refractory perianal fistulas in patients with Crohn’s disease. Gastroenterology 149(918–927):e916Google Scholar
  18. Monteleone G, Pallone F (2015) Mongersen, an Oral SMAD7 antisense oligonucleotide, and Crohn’s disease. N Engl J Med 372:2461CrossRefGoogle Scholar
  19. Panes J, Garcia-Olmo D, Van Assche G, Colombel JF, Reinisch W, Baumgart DC, Dignass A, Nachury M, Ferrante M, Kazemi-Shirazi L et al (2016) Expanded allogeneic adipose-derived mesenchymal stem cells (Cx601) for complex perianal fistulas in Crohn’s disease: a phase 3 randomised, double-blind controlled trial. Lancet 388:1281–1290CrossRefGoogle Scholar
  20. Ren G, Zhang L, Zhao X, Xu G, Zhang Y, Roberts AI, Zhao RC, Shi Y (2008) Mesenchymal stem cell-mediated immunosuppression occurs via concerted action of chemokines and nitric oxide. Cell Stem Cell 2:141–150CrossRefGoogle Scholar
  21. Sala E, Genua M, Petti L, Anselmo A, Arena V, Cibella J, Zanotti L, D'Alessio S, Scaldaferri F, Luca G et al (2015) Mesenchymal stem cells reduce colitis in mice via release of TSG6, Independently of Their Localization to the Intestine. Gastroenterology 149:163–176.e20CrossRefGoogle Scholar
  22. Stein M, Keshav S, Harris N, Gordon S (1992) Interleukin 4 potently enhances murine macrophage mannose receptor activity: a marker of alternative immunologic macrophage activation. J Exp Med 176:287–292CrossRefGoogle Scholar
  23. Terai S, Tsuchiya A (2017) Status of and candidates for cell therapy in liver cirrhosis: overcoming the "point of no return" in advanced liver cirrhosis. J Gastroenterol 52:129–140CrossRefGoogle Scholar
  24. Tsuchiya A, Kojima Y, Ikarashi S, Seino S, Watanabe Y, Kawata Y, Terai S (2017) Clinical trials using mesenchymal stem cells in liver diseases and inflammatory bowel diseases. Inflamm Regen 37Google Scholar
  25. Wang Y, Chen X, Cao W, Shi Y (2014) Plasticity of mesenchymal stem cells in immunomodulation: pathological and therapeutic implications. Nat Immunol 15:1009–1016CrossRefGoogle Scholar
  26. Watanabe S, Arimura Y, Nagaishi K, Isshiki H, Onodera K, Nasuno M, Yamashita K, Idogawa M, Naishiro Y, Murata M et al (2014) Conditioned mesenchymal stem cells produce pleiotropic gut trophic factors. J Gastroenterol 49:270–282CrossRefGoogle Scholar
  27. Williams KL, Fuller CR, Dieleman LA, DaCosta CM, Haldeman KM, Sartor RB, Lund PK (2001) Enhanced survival and mucosal repair after dextran sodium sulfate-induced colitis in transgenic mice that overexpress growth hormone. Gastroenterology 120:925–937CrossRefGoogle Scholar
  28. Yamada A, Arakaki R, Saito M, Tsunematsu T, Kudo Y, Ishimaru N (2016) Role of regulatory T cell in the pathogenesis of inflammatory bowel disease. World J Gastroenterol 22:2195–2205CrossRefGoogle Scholar
  29. Yui S, Nakamura T, Sato T, Nemoto Y, Mizutani T, Zheng X, Ichinose S, Nagaishi T, Okamoto R, Tsuchiya K et al (2012) Functional engraftment of colon epithelium expanded in vitro from a single adult Lgr5(+) stem cell. Nat Med 18:618–623CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Yuzo Kawata
    • 1
  • Atsunori Tsuchiya
    • 1
    Email author
  • Satoshi Seino
    • 1
  • Yusuke Watanabe
    • 1
  • Yuichi Kojima
    • 1
  • Shunzo Ikarashi
    • 1
  • Kentaro Tominaga
    • 1
  • Junji Yokoyama
    • 1
  • Satoshi Yamagiwa
    • 1
  • Shuji Terai
    • 1
    Email author
  1. 1.Division of Gastroenterology and Hepatology, Graduate school of medical and dental sciencesNiigata UniversityNiigataJapan

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