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Open Access 01-12-2021 | Respiratory Microbiota | Research

Microbiome diversity declines while distinct expansions of Th17, iNKT, and dendritic cell subpopulations emerge after anastomosis surgery

Authors: Emilie E. Vomhof-DeKrey, Allie Stover, Marc D. Basson

Published in: Gut Pathogens | Issue 1/2021

Abstract

Background

Anastomotic failure causes morbidity and mortality even in technically correct anastomoses. Initial leaks must be prevented by mucosal reapproximation across the anastomosis. Healing is a concerted effort between intestinal epithelial cells (IECs), immune cells, and commensal bacteria. IEC TLR4 activation and signaling is required for mucosal healing, leading to inflammatory factor release that recruits immune cells to limit bacteria invasion. TLR4 absence leads to mucosal damage from loss in epithelial proliferation, attenuated inflammatory response, and bacteria translocation. We hypothesize after anastomosis, an imbalance in microbiota will occur due to a decrease in TLR4 expression and will lead to changes in the immune milieu.

Results

We isolated fecal content and small intestinal leukocytes from murine, Roux-en-Y and end-to-end anastomoses, to identify microbiome changes and subsequent alterations in the regulatory and pro-inflammatory immune cells 3 days post-operative. TLR4⁺ IECs were impaired after anastomosis. Microbiome diversity was reduced, with Firmicutes, Bacteroidetes, and Saccharibacteria decreased and Proteobacteria increased. A distinct TCRβ^hi CD4⁺ T cells subset after anastomosis was 10–20-fold greater than in control mice. 84% were Th17 IL-17A/F⁺ IL-22⁺ and/or TNFα⁺. iNKT cells were increased and TCRβ^hi. 75% were iNKT IL-10⁺ and 13% iNKTh17 IL-22⁺. Additionally, Treg IL-10⁺ and IL-22⁺ cells were increased. A novel dendritic cell subset was identified in anastomotic regions that was CD11b^hi CD103^mid and was 93% IL-10⁺.

Conclusions

This anastomotic study demonstrated a decrease in IEC TLR4 expression and microbiome diversity which then coincided with increased expansion of regulatory and pro-inflammatory immune cells and cytokines. Defining the anastomotic mucosal environment could help inform innovative therapeutics to target excessive pro-inflammatory invasion and microbiome imbalance.

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Abreu MT. Toll-like receptor signalling in the intestinal epithelium: How bacterial recognition shapes intestinal function. Nat Rev Immunol. 2010;10:131–43.CrossRefPubMed

Alam A, Neish A. Role of gut microbiota in intestinal wound healing and barrier function. Tissue Barriers. 2018. https://doi.org/10.1080/21688370.2018.1539595.CrossRefPubMedPubMedCentral

Amir A, McDonald D, Navas-Molina JA, Kopylova E, Morton JT, Zech XuZ, Kightley EP, Thompson LR, Hyde ER, Gonzalez A, Knight R. Deblur rapidly resolves single-nucleotide community sequence patterns. mSystems. 2017. https://doi.org/10.1128/msystems.00191-16.CrossRefPubMedPubMedCentral

An D, Oh SF, Olszak T, Neves JF, Avci FY, Erturk-Hasdemir D, Lu X, Zeissig S, Blumberg RS, Kasper DL. Sphingolipids from a symbiotic microbe regulate homeostasis of host intestinal natural killer T cells. Cell. 2014;156:123–33. https://doi.org/10.1016/j.cell.2013.11.042.CrossRefPubMedPubMedCentral

Bain CC, Scott CL, Uronen-Hansson H, Gudjonsson S, Jansson O, Grip O, Guilliams M, Malissen B, Agace WW, Mowat AMI. Resident and pro-inflammatory macrophages in the colon represent alternative context-dependent fates of the same Ly6C hi monocyte precursors. Mucosal Immunol. 2013;6:498–510. https://doi.org/10.1038/mi.2012.89.CrossRefPubMed

Bokulich NA, Kaehler BD, Rideout JR, Dillon M, Bolyen E, Knight R, Huttley GA, Gregory CJ. Optimizing taxonomic classification of marker-gene amplicon sequences with QIIME 2’s q2-feature-classifier plugin. Microbiome. 2018. https://doi.org/10.1186/s40168-018-0470-z.CrossRefPubMedPubMedCentral

Caporaso JG, Kuczynski J, Stombaugh J, Bittinger K, Bushman FD, Costello EK, Fierer N, Pẽa AG, Goodrich JK, Gordon JI, Huttley GA, Kelley ST, Knights D, Koenig JE, Ley RE, Lozupone CA, McDonald D, Muegge BD, Pirrung M, Reeder J, Sevinsky JR, Turnbaugh PJ, Walters WA, Widmann J, Yatsunenko T, Zaneveld J, Knight R. QIIME allows analysis of high-throughput community sequencing data. Nat Methods. 2010;7:335–6.CrossRefPubMedPubMedCentral

Cerovic V, Bain CC, Mowat AM, Milling SWF. Intestinal macrophages and dendritic cells: what’s the difference? Trends Immunol. 2014;35:270–7.CrossRefPubMed

Chistiakov DA, Bobryshev YV, Kozarov E, Sobenin IA, Orekhov AN. Intestinal mucosal tolerance and impact of gut microbiota to mucosal tolerance. Front Microbiol. 2014. https://doi.org/10.3389/fmicb.2014.00781.CrossRefPubMed

10.

Coombes JL, Powrie F. Dendritic cells in intestinal immune regulation. Nat Rev Immunol. 2008;8:435–46.CrossRefPubMedPubMedCentral

11.

Coombes JL, Siddiqui KRR, Arancibia-Cárcamo CV, Hall J, Sun CM, Belkaid Y, Powrie F. A functionally specialized population of mucosal CD103+ DCs induces Foxp3+ regulatory T cells via a TGF-β -and retinoic acid-dependent mechanism. J Exp Med. 2007;204:1757–64. https://doi.org/10.1084/jem.20070590.CrossRefPubMedPubMedCentral

12.

Devine AA, Gonzalez A, Speck KE, Knight R, Helmrath M, Lund PK, Azcarate-Peril MA. Impact of ileocecal resection and concomitant antibiotics on the microbiome of the murine jejunum and colon. PLoS ONE. 2013. https://doi.org/10.1371/journal.pone.0073140.CrossRefPubMedPubMedCentral

13.

Dowds CM, Blumberg RS, Zeissig S. Control of intestinal homeostasis through crosstalk between natural killer T cells and the intestinal microbiota. Clin Immunol. 2014;159:128–33. https://doi.org/10.1016/j.clim.2015.05.008.CrossRef

14.

Frosali S, Pagliari D, Gambassi G, Landolfi R, Pandolfi F, Cianci R. How the intricate interaction among toll-like receptors, microbiota, and intestinal immunity can influence gastrointestinal pathology. J Immunol Res. 2015. https://doi.org/10.1155/2015/489821.CrossRefPubMedPubMedCentral

15.

Fujimoto K, Karuppuchamy T, Takemura N, Shimohigoshi M, Machida T, Haseda Y, Aoshi T, Ishii KJ, Akira S, Uematsu S. A new subset of CD103+CD8α+ dendritic cells in the small intestine expresses TLR3, TLR7, and TLR9 and induces Th1 response and CTL activity. J Immunol. 2011;186:6287–95. https://doi.org/10.4049/jimmunol.1004036.CrossRefPubMed

16.

Goodyear AW, Kumar A, Dow S, Ryan EP. Optimization of murine small intestine leukocyte isolation for global immune phenotype analysis. J Immunol Methods. 2014;405:97–108. https://doi.org/10.1016/j.jim.2014.01.014.CrossRefPubMed

17.

Hoehn RS, Seitz AP, Jernigan PL, Gulbins E, Edwards MJ. Ischemia/reperfusion injury alters sphingolipid metabolism in the gut. Cell Physiol Biochem. 2016;39:1262–70. https://doi.org/10.1159/000447831.CrossRefPubMed

18.

Hyman N, Manchester TL, Osler T, Burns B, Cataldo PA. Anastomotic leaks after intestinal anastomosis: it’s later than you think. Ann Surg. 2007;245:254–8. https://doi.org/10.1097/01.sla.0000225083.27182.85.CrossRefPubMedPubMedCentral

19.

Ivanov II, Atarashi K, Manel N, Brodie EL, Shima T, Karaoz U, Wei D, Goldfarb KC, Santee CA, Lynch SV, Tanoue T, Imaoka A, Itoh K, Takeda K, Umesaki Y, Honda K, Littman DR. Induction of intestinal Th17 cells by segmented filamentous bacteria. Cell. 2009;139:485–98. https://doi.org/10.1016/j.cell.2009.09.033.CrossRefPubMedPubMedCentral

20.

Jakubzick C, Bogunovic M, Bonito AJ, Kuan EL, Merad M, Randolph GJ. Lymph-migrating, tissue-derived dendritic cells are minor constituents within steady-state lymph nodes. J Exp Med. 2008;205:2839–50. https://doi.org/10.1084/jem.20081430.CrossRefPubMedPubMedCentral

21.

Katoh K, Standley DM. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol. 2013;30:772–80. https://doi.org/10.1093/molbev/mst010.CrossRefPubMedPubMedCentral

22.

Koch S, Nusrat A. The life and death of epithelia during inflammation: lessons learned from the gut. Annu Rev Pathol Mech Dis. 2012;7:35–60.CrossRef

23.

Lapthorne S, Pereira-Fantini PM, Fouhy F, Wilson G, Thomas SL, Dellios NL, Scurr M, O’Sullivan O, Paul Ross R, Stanton C, Fitzgerald GF, Cotter PD, Bines JE. Gut microbial diversity is reduced and is associated with colonic inflammation in a piglet model of short bowel syndrome. Gut Microbes. 2013. https://doi.org/10.4161/gmic.24372.CrossRefPubMedPubMedCentral

24.

Lipska MA, Bissett IP, Parry BR, Merrie AEH. Anastomotic leakage after lower gastrointestinal anastomosis: men are at a higher risk. ANZ J Surg. 2006;76:579–85. https://doi.org/10.1111/j.1445-2197.2006.03780.x.CrossRefPubMed

25.

Longman RS, Littman DR. The functional impact of the intestinal microbiome on mucosal immunity and systemic autoimmunity. Curr Opin Rheumatol. 2015;27:381–7. https://doi.org/10.1097/BOR.0000000000000190.CrossRefPubMedPubMedCentral

26.

Love MI, Huber W, Anders S. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 2014. https://doi.org/10.1186/s13059-014-0550-8.CrossRefPubMedPubMedCentral

27.

Mamalaki C, Norton T, Tanaka Y, Townsend AR, Chandler P, Simpson E, Kioussis D. Thymic depletion and peripheral activation of class I major histocompatibility complex-restricted T cells by soluble peptide in T-cell receptor transgenic mice. Proc Natl Acad Sci USA. 1992;89:11342–6. https://doi.org/10.1073/pnas.89.23.11342.CrossRefPubMedPubMedCentral

28.

Manichanh C, Rigottier-Gois L, Bonnaud E, Gloux K, Pelletier E, Frangeul L, Nalin R, Jarrin C, Chardon P, Marteau P, Roca J, Dore J. Reduced diversity of faecal microbiota in Crohn’s disease revealed by a metagenomic approach. Gut. 2006;55:205–11. https://doi.org/10.1136/gut.2005.073817.CrossRefPubMedPubMedCentral

29.

McMurdie PJ, Holmes S. Phyloseq: an R package for reproducible interactive analysis and graphics of microbiome census data. PLoS ONE. 2013. https://doi.org/10.1371/journal.pone.0061217.CrossRefPubMedPubMedCentral

30.

Morse BC, Simpson JP, Jones YR, Johnson BL, Knott BM, Kotrady JA. Determination of independent predictive factors for anastomotic leak: analysis of 682 intestinal anastomoses. Am J Surg. 2013;206:950–6. https://doi.org/10.1016/j.amjsurg.2013.07.017.CrossRefPubMed

31.

Murrell ZA, Stamos MJ. Reoperation for anastomotic failure. Clin Colon Rectal Surg. 2006;19:213–6. https://doi.org/10.1055/s-2006-956442.CrossRefPubMedPubMedCentral

32.

Nieuwenhuis EES, Matsumoto T, Lindenbergh D, Willemsen R, Kaser A, Simons-Oosterhuis Y, Brugman S, Yamaguchi K, Ishikawa H, Aiba Y, Koga Y, Samsom JN, Oshima K, Kikuchi M, Escher JC, Hattori M, Onderdonk AB, Blumberg RS. Cd1d-dependent regulation of bacterial colonization in the intestine of mice. J Clin Invest. 2009;119:1241–50. https://doi.org/10.1172/JCI36509.CrossRefPubMedPubMedCentral

33.

Olszak T, An D, Zeissig S, Vera MP, Richter J, Franke A, Glickman JN, Siebert R, Baron RM, Kasper DL, Blumberg RS. Microbial exposure during early life has persistent effects on natural killer T cell function. Science. 2012;336:489–93. https://doi.org/10.1126/science.1219328.CrossRefPubMedPubMedCentral

34.

Parks OB, Pociask DA, Hodzic Z, Kolls JK, Good M. Interleukin-22 signaling in the regulation of intestinal health and disease. Front Cell Dev Biol. 2016. https://doi.org/10.3389/fcell.2015.00085.CrossRefPubMedPubMedCentral

35.

Persson EK, Scott CL, Mowat AM, Agace WW. Dendritic cell subsets in the intestinal lamina propria: ontogeny and function. Eur J Immunol. 2013;43:3098–107. https://doi.org/10.1002/eji.201343740.CrossRefPubMedPubMedCentral

36.

Price MN, Dehal PS, Arkin AP. FastTree 2—approximately maximum-likelihood trees for large alignments. PLoS ONE. 2010. https://doi.org/10.1371/journal.pone.0009490.CrossRefPubMedPubMedCentral

37.

Rakoff-Nahoum S, Paglino J, Eslami-Varzaneh F, Edberg S, Medzhitov R. Recognition of commensal microflora by toll-like receptors is required for intestinal homeostasis. Cell. 2004;118:229–41. https://doi.org/10.1016/j.cell.2004.07.002.CrossRefPubMed

38.

Rivollier A, He J, Kole A, Valatas V, Kelsall BL. Inflammation switches the differentiation program of Ly6chi monocytes from antiinflammatory macrophages to inflammatory dendritic cells in the colon. J Exp Med. 2012;209:139–55. https://doi.org/10.1084/jem.20101387.CrossRefPubMedPubMedCentral

39.

Rognes T, Flouri T, Nichols B, Quince C, Mahé F. VSEARCH: a versatile open source tool for metagenomics. PeerJ. 2016. https://doi.org/10.7717/peerj.2584.CrossRefPubMedPubMedCentral

40.

Round JL, Mazmanian SK. The gut microbiota shapes intestinal immune responses during health and disease. Nat Rev Immunol. 2009;9:313–23.CrossRefPubMedPubMedCentral

41.

Round JL, Mazmanian SK. Inducible Foxp3+ regulatory T-cell development by a commensal bacterium of the intestinal microbiota. Proc Natl Acad Sci USA. 2010;107:12204–9. https://doi.org/10.1073/pnas.0909122107.CrossRefPubMedPubMedCentral

42.

de Guinoa SJ, Jimeno R, Gaya M, Kipling D, Garzón MJ, Dunn-Walters D, Ubeda C, Barral P. CD 1d-mediated lipid presentation by CD 11c+ cells regulates intestinal homeostasis. EMBO J. 2018. https://doi.org/10.15252/embj.201797537.CrossRef

43.

Sakr A, Emile SH, Abdallah E, Thabet W, Khafagy W. Predictive factors for small intestinal and colonic anastomotic leak: a multivariate analysis. Indian J Surg. 2017;79:555–62. https://doi.org/10.1007/s12262-016-1556-0.CrossRefPubMed

44.

Salio M, Silk JD, Yvonne Jones E, Cerundolo V. Biology of CD1- and MR1-restricted T cells. Annu Rev Immunol. 2014;32:323–66.CrossRefPubMed

45.

Schrum AG, Turka LA, Palmer E. Surface T-cell antigen receptor expression and availability for long-term antigenic signaling. Immunol Rev. 2003;196:7–24.CrossRefPubMed

46.

Schrum AG, Wells AD, Turka LA. Enhanced surface TCR replenishment mediated by CD28 leads to greater TCR engagement during primary stimulation. Int Immunol. 2000;12:833–42. https://doi.org/10.1093/intimm/12.6.833.CrossRefPubMed

47.

Selvanantham T, Lin Q, Guo CX, Surendra A, Fieve S, Escalante NK, Guttman DS, Streutker CJ, Robertson SJ, Philpott DJ, Mallevaey T. NKT cell-deficient mice harbor an altered microbiota that fuels intestinal inflammation during chemically induced colitis. J Immunol. 2016;197:4464–72. https://doi.org/10.4049/jimmunol.1601410.CrossRefPubMed

48.

Sommovilla J, Zhou Y, Sun RC, Choi PM, Diaz-Miron J, Shaikh N, Sodergren E, Warner BB, Weinstock GM, Tarr PI, Warner BW. Small bowel resection induces long-term changes in the enteric microbiota of mice. J Gastrointest Surg. 2014;19:56–64. https://doi.org/10.1007/s11605-014-2631-0.CrossRefPubMedPubMedCentral

49.

Sun CM, Hall JA, Blank RB, Bouladoux N, Oukka M, Mora JR, Belkaid Y. Small intestine lamina propria dendritic cells promote de novo generation of Foxp3 T reg cells via retinoic acid. J Exp Med. 2007;204:1775–85. https://doi.org/10.1084/jem.20070602.CrossRefPubMedPubMedCentral

50.

Tamoutounour S, Henri S, Lelouard H, de Bovis B, de Haar C, van der Woude CJ, Woltman AM, Reyal Y, Bonnet D, Sichien D, Bain CC, Mowat AM, Reis e Sousa C, Poulin LF, Malissen B, Guilliams M. CD64 distinguishes macrophages from dendritic cells in the gut and reveals the Th1-inducing role of mesenteric lymph node macrophages during colitis. Eur J Immunol. 2012;42:3150–66. https://doi.org/10.1002/eji.201242847.CrossRefPubMed

51.

Vargas-Caraveo A, Sayd A, Robledo-Montaña J, Caso JR, Madrigal JLM, García-Bueno B, Leza JC. Toll-like receptor 4 agonist and antagonist lipopolysaccharides modify innate immune response in rat brain circumventricular organs. J Neuroinflammation. 2020;17:1–17. https://doi.org/10.1186/s12974-019-1690-2.CrossRef

52.

Vomhof-DeKrey EE, Lansing JT, Darland DC, Umthun J, Stover AD, Brown C, Basson MD. Loss of Slfn3 induces a sex-dependent repair vulnerability after 50% bowel resection. Am J Physiol Liver Physiol. 2021;320:G136–52. https://doi.org/10.1152/ajpgi.00344.2020.CrossRef

53.

Wingender G, Stepniak D, Krebs P, Lin L, McBride S, Wei B, Braun J, Mazmanian SK, Kronenberg M. Intestinal microbes affect phenotypes and functions of invariant natural killer T cells in mice. Gastroenterology. 2012;143:418–28. https://doi.org/10.1053/j.gastro.2012.04.017.CrossRefPubMed

54.

Zhang Y, Li X, Carpinteiro A, Gulbins E. Acid sphingomyelinase amplifies redox signaling in pseudomonas aeruginosa-induced macrophage apoptosis. J Immunol. 2008;181:4247–54. https://doi.org/10.4049/jimmunol.181.6.4247.CrossRefPubMed

55.

Zhang Y, Li X, Grassmé H, Döring G, Gulbins E. Alterations in ceramide concentration and pH determine the release of reactive oxygen species by Cftr-deficient macrophages on infection. J Immunol. 2010;184:5104–11. https://doi.org/10.4049/jimmunol.0902851.CrossRefPubMed

56.

Zundler S, Tauschek V, Neurath MF. Immune cell circuits in mucosal wound healing: clinical implications. Visc Med. 2020;36:129–36.CrossRefPubMedPubMedCentral

Title: Microbiome diversity declines while distinct expansions of Th17, iNKT, and dendritic cell subpopulations emerge after anastomosis surgery
Authors: Emilie E. Vomhof-DeKrey
Allie Stover
Marc D. Basson
Publication date: 01-12-2021
Publisher: BioMed Central
Keyword: Respiratory Microbiota
Published in: Gut Pathogens / Issue 1/2021
Electronic ISSN: 1757-4749
DOI: https://doi.org/10.1186/s13099-021-00447-z

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