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Small intestinal intraepithelial TCRγδ+ T lymphocytes are present in the premature intestine but selectively reduced in surgical necrotizing enterocolitis.

Weitkamp JH, Rosen MJ, Zhao Z, Koyama T, Geem D, Denning TL, Rock MT, Moore DJ, Halpern MD, Matta P, Denning PW - PLoS ONE (2014)

Bottom Line: In human preterm infants, the proportion of IEL was reduced by 66% in 11 NEC ileum resections compared to 30 non-NEC controls (p<0.001).While γδ IEL dominated over conventional αβ IEL early in gestation in mice and in humans, γδ IEL were preferential decreased in the ileum of surgical NEC patients compared to non-NEC controls (50% reduction, p<0.05).Loss of IEL in human NEC was associated with downregulation of the Th17 transcription factor retinoic acid-related orphan nuclear hormone receptor C (RORC, p<0.001).

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Vanderbilt University School of Medicine and Monroe Carell Jr. Children's Hospital at Vanderbilt, Nashville, Tennessee, United States of America.

ABSTRACT

Background: Gastrointestinal barrier immaturity predisposes preterm infants to necrotizing enterocolitis (NEC). Intraepithelial lymphocytes (IEL) bearing the unconventional T cell receptor (TCR) γδ (γδ IEL) maintain intestinal integrity and prevent bacterial translocation in part through production of interleukin (IL) 17.

Objective: We sought to study the development of γδ IEL in the ileum of human infants and examine their role in NEC pathogenesis. We defined the ontogeny of γδ IEL proportions in murine and human intestine and subjected tcrδ-/- mice to experimental gut injury. In addition, we used polychromatic flow cytometry to calculate percentages of viable IEL (defined as CD3+ CD8+ CD103+ lymphocytes) and the fraction of γδ IEL in surgically resected tissue from infants with NEC and gestational age matched non-NEC surgical controls.

Results: In human preterm infants, the proportion of IEL was reduced by 66% in 11 NEC ileum resections compared to 30 non-NEC controls (p<0.001). While γδ IEL dominated over conventional αβ IEL early in gestation in mice and in humans, γδ IEL were preferential decreased in the ileum of surgical NEC patients compared to non-NEC controls (50% reduction, p<0.05). Loss of IEL in human NEC was associated with downregulation of the Th17 transcription factor retinoic acid-related orphan nuclear hormone receptor C (RORC, p<0.001). TCRδ-deficient mice showed increased severity of experimental gut injury (p<0.05) with higher TNFα expression but downregulation of IL17A.

Conclusion: Complimentary mouse and human data suggest a role of γδ IEL in IL17 production and intestinal barrier production early in life. Specific loss of the γδ IEL fraction may contribute to NEC pathogenesis. Nutritional or pharmacological interventions to support γδ IEL maintenance in the developing small intestine could serve as novel strategies for NEC prevention.

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γδ IEL are the predominant IEL subtype in the immature murine small intestine.A) Flow cytometry of distal small intestinal intraepithelial cells from 1 week old (1 wk, n = 4), 2 weeks old (2 wk, n = 3), 3 weeks old (3 wk, n = 2) and adult (n = 1) C57BL/6J mice stained for CD3, CD8α, CD103, TCRγδ and TCRβ as described. Intraepithelial cells were pregated on CD103+, CD3+ to depict IEL and then further gated on TCRγδ and TCRβ as shown. B) Percent (mean ±SE) γδ IEL (defined as percent of total IEL that were TCRγδ+, TCRβ− IEL) in the distal small intestines of 1 wk, 2 wk, 3 wk, and adult mice. Data are representative of 3 independent experiments (*p<0.05 when compared to 1 wk samples).
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pone-0099042-g004: γδ IEL are the predominant IEL subtype in the immature murine small intestine.A) Flow cytometry of distal small intestinal intraepithelial cells from 1 week old (1 wk, n = 4), 2 weeks old (2 wk, n = 3), 3 weeks old (3 wk, n = 2) and adult (n = 1) C57BL/6J mice stained for CD3, CD8α, CD103, TCRγδ and TCRβ as described. Intraepithelial cells were pregated on CD103+, CD3+ to depict IEL and then further gated on TCRγδ and TCRβ as shown. B) Percent (mean ±SE) γδ IEL (defined as percent of total IEL that were TCRγδ+, TCRβ− IEL) in the distal small intestines of 1 wk, 2 wk, 3 wk, and adult mice. Data are representative of 3 independent experiments (*p<0.05 when compared to 1 wk samples).

Mentions: Young mice are frequently used for NEC-like injury models and correlating the maturity of the mucosal immune system between neonatal mice and humans is complex [33]. In addition, the human data on postnatal development may have been skewed, as neonatal intestinal tissue samples cannot be obtained from healthy neonates. Therefore we isolated epithelial-associated immune cells from the small intestines of wild type neonatal mice ages 1 week to adult (Figure 4A). γδ IEL were the predominant IEL subtype in younger mice (73% in 1 week old mice versus 59% in adult mice, p<0.05), with frequency approaching adult levels by 3 weeks of life (60%, p<0.05 vs. 1 week old) (Figure 4B).


Small intestinal intraepithelial TCRγδ+ T lymphocytes are present in the premature intestine but selectively reduced in surgical necrotizing enterocolitis.

Weitkamp JH, Rosen MJ, Zhao Z, Koyama T, Geem D, Denning TL, Rock MT, Moore DJ, Halpern MD, Matta P, Denning PW - PLoS ONE (2014)

γδ IEL are the predominant IEL subtype in the immature murine small intestine.A) Flow cytometry of distal small intestinal intraepithelial cells from 1 week old (1 wk, n = 4), 2 weeks old (2 wk, n = 3), 3 weeks old (3 wk, n = 2) and adult (n = 1) C57BL/6J mice stained for CD3, CD8α, CD103, TCRγδ and TCRβ as described. Intraepithelial cells were pregated on CD103+, CD3+ to depict IEL and then further gated on TCRγδ and TCRβ as shown. B) Percent (mean ±SE) γδ IEL (defined as percent of total IEL that were TCRγδ+, TCRβ− IEL) in the distal small intestines of 1 wk, 2 wk, 3 wk, and adult mice. Data are representative of 3 independent experiments (*p<0.05 when compared to 1 wk samples).
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Related In: Results  -  Collection

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pone-0099042-g004: γδ IEL are the predominant IEL subtype in the immature murine small intestine.A) Flow cytometry of distal small intestinal intraepithelial cells from 1 week old (1 wk, n = 4), 2 weeks old (2 wk, n = 3), 3 weeks old (3 wk, n = 2) and adult (n = 1) C57BL/6J mice stained for CD3, CD8α, CD103, TCRγδ and TCRβ as described. Intraepithelial cells were pregated on CD103+, CD3+ to depict IEL and then further gated on TCRγδ and TCRβ as shown. B) Percent (mean ±SE) γδ IEL (defined as percent of total IEL that were TCRγδ+, TCRβ− IEL) in the distal small intestines of 1 wk, 2 wk, 3 wk, and adult mice. Data are representative of 3 independent experiments (*p<0.05 when compared to 1 wk samples).
Mentions: Young mice are frequently used for NEC-like injury models and correlating the maturity of the mucosal immune system between neonatal mice and humans is complex [33]. In addition, the human data on postnatal development may have been skewed, as neonatal intestinal tissue samples cannot be obtained from healthy neonates. Therefore we isolated epithelial-associated immune cells from the small intestines of wild type neonatal mice ages 1 week to adult (Figure 4A). γδ IEL were the predominant IEL subtype in younger mice (73% in 1 week old mice versus 59% in adult mice, p<0.05), with frequency approaching adult levels by 3 weeks of life (60%, p<0.05 vs. 1 week old) (Figure 4B).

Bottom Line: In human preterm infants, the proportion of IEL was reduced by 66% in 11 NEC ileum resections compared to 30 non-NEC controls (p<0.001).While γδ IEL dominated over conventional αβ IEL early in gestation in mice and in humans, γδ IEL were preferential decreased in the ileum of surgical NEC patients compared to non-NEC controls (50% reduction, p<0.05).Loss of IEL in human NEC was associated with downregulation of the Th17 transcription factor retinoic acid-related orphan nuclear hormone receptor C (RORC, p<0.001).

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Vanderbilt University School of Medicine and Monroe Carell Jr. Children's Hospital at Vanderbilt, Nashville, Tennessee, United States of America.

ABSTRACT

Background: Gastrointestinal barrier immaturity predisposes preterm infants to necrotizing enterocolitis (NEC). Intraepithelial lymphocytes (IEL) bearing the unconventional T cell receptor (TCR) γδ (γδ IEL) maintain intestinal integrity and prevent bacterial translocation in part through production of interleukin (IL) 17.

Objective: We sought to study the development of γδ IEL in the ileum of human infants and examine their role in NEC pathogenesis. We defined the ontogeny of γδ IEL proportions in murine and human intestine and subjected tcrδ-/- mice to experimental gut injury. In addition, we used polychromatic flow cytometry to calculate percentages of viable IEL (defined as CD3+ CD8+ CD103+ lymphocytes) and the fraction of γδ IEL in surgically resected tissue from infants with NEC and gestational age matched non-NEC surgical controls.

Results: In human preterm infants, the proportion of IEL was reduced by 66% in 11 NEC ileum resections compared to 30 non-NEC controls (p<0.001). While γδ IEL dominated over conventional αβ IEL early in gestation in mice and in humans, γδ IEL were preferential decreased in the ileum of surgical NEC patients compared to non-NEC controls (50% reduction, p<0.05). Loss of IEL in human NEC was associated with downregulation of the Th17 transcription factor retinoic acid-related orphan nuclear hormone receptor C (RORC, p<0.001). TCRδ-deficient mice showed increased severity of experimental gut injury (p<0.05) with higher TNFα expression but downregulation of IL17A.

Conclusion: Complimentary mouse and human data suggest a role of γδ IEL in IL17 production and intestinal barrier production early in life. Specific loss of the γδ IEL fraction may contribute to NEC pathogenesis. Nutritional or pharmacological interventions to support γδ IEL maintenance in the developing small intestine could serve as novel strategies for NEC prevention.

Show MeSH
Related in: MedlinePlus