Inhibition of HIF-prolyl hydroxylases improves healing of intestinal anastomoses
Moritz J. Strowitzki, Gwendolyn Kimmer, Julian Wehrmann, Alina S. Ritter, Praveen Radhakrishnan, Vanessa M. Opitz, Christopher Tuffs, Marvin Biller, Julia Kugler, Ulrich Keppler, Jonathan M. Harnoss, Johannes Klose, Thomas Schmidt, Alfonso Blanco, Cormac T. Taylor, Martin Schneider
Moritz J. Strowitzki, Gwendolyn Kimmer, Julian Wehrmann, Alina S. Ritter, Praveen Radhakrishnan, Vanessa M. Opitz, Christopher Tuffs, Marvin Biller, Julia Kugler, Ulrich Keppler, Jonathan M. Harnoss, Johannes Klose, Thomas Schmidt, Alfonso Blanco, Cormac T. Taylor, Martin Schneider
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Research Article Gastroenterology Inflammation

Inhibition of HIF-prolyl hydroxylases improves healing of intestinal anastomoses

Abstract

Anastomotic leakage (AL) accounts for a major part of in-house mortality in patients undergoing colorectal surgery. Local ischemia and abdominal sepsis are common risk factors contributing to AL and are characterized by upregulation of the hypoxia-inducible factor (HIF) pathway. The HIF pathway is critically regulated by HIF-prolyl hydroxylases (PHDs). Here, we investigated the significance of PHDs and the effects of pharmacologic PHD inhibition (PHI) during anastomotic healing. Ischemic or septic colonic anastomoses were created in mice by ligation of mesenteric vessels or lipopolysaccharide-induced abdominal sepsis, respectively. Genetic PHD deficiency (Phd1–/–, Phd2+/–, and Phd3–/–) or PHI were applied to manipulate PHD activity. Pharmacologic PHI and genetic PHD2 haplodeficiency (Phd2+/–) significantly improved healing of ischemic or septic colonic anastomoses, as indicated by increased bursting pressure and reduced AL rates. Only Phd2+/– (but not PHI or Phd1–/–) protected from sepsis-related mortality. Mechanistically, PHI and Phd2+/– induced immunomodulatory (M2) polarization of macrophages, resulting in increased collagen content and attenuated inflammation-driven immune cell recruitment. We conclude that PHI improves healing of colonic anastomoses in ischemic or septic conditions by Phd2+/–-mediated M2 polarization of macrophages, conferring a favorable microenvironment for anastomotic healing. Patients with critically perfused colorectal anastomosis or abdominal sepsis could benefit from pharmacologic PHI.

Authors

Moritz J. Strowitzki, Gwendolyn Kimmer, Julian Wehrmann, Alina S. Ritter, Praveen Radhakrishnan, Vanessa M. Opitz, Christopher Tuffs, Marvin Biller, Julia Kugler, Ulrich Keppler, Jonathan M. Harnoss, Johannes Klose, Thomas Schmidt, Alfonso Blanco, Cormac T. Taylor, Martin Schneider

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Figure 2

Haplodeficiency of HIF-prolyl hydroxylase 2 augments healing of ischemic colonic anastomoses.

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Haplodeficiency of HIF-prolyl hydroxylase 2 augments healing of ischemic...
(A) Experimental schedule. Surgical anastomosis and blood vessel ligation were performed in WT, Phd1-deficient (Phd1–/–), and Phd2-haplodeficient (Phd2+/–) mice, and evaluation was performed 3 days thereafter. (B) Representative images of ischemic colonic anastomoses (dotted white lines) from WT and Phd2+/– mice. Note indirect signs of AL, such as severe adhesion (§) and abscess (white arrowheads). (C) Bursting pressure analysis of ischemic colonic anastomoses harvested from WT, Phd1–/–, and Phd2+/– mice (n = 10–11 animals per group; ANOVA with post hoc test). (D) Left: Representative MTG stainings, revealing the thickness of collagen capsules (black dashed lines) close to sutures (§) in ischemic anastomoses harvested from WT and Phd2+/– mice (scale bar represents 100 μm). Right: Histomorphometric quantification of collagen capsule thickness in ischemic colonic anastomoses from WT and Phd2+/– mice (n = 4 animals per group; Mann-Whitney U test). (E) Representative immunolabeling of leukocytes (CD45, left upper panel) and macrophages (F4/80, left lower panel) in ischemic anastomoses from WT and Phd2+/– mice and histomorphometric quantification (right) (n = 3–4 animals per group; scale bar represents 200 μm; § and # indicate positions of [extracted] sutures and gut lumen, respectively). HPF, high-power field.