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Frontiers in veterinary science2023; 10; 1110019; doi: 10.3389/fvets.2023.1110019

Hypoxia signaling in the equine small intestine: Expression and distribution of hypoxia inducible factors during experimental ischemia.

Abstract: Hypoxia inducible factors (HIF) are widely researched in human medicine for their role in different disease processes. The aim of this study was to investigate the expression and distribution of HIF in experimental small intestinal ischemia in the horse. Unassigned: In 14 horses under general anesthesia, segmental jejunal ischemia with 90% reduction in blood flow was induced. The horses were randomly divided into two groups of seven horses, one subjected to ischemic postconditioning (IPoC) by delayed reperfusion, and a control group (group C) undergoing undelayed reperfusion. Intestinal samples were taken pre-ischemia, after ischemia and after reperfusion. Following immunohistochemical staining for HIF1α and -2α, the immunoreactivity pattern in the small intestine was evaluated by light microscopy, and the mucosal enterocyte and muscularis staining were semi-quantitatively scored. Additionally, mucosal HIF1α protein levels were determined by an Enzyme Linked Immunosorbent Assay (ELISA), and mRNA levels of HIF1α and its target genes by a two-step real-time Reverse Transcriptase Polymerase Chain Reaction. Statistical comparison was performed between the groups and time points using parametric and non-parametric tests (p < 0.05). Unassigned: All cell types exhibited cytoplasmic and nuclear immunoreactivity for HIF1α. After reperfusion, the cytoplasmic staining of the crypt and villus enterocytes as well as the villus nuclear staining significantly increased, whereas the perinuclear granules in the crypts decreased. The protein levels showed a significant decrease in group C at reperfusion, with lower HIF1α levels in group C compared to group IPoC during ischemia and reperfusion. No other group differences could be detected. In the HIF2α stained slides, mild to moderate cytoplasmic staining yet no nuclear immunoreactivity of the enterocytes was observed, and no significant changes over time were noted. Unassigned: the changes in HIF1α immunoreactivity pattern and expression over time suggest that this transcription factor plays a role in the intestinal response to ischemia in horses. However, the current study could not identify an effect of IPoC on HIF distribution or expression.
Publication Date: 2023-02-24 PubMed ID: 36908508PubMed Central: PMC9998946DOI: 10.3389/fvets.2023.1110019Google Scholar: Lookup
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  • Journal Article

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This research investigates how hypoxia inducible factors (HIF), proteins that respond to low oxygen levels, are expressed and distributed in the small intestine of horses under experimental conditions of ischemia, a reduced blood flow. The results indicate that HIF1α, a type of HIF, might have a role in how the horse’s intestine reacts to ischemia, but the impact of a technique called ischemic postconditioning (IPoC) is unclear.

Introduction to the research

  • The research aimed to study the expression and distribution patterns of HIF in the context of small intestinal ischemia in horses. Ischemia refers to a condition that results from reduced blood supply to a particular body part.
  • The study was performed on a total of 14 horses which were subjected to general anesthesia. Their intestines were subjected to ischemia with a 90% reduction in blood flow.
  • These horses were segregated into two groups for experimentation – group C (control group) and a group subjected to IPoC by delayed reperfusion. Reperfusion means restoring the blood flow to the part of the body where blood supply was reduced or blocked.

Methodology and Results

  • Intestinal samples were collected pre-ischemia, post-ischemia, and post-reperfusion. The samples underwent immunohistochemical staining for HIF1α and HIF2α, and were analyzed under a light microscope.
  • The researchers found that all cell types exhibited a combination of cytoplasmic and nuclear immunoreactivity for HIF1α. Following reperfusion, the immunoreactivity patterns of specific enterocytes exhibit significant changes.
  • A significant decrease in the protein levels was evident in group C (control group) at reperfusion, and the HIF1α levels were found to be lower in comparison to the IPoC group during ischemia and reperfusion.
  • However, no significant changes over time were observed in HIF2α-stained slides, indicating its potential lesser role in the reaction to ischemia.

Conclusion

  • The study concludes that the observed alterations in the immunoreactivity pattern and expression of HIF1α over time suggest its role in the intestinal response to ischemia in horses.
  • Nevertheless, the research couldn’t identify any significant impact of IPoC on the distribution or expression of HIF.

The findings of this research taken in context with ongoing studies on HIF can potentially contribute to a better understanding of disease processes in humans and animals alike.

Cite This Article

APA
Verhaar N, de Buhr N, von Köckritz-Blickwede M, Dümmer K, Hewicker-Trautwein M, Pfarrer C, Dengler F, Kästner S. (2023). Hypoxia signaling in the equine small intestine: Expression and distribution of hypoxia inducible factors during experimental ischemia. Front Vet Sci, 10, 1110019. https://doi.org/10.3389/fvets.2023.1110019

Publication

ISSN: 2297-1769
NlmUniqueID: 101666658
Country: Switzerland
Language: English
Volume: 10
Pages: 1110019

Researcher Affiliations

Verhaar, Nicole
  • Clinic for Horses, University of Veterinary Medicine Hannover, Hannover, Germany.
de Buhr, Nicole
  • Department of Biochemistry, University of Veterinary Medicine Hannover, Hannover, Germany.
  • Research Center for Emerging Infections and Zoonoses, University of Veterinary Medicine Hannover, Hannover, Germany.
von Köckritz-Blickwede, Maren
  • Department of Biochemistry, University of Veterinary Medicine Hannover, Hannover, Germany.
  • Research Center for Emerging Infections and Zoonoses, University of Veterinary Medicine Hannover, Hannover, Germany.
Dümmer, Katrin
  • Department of Biochemistry, University of Veterinary Medicine Hannover, Hannover, Germany.
  • Research Center for Emerging Infections and Zoonoses, University of Veterinary Medicine Hannover, Hannover, Germany.
Hewicker-Trautwein, Marion
  • Institute of Pathology, University of Veterinary Medicine Hannover, Hannover, Germany.
Pfarrer, Christiane
  • Institute for Anatomy, University of Veterinary Medicine Hannover, Hannover, Germany.
Dengler, Franziska
  • Institute of Physiology, Pathophysiology and Biophysics, Department of Biomedical Sciences, University of Veterinary Medicine Vienna, Vienna, Austria.
Kästner, Sabine
  • Clinic for Horses, University of Veterinary Medicine Hannover, Hannover, Germany.
  • Small Animal Clinic, University of Veterinary Medicine Hannover, Hannover, Germany.

Conflict of Interest Statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

References

This article includes 52 references
  1. Tinker MK, White N, Lessard P, Thatcher C, Pelzer K, Davis B. Prospective study of equine colic incidence and mortality. Equine Vet J (1997) 29:448–53.
  2. Tirpe AA, Gulei D, Ciortea SM, Crivii C, Berindan-Neagoe I. Hypoxia: overview on hypoxia-mediated mechanisms with a focus on the role of HIF genes. Int J Mol Sci (2019) 20:6140.
    doi: 10.3390/ijms20246140pmc: PMC6941045pubmed: 31817513google scholar: lookup
  3. Semenza GL. Regulation of mammalian O2 homeostasis by hypoxia-inducible factor 1. Annu Rev Cell Dev Biol (1999) 15:551–78.
  4. Chan DA, Sutphin PD, Yen SE, Giaccia AJ. Coordinate regulation of the oxygen-dependent degradation domains of hypoxia-inducible factor 1 alpha. Mol Cell Biol (2005) 25:6415–26.
  5. Wang GL, Jiang BH, Rue EA, Semenza GL. Hypoxia-inducible factor 1 is a basic-helix-loop-helix-PAS heterodimer regulated by cellular O2 tension. Proc Natl Acad Sci U S A (1995) 92:5510–4.
    doi: 10.1073/pnas.92.12.5510pmc: PMC41725pubmed: 7539918google scholar: lookup
  6. Singhal R, Shah YM. Oxygen battle in the gut: hypoxia and hypoxia-inducible factors in metabolic and inflammatory responses in the intestine. J Biol Chem (2020) 295:10493–505.
    doi: 10.1074/jbc.REV120.011188pmc: PMC7383395pubmed: 32503843google scholar: lookup
  7. Jing X, Yang F, Shao C, Wei K, Xie M, Shen H. Role of hypoxia in cancer therapy by regulating the tumor microenvironment. Mol Cancer (2019) 18:157.
    doi: 10.1186/s12943-019-1089-9pmc: PMC6844052pubmed: 31711497google scholar: lookup
  8. Akhtar M, Sutherland A, Huang H, Ploeg R, Pugh C. The role of hypoxia-inducible factors in organ donation and transplantation: the current perspective and future opportunities. Am J Transplant (2014) 14:1481–7.
    doi: 10.1111/ajt.12737pubmed: 24909061google scholar: lookup
  9. Saeedi BJ, Kao DJ, Kitzenberg DA, Dobrinskikh E, Schwisow KD, Masterson JC. HIF-dependent regulation of claudin-1 is central to intestinal epithelial tight junction integrity. Mol Biol Cell (2015) 26:2252–62.
    doi: 10.1091/mbc.E14-07-1194pmc: PMC4462943pubmed: 25904334google scholar: lookup
  10. Synnestvedt K, Furuta GT, Comerford KM, Louis N, Karhausen J, Eltzschig HK. Ecto-5'-nucleotidase (CD73) regulation by hypoxia-inducible factor-1 mediates permeability changes in intestinal epithelia. J Clin Invest (2002) 110:993–1002.
    doi: 10.1172/JCI0215337pmc: PMC151145pubmed: 12370277google scholar: lookup
  11. Yang S, Yu M, Sun L, Xiao W, Yang X, Sun L. Interferon-γ-induced intestinal epithelial barrier dysfunction by NF-κB/HIF-1α pathway. J Interferon Cytokine Res (2014) 34:195–203.
    doi: 10.1089/jir.2013.0044pubmed: 24237301google scholar: lookup
  12. Bäcker V, Cheung F-Y, Siveke JT, Fandrey J, Winning S. Knockdown of myeloid cell hypoxia-inducible factor-1α ameliorates the acute pathology in DSS-induced colitis. PLoS ONE (2017) 12:e0190074.
  13. Karhausen J, Furuta GT, Tomaszewski JE, Johnson RS, Colgan SP, Haase VH. Epithelial hypoxia-inducible factor-1 is protective in murine experimental colitis. J Clin Invest (2004) 114:1098–106.
    doi: 10.1172/JCI200421086pmc: PMC522241pubmed: 15489957google scholar: lookup
  14. Grenz A, Clambey E, Eltzschig HK. Hypoxia signaling during intestinal ischemia and inflammation. Curr Opin Crit Care (2012) 18:178–85.
  15. Koury J, Deitch EA, Homma H, Abungu B, Gangurde P, Condon MR. Persistent HIF-1α activation in gut ischemia/reperfusion injury: potential role of bacteria and lipopolysaccharide. Shock (2004) 22:270–7.
  16. Kannan KB, Colorado I, Reino D, Palange D, Lu Q, Qin X. Hypoxia-inducible factor plays a gut-injurious role in intestinal ischemia reperfusion injury. Am J Physiol Gastrointest Liver Physiol (2011) 300:G853–61.
    doi: 10.1152/ajpgi.00459.2010pmc: PMC3094138pubmed: 21183660google scholar: lookup
  17. Feinman R, Deitch EA, Watkins AC, Abungu B, Colorado I, Kannan KB. HIF-1 mediates pathogenic inflammatory responses to intestinal ischemia-reperfusion injury. Am J Physiol Gastrointest Liver Physiol (2010) 299:G833–43.
    doi: 10.1152/ajpgi.00065.2010pmc: PMC2957330pubmed: 20689059google scholar: lookup
  18. Bauck AG, Grosche A, Morton AS, Vickroy TW, Freeman DE. Effect of lidocaine on in ammation in equine jejunum subjected to manipulation only and remote to intestinal segments subjected to ischemia. Am J Vet Res (2017) 78:977–89.
    doi: 10.2460/ajvr.78.8.977pubmed: 28738006google scholar: lookup
  19. De Ceulaer K, Delesalle C, Van Elzen R, Van Brantegem L, Weyns A, Van Ginneken C. Morphological data indicate a stress response at the oral border of strangulated small intestine in horses. Res Vet Sci (2011) 91:294–300.
    doi: 10.1016/j.rvsc.2010.11.020pubmed: 21216416google scholar: lookup
  20. Mastrogiannaki M, Matak P, Keith B, Simon MC, Vaulont S, Peyssonnaux C. HIF-2α, but not HIF-1α, promotes iron absorption in mice. J Clin Invest (2009) 119:1159–66.
    doi: 10.1172/JCI38499pmc: PMC2673882pubmed: 19352007google scholar: lookup
  21. Giatromanolaki A, Sivridis E, Maltezos E, Papazoglou D, Simopoulos C, Gatter KC. Hypoxia inducible factor 1alpha and 2alpha overexpression in inflammatory bowel disease. J Clin Pathol (2003) 56:209–13.
    doi: 10.1136/jcp.56.3.209pmc: PMC1769899pubmed: 12610101google scholar: lookup
  22. Wiesener MS, Jürgensen JS, Rosenberger C, Scholze C, Hörstrup JH, Warnecke C. Widespread, hypoxia-inducible expression of HIF-2α in distinct cell populations of different organs. FASEB J (2003) 17:271–3.
    doi: 10.1096/fj.02-0445fjepubmed: 12490539google scholar: lookup
  23. Makino Y, Cao R, Svensson K, Bertilsson G, Asman M, Tanaka H. Inhibitory PAS domain protein is a negative regulator of hypoxia-inducible gene expression. Nature (2001) 414:550–4.
    doi: 10.1038/35107085pubmed: 11734856google scholar: lookup
  24. Duan C. Hypoxia-inducible factor 3 biology: complexities and emerging themes. Am J Physiol Cell Physiol (2016) 310:C260–9.
    doi: 10.1152/ajpcell.00315.2015pubmed: 26561641google scholar: lookup
  25. Zhao ZQ, Corvera JS, Halkos ME, Kerendi F, Wang NP, Guyton RA. Inhibition of myocardial injury by ischemic postconditioning during reperfusion: comparison with ischemic preconditioning. Am J Physiol Heart Circ Physiol (2003) 285:579–88.
    doi: 10.1152/ajpheart.01064.2002pubmed: 12860564google scholar: lookup
  26. Jia Z, Lian W, Shi H, Cao C, Han S, Wang K. Ischemic postconditioning protects against intestinal ischemia/reperfusion injury via the HIF-1alpha/miR-21 axis. Sci Rep (2017) 7:16190.
    doi: 10.1038/s41598-017-16366-6pmc: PMC5700993pubmed: 29170412google scholar: lookup
  27. Verhaar N, Breves G, Hewicker-Trautwein M, Pfarrer C, Rohn K, Burmester M. The effect of ischaemic postconditioning on mucosal integrity and function in equine jejunal ischaemia. Equine Vet J (2022) 54:427–37.
    doi: 10.1111/evj.13450pubmed: 34003501google scholar: lookup
  28. Zhao H-X, Wang X-L, Wang Y-H, Wu Y, Li X-Y, Lv X-P. Attenuation of myocardial injury by postconditioning: role of hypoxia inducible factor-1α. Basic Res Cardiol (2010) 105:109.
    doi: 10.1007/s00395-009-0044-0pubmed: 19597757google scholar: lookup
  29. Liu Y, Nie H, Zhang K, Ma D, Yang G, Zheng Z. A feedback regulatory loop between HIF-1α and miR-21 in response to hypoxia in cardiomyocytes. FEBS Lett (2014) 588:3137–46.
    doi: 10.1016/j.febslet.2014.05.067pubmed: 24983504google scholar: lookup
  30. Wan D, Zhang Z, Yang H. Cardioprotective effect of miR-214 in myocardial ischemic postconditioning by down-regulation of hypoxia inducible factor 1, alpha subunit inhibitor. Cell Mol Biol (2015) 61:1.
    pubmed: 26025394
  31. Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem (1976) 72:248–54.
    doi: 10.1016/0003-2697(76)90527-3pubmed: 942051google scholar: lookup
  32. Pfaffl MW, Horgan GW, Dempfle L. Relative expression software tool (REST) for group-wise comparison and statistical analysis of relative expression results in real-time PCR. Nucleic Acids Res (2002) 30:e36.
    doi: 10.1093/nar/30.9.e36pmc: PMC113859pubmed: 11972351google scholar: lookup
  33. Tuboly E, Futakuchi M, Varga G, Érces D, Tokés T, Mészáros A. C5a inhibitor protects against ischemia/reperfusion injury in rat small intestine. Microbiol Immunol (2016) 60:35–46.
    doi: 10.1111/1348-0421.12338pmc: PMC4819679pubmed: 26576826google scholar: lookup
  34. Blikslager AT. The Equine Acute Abdomen. Hoboken, NJ: John Wiley and Sons. (2017).
    doi: 10.1002/9781119063254google scholar: lookup
  35. Krock BL, Skuli N, Simon MC. Hypoxia-induced angiogenesis: good and evil. Genes Cancer (2011) 2:1117–33.
    doi: 10.1177/1947601911423654pmc: PMC3411127pubmed: 22866203google scholar: lookup
  36. Ji Z-P, Li Y-X, Shi B-X, Zhuang Z-N, Yang J-Y, Guo S. preconditioning protects Ca2+-ATPase activation of intestinal mucosal cells against R/I injury in a rat liver transplantation model. World J Gastroenterol (2018) 24:360.
    doi: 10.3748/wjg.v24.i3.360pmc: PMC5776397pubmed: 29391758google scholar: lookup
  37. Scharte M, Han X, Bertges DJ, Fink MP, Delude RL. Cytokines induce HIF-1 DNA binding and the expression of HIF-1-dependent genes in cultured rat enterocytes. Am J Physiol Gastrointest Liver Physiol (2003) 284:G373–84.
    doi: 10.1152/ajpgi.00076.2002pubmed: 12388200google scholar: lookup
  38. Garcia-Vasquez C, Fernandez-Acenero MJ, Garcia Gomez-Heras S, Pastor C. Fibrin patch influences the expression of hypoxia-inducible factor-1alpha and nuclear factor-kappaBp65 factors on ischemic intestinal anastomosis. Exp Biol Med (2018) 243:803–8.
    doi: 10.1177/1535370218777216pmc: PMC6022912pubmed: 29932372google scholar: lookup
  39. Hellwig-Bürgel T, Rutkowski K, Metzen E, Fandrey J, Jelkmann W. Interleukin-1β and tumor necrosis factor-α stimulate DNA binding of hypoxia-inducible factor-1. Blood (1999) 94:1561–7.
  40. Verhaar N, de Buhr N, von Köckritz-Blickwede M, Hewicker-Trautwein M, Pfarrer C, Mazzuoli-Weber G. Ischaemic postconditioning reduces apoptosis in experimental jejunal ischaemia in horses. BMC Vet Res (2021) 17:1–14.
    doi: 10.1186/s12917-021-02877-ypmc: PMC8077964pubmed: 33902575google scholar: lookup
  41. Ye Z, Guo Q, Xia P, Wang N, Wang E, Yuan Y. Sevoflurane postconditioning involves an up-regulation of HIF-1α and HO-1 expression via PI3K/Akt pathway in a rat model of focal cerebral ischemia. Brain Res (2012) 1463:63–74.
  42. Raphael J, Zuo Z, Abedat S, Beeri R, Gozal Y. Isoflurane preconditioning decreases myocardial infarction in rabbits via up-regulation of hypoxia inducible factor 1 that is mediated by mammalian target of rapamycin. J Am Soc Anesthesiol (2008) 108:415–25.
    doi: 10.1097/ALN.0b013e318164cab1pubmed: 18292679google scholar: lookup
  43. Wang C, Weihrauch D, Schwabe DA, Bienengraeber M, Warltier DC, Kersten JR. Extracellular signal-regulated kinases trigger isoflurane preconditioning concomitant with upregulation of hypoxia-inducible factor-1α and vascular endothelial growth factor expression in rats. Anesth Analg (2006) 103:281–8.
  44. Ngamsri K-C, Fabian F, Fuhr A, Gamper-Tsigaras J, Straub A, Fecher D. Sevoflurane exerts protective effects in murine peritonitis-induced sepsis via hypoxia-inducible factor 1α/adenosine A2B receptor signaling. Anesthesiology (2021) 135:136–50.
    doi: 10.1097/ALN.0000000000003788pubmed: 33914856google scholar: lookup
  45. Mariani F, Sena P, Marzona L, Riccio M, Fano R, Manni P. Cyclooxygenase-2 and hypoxia-inducible factor-1α protein expression is related to inflammation, and up-regulated since the early steps of colorectal carcinogenesis. Cancer Lett (2009) 279:221–9.
    doi: 10.1016/j.canlet.2009.02.001pubmed: 19268443google scholar: lookup
  46. Li S, Yao D, Wang L, Wu W, Qiu L, Yao M. characteristics of hypoxia-inducible factor-1α and its clinical values in diagnosis and prognosis of hepatocellular carcinoma. Hepat Mon (2011) 11:821–8.
  47. Armando F, Gambini M, Corradi A, Giudice C, Pfankuche VM, Brogden G. Oxidative stress in canine histiocytic sarcoma cells induced by an infection with canine distemper virus led to a dysregulation of HIF-1α downstream pathway resulting in a reduced expression of VEGF-B in vitro. Viruses (2020) 12:200.
    doi: 10.3390/v12020200pmc: PMC7077254pubmed: 32054075google scholar: lookup
  48. Nishie H, Takahashi T, Inoue K, Shimizu H, Morimatsu H, Toda Y. Site-specific induction of intestinal hypoxia-inducible factor-1α after hemorrhagic shock. Mol Med Rep (2009) 2:149–52.
    doi: 10.3892/mmr_00000075pubmed: 21475804google scholar: lookup
  49. Dengler F, Sternberg F, Grages M, Kästner SB, Verhaar N. Adaptive mechanisms in no flow vs. low flow ischemia in equine jejunum epithelium: different paths to the same destination. Front Vet Sci 9:947482. (2022).
    doi: 10.3389/fvets.2022.947482pmc: PMC9493374pubmed: 36157182google scholar: lookup
  50. Ebersole JL, Novak MJ, Orraca L, Martinez-Gonzalez J, Kirakodu S, Chen KC. Hypoxia-inducible transcription factors, HIF1A and HIF2A, increase in aging mucosal tissues. Immunology (2018) 154:452–64.
    doi: 10.1111/imm.12894pmc: PMC6002220pubmed: 29338076google scholar: lookup
  51. Chen Y, Lee S-H, Tsai Y-H, Tseng S-H. Ischemic preconditioning increased the intestinal stem cell activities in the intestinal crypts in mice. J Surg Res (2014) 187:85–93.
    doi: 10.1016/j.jss.2013.10.001pubmed: 24176207google scholar: lookup
  52. Movafagh S, Crook S, Vo K. Regulation of hypoxia-inducible factor-1a by reactive oxygen species: new developments in an old debate. J Cell Biochem (2015) 116:696–703.
    doi: 10.1002/jcb.25074pubmed: 25546605google scholar: lookup