Skip to main content
SpringerLink
Log in

Bile acids inhibit Na+/H+ exchanger and Cl−/HCO3 − exchanger activities via cellular energy breakdown and Ca2+ overload in human colonic crypts

  • Molecular and cellular mechanisms of disease
  • Published:
Pflügers Archiv - European Journal of Physiology Aims and scope Submit manuscript

Abstract

Bile acids play important physiological role in the solubilisation and absorption of dietary lipids. However, under pathophysiological conditions, such as short bowel syndrome, they can reach the colon in high concentrations inducing diarrhoea. In this study, our aim was to characterise the cellular pathomechanism of bile-induced diarrhoea using human samples. Colonic crypts were isolated from biopsies of patients (controls with negative colonoscopic findings) and of cholecystectomised/ileum-resected patients with or without diarrhoea. In vitro measurement of the transporter activities revealed impaired Na+/H+ exchanger (NHE) and Cl−/HCO3 − exchanger (CBE) activities in cholecystectomised/ileum-resected patients suffering from diarrhoea, compared to control patients. Acute treatment of colonic crypts with 0.3 mM chenodeoxycholate caused dose-dependent intracellular acidosis; moreover, the activities of acid/base transporters (NHE and CBE) were strongly impaired. This concentration of chenodeoxycholate did not cause morphological changes in colonic epithelial cells, although significantly reduced the intracellular ATP level, decreased mitochondrial transmembrane potential and caused sustained intracellular Ca2+ elevation. We also showed that chenodeoxycholate induced Ca2+ release from the endoplasmic reticulum and extracellular Ca2+ influx contributing to the Ca2+ elevation. Importantly, our results suggest that the chenodeoxycholate-induced inhibition of NHE activities was ATP-dependent, whereas the inhibition of CBE activity was mediated by the sustained Ca2+ elevation. We suggest that bile acids inhibit the function of ion transporters via cellular energy breakdown and Ca2+ overload in human colonic epithelial cells, which can reduce fluid and electrolyte absorption in the colon and promote the development of diarrhoea.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

Abbreviations

BA:

Bile acids

BAM:

Bile acid malabsorption

BAPTA-AM:

1,2-Bis(o-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid

[Ca2+]i :

Intracellular Ca2+ concentration

CBE:

Cl−/HCO3 − exchanger

CCCP:

Carbonyl cyanide m-chlorophenyl hydrazone

CDC:

Chenodeoxycholate

ER:

Endoplasmic reticulum

GCDC:

Glycochenodeoxycholate

H2DIDS:

Dihydro-4,4′-diisothiocyanostilbene-2,2′-disulphonic acid

HOE-642:

4-Isopropyl-3-methylsulphonylbenzoyl-guanidin methanesulphonate

IP3R:

Inositol-triphosphate receptor

(ΔΨ)m :

Mitochondrial transmembrane potential

NHE:

Na+/H+ exchanger

pHi :

Intracellular pH

RR:

Ruthenium red

RyR:

Ryanodin receptor

SERCA:

Sarcoplasmic/endoplasmic reticulum calcium ATPase

SLC26:

Solute carrier family 26

TEM:

Transmission electron microscopy

Tg:

Thapsigargin

References

  1. Arlow FL, Dekovich AA, Priest RJ, Beher WT (1987) Bile acid-mediated postcholecystectomy diarrhea. Arch Intern Med 147(7):1327–1329

    Article  CAS  PubMed  Google Scholar 

  2. Bachmann O, Riederer B, Rossmann H, Groos S, Schultheis PJ, Shull GE, Gregor M, Manns MP, Seidler U (2004) The Na+/H+ exchanger isoform 2 is the predominant NHE isoform in murine colonic crypts and its lack causes NHE3 upregulation. Am J Physiol Gastrointest Liver Physiol 287(1):G125–G133

  3. Borghede MK, Schlutter JM, Agnholt JS, Christensen LA, Gormsen LC, Dahlerup JF (2011) Bile acid malabsorption investigated by selenium-75-homocholic acid taurine ((75)SeHCAT) scans: causes and treatment responses to cholestyramine in 298 patients with chronic watery diarrhoea. Eur J Int Med 22(6):e137–e140

    Article  CAS  Google Scholar 

  4. Byrne AM, Foran E, Sharma R, Davies A, Mahon C, O'Sullivan J, O’Donoghue D, Kelleher D (2010) Long A bile acids modulate the Golgi membrane fission process via a protein kinase Ceta and protein kinase D-dependent pathway in colonic epithelial cells. Carcinogenesis 31(4):737–744

    Article  CAS  PubMed  Google Scholar 

  5. Cabado AG, Yu FH, Kapus A, Lukacs G, Grinstein S, Orlowski J (1996) Distinct structural domains confer cAMP sensitivity and ATP dependence to the Na+/H+ exchanger NHE3 isoform. J Biol Chem 271(7):3590–3599

  6. Cinar A, Chen M, Riederer B, Bachmann O, Wiemann M, Manns M, Kocher O, Seidler U (2007) NHE3 inhibition by cAMP and Ca2+ is abolished in PDZ-domain protein PDZK1-deficient murine enterocytes. J Physiol 581(Pt 3):1235–1246

  7. Criddle DN, Murphy J, Fistetto G, Barrow S, Tepikin AV, Neoptolemos JP, Sutton R, Petersen OH (2006) Fatty acid ethyl esters cause pancreatic calcium toxicity via inositol trisphosphate receptors and loss of ATP synthesis. Gastroenterology 130(3):781–793

    Article  CAS  PubMed  Google Scholar 

  8. Farkas K, Yeruva S, Rakonczay Z Jr, Ludolph L, Molnar T, Nagy F, Szepes Z, Schnur A, Wittmann T, Hubricht J, Riederer B, Venglovecz V, Lazar G, Kiraly M, Zsembery A, Varga G, Seidler U, Hegyi P (2011) New therapeutic targets in ulcerative colitis: the importance of ion transporters in the human colon. Inflamm Bowel Dis 17(4):884–898

    Article  PubMed  Google Scholar 

  9. Ford GA, Preece JD, Davies IH, Wilkinson SP (1992) Use of the SeHCAT test in the investigation of diarrhoea. Postgrad Med J 68(798):272–276

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  10. Fromm H, Malavolti M (1986) Bile acid-induced diarrhoea. Clin Gastroenterol 15(3):567–582

    CAS  PubMed  Google Scholar 

  11. Gawenis LR, Stien X, Shull GE, Schultheis PJ, Woo AL, Walker NM, Clarke LL (2002) Intestinal NaCl transport in NHE2 and NHE3 knockout mice. Am J Physiol Gastrointest Liver Physiol 282(5):G776–G784

    Article  CAS  PubMed  Google Scholar 

  12. Gerasimenko JV, Flowerdew SE, Voronina SG, Sukhomlin TK, Tepikin AV, Petersen OH, Gerasimenko OV (2006) Bile acids induce Ca2+ release from both the endoplasmic reticulum and acidic intracellular calcium stores through activation of inositol trisphosphate receptors and ryanodine receptors. J Biol Chem 281(52):40154–40163

    Article  CAS  PubMed  Google Scholar 

  13. Gerasimenko JV, Sherwood M, Tepikin AV, Petersen OH, Gerasimenko OV (2006) NAADP, cADPR and IP3 all release Ca2+ from the endoplasmic reticulum and an acidic store in the secretory granule area. J Cell Sci 119(Pt 2):226–238

  14. Hamilton JP, Xie G, Raufman JP, Hogan S, Griffin TL, Packard CA, Chatfield DA, Hagey LR, Steinbach JH, Hofmann AF (2007) Human cecal bile acids: concentration and spectrum. Am J Physiol Gastrointest Liver Physiol 293(1):G256–G263

    Article  CAS  PubMed  Google Scholar 

  15. Hegyi P, Gray MA, Argent BE (2003) Substance P inhibits bicarbonate secretion from guinea pig pancreatic ducts by modulating an anion exchanger. Am J Physiol Cell Physiol 285(2):C268–C276

    Article  CAS  PubMed  Google Scholar 

  16. Hegyi P, Rakonczay Z Jr, Gray MA, Argent BE (2004) Measurement of intracellular pH in pancreatic duct cells: a new method for calibrating the fluorescence data. Pancreas 28(4):427–434

    Article  PubMed  Google Scholar 

  17. Hofmann AF (2009) The enterohepatic circulation of bile acids in mammals: form and functions. Front Biosci (Landmark Ed) 14:2584–2598

    Article  CAS  Google Scholar 

  18. Hoglund P, Haila S, Scherer SW, Tsui LC, Green ED, Weissenbach J, Holmberg C, de la Chapelle A, Kere J (1996) Positional candidate genes for congenital chloride diarrhea suggested by high-resolution physical mapping in chromosome region 7q31. Genome Res 6(3):202–210

    Article  CAS  PubMed  Google Scholar 

  19. Hoogerwerf WA, Tsao SC, Devuyst O, Levine SA, Yun CH, Yip JW, Cohen ME, Wilson PD, Lazenby AJ, Tse CM, Donowitz M (1996) NHE2 and NHE3 are human and rabbit intestinal brush-border proteins. Am J Physiol 270(1 Pt 1):G29–G41

    CAS  PubMed  Google Scholar 

  20. Hylemon PB, Zhou H, Pandak WM, Ren S, Gil G, Dent P (2009) Bile acids as regulatory molecules. J Lipid Res 50(8):1509–1520

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  21. Inoue M, Fujishiro N, Imanaga I, Sakamoto Y (2002) Role of ATP decrease in secretion induced by mitochondrial dysfunction in guinea-pig adrenal chromaffin cells. J Physiol 539(Pt 1):145–155

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  22. Keely SJ (2010) Missing link identified: GpBAR1 is a neuronal bile acid receptor. Neurogastroenterol Motil 22(7):711–717

    Article  CAS  PubMed  Google Scholar 

  23. Kiela PR, Xu H, Ghishan FK (2006) Apical Na+/H + exchangers in the mammalian gastrointestinal tract. J Physiol Pharmacol 57(Suppl 7):51–79

  24. Kunzelmann K, Mall M (2002) Electrolyte transport in the mammalian colon: mechanisms and implications for disease. Physiol Rev 82(1):245–289

    CAS  PubMed  Google Scholar 

  25. Lamprecht G, Hsieh CJ, Lissner S, Nold L, Heil A, Gaco V, Schafer J, Turner JR, Gregor M (2009) Intestinal anion exchanger down-regulated in adenoma (DRA) is inhibited by intracellular calcium. J Biol Chem 284(29):19744–19753

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  26. Leyssens A, Nowicky AV, Patterson L, Crompton M, Duchen MR (1996) The relationship between mitochondrial state, ATP hydrolysis, [Mg2+]i and [Ca2+]i studied in isolated rat cardiomyocytes. J Physiol 496(Pt 1):111–128

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  27. Makela S, Kere J, Holmberg C, Hoglund P (2002) SLC26A3 mutations in congenital chloride diarrhea. Hum Mutat 20(6):425–438

    Article  CAS  PubMed  Google Scholar 

  28. Maleth J, Venglovecz V, Razga Z, Tiszlavicz L, Rakonczay Z Jr, Hegyi P (2011) Non-conjugated chenodeoxycholate induces severe mitochondrial damage and inhibits bicarbonate transport in pancreatic duct cells. Gut 60(1):136–138

  29. Musch MW, Arvans DL, Wu GD, Chang EB (2009) Functional coupling of the downregulated in adenoma Cl-/base exchanger DRA and the apical Na+/H + exchangers NHE2 and NHE3. Am J Physiol Gastrointest Liver Physiol 296(2):G202–G210

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  30. Payne CM, Crowley-Weber CL, Dvorak K, Bernstein C, Bernstein H, Holubec H, Crowley C, Garewal H (2005) Mitochondrial perturbation attenuates bile acid-induced cytotoxicity. Cell Biol Toxicol 21(5–6):215–231

    Article  CAS  PubMed  Google Scholar 

  31. Petersen OH, Tepikin AV (2008) Polarized calcium signaling in exocrine gland cells. Annu Rev Physiol 70:273–299

    Article  CAS  PubMed  Google Scholar 

  32. Philipp B (2011) Bacterial degradation of bile salts. Appl Microbiol Biotechnol 89(4):903–915

    Article  CAS  PubMed  Google Scholar 

  33. Rossel P, Sortsoe Jensen H, Qvist P, Arveschoug A (1999) Prognosis of adult-onset idiopathic bile acid malabsorption. Scand J Gastroenterol 34(6):587–590

    Article  CAS  PubMed  Google Scholar 

  34. Schultheis PJ, Clarke LL, Meneton P, Miller ML, Soleimani M, Gawenis LR, Riddle TM, Duffy JJ, Doetschman T, Wang T, Giebisch G, Aronson PS, Lorenz JN, Shull GE (1998) Renal and intestinal absorptive defects in mice lacking the NHE3 Na+/H+ exchanger. Nat Genet 19(3):282–285

  35. Schweinfest CW, Spyropoulos DD, Henderson KW, Kim JH, Chapman JM, Barone S, Worrell RT, Wang Z, Soleimani M (2006) slc26a3 (dra)-deficient mice display chloride-losing diarrhea, enhanced colonic proliferation, and distinct up-regulation of ion transporters in the colon. J Biol Chem 281(49):37962–37971

    Article  CAS  PubMed  Google Scholar 

  36. Sciarretta G, Furno A, Mazzoni M, Malaguti P (1992) Post-cholecystectomy diarrhea: evidence of bile acid malabsorption assessed by SeHCAT test. Am J Gastroenterol 87(12):1852–1854

    CAS  PubMed  Google Scholar 

  37. Shimada-Shimizu N, Hisamitsu T, Nakamura TY, Wakabayashi S (2013) Evidence that Na+/H+ exchanger 1 is an ATP-binding protein. Febs J 280(6):1430–1442

  38. Shiue H, Musch MW, Wang Y, Chang EB, Turner JR (2005) Akt2 phosphorylates ezrin to trigger NHE3 translocation and activation. J Biol Chem 280(2):1688–1695

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  39. Smith MJ, Cherian P, Raju GS, Dawson BF, Mahon S, Bardhan KD (2000) Bile acid malabsorption in persistent diarrhoea. J R Coll Physicians Lond 34(5):448–451

    CAS  PubMed  Google Scholar 

  40. Surawicz CM (2010) Mechanisms of diarrhea. Curr Gastroenterol Rep 12(4):236–241

    Article  PubMed  Google Scholar 

  41. Thomas JA, Buchsbaum RN, Zimniak A, Racker E (1979) Intracellular pH measurements in Ehrlich ascites tumor cells utilizing spectroscopic probes generated in situ. Biochemistry 18(11):2210–2218

    Article  CAS  PubMed  Google Scholar 

  42. Venglovecz V, Rakonczay Z Jr, Ozsvari B, Takacs T, Lonovics J, Varro A, Gray MA, Argent BE, Hegyi P (2008) Effects of bile acids on pancreatic ductal bicarbonate secretion in guinea pig. Gut 57(8):1102–1112

    Article  CAS  PubMed  Google Scholar 

  43. Voronina SG, Barrow SL, Gerasimenko OV, Petersen OH, Tepikin AV (2004) Effects of secretagogues and bile acids on mitochondrial membrane potential of pancreatic acinar cells: comparison of different modes of evaluating DeltaPsim. J Biol Chem 279(26):27327–27338

    Article  CAS  PubMed  Google Scholar 

  44. Walker NM, Simpson JE, Yen PF, Gill RK, Rigsby EV, Brazill JM, Dudeja PK, Schweinfest CW, Clarke LL (2008) Down-regulated in adenoma Cl-/HCO3 - exchanger couples with Na/H exchanger 3 for NaCl absorption in murine small intestine. Gastroenterology 135(5):1645–653 e1643

  45. Walters JR, Pattni SS (2010) Managing bile acid diarrhoea. Ther Adv Gastroenterol 3(6):349–357

    Article  Google Scholar 

  46. Weintraub WH, Machen TE (1989) pH regulation in hepatoma cells: roles for Na-H exchange, Cl-HCO3 exchange, and Na-HCO3 cotransport. Am J Physiol 257(3 Pt 1):G317–G327

    CAS  PubMed  Google Scholar 

  47. Wilcox C, Turner J, Green J (2014) Systematic review: the management of chronic diarrhoea due to bile acid malabsorption. Aliment Pharmacol Ther 39(9):923–939. doi:10.1111/apt.12684

    Article  CAS  PubMed  Google Scholar 

  48. Xu H, Zhang B, Li J, Wang C, Chen H, Ghishan FK (2012) Impaired mucin synthesis and bicarbonate secretion in the colon of NHE8 knockout mice. Am J Physiol Gastrointest Liver Physiol 303(3):G335–G343

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  49. Yeruva S, Farkas K, Hubricht J, Rode K, Riederer B, Bachmann O, Cinar A, Rakonczay Z, Molnar T, Nagy F, Wedemeyer J, Manns M, Raddatz D, Musch MW, Chang EB, Hegyi P, Seidler U (2010) Preserved Na(+)/H(+) exchanger isoform 3 expression and localization, but decreased NHE3 function indicate regulatory sodium transport defect in ulcerative colitis. Inflamm Bowel Dis 16(7):1149–1161

    Article  PubMed  Google Scholar 

Download references

Grant support

Our research is supported by Hungarian National Development Agency grants (TÁMOP-4.2.2.A-11/1/KONV-2012-0035, TÁMOP-4.2.2-A-11/1/KONV-2012-0052, TÁMOP-4.2.2.A-11/1/KONV-2012-0073), the Hungarian Scientific Research Fund (OTKA NF105758, NF100677, K109756, PD105948) and the Hungarian Academy of Sciences (BO/00531/11/5, BO/00632/14/5). This research was also supported by the European Union and the State of Hungary, co-financed by the European Social Fund in the framework of TÁMOP 4.2.4.A/2-11-1-2012-0001 and TÁMOP-4.2.4.A2- 710-SZJÖ-TOK-13-0017 ‘National Excellence Program’ and MTA-SZTE Momentum Grant (LP2014-10/2014).

Author information

Authors and Affiliations

  1. Faculty of Medicine, First Department of Medicine, University of Szeged, Korányi fasor 8-10, H-6720, Szeged, Hungary

    É. Pallagi-Kunstár, K. Farkas, J. Maléth, Z. Rakonczay Jr, F. Nagy, T. Molnár, Z. Szepes, J. Lonovics, T. Wittmann & P. Hegyi

  2. Department of Pharmacology and Pharmacotherapy, University of Szeged, Szeged, Hungary

    V. Venglovecz

  3. Department of Pathology, University of Szeged, Szeged, Hungary

    Z. Rázga

  4. MTA-SZTE Translational Gastroenterology Research Group, Szeged, Hungary

    P. Hegyi

Authors
  1. É. Pallagi-Kunstár
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. K. Farkas
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. J. Maléth
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Z. Rakonczay Jr
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. F. Nagy
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. T. Molnár
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Z. Szepes
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. V. Venglovecz
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. J. Lonovics
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Z. Rázga
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. T. Wittmann
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. P. Hegyi
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to P. Hegyi.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Pallagi-Kunstár, É., Farkas, K., Maléth, J. et al. Bile acids inhibit Na+/H+ exchanger and Cl−/HCO3 − exchanger activities via cellular energy breakdown and Ca2+ overload in human colonic crypts. Pflugers Arch - Eur J Physiol 467, 1277–1290 (2015). https://doi.org/10.1007/s00424-014-1560-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00424-014-1560-9

Keywords

Search

Navigation