Abstract
The present study aimed to evaluate the potential of cell surface and extracellular proteins in regulation of intestinal epithelial barrier (IEB) function. Eight potentially probiotic L. reuteri strains were evaluated for presence of mapA gene and its expression on co-culturing with the Caco-2 cells. The ability of untreated (Viable), heat-inactivated, 5 M LiCL treated L. reuteri strains as well as their cell-free supernatant (CFS) to modulate expression of IEB function genes (hBD-2, hBD-3, claudin-1 and occludin) was also evaluated. Caco-2 cells were treated with cell surface and extracellular protein extracts and investigated for change in expression of targeted IEB function genes. The results showed that mapA gene is present in all the tested L. reuteri strains and expression of mapA and its receptors (anxA13 and palm) increase significantly on co-culturing of L. reuteri and Caco-2 cells. Also, up-regulated expression of IEB function genes was observed on co-culturing of L. reuteri (viable, heat-inactivated and CFS) and their protein extracts with Caco-2 cells in contrast to down-regulation observed with the pathogenic strain of Salmonella typhi. Therefore, this study concludes that the cell surface and extracellular protein from L. reuteri act as an effective mediator molecules to regulate IEB function.
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References
Aleljung P, Shen W, Rozalska B, Hellman U, Ljungh A, Wadstrom T (1994) Purification of collagen-binding proteins of Lactobacillus reuteri NCIB 11951. Curr Microbiol 28:231–236. https://doi.org/10.1007/BF01575966
Antoni L, Nuding S, Wehkamp J, Stange EF (2014) Intestinal barrier in inflammatory bowel disease. World J Gastroenterol 20(5):1165–1179. https://doi.org/10.3748/wjg.v20.i5.1165
Berkes J, Viswanathan VK, Savkovic SD, Hecht G (2003) Intestinal epithelial responses to enteric pathogens: effects on the tight junction barrier, ion transport, and inflammation. Gut 52(3):439–51. https://doi.org/10.1136/gut.52.3.439
Casas IA, Dobrogosz WJ (2000) Validation of the probiotic concept: Lactobacillus reuteri confers broad spectrum protection against disease in humans and animals. Microb Ecol Health Dis 12:247–285. https://doi.org/10.1080/08910600050216246-1
Chen CN, Sung HW, Liang HF, Chang WH (2002) Feasibility study using a natural compound (reuterin) produced by Lactobacillus reuteri in sterilizing and cross-linking biological tissues. J Biomed Mater Res 61(3):360–369. https://doi.org/10.1002/jbm.10153
Coconnier MH, Lievin V, Lorrot M, Servin AL (2000) Antagonistic activity of Lactobacillus acidophilus LB against intracellular Salmonella enteric serovar Typhimurium infecting human enterocyte-like Caco-2/TC-7 cells. Appl Environ Microbiol 66:1152–1157. https://doi.org/10.1128/AEM.66.3.1152-1157.2000
Collado MC, Jalonen L, Meriluoto J, Salminen S (2006) Protection mechanism of probiotic combination against human pathogens: in vitro adhesion to human intestinal mucus. Asia Pac J Clin Nutr 15:570–575
Colum D, Liam OM, Lisa M, Gerardine T, Darrin M, Sile OH, Maria F, Sarah F (2001) In vitro selection criteria for probiotic bacteria of human origin: correlation with in vivo findings. Am J Clin Nutr 73:386S-392S. https://doi.org/10.1093/ajcn/73.2.386s
Cui W, Li LX, Sun CM, Wen Y, Zhou Y, Dong YL, Liu P (2010) Tumor necrosis factor alpha increases epithelial barrier permeability by disrupting tight junctions in Caco-2 cells. Braz J Med Biol Res 43(4):330–337. https://doi.org/10.1590/S0100-879X2010007500020
Duary RK, Rajput YS, Batish VK, Grover S (2011) Assessing the adhesion of putative indigenous probiotic lactobacilli to human colonic epithelial cells. Ind J Med Res 134:664–671. https://doi.org/10.4103/0971-5916.90992
Fayol-Messaoudi D, Berger CN, Coconnier-Polter MH, Lievin-Le Moal V, Servin AL (2005) pH-, Lactic acid-, and non-lactic acid-dependent activities of probiotic Lactobacilli against Salmonella enterica Serovar Typhimurium. Appl Environ Microbiol 71(10):6008–6013. https://doi.org/10.1128/AEM.71.10.6008-6013.2005
Garg S, Singh TP, Reddi S, Malik RK, Kapila S (2017) Intervention of probiotic Lb. reuteri fermented milk as an adjuvant to combat protein energy malnourishment induced gut disturbances in albino mice. J Funct Foods 36:467–479. https://doi.org/10.1016/j.jff.2017.07.017
Garg S, Singh TP, Malik RK (2019) In Vivo Implications of Potential Probiotic Lactobacillus reuteri LR6 on the Gut and Immunological Parameters as an Adjuvant against Protein Energy Malnutrition. Probiotics & Antimicro Prot. https://doi.org/10.1007/s12602-019-09563-4
Henriksson A, Conway PL (1996) Adhesion of Lactobacillus fermentum 104-S to porcine stomach mucus. Curr Microbiol 33:31–34. https://doi.org/10.1007/s002849900069
Henriksson A, Szewzyk R, Conway PL (1991) Characteristics of the adhesive determinants of Lactobacillus fermentum 104. Appl Environ Microbiol 57:499–502
Iacob S, Iacob DG (2019) Infectious Threats, the Intestinal Barrier, and Its Trojan Horse: Dysbiosis. Front Microbiol 10:1676. https://doi.org/10.3389/fmicb.2019.01676
Juntunen M, Kirjavainen PV, Ouwehand AC, Salminen SJ, Isolauri E (2001) Adherence of probiotic bacteria to human intestinal mucus in healthy infants and during rotavirus infection. Clin Diagn Lab Immunol 8(2):293–296. https://doi.org/10.1128/CDLI.8.2.293-296.2001
Kos B, Suskovic J, Vukovic S, Simpraga M, Frece J, Matosic S (2003) Adhesion and aggregation ability of probiotic strain Lactobacillus acidophilus M92. J Appl Microbiol 94:981–987. https://doi.org/10.1046/j.1365-2672.2003.01915.x
Lee YK, Puong KY, Ouwehand AC, Salminen S (2003) Displacement of bacterial pathogens from mucus and Caco-2 cell surface by lactobacilli. J Med Microbiol 52:925–930. https://doi.org/10.1099/jmm.0.05009-0
Maassen CB, van Holten-Neelen C, Balk F, den Bak-Glashouwer MJ, Leer RJ, Laman JD, Boersma WJ, Claassen E (2000) Strain-dependent induction of cytokine profiles in the gut by orally administered Lactobacillus strains. Vaccine 18:2613–2623. https://doi.org/10.1016/S0264-410X(99)00378-3
Mathipa MG, Thantsha MS (2017) Probiotic engineering: towards development of robust probiotic strains with enhanced functional properties and for targeted control of enteric pathogens. Gut Pathog 9:28. https://doi.org/10.1186/s13099-017-0178-9
Matsuo Y, Miyoshi Y, Okada S, Satoh E (2012) Receptor-like Molecules on human intestinal epithelial cells interact with an adhesion factor from Lactobacillus reuteri. Biosci Microbiota Food Health 31(4):93–102. https://doi.org/10.12938/bmfh.31.93
McGuckin MA, Eri R, Simms LA, Florin TH, Radford-Smith G (2009) Intestinal barrier dysfunction in inflammatory bowel diseases. Inflamm Bowel Dis 15:100–113. https://doi.org/10.1002/ibd.20539
McIntosh RS, Cade JE, Al-Abed M, Shanmuganathan V, Gupta R, Bhan A, Tighe PJ, Dua HS (2005) The spectrum of antimicrobial peptide expression at the ocular surface. IOVS 46(4):1379–1385. https://doi.org/10.1167/iovs.04-0607
Miyoshi Y, Okada S, Uchimura T, Satoh E (2006) A mucus adhesion promoting protein, MapA, mediates the adhesion of Lactobacillus reuteri to Caco-2 human intestinal epithelial cells. Biosci Biotechnol Biochem 70:1622–1628. https://doi.org/10.1271/bbb.50688
Monteagudo-Mera A, Rastall RA, Gibson GR, Charalampopoulos D, Chatzifragkou A (2019) Adhesion mechanisms mediated by probiotics and prebiotics and their potential impact on human health. App Microbiol Biotechnol 103:6463–6472. https://doi.org/10.1007/s00253-019-09978-7
Mu Q, Kirby J, Reilly CM, Luo XM (2017) Leaky gut as a danger signal for autoimmune diseases. Front Immunol 8:598. https://doi.org/10.3389/fimmu.2017.00598
Munoz-Provencio D, Monedero V (2011) Shotgun phage display of Lactobacillus casei BL23 against collagen and fibronectin. J Microbiol Biotechnol 21:197–203. https://doi.org/10.4014/jmb.1009.09011
Ohland CL, MacNaughton WK (2010) Probiotic bacteria and intestinal epithelial barrier function. Am J Physiol Gastrointest Liver Physiol 298:G807–G819. https://doi.org/10.1152/ajpgi.00243.2009
Orlando A, Linsalata M, Notarnicola M, Tutino V, Russo F (2014) Lactobacillus GG restoration of the gliadin induced epithelial barrier disruption: the role of cellular polyamines. BMC Microbiol 14:19. https://doi.org/10.1186/1471-2180-14-19
Ouwehand AC, Salminen SJ (1998) The health effects of cultured milk products with viable and non-viable bacteria. Int Dairy J 8:749–758. https://doi.org/10.1016/S0958-6946(98)00114-9
Ouwehand AC, ToIkko S, Kulmala J, Salminen S, Salminen E (2000) Adhesion of inactivated probiotic strains to intestinal mucus. Lett Appl Microbiol 31:82–86. https://doi.org/10.1046/j.1472-765x.2000.00773.x
Ramiah K, van Reenen CA, Dicks LM (2008) Surface-bound proteins of Lactobacillus plantarum 423 that contribute to adhesion of Caco-2 cells and their role in competitive exclusion and displacement of Clostridium sporogenes and Enterococcus faecalis. Res Microbiol 159:470–475. https://doi.org/10.1016/j.resmic.2008.06.002
Resta-Lenert S, Barrett KE (2003) Live probiotics protect intestinal epithelial cells from the effects of infection with enteroinvasive Escherichia coli (EIEC). Gut 52:988–997. https://doi.org/10.1136/gut.52.7.988
Roos S, Aleljung P, Robert N, Lee B, Wadstrom T, Lindberg M, Jonsson H (1996) A collagen binding protein from Lactobacillus reuteri is part of an ABC transporter system? FEMS Microbiol Lett 144:33–38. https://doi.org/10.1016/0378-1097(96)00334-5
Sˇusˇkovic´ J, Kos B, Beganovic´ J, Lebosˇ Pavunc A, Habjanicˇ K, Matosˇic´ S (2010) Antimicrobial activity—the most important property of probiotic and starter lactic acid bacteria. Food Technol Biotechnol 48(3):296–307
Sa´nchez B, Urdaci MC, Margolles A (2010) Extracellular proteins secreted by probiotic bacteria as mediators of effects that promote mucosa– bacteria interactions. Microbiology 156:3232–3242. https://doi.org/10.1099/mic.0.044057-0
Satoh E, Leer RJ, Conway PL, Pouwels PH (1999) Mucus adhesion promoting protein of Lactobacillus reuteri 104R. In Sixth Symposium on Lactic Acid Bacteria; Genetics, Metabolism and Applications, Veldhoven, Book of Abstracts, J39
Schlee M, Wehkamp J, Altenhoefer A, Oelschlaeger TA, Stange EF, Fellermann K (2007) Induction of human β-defensin 2 by the probiotic Escherichia coli Nissle 1917 is mediated through flagellin. Infect Immun 75:2399–2407. https://doi.org/10.1128/IAI.01563-06
Servin AL, Coconnier MH (2003) Adhesion of probiotic strains to the intestinal mucosa and interaction with pathogens. Best Pract Res Clin Gastroenterol 17:741–754. https://doi.org/10.1016/S1521-6918(03)00052-0
Sillanpää J, Martínez B, Antikainen J, Toba T, Kalkkinen N, Tankka S, Lounatmaa K, Keränen J, Höök M, Westerlund-Wikström B, Pouwels PH, Korhonen TK (2000) Characterization of the collagen-binding S-layer protein CbsA of Lactobacillus crispatus. J Bacteriol 182:6440–6450. https://doi.org/10.1128/JB.182.22.6440-6450.2000
Singh TP, Kaur G, Malik RK, Schillinger U, Guigas C, Kapila S (2012) Characterization of intestinal Lactobacillus reuteri strains as potential probiotics. Probiot Antimicro Prot 4(1):47–58. https://doi.org/10.1007/s12602-012-9090-2
Singh TP, Malik RK, Kaur G, Renuka, (2014) Safety assessment and evaluation of probiotic potential of Lactobacillus reuteri strains under in vitro conditions. Int J Curr Microbiol App Sci 3(2):335–348
Singh TP, Malik RK, Katkamwar SG, Kaur G (2015) Hypocholesterolemic effects of Lactobacillus reuteri LR6 in rats fed on high-cholesterol diet. Int J Food Sci Nutr 66(1):71–75. https://doi.org/10.3109/09637486.2014.953450
Singh TP, Malik RK, Kaur G (2016) Cell surface proteins play an important role in probiotic activities of Lactobacillus reuteri. Nutrire 41:5. https://doi.org/10.1186/s41110-016-0007-9
Singh TP, Kaur G, Kapila S, Malik RK (2017) Antagonistic activity of Lactobacillus reuteri strains on the adhesion characteristics of selected pathogens. Front Microbiol 8:486. https://doi.org/10.3389/fmicb.2017.00486
Valeur N, Engel P, Carbajal N, Connolly E, Ladefoged K (2004) Colonisation and immunomodulation by Lactobacillus reuteri ATCC 55730 in the human gastrointestinal tract. Appl Environ Microbiol 70:1176–1181. https://doi.org/10.1128/AEM.70.2.1176-1181.2004
Vancamelbeke M, Vermeire S (2017) The intestinal barrier: a fundamental role in health and disease. Expert Rev Gastroenterol Hepatol 11(9):821–834. https://doi.org/10.1080/17474124.2017.1343143
Vesterlund S, Paltta J, Karp M, Ouwehand AC (2005) Measurement of bacterial adhesion- in vitro evaluation of different methods. J Microbiol Methods 60:225–233. https://doi.org/10.1016/j.mimet.2004.09.013
Walter J, Tannock G, Tilsala-Timisjarvi W, Rodtong AS, Loach DM, Munro K, Alatossava T (2000) Detection and identification of gastrointestinal Lactobacillus species by using denaturing gradient gel electrophoresis and species-specific PCR primers. Appl Environ Microbiol 66:297–303. https://doi.org/10.1128/AEM.66.1.297-303.2000
Wessel D, Flugge UI (1984) A method for the quantitative recovery of protein in dilute solution in the presence of detergents and lipids. Anal Biochem 138:141–143. https://doi.org/10.1016/0003-2697(84)90782-6
Zhang W, Wang H, Liu J, Zhao Y, Gao K, Zhang J (2013) Adhesive ability means inhibition activities for Lactobacillus against pathogens and S-layer protein plays an important role in adhesion. Anaerobe 22:97–103. https://doi.org/10.1016/j.anaerobe.2013.06.005
Acknowledgements
We are thankful to Dr. Suman Kapila, Principal Scientist, Department of Animal Biochemistry and Dr. Sachinandan De, Principal Scientist, Department of Animal Biotechnology, National Dairy Research Institute, Karnal, India for providing lab facilities to perform cell culture and RT-PCR work, respectively.
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Singh, T.P., Tehri, N., Kaur, G. et al. Cell surface and extracellular proteins of potentially probiotic Lactobacillus reuteri as an effective mediator to regulate intestinal epithelial barrier function. Arch Microbiol 203, 3219–3228 (2021). https://doi.org/10.1007/s00203-021-02318-2
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DOI: https://doi.org/10.1007/s00203-021-02318-2