title: Oral administration of recombinant Mycobacterium smegmatis expressing a tripeptide construct derived from endogenous and microbial antigens prevents atherosclerosis in ApoE(-/-) mice.
abstract: BACKGROUND
  Immunotherapy by inducing oral tolerance to atherogenic self-antigens is gaining importance as an alternative treatment modality for atherosclerosis. The use of live bacterial vectors to express the recombinant antigen in vivo will obviate the need for large-scale purification of recombinant protein and may also augment the efficacy of oral tolerance induction.
  
  
  OBJECTIVE
  The objective of the study was to explore the use of recombinant Mycobacterium smegmatis as a live vector for oral delivery of antigens to induce immune tolerance.
  
  
  RESULTS
  We developed a M. smegmatis vector to secrete a recombinant tripeptide construct (AHC; peptides from Apolipoprotein B, Heat-shock protein 60 and Chlamydia pneumoniae outer membrane protein) expressed in a dendroaspin protein scaffold in pJH154 background. Immune response and oral tolerance to the cloned peptides were studied in C57/BL6 mice. The efficacy of this live vaccine to control atherosclerosis was studied in ApoE(-/-) knockout mice in C57/BL6 background. Oral administration of M. smegmatis secreting the cloned AHC antigen was found to induce tolerance to cloned protein and reduce the development of atherosclerosis by 24.0% compared to control. Protection against atherosclerosis was associated with increase in expression of regulatory T cell-associated markers including CTLA4 (1.8-fold), Foxp3 (2.6-fold), TGF-β (2.8-fold), IL10 (2.9-fold), and reduction in lipids, macrophage infiltration, and expression of inflammatory mediators in aorta.
  
  
  CONCLUSIONS
  Our results suggest that M. smegmatis can be developed as an oral carrier of recombinant proteins to treat inflammatory autoimmune diseases.
authors.affiliation: Molecular Immunology, Thrombosis Research Institute, Bangalore, India.
authors.forename: Vrushali
authors.initials: V
authors.lastname: Deshpande
authors.affiliation: Molecular Immunology, Thrombosis Research Institute, Bangalore, India.
authors.forename: Rameshkumar
authors.initials: R
authors.lastname: Krishnan
authors.affiliation: Molecular Immunology, Thrombosis Research Institute, Bangalore, India.
authors.forename: Sheena
authors.initials: S
authors.lastname: Philip
authors.affiliation: Department of Medical Microbiology and Immunobiology, University of Szeged, Szeged, Hungary.
authors.forename: Ildiko
authors.initials: I
authors.lastname: Faludi
authors.affiliation: Manipal University at Thrombosis Research Institute, Bangalore, India.
authors.forename: Thiruvelselvan
authors.initials: T
authors.lastname: Ponnusamy
authors.affiliation: Manipal University at Thrombosis Research Institute, Bangalore, India.
authors.forename: Lakshmi Narasimha Rao
authors.initials: LN
authors.lastname: Thota
authors.affiliation: Department of Medical Microbiology and Immunobiology, University of Szeged, Szeged, Hungary.
authors.forename: Valeria
authors.initials: V
authors.lastname: Endresz
authors.affiliation: Molecular Immunology, Thrombosis Research Institute, London, UK.
authors.forename: Xinjie
authors.initials: X
authors.lastname: Lu
authors.affiliation: Molecular Immunology, Thrombosis Research Institute, Bangalore, India.|Molecular Immunology, Thrombosis Research Institute, London, UK.
authors.forename: Vijay V
authors.initials: VV
authors.lastname: Kakkar
authors.affiliation: Molecular Immunology, Thrombosis Research Institute, Bangalore, India. lakshmi.mundkur@triindia.org.in.
authors.forename: Lakshmi A
authors.identifier: http://orcid.org/0000-0003-2928-1726
authors.initials: LA
authors.lastname: Mundkur
mesh_terms: D000284:Administration, Oral; D000818:Animals; D000941:Antigens / Q000008:administration & dosage* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D001018:Aortic Diseases / Q000235:genetics / Q000276:immunology / Q000378:metabolism / Q000517:prevention & control*; D001057:Apolipoproteins E / Q000172:deficiency* / Q000235:genetics; D050197:Atherosclerosis / Q000235:genetics / Q000276:immunology / Q000378:metabolism / Q000517:prevention & control*; D060908:CTLA-4 Antigen / Q000276:immunology / Q000378:metabolism; D004195:Disease Models, Animal; D005260:Female; D051858:Forkhead Transcription Factors / Q000276:immunology / Q000378:metabolism; D020022:Genetic Predisposition to Disease; D005822:Genetic Vectors; D007108:Immune Tolerance; D007114:Immunization; D007167:Immunotherapy / Q000379:methods*; D018836:Inflammation Mediators / Q000276:immunology / Q000378:metabolism; D016753:Interleukin-10 / Q000276:immunology / Q000378:metabolism; D050356:Lipid Metabolism; D008264:Macrophages / Q000276:immunology / Q000378:metabolism; D008297:Male; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D020102:Mycobacterium smegmatis / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D009842:Oligopeptides / Q000008:administration & dosage* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D010641:Phenotype; D050378:T-Lymphocytes, Regulatory / Q000276:immunology / Q000378:metabolism; D013997:Time Factors; D016212:Transforming Growth Factor beta / Q000276:immunology / Q000378:metabolism; D014614:Vaccines, Synthetic / Q000008:administration & dosage
chemical_list: D000941:Antigens; D001057:Apolipoproteins E; D060908:CTLA-4 Antigen; C556707:Ctla4 protein, mouse; D051858:Forkhead Transcription Factors; C498833:Foxp3 protein, mouse; C556062:IL10 protein, mouse; D018836:Inflammation Mediators; D009842:Oligopeptides; D016212:Transforming Growth Factor beta; D014614:Vaccines, Synthetic; D016753:Interleukin-10
keywords: Apolipoprotein; Atherosclerosis; Autoimmune diseases; Heat-shock proteins; Immune tolerance; Live vaccine; Mycobacterium smegmatis; Regulatory T cells