Genetic engineering Streptococcus thermophilus to produce antioxidant enzymes enhances their anti-inflammatory activity

S. DEL CARMEN; A. DE MORENO DE LEBLANC; R. MARTIN; F. CHAIN; P. KHARRAT; P. LANGELLA; L. BERMUDEZ-HUMARAN; J.G. LEBLANC

San Miguel de Tucuman

Simposio; IV International Symposium on Lactic Acid Bacteria: Food, Health and Applications; 2013

Centro de Referencia para Lactobacilos (CERELA)

Previous studies have shown that yoghurt prepared with a mixture of Lactobacillus bulgaricus and Streptococcus thermophilus strains from CERELA Culture Collection, prevented intestinal inflammation through an immunomodulating mechanism. It has also been recently reported that catalase (Cat) and superoxide dismutase (SOD) producing lactic acid bacteria reduced inflammation by an antioxidant mechanism.The aim of this study was to develop a lactic acid bacteria strain with immunomodulating as well as antioxidant properties and evaluate its anti-inflammatory effect in vivo.Lactobacillus bulgaricus (CRL861, 863, 864, 866, 869, 871, 872 and 887) and Streptococcus thermophilus (CRL806 and 807) strains (used previously for yoghurt preparation) were co-cultured with human HT29 or RAW cells lines and with a primary culture of mononuclear cells isolated from human peripheral blood or from mice Peyer?s Patches and the production of pro- and anti-inflammatroy cytokines were measured in the cell supernatants by ELISA. The strain selected was transformed with plasmids pIL253-mnkat or pIL253-sodA encoding for Cat or SOD enzymes respectively. The anti-inflammatory effect of these strains was evaluated in a trinitrobenzenesulfonic acid (TNBS) induced colitis mouse model. The screening assays for the strains showed that cells stimulated with Streptococcus thermophilus CRL807 were able to maintain an elevated anti-/pro-inflammatory cytokine ratio in vitro and in vivo. Streptococcus thermophilus CRL807 was transformed and the enzyme activities were demonstrated in the genetically modified (GM) strains. Finally the anti-inflammatory efficiency of the GM strains was evaluated in vivo. Mice that received Cat or SOD producing streptococci showed decreased weight lost, lower microbial translocation to liver, lower macroscopic and microscopic damage scores and increased enzymatic activities in their large intestines compared to those that received the wild-type strain or that did not receive any treatment. The analysis of cytokine producing cells showed that GM-strain maintained the immunomodulatory capacity of the wild-type strain with increased IL-10+/IL-17+ cell ratios. The greatest anti-inflammatory activity was observed with the mixture of Cat and SOD producing streptococci strains. Our findings show that the treatment with recombinant streptococci producing antioxidant enzymes attenuates inflammation in a murine model of colitis and that this genetic manipulation significantly enhanced the innate anti-inflammatory potential of the strain.