ASSESSMENT OF ANTI-INFLAMMATORY ACTIVITY OF LACTIC ACID BACTERIA FROM 'NUNU' ON IOTA-CARRAGEENAN INDUCED PAW OEDEMA IN WISTAR RATS

Abstract

Inflammation is a normal biological response of the body’s immune system which can be triggered by factors such as pathogens, stress, toxic compounds and some pharmaceuticals. Unresolved inflammation is a leading cause of several devastating health conditions globally. Hence, this study was designed to evaluate and compare the anti-inflammatory activity of Lactic Acid Bacteria (LAB) isolated from a locally fermented dairy product (“Nunu”) in Nigeria, as an alternative to the conventional therapies which produce deleterious effects in some individuals. Lactic acid bacteria were enumerated and isolated on MRS agar using pour plate method, incubated anaerobically at 37oc for 48 hours. Preliminary identification of isolates was carried out using morphological and conventional biochemical characterization. Further identification of isolates to strain level was carried out using molecular technique. The isolated LAB was used to treat acute inflammation in albino rats induced with 1% Iotacarrageenan. Apparently healthy rats (n = 84) were distributed into six groups (A-F), fourteen rats per group. Rats in Groups A were neither administered carrageenan nor treated with LAB (general control), while rats Group B were administered carrageenan injection only without any treatment (negative control). Rats in Groups C-E were treated with the various strains of LAB and rats in Group F were administered diclofenac sodium following the administration of carrageenan (positive control). The dose of LAB used for the oral treatment was 5 × 107 CFU/mL, while the dose of diclofenac sodium used was 150mg/kg body weight of the rats. The rat paw sizes (mm) was checked at t = 0, 1, 4, 8h, 1day, 3days, 1week and 2weeks. Heamatological parameters such as Erythrocyte sedimentation rate (ESR), total red blood cell (RBC) and White blood cell (WBC) was performed on rat blood samples to assess inflammation. Differential leukocyte counts were also carried out using microscopy. Myeloperoxidase activity (MPO) was carried out to assess the influx of neutrophils to the paw of inflamed and treated rats. Assay for C-reactive protein (CR-P), Interleukin 10 (IL-10), and Inflammation is a normal biological response of the body’s immune system which can be triggered by factors such as pathogens, stress, toxic compounds and some pharmaceuticals. Unresolved inflammation is a leading cause of several devastating health conditions globally. Hence, this study was designed to evaluate and compare the anti-inflammatory activity of Lactic Acid Bacteria (LAB) isolated from a locally fermented dairy product (“Nunu”) in Nigeria, as an alternative to the conventional therapies which produce deleterious effects in some individuals. Lactic acid bacteria were enumerated and isolated on MRS agar using pour plate method, incubated anaerobically at 37oc for 48 hours. Preliminary identification of isolates was carried out using morphological and conventional biochemical characterization. Further identification of isolates to strain level was carried out using molecular technique. The isolated LAB was used to treat acute inflammation in albino rats induced with 1% Iotacarrageenan. Apparently healthy rats (n = 84) were distributed into six groups (A-F), fourteen rats per group. Rats in Groups A were neither administered carrageenan nor treated with LAB (general control), while rats Group B were administered carrageenan injection only without any treatment (negative control). Rats in Groups C-E were treated with the various strains of LAB and rats in Group F were administered diclofenac sodium following the administration of carrageenan (positive control). The dose of LAB used for the oral treatment was 5 × 107 CFU/mL, while the dose of diclofenac sodium used was 150mg/kg body weight of the rats. The rat paw sizes (mm) was checked at t = 0, 1, 4, 8h, 1day, 3days, 1week and 2weeks. Heamatological parameters such as Erythrocyte sedimentation rate (ESR), total red blood cell (RBC) and White blood cell (WBC) was performed on rat blood samples to assess inflammation. Differential leukocyte counts were also carried out using microscopy. Myeloperoxidase activity (MPO) was carried out to assess the influx of neutrophils to the paw of inflamed and treated rats. Assay for C-reactive protein (CR-P), Interleukin 10 (IL-10), andInflammation is a normal biological response of the body’s immune system which can be triggered by factors such as pathogens, stress, toxic compounds and some pharmaceuticals. Unresolved inflammation is a leading cause of several devastating health conditions globally. Hence, this study was designed to evaluate and compare the anti-inflammatory activity of Lactic Acid Bacteria (LAB) isolated from a locally fermented dairy product (“Nunu”) in Nigeria, as an alternative to the conventional therapies which produce deleterious effects in some individuals. Lactic acid bacteria were enumerated and isolated on MRS agar using pour plate method, incubated anaerobically at 37oc for 48 hours. Preliminary identification of isolates was carried out using morphological and conventional biochemical characterization. Further identification of isolates to strain level was carried out using molecular technique. The isolated LAB was used to treat acute inflammation in albino rats induced with 1% Iotacarrageenan. Apparently healthy rats (n = 84) were distributed into six groups (A-F), fourteen rats per group. Rats in Groups A were neither administered carrageenan nor treated with LAB (general control), while rats Group B were administered carrageenan injection only without any treatment (negative control). Rats in Groups C-E were treated with the various strains of LAB and rats in Group F were administered diclofenac sodium following the administration of carrageenan (positive control). The dose of LAB used for the oral treatment was 5 × 107 CFU/mL, while the dose of diclofenac sodium used was 150mg/kg body weight of the rats. The rat paw sizes (mm) was checked at t = 0, 1, 4, 8h, 1day, 3days, 1week and 2weeks. Heamatological parameters such as Erythrocyte sedimentation rate (ESR), total red blood cell (RBC) and White blood cell (WBC) was performed on rat blood samples to assess inflammation. Differential leukocyte counts were also carried out using microscopy. Myeloperoxidase activity (MPO) was carried out to assess the influx of neutrophils to the paw of inflamed and treated rats. Assay for C-reactive protein (CR-P), Interleukin 10 (IL-10), and Transforming growth factor-Beta (TGF-β) was performed on serum samples of the rats using ELISA. The LAB isolated from the ‘Nunu’ samples were molecularly identified as L. fermentum CIP 102980, L. fermentum NBRC 15885 and L. fermentum NCDO 1750. Lactobacillus fermentum CIP 102980 and L. fermentum NBRC 15885 both showed good ability to minimize the effect of erythropenia caused by inflammation by gradually restoring the circulating levels of RBC than in diclofenac sodium treated rats. Lactobacillus fermentum CIP 102980 also showed the best ability to regulate leukocyte (lymphocytes, neutrophil and monocytes) infiltration in the blood circulation of the acutely inflamed rats by 24 hours. On the other hand, L. fermentum NBRC 15885 exerted the highest effect in controlling neutrophil accumulation in the paw of rats. It also exerted the highest production of serum antiinflammatory cytokines, IL-10 and TGF-β after 24 and 8 hours at 167.80±10.01 pg/ml and 452.60±104.03pg/ml respectively, while decreasing the level of serum inflammatory biomarker CR-P to below standard (2000 ng/ml). All the isolated LABs exerted significant activity than diclofenac sodium, with L. fermentum NBRC 15885 showing the highest effect. Therefore, the isolated LABs can be used as a better alternative to conventional therapy to alleviate complications associated with unresolved inflammatory response.