Role of Bacillus subtilis VSG4-derived biosurfactant in mediating immune responses in Labeo rohita

Giri, S.S.,Sen, S.S.,Jun, J.W.,Sukumaran, V.,Park, S.C. Academic Press 2016 FISH AND SHELLFISH IMMUNOLOGY Vol.54 No.-

<P>This study aimed to isolate biosurfactant from CO2-sequestering Bacillus subtilis VSG4 and to evaluate its immunostimulatory effect in Labeo rohita fingerlings. Fish were injected intraperitoneally (i.p.) with 0.1 mL of phosphate-buffered saline (PBS) containing the water-soluble fraction of purified biosurfactant at 50 (S50), 100 (S100), 200 (S200), or 300 (S300) mu g mL(-1). Fish injected with PBS served as controls. Various immunological parameters, including immune-related gene expression, were measured at 14, 21, and 28 days post administration (dpa). At 28 dpa, the.fish were challenged with Aeromonas hydrophila and mortality was recorded up to 14 days. Among the immune parameters tested, lysozyme levels (36.32 +/- 1.79 U mL(-1)), alternative complement pathway activity (76.26 +/- 2.18 U mL(-1)), phagocytic activity (32.18 +/- 0.67%), and serum bactericidal activity (73.2 +/- 4.7%) were significantly higher (P < 0.05) in the 5200 group at 21 dpa than in the controls. Respiratory burst activity (0.386 +/- 0.008 OD630nm) was the highest in the 5200 group at 28 dpa. Of the immune-related genes examined, pro-inflammatory cytokines (TNF-alpha and IL-1 beta) were significantly down-regulated in the S200 and 5300 groups. Expression of anti-inflammatory cytokines (IL-10 and TGF-beta) as well as IKB-alpha was higher (P < 0.05) in the S100-S300 groups at 21 dpa. The expression of NF-kappa B p65, IKK-beta, MAPKp38, and Myd88 was down-regulated in the treated groups when compared to the controls. Fish in the 5200 group exhibited the highest post challenge relative survival rate (67.88%). Collectively, these results suggest that secondary metabolite (biosurfactant) isolated from B. subtilis VSG4 at 200 mu g mL(-1) can positively influence immune responses, enhance disease resistance, and stimulate immune-related gene expression in L rohita. (C) 2016 Elsevier Ltd. All rights reserved.</P>

Protective effects of leucine against lipopolysaccharide-induced inflammatory response in Labeo rohita fingerlings

Giri, S.S.,Sen, S.S.,Jun, J.W.,Sukumaran, V.,Park, S.C. Academic Press 2016 Fish & shellfish immunology Vol.52 No.-

<P>The present study investigated the protective effects of leucine against lipopolysaccharide (LPS)-induced inflammatory responses in Labeo rohita (rohu) in vivo and in vitro. Primary hepatocytes, isolated from the hepatopancreas, were exposed to different concentrations of LPS for 24 h to induce an inflammatory response, and the protective effects of leucine against LPS-induced inflammation were studied. Finally, we investigated the efficiency of dietary leucine supplementation in attenuating an immune challenge induced by LPS in vivo. Exposure of cells to 10-25 mu g mL(-1) of LPS for 24 h resulted in a significant production of nitric oxide and release of lactate dehydrogenase to the medium, whereas cell viability and protein content were reduced (p < 0.05). LPS exposure (10 mu g mL(-1)) increased mRNA levels of the pro inflammatory cytokines TNF-alpha, IL-1 beta and IL-8 in vitro (p < 0.05). However, pretreatment with leucine prevented the LPS-induced upregulation of TNF-alpha, IL-1 beta and IL-8 mRNAs by downregulating TLR4, MyD88, NF-kappa Bp65, and MAPKp38 mRNA expression. Interestingly, mRNA expression of the anti-inflammatory cytokine, IL-10, which was increased by LPS treatment, was further enhanced (p < 0.05) by leucine pretreatment. The enhanced expression of IL-10 might inhibit the production of other pro inflammatory cytokines. It was found that leucine pretreatment attenuated the excessive activation of LPS-induced TLR4-MyD88 signaling as manifested by lower level of TLR4, MyD88, MAPKp38, NF-kappa Bp65 and increased level of I kappa B-alpha protein in leucine pre-treatment group. In vivo experiments demonstrated that leucine pre-supplementation could protect fish against LPS-induced inflammation through an attenuation of TLR4-MyD88 signaling pathway. Taken together, we propose that leucine pre-supplementation decreases LPS-induced immune damage in rohu by enhancing the expression of IL-10 and by regulating the TLR4-MyD88 signaling pathways. (C) 2016 Elsevier Ltd. All rights reserved.</P>

Effects of intracellular products of Bacillus subtilis VSG1 and Lactobacillus plantarum VSG3 on cytokine responses in the head kidney macrophages of Labeo rohita

Giri, S.S.,Sen, S.S.,Chi, C.,Kim, H.J.,Yun, S.,Park, S.C.,Sukumaran, V. Academic Press 2015 FISH AND SHELLFISH IMMUNOLOGY Vol.47 No.2

The efficiency of intracellular products (ICPs) of the probiotics Bacillus subtilis VSG1 and Lactobacillus plantarum VSG3 in stimulating cytokine responses in the head kidney (HK) macrophages of Labeo rohita was investigated. The HK macrophages were incubated with ICPs and lipopolysaccharide (LPS), and the responses of cytokine genes, namely interleukin-10 (IL-10), IL-1β, IL-12p35, IL-12p40, IL-18, tumour necrosis factor-α (TNF-α), nuclear factor kappa B (NF-κB), cyclo-oxygenase-2 (COX-2), interferon-1 (IFN-1), and IFN-γ were assessed by quantitative real-time PCR (qRT-PCR) at 2, 6, 12, 24, and 48 h post-stimulation (hps). Among the proinflammatory cytokines, a strong increase in the gene expression of IL-1β and TNF-α was displayed mainly at 2-6 hps with ICPs, as compared to that of the positive control (LPS) or the negative control (PBS) (P < 0.05). However, COX-2 and NF-κB showed higher expression at 2 and 24 hps, and 6-24 hps with ICPs, respectively. Antiviral cytokines IFN-1 and IFN-γ displayed strong expressions (P < 0.05) at 6-12 hps, and 12-24 hps with ICPs, respectively. Upregulation of the anti-inflammatory cytokine, IL-10, was recorded at 6-24 hps with ICPs, as compared to that controls. Expressions of cell-mediated immune factor genes (IL-12p35, IL-12p40, and IL-18) were also significantly upregulated at different time points, except 48 hps, in HK macrophages stimulated with ICPs. Furthermore, enhanced cellular (phagocytic activity and nitroblue tetrazolium assay) and humoral (lysozyme) immune parameters in stimulated cells confirmed the induction of the inflammatory response. Therefore, the results of this in vitro study indicate that the ICPs of B. subtilis VSG1 or L. plantarum VSG3 have great potential for stimulating the cytokine responses in fish, and are thereby potential immunostimulants to fish. Further studies could be conducted to explore its suitability as an adjuvant vaccine in aquaculture.

Immunotoxicological effects of cadmium on Labeo rohita, with emphasis on the expression of HSP genes

Giri, S.S.,Sen, S.S.,Jun, J.W.,Sukumaran, V.,Park, S.C. Academic Press 2016 Fish & shellfish immunology Vol.54 No.-

<P>The present study evaluated the effects of exposure (28 days) to a sub-lethal concentration of cadmium (Cd) (0.65 mg CdCl2 L-1) on the immune responses and expression of immune-related and heat shock protein (HSP) genes in Labeo rohita, an important aquacultured fish species. Among the immune parameters studied, significantly lower lysozyme activity was observed in fish 28 days post-exposure (dpe) to Cd as compared to control fish. Alternative complement pathway activity was slightly higher in the Cd-exposed group at 2 dpe than in controls, and this activity declined gradually thereafter. The phagocytic activity and serum immunoglobulin M (IgM) levels were insignificantly lower in the Cd-exposed group at all assessed time points than in controls. Among serum enzymatic activities, peroxidase activity was always higher in the Cd-exposed group than in controls, but the increase was insignificant at all assessed time points. Additionally, serum glutamic pyruvic transaminase and alkaline phosphatase activities were significantly higher in the Cd-exposed group at 14 and 28 dpe. Immune and HSP gene expression patterns were observed in kidney and liver tissues, respectively, by RT-PCR, and HSP5 were further analysed by immunoblotting. Cd had an immunosuppressive effect, leading to down regulation of TNF-alpha, IL-1 beta, IL-10, and IFN-gamma. However, Cd exposure led to the up-regulation of HSP47, HSP60, HSP70, HSP78, and HSP90, indicating Cd-induced cellular stress. Taken together, the results of this study demonstrate the immunotoxic effect of Cd. Cd exposure makes Labeo rohita immunocompromised, and this could subsequently increase the disease susceptibility of Labeo rohita. (C) 2016 Elsevier Ltd. All rights reserved.</P>

Pinocembrin attenuates lipopolysaccharide-induced inflammatory responses in Labeo rohita macrophages via the suppression of the NF-κB signalling pathway

Giri, S.S.,Sen, S.S.,Sukumaran, V.,Park, S.C. Academic Press 2016 FISH AND SHELLFISH IMMUNOLOGY Vol.56 No.-

<P>Pinocembrin is a flavonoid that has been reported to exhibit various pharmacological and biological activities including antimicrobial, antioxidant, and anti-inflammatory. To explore the anti-inflammatory activity of pinocembrin in a fish cell line, we investigated its ability to regulate the inflammatory mediators elevated by lipopolysaccharide (LPS) in Labeo rohita head-kidney (HK) macrophages. HK macrophages of L. rohita were treated with LPS (1 (mu g mL(-1)) in the presence or absence of pinocembrin. We examined the inhibitory effect of pinocembrin on LPS-induced nitric oxide (NO) and prostaglandin E-2 (PGE(2)) production. The inhibitory effect of pinocembrin on nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) was investigated by RT-PCR and western blot. The effect of pinocembrin on pro-inflammatory cytokines (tumour necrosis factor alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta)) and anti-inflammatory cytokine IL-10 was investigated by ELISA and RT-PCR. The phosphorylation of three mitogen activated protein kinases (MAPKs) ERK, JNK, and p38 was analysed by western blot. Pinocembrin inhibited LPS-induced productions of NO and PGE2, and also markedly inhibited TNF-alpha, IL-1 beta, iNOS, and COX-2 production in a concentration-dependent manner. In addition, TNF-alpha and IL-1 beta mRNA expression levels decreased significantly, while IL-10 mRNA expression increased (P < 0.05) with pinocembrin pre-treatment. RT-PCR and western blot analysis showed that pinocembrin decreased both the mRNA and protein expression levels of LPS-induced iNOS and COX-2 in HK macrophages. Pinocembrin suppressed the phosphorylation of MAPK in LPS-stimulated HK macrophages. Further, pinocembrin significantly inhibited LPS-induced NF-kappa B transcriptional activity via the attenuation of I kappa B alpha degradation. Taken together, pinocembrin reduced the levels of pro-inflammatory mediators, such as iNOS, COX-2, TNF-alpha, and IL-1 beta, by inhibiting NF-kappa B activation via the suppression of ERK and p38 phosphorylation, and by attenuating the degradation of I kappa B alpha. These results suggest that pinocembrin is a potential novel candidate for the treatment of inflammatory conditions in L. rohita macrophages. (C) 2016 Elsevier Ltd. All rights reserved.</P>

Dynamic response of heat and mass transfer in blood flow through stenosed bifurcated arteries

S. Chakravarty,S. Sen 한국유변학회 2005 Korea-Australia rheology journal Vol.17 No.2

The present study deals with a mathematical model describing the dynamic response of heat and mass transfer in blood flow through bifurcated arteries under stenotic condition. The geometry of the bifurcated arterial segment possessing constrictions in both the parent and the daughter arterial lumen frequently appearing in the diseased arteries causing malfunction of the cardiovascular system, is formulated mathematically with the introduction of the suitable curvatures at the lateral junction and the flow divider. The blood flowing through the artery is treated to be Newtonian. The nonlinear unsteady flow phenomena is governed by the Navier-Stokes equations while those of heat and mass transfer are controlled by the heat conduction and the convection-diffusion equations respectively. All these equations together with the appropriate boundary conditions describing the present biomechanical problem following the radial coordinate transformation are solved numerically by adopting finite difference technique. The respective profiles of the flow field, the temperature and the concentration and their distributions as well are obtained. The influences of the stenosis, the arterial wall motion and the unsteady behaviour of the system in terms of the heat and mass transfer on the blood stream in the entire arterial segment are highlighted through several plots presented at the end of the paper in order to illustrate the applicability of the present model under study.

Effect of Supplementation of Bacillus subtilis LS 1-2 Grown on Citrus-juice Waste and Corn-soybean Meal Substrate on Growth Performance, Nutrient Retention, Caecal Microbiology and Small Intestinal Morphology of Broilers

Sen, Sinol,Ingale, S.L.,Kim, J.S.,Kim, K.H.,Kim, Y.W.,Khong, Chou,Lohakare, J.D.,Kim, E.K.,Kim, H.S.,Kwon, I.K.,Chae, B.J. Asian Australasian Association of Animal Productio 2011 Animal Bioscience Vol.24 No.8

A feeding trial was conducted to investigate the effect of dietary supplementation of Bacillus subtilis LS 1-2 grown on citrus-juice waste and corn-soybean substrate on growth performance, nutrient retention, caecal microbial population and intestinal morphology in broilers. Three hundred twenty d-old Ross chicks were randomly allotted to 4 treatments on the basis of BW in a randomized complete block design. Each treatment had 4 replicates of 20 chicks in each. Experimental diets were fed in 2 phases, starter (d 0 to 21) and finisher (d 21 to 35). Dietary treatments were; negative control (NC: basal diet without any antimicrobial), positive control (PC: basal diet added with 20 mg/kg Avilamycin), basal diet added with 0.30% Bacillus subtilis LS 1-2 grown on corn-soybean substrate (P1), and basal diet added with 0.30% Bacillus subtilis LS 1-2 grown on citrus-juice waste substrate (P2). Overall BW gain, feed intake and FCR were better (p<0.05) in PC, P1 and P2 treatments as compared to NC. Moreover, overall BW gain and FCR in PC and P2 treatments were greater than P1. Retention of CP, GE (d 21, d 35) and DM (d 35) were increased (p<0.05) in treatments PC, P1 and P2 compared with NC. At d 35, caecal Clostridium and Coliform counts were lower (p<0.05) in treatments PC, P1 and P2 than NC. Moreover, Clostridium and Coliform counts in treatment PC was lower (p<0.05) than P1. Villus height and villus height to crypt depth ratio in both duodenum and ileum were increased (p<0.05) in treatments PC, P1, P2 as compared to NC. However, retention of nutrients, caecal microbial population and intestinal morphology remained comparable among treatments P1 and P2. It is concluded that Bacillus subtilis LS 1-2 inclusion at 0.30% level had beneficial effects on broilers' growth performance, nutrient retention, caecal microflora and intestinal morphology. Additionally, citrus-juice waste can be used as substrate for growth of probiotic Bacillus subtilis LS 1-2.

Ill- versus well-posed singular linear systems: scope of randomized algorithms

S.K. Sen,Ravi P. Agarwal,Gholam Ali Shaykhian 한국전산응용수학회 2009 Journal of applied mathematics & informatics Vol.27 No.3

The linear system Ax = b will have (i) no solution, (ii) only one non-trivial (trivial) solution, or (iii) infinity of solutions. Our focus will be on cases (ii) and (iii). The mathematical models of many real- world problems give rise to (a) ill-conditioned linear systems, (b) singular linear systems (A is singular with all its linearly independent rows are suffi- ciently linearly independent), or (c) ill-conditioned singular linear systems (A is singular with some or all of its strictly linearly independent rows are near-linearly dependent). This article highlights the scope and need of a randomized algorithm for ill-conditioned/singular systems when a reason- ably narrow domain of a solution vector is specified. Further, it stresses that with the increasing computing power, the importance of randomized algorithms is also increasing. It also points out that, for many optimization linear/nonlinear problems, randomized algorithms are increasingly domi- nating the deterministic approaches and, for some problems such as the traveling salesman problem, randomized algorithms are the only alterna- tives. The linear system Ax = b will have (i) no solution, (ii) only one non-trivial (trivial) solution, or (iii) infinity of solutions. Our focus will be on cases (ii) and (iii). The mathematical models of many real- world problems give rise to (a) ill-conditioned linear systems, (b) singular linear systems (A is singular with all its linearly independent rows are suffi- ciently linearly independent), or (c) ill-conditioned singular linear systems (A is singular with some or all of its strictly linearly independent rows are near-linearly dependent). This article highlights the scope and need of a randomized algorithm for ill-conditioned/singular systems when a reason- ably narrow domain of a solution vector is specified. Further, it stresses that with the increasing computing power, the importance of randomized algorithms is also increasing. It also points out that, for many optimization linear/nonlinear problems, randomized algorithms are increasingly domi- nating the deterministic approaches and, for some problems such as the traveling salesman problem, randomized algorithms are the only alterna- tives.

ILL-VERSUS WELL-POSED SINGULAR LINEAR SYSTEMS: SCOPE OF RANDOMIZED ALGORITHMS

Sen, S.K.,Agarwal, Ravi P.,Shaykhian, Gholam Ali The Korean Society for Computational and Applied M 2009 Journal of applied mathematics & informatics Vol.27 No.3

The linear system Ax = b will have (i) no solution, (ii) only one non-trivial (trivial) solution, or (iii) infinity of solutions. Our focus will be on cases (ii) and (iii). The mathematical models of many real-world problems give rise to (a) ill-conditioned linear systems, (b) singular linear systems (A is singular with all its linearly independent rows are sufficiently linearly independent), or (c) ill-conditioned singular linear systems (A is singular with some or all of its strictly linearly independent rows are near-linearly dependent). This article highlights the scope and need of a randomized algorithm for ill-conditioned/singular systems when a reasonably narrow domain of a solution vector is specified. Further, it stresses that with the increasing computing power, the importance of randomized algorithms is also increasing. It also points out that, for many optimization linear/nonlinear problems, randomized algorithms are increasingly dominating the deterministic approaches and, for some problems such as the traveling salesman problem, randomized algorithms are the only alternatives.

Theoretical study on the constricted flow phenomena in arteries

S. Sen,S. Chakravarty 한국유변학회 2012 Korea-Australia rheology journal Vol.24 No.4

The present study is dealt with the constricted flow characteristics of blood in arteries by making use of an appropriate mathematical model. The constricted artery experiences the generated wall shear stress due to flow disturbances in the presence of constriction. The disturbed flow in the stenosed arterial segment causes malfunction of the cardiovascular system leading to serious health problems in the form of heart attack and stroke. The flowing blood contained in the stenosed artery is considered to be non-Newtonian while the flow is treated to be two-dimensional. The present pursuit also accounts for the motion of the arterial wall and its effect on local fluid mechanics. The flow analysis applies the time-dependent, two-dimensional incompressible nonlinear Navier-Stokes equations for non-Newtonian fluid representing blood. An extensive quantitative analysis presented at the end of the paper based on large scale numerical computations of the quantities of major physiological significance enables one to estimate the constricted flow characteristics in the arterial system under consideration which deviates significantly from that of normal physiological flow conditions.