Immunomodulatory Effect of a Polyherbal Formulation (Imusil) on Cyclophosphamide Induced Experimental Animal Model

Abstract

Objective: In the present study, we investigated the immunomodulatory effect of a polyherbal formulation referred to as Imusil (IM) on cyclophosphamide (CP) induced immunosuppression model.

Methods: CP induced experimental animal model was used for evaluating the immunomodulatory effect of IM. For the study, animals were divided into four groups. Group I is served as the normal control, group II is treated only with CP, group III is treated with the standard drug, levamisole and group IV is treated with IM. The experimental duration was 30 days. At the end of the study, we had evaluated various parameters such as immune organ index, liver marker enzymes, antioxidants, haematological analysis, Th1/Th2 cytokine balance and humoral immune responses were examined using ELISA kits, T-lymphocyte subsets by flow cytometry, and histopathological analysis of the liver, spleen and thymus by H&E staining.

Results: The results obtained from the study revealed that the treatment of immunosuppressed animals with IM significantly (p<0.05) reversed the immune response in a positive manner. Treatment with IM properly shields the immune organs and triggers the cell-mediated and humoral immune responses accordingly. Thus, no significant changes were observed in the haematological parameters. Moreover, IM supplementation helps to boost up the antioxidant activity, thereby preventing oxidative stress-mediated damage, and also protects the liver from the toxicity induced by CP.

Conclusion: The results suggest that IM has the ability to counteract the immunosuppressive effect of chemotherapeutic drugs by stimulating the immune system, along with its potent antioxidant and hepatoprotective properties.

Keywords: Cytokines; Herbal medicine; Immune responses; Immunomodulators; antioxidants.

Figure 1

The Effect of IM on the Immune Organ Index. Values expressed as average of 6 samples ± SEM in each group. ‘a’ -Statistical difference with normal group at P ≤ 0.05. ‘b’ -Statistical difference with CP treated rats at P ≤ 0.05

Figure 2

Effect of IM on Humoral Immune Response. Values expressed as average of 6 samples ± SEM in each group. ‘a’ -Statistical difference with normal group at P ≤ 0.05. ‘b’ -Statistical difference with CP treated rats at P ≤ 0.05

Figure 3

Effect of IM on Th1/Th2 Cytokines. Values expressed as average of 6 samples ± SEM .in each group. ‘a’-Statistical difference with normal group at P ≤ 0.05. ‘b’-Statistical difference with CP treated rats at P ≤ 0.05

Figure 4

Effect of IM on Liver Enzyme Markers. The values are expressed as mean± SEM of six rats in each group. a - Statistical difference with control group at P< 0.05. b – Statistical difference with CP treated rats at P< 0.05. U: SGOT- µmol of oxaloacetate liberated /min/mg protein. U: SGPT- µmol of pyruvate formed /min/mg protein. U: ALP- amount of enzyme to decompose 1 µmole of P-NPP/minute at 25° C.

Figure 5

Effect of IM on the Antioxidant Activities of SOD and CAT and the Level of GSH. The Values expressed as average of 6 samples ± SEM in each group. ‘a’-Statistical difference with normal group at P ≤ 0.05. ‘b’-Statistical difference with CP treated rats at P ≤ 0.05

Figure 6

Effect of IM on the Population Change in T Lymphocyte Subset in CP Induced Immunosuppressed Rats. T lymphocytes were detected using PE-, APC- and FITC- conjugated monoclonal antibodies specific for the rat’s T cell markers CD3 (A), CD4 (B), and CD8 (C) respectively. Stained cells were analysed by flow cytometry using a BD FACSDiva software

Figure 7

Histopathological Analysis of Rat Spleen, Thymus and Liver. Tissues were stained with H&E and examined by optical microscopy. WP- White pulp; RP- Red pulp; CA- Central artery; DS- Dilated sinusoid; A-Atrophy; M- Medulla; C- Cortex; H- Hepatocytes; K-Kupffer cells; PT- Portal triad; S- Sinusoidal space; V-Vacuole; DH- Degenerated hepatocytes; CS-Congested sinusoidal space