Immunomodulatory and therapeutic activity of curcumin

International Immunopharmacology 11 (2011) 331–341 Contents lists available at ScienceDirect International Immunopharmacology j o u r n a l h o m e p a g e : w w w. e l s ev i e r. c o m / l o c a t e / i n t i m p Review Immunomodulatory and therapeutic activity of curcumin Raghvendra M. Srivastava a, Sarvjeet Singh b, Shiv K. Dubey c, Krishna Misra d, Ashok Khar e,⁎ a Department of Otolaryngology, Hillman Cancer Centre, University of Pittsburgh Cancer Institute, Pittsburgh, PA 15213, USA Department of Internal Medicine, Division of Cardiology, University of Texas Southwestern Medical Center, Dallas, Texas 75390, USA c Department of Internal Medicine, Division of Hematology-Oncology, University of Michigan Medical School, Ann Arbor, Michigan 48109, USA d Division of Bioinformatics, Indian Institute of Information Technology, Allahabad, India e CMBRC, Apollo Hospitals Educational and Research Foundation, Apollo Health City, Jubilee Hills, Hyderabad 500033, India b a r t i c l e i n f o Article history: Received 1 July 2010 Accepted 22 August 2010 Available online 8 September 2010 Keywords: Curcumin Anti-inflammatory Anti-cancerous Immune and metabolic diseases a b s t r a c t Inflammation is a disease of vigorous uncontrolled activated immune responses. Overwhelming reports have suggested that the modulation of immune responses by curcumin plays a dominant role in the treatment of inflammation and metabolic diseases. Observations from both in-vitro and in-vivo studies have provided strong evidence towards the therapeutic potential of curcumin. These studies have also identified a plethora of biological targets and intricate mechanisms of action that characterize curcumin as a potent ‘drug’ for numerous ailments. During inflammation the functional influence of lymphocytes and the related cross-talk can be modulated by curcumin to achieve the desired immune status against diseases. This review describes the regulation of immune responses by curcumin and effectiveness of curcumin in treatment of diseases of diverse nature. © 2010 Elsevier B.V. All rights reserved. Contents 1. 2. 3. 4. 5. 6. 7. 8. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Immunomodulatory action of curcumin on T lymphocytes . . . . . . . . . . . . . . . . . . . . . . . Immunoinhibitory action of curcumin on dendritic cells (DCs) . . . . . . . . . . . . . . . . . . . . . Immunomodulatory effect of curcumin on natural killer (NK) cells . . . . . . . . . . . . . . . . . . . Immunomodulatory effect of curcumin on monocytes and macrophages (Mϕ) . . . . . . . . . . . . . . Immunomodulatory effect of Curcumin on B cells . . . . . . . . . . . . . . . . . . . . . . . . . . . Immunomodulatory effect of curcumin on neutrophils and eosinophils and mast cells and its anti-oxidant Curcumin in health and disease . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8.1. Role of curcumin in the neoplastic diseases . . . . . . . . . . . . . . . . . . . . . . . . . . . 8.2. Curcumin in cardiovascular disease . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8.3. Curcumin in neurodegenerative disease. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8.4. Immunomodulatory action of curcumin in the prevention of inflammatory diseases . . . . . . . . 9. Concluding remarks and future perspectives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . properties . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 331 332 333 334 334 335 336 336 336 337 338 338 339 339 1. Introduction Abbreviations: Ag, antigen; Ab, antibody; NO, nitric oxide; LPS, lipopolysaccharide; ConA, concanavalin A; AP-1, activator protein 1; NF-κB, nuclear factor-kappaB; NF-AT, nuclear factor of activated T cells; PMA, phorbol 12-myristate 13-acetate; PHA, phytohaemagglutinin; ROS, reactive oxygen species; ROIs, reactive oxygen intermediates; COX-2, cyclooxygenase-2; APC, antigen presenting cells; DCs, dendritic cells; IDO, indoleamine 2,3-dioxygenase. ⁎ Corresponding author. E-mail address: ashok.khar@aherf.net (A. Khar). 1567-5769/$ – see front matter © 2010 Elsevier B.V. All rights reserved. doi:10.1016/j.intimp.2010.08.014 Turmeric is a mixture of compounds related to curcumin known as curcuminoids consisting of curcumin [i.e.diferuloylmethane or 1,7-bis (4-hydroxy-3-methoxy-phenyl) hepta-1, 6-diene-3, 5-dione)] as the major component, demethoxycurcumin, bisdemethoxycurcumin and cyclocurcumin [1] (Fig. 1). Curcumin has been in use for its medicinal benefits since centuries but the first documented case of its use as a drug emerged only in 1937 when it was utilized to treat biliary disease. Since then its therapeutic potential has been explored in 332 R.M. Srivastava et al. / International Immunopharmacology 11 (2011) 331–341 even minor fluctuations in the cellular redox milieu [10,11]. These transcription factors in turn control cell cycle, differentiation, stress response and other physiological processes [12–15]. The intricate mechanism of action of curcumin involves various biological targets viz transcription factors: NF-AT, AP-1, signal transducers and activator of transcription (STAT), p53 and kinases: mitogen-activated protein kinases, cytokines release, and the receptors found on different immune cell type. These actions of curcumin greatly affect the innate and adaptive arms of immunity, especially in the pathological conditions. Curcumin effectively modulates the function of T cells, B cells, dendritic cells (DCs), monocytes, macrophages (mφ) and neutrophils. Overwhelming reports have supported the anti-inflammatory action of curcumin and its potential role in the therapy of numerous immune cell related diseases. Although curcumin does not have a drug profile yet, the safety and non-toxic effect of oral curcumin (12 g/day) which is much higher than its regular in-take as food supplement have been established by the drug governing agency [16]. Recently, the pre-clinical and clinical studies that were conducted at different places have been reviewed [17]. However, there are certain limitations concerning the use of curcumin as a drug. Due to its insolublility in water, curcumin has very poor bioavailability, its cellular uptake is slow and it gets metabolized very fast once inside the cell. Therefore it requires repetitive oral doses in order to achieve significant concentration inside the cells for any physiological effects. To address these limitations a large number of curcumin analogues have been prepared that have shown improved uptake, metabolism and activity. In this review we discuss the effect and applications of curcumin across a spectrum of pathological conditions involving immune cells, metabolic targets and diseases. Fig. 1. Curcuminoids present in turmeric. 2. Immunomodulatory action of curcumin on T lymphocytes inflammatory diseases, neoplastic disease, cardiovascular and neurodegenerative disease, diabetes, cystic fibrosis and other disorders. Due to a vast number of biological targets and virtually no side effects, curcumin has achieved the potential therapeutic interest to cure immune related, metabolic diseases and cancer [2–7] (Table 1). Majority of the studies suggested that the biological effects of curcumin are mainly derived from its ability to either bind directly to various proteins such as cyclooxygenase-2 (COX-2), lipoxygenase, GSK3b and several other regulatory enzymes or by its ability to modulate intracellular redox state [1,8,9]. Modulation of cellular redox homeostasis exerts an indirect but more global effect on a number of cellular processes, since several critical transcription factors such as activator protein 1 (AP1), nuclear factor-kappaB (NFκB), nuclear factor of activated T cells (NF-AT), p53 etc. are sensitive to Sikora et al. demonstrated that the mitogen concanavalin A (ConA) stimulated and the spontaneous proliferation of rat thymocytes could be inhibited by curcumin (50 μM) and similar anti-proliferative effects of curcumin on ConA-stimulated Jurkat T cell line were also reported. In contrast, the similar dose of curcumin could protect rat thymocytes and Jurkat T cells from dexamethasone and ultra-violet irradiation induced apoptosis, respectively. These bimodal effects of curcumin were correlated with the suppressive effects of curcumin on AP-1 transcription factor activation; however no effect of curcumin was seen on AP-1 under normal conditions [18]. In contrast to the study of Sikora et al., an independent study showed that curcumin (50 μM) could induce cell death in the normal quiescent and proliferating human lymphocytes through caspase-3 activation but Table 1 The potential of curcumin was shown in the various diseases involving multiple mechanisms in the respective cell types. Diseases Cell types Mechanisms of action References Alzheimer disease Monocytic THP-1 cell line, peripheral blood monocytes Th-17 producing T cells, TLR4 and TLR 9 expressing T cells. Anti-inflammtory: by blocking the amyloid peptide induced expression of TNF-α, IL-1β, MCP-1, IL-8, MIP-1β and CCR5 Anti-inflammatory: blocking of EAE incidences by blocking IL-6; IL-21 signaling, and the differentiation of Th-17 producing T cells, modulation of the function of TLR-4 and TLR-9 on T cells, blocking of IL-12 signaling Anti-inflammatory: decreased the frequency of eosinophils and the inflammatory cells by the regulation iNOS, inhibition of the IgE and Ag-induced degranulation of mast cells. Anti-inflammatory: suppressed ROIs generation, Blocks crystal induced neutrophil activation, suppressed arthritis Ag-induced T cell proliferation Suppressed pp38, suppressed pro-inflammatory IL-1β and enhanced IL-10 level [59,61] Multiple sclerosis Allergy Arthritis Inflammatory bowel disease Eosinophils, bronchoalveolar inflammatory cells, mast cells Neutrophils, T cells Intestinal mucosal biopsies of patients. Psoriasis Keratinocytes Inflammatory cardiovascular Myocardial tissue, endothelial disorders cell line Wound healing Infilitrating Mϕs, keratinocytes, fibroblasts Inflammatory type II Infiltrating Mϕs, adipose tissue, diabetes hepatic tissue [31–33] [75–77] [34,70,71] [151,152] Blocks TNF-α mediated activation of cells [153] Blocks neutrophils activation. Attenuate the plasma level of IL-10, IL-8 and TNF-α. Blocks TNF-α [78,153] induced pro-inflammatory responses in cell line Anti-androgen receptor signaling activity, decreased local TNF-α level [58] Reduced Mϕs frequency in the adipose tissue, reduced expression of TNF-α, MCP-1 and reduced [7] NF-κB activity in the hepatic tissue R.M. Srivastava et al. / International Immunopharmacology 11 (2011) 331–341 without DNA degradation. This study also highlighted that curcumin affects the viability of proliferating T cells much severely than quiescent T cells [19,20]. Furthermore Deters et al. demonstrated that curcumin (2.8–10 μM) can significantly abrogate the proliferation of peripheral blood mononuclear cells (PBMC) induced by OKT3 mAb (a human TCR/CD3 complex Ab) [21]. The concentration range of curcumin used by Deters et al. was similar to the concentrations that was shown to significantly affect human T cell proliferation induced by various distinct stimuli viz. phorbol 12-myristate 13-acetate (PMA), CD28, phytohaemagglutinin (PHA) [22]. Although direct suppressive effects of curcumin on superantigen induced proliferation of T cells were demonstrated in several studies as described before, few studies also demonstrated that T cells fail to get appropriate amount of co-stimulatory signals from curcumin-treated Ag presenting cells (APCs), as curcumin (20–30 μM) aborted the upregulation of CD86 and CD83 in response to the APC maturation stimuli. This inhibitory effect of curcumin on T cells was independent of the HLADR levels on the respective APCs as the HLA-DR level was not downregulated by the curcumin. Curcumin, however, could also reduce the DCs' dependent allogenic CD4+ T cell proliferation in a mixed lymphocyte reaction assay at 1:16 ratio of DCs to T cells. In this study a probable affect of curcumin on the cytoskeleal elements of DCs was argued in this context, which may be attributed to its inhibitory anti-proliferative effects [23]. Another study demonstrated a significant increase in ConA-stimulated proliferation of splenic cells at 6.25 μM curcumin and a significant decrease in proliferation at 12.5 μM and a complete blockage of proliferation with 25 μM curcumin, which confirmed the distinct function of curcumin at variable concentrations. Also, curcumin irreversibly inhibited the induction of lymphoproliferation by other mitogens and alloantigens. As the in-vivo effects of curcumin are highly dependent on the bioavailable concentration of curcumin, it is indeed a daunting agenda to correlate the in-vitro activities of curcumin and in-vivo responses in the pathological conditions, especially in the localized pathological conditions like non-metastatic tumors of different origins [24]. As highlighted in the review so far, a variety of results indicated the invitro T cell immunosuppressive properties of curcumin in terms of T cell death, as well as blocking the proliferation capacity of T cells. Nevertheless curcumin has been in use since centuries and its consumption has not been associated with any immunocompromised disorders; seriously arguing the significance of its immunosuppressive properties on T cells that have been reported under in-vitro conditions. We had demonstrated that T cells that were harvested from the curcumin-injected (40 mg/kg/day; i.p) animals showed enhanced lymphoproliferation and a similar proliferative effect of curcumin was also observed when T cells were stimulated with ConA and PHA in conjunction with curcumin. Our study also provided evidence of specific lymphoproliferative effect of curcumin in-vivo, by using cyclosporine A, a potent immunosuppressant drug. Interestingly, the enhanced Antigen (Ag)-specific T cell proliferation was also observed in curcumin-injected rats that had received a highly immunogenic AK-5 histiocytoma cells as a source of tumor Ag [25]. The evidence for the enhanced frequency of CD4+T cells was furthermore reported in another spontaneously generated tumor model of adenoma in C57BL/6J-Min/+ (Min/+) mouse that were fed with 0.1% dietary curcumin. In a statistically controlled lymphocyte infiltration setup, this study reported an enhanced number of CD8+, CD4+ and CD3+ T cells in the curcumin fed animals. In this model the spontaneous polyp formation in the mucosa was significantly reduced by the curcumin administration and the anti-tumor mechanism was correlated with the enhanced cytokine level due to the increased number of activated CD4+ T cells, although no direct evidence for such a conclusion was described in this study. Also, this study showed enhanced level of B cells in the intestinal mucosa, but no role or increase in the number of monocytes was found in this spontaneous tumor model system [26]. In another elegant tumor model, in which 333 tumor growth could disintegrate the thymus morphology, curcumin (50 mg/kg body weight) restored the thymic integrity including CD3+ T cell frequency and served as immunoprotective compound during carcinogenesis. This effect of curcumin was attributed mechanistically to the anti-oxidant properties of curcumin because this tumor induced oxidative stress in thymic T cells [27]. As an extension of this work the same group demonstrated that curcumin could prevent the tumor induced apoptosis of thymocytes as well as restoration of the frequency of CD4+ T/CD8+ T cells in the same tumor model system with the same dose of curcumin. Mechanistically it was also shown that curcumin modulated Jak-3/Stat-5 activity to restore the immune cell frequency and activity [28,29]. Curcumin decreased IL12-induced STAT4 phosphorylation but enhanced the (interferon) IFN-β-induced STAT4 phosphorylation; curcumin decreased IL-12 induced IFN-γ production and IL-12 Rβ1 and β2 expression, whereas it enhanced IL-10 production and IFN receptor (IFNAR) subunits 1 and 2 expression. Curcumin also increased IFN-α-induced IL-10 and IFNAR1 expression. Pretreatment with curcumin decreased IFN-αinduced IFNAR2 expression and failed to modify the level of IFN-αinduced phospho-STAT4 activation. These findings favour the distinct mode of action of curcumin when T cells get activated with different stimuli and also confirmed the multifarious targets of curcumin in the activated T cells [30]. It was recently acknowledged that IL-17 producing Th1 cells play an instrumental role in the established model of experimental autoimmune encephalomyelitis (EAE), which mimics multiple sclerosis. Oral curcumin (100 or 200 mg/kg body weight) administration in the rats suppressed the frequency of inflammatory cells in the spinal cord along with lowering the frequency of paralytic incidences, which was the disease marker in this model. The decreased level of IL-17, transforming growth factor beta (TGF-β), IL-6, IL-21, STAT3 expression and STAT3-phosphorylation was reported in curcumin-treated groups. Also, it was shown that curcumin blocks the differentiation of Th-17 cells by blocking STAT-3 transcription in T cells. Moreover curcumin inhibited neural AgMBP68–86 peptide specific lymphocytes responses and IL-17 mRNA expression. These recent evidences furthermore proved the significance of curcumin in the IL-17 mediated disorders [31]. Bright and colleague reported that T cells expressing Toll-like receptors-4 and 9 (TLR4 and TLR9) play an instrumental role in the pathogenesis of EAE model. Curcumin treatment led to the decrease in the expression of PLPp139–151 and MOGp35–55 Ag-induced TLR4 and TLR9 on the CD4+ T cells and CD8+ T cells, which also ameliorated this disease. It was found that TLR 4 and TLR9 acted as co-stimulatory receptors to enhance the proliferation and cytokine production in response to the specific agonists [32]. Previously Bright's group had also reported that curcumin can inhibit IL-12 production in spleen cells, Mϕ and microglia and curcumin can inhibit EAE by blocking IL-12 signaling in T lymphocytes [33]. Curcumin also inhibited the proliferation of mouse splenic T cells that were stimulated with ConA and in a model of type II collagen (CII)-induced arthritis (CIA) in which T cell proliferation was induced by bovine type II collagen Ag. Moreover, curcumin reduced anti-CII IgG2a Ab in the serum of CIA mouse [34]. 3. Immunoinhibitory action of curcumin on dendritic cells (DCs) Being at the centre of various immunological responses, DCs control various pathogenic conditions and recently several groups have investigated the action of curcumin on DCs' function. In a detailed study Kim et al. reported for the first time that curcumin, at a dose of up to 25 μM, inhibits DC maturation and the related immunostimulatory function. They also showed that more than 50 μM concentration was toxic for DCs. Surprisingly however various studies have used 50 μM concentration in different immune cells as described elsewhere in this review and the discrepancy between the uses of different concentration of curcumin reflects the variable dose sensitivities of different immune cells and cell lines to curcumin. 334 R.M. Srivastava et al. / International Immunopharmacology 11 (2011) 331–341 However, it remains to be investigated if curcumin dose sensitivity of immature and mature DCs results in distinct biological outcomes. Kim et al. also showed that curcumin could suppress the lipopolysaccaride (LPS) mediated surface overexpression of CD86, CD80 and MHCII expression in murine DCs during maturation but at the same time curcumin treatment increased the FITC-dextran particle uptake significantly. These observations provided the evidence to modulate DC mediated specific immune response in the autoimmune disorders for regulating the function of T cells [35]. Park and colleagues reported that the pretreatment with curcumin (1–25 μM) could also suppress the LPS (200 ng/ml) induced indoleamine 2,3-dioxygenase (IDO) production in bone marrow derived-DCs (BMDCs). However, curcumin enhanced the COX-2 expression by three fold and prostaglandin E2 production by two fold in the LPS treated DCs. The enhanced prostaglandin E2 level by curcumin was attributed to the suppression of LPS-induced IDO production in this study. The curcumin or prostaglandin E2 treated DCs showed reduced proliferation of OVAspecific CD8+ T cells that was induced by LPS [36]. Although intravenous LPS (3 μg) injection reduces the splenic blood flow by 31% and reduces the access of Ag to the mouse spleen [37], a high dose of LPS (1.5 mg/kg body weight, i.p.) induced heavy IDO in splenic DCs and pre-injection of curcumin (50 mg/kg body weight, i.p.) inhibited the LPS-induced IDO production in the splenic DCs [36]. In contrast to the effect of curcumin on BMDCs showing enhanced COX-2 expression, another study showed the dose dependent (2–16 μM) inhibiton of the production of LPS (0.2 ng/ml) induced COX-2 in the BV2 microglial cells. This contrasting results on the COX-2 level in the BV2 microglial cells and BMDCs may be due to a very high difference in the LPS concentration (0.2 vs 200 ng/ml) or LPS serotype that were used in these studies. Moreover, a distinct cellular response to LPS could not be ruled out in different cell types that may greatly differ in the density of LPS receptor or co-receptors [38]. IFN-γ regulates multiple elements of DCs response and since it is being used for monocyte derived-DCs (MoDCs) conditioning in cancer therapy, it can drive DCs for potent Th1 polarizing activities. Although IFN-γ (5 to 500 IU/ml) treatment was shown to upregulate CD86, CD38, CCR7 on MoDCs in 48 h [39], no significant increase in CD86 and CD80 level was found at 24 h after 200 IU/ml of IFN-γ treatment in murine BMDCs. However, IFN-γ (100 IU/ml) upregulated IDO production in BMDCs and curcumin (1–25 μM) inhibited the functions as well as level of IDO in IFN-γ stimulated murine BMDCs. Thus curcumin reversed the IDO-mediated reduced T cell proliferation function. This study also showed that curcumin can modulate IFN-γ induced IDO expression by affecting Janus kinase 1 (JAK) and protein kinase C δ (PKC) signaling [40]. In a similar direction, additional data showed that curcumin-treated DCs led to the development of anergic CD4+T cells and curcumin-treated DCs could also induce regulatory T cells (Tregs) development. More interestingly, curcumin-treated DCs also promoted the production of IL-10 and αAlDHAa1 (α retinal dehydrogensae). These retinoids function as the regulators of mucosal immune responses. Curcumin induced Treg cells inhibited Ag-specific T cell activation in-vitro and could inhibit colitis caused by Ag-specific pathogenic T cells in-vivo. These findings supported the important role of curcumin in the modulation of DCs' function to achieve tolerogenic responses [41]. Curcumin (1 μM) itself could suppress the LPS-induced IL-12/23p40 production in-vitro, whereas IL-10 (2.5 ng/ ml) and curcumin (0.1 μM) at their suboptimal concentrations acted synergistically to suppress the LPS-induced IL-12/23p40 production from DCs. However, no appreciable therapeutic effect of the dietery curcumin was noticed in the IL10-deficient mice having Th1 mediated colitis. Also, no significant improvement in the colitis was observed as curcumin failed to modify the pathogenic T cells in IL-10 deficient mouse. These results comprehensively confirmed the dependence of curcumin on IL-10 for its immunoinhibitory actions. Nevertheless it provided the evidence that a very little bioavailable concentration of curcumin might effectively modify the overall immune response in combination with IL-10. Curcumin and IL-10 also acted synergistically to inhibit the NF-κB activity in intestinal epithelial cells and thus could provide the additional benefits without influencing the function of immune cells. These results also provoke the possibility of a combinatorial approach to investigate the immunohibitory action of curcumin with other immunohibitory molecules viz. TGF-β, prostaglandins etc. [42]. Such investigations will be valuable to specify the action of curcumin in various pathogenic conditions in future. 4. Immunomodulatory effect of curcumin on natural killer (NK) cells NK cells directly participate in the killing of tumor cells after the recognition of stress inducible ligands and killing involves the induction of cell death by perforin and granzyme B. Various investigators have directly measured the NK cell activity against tumor cells both, in-vitro and in-vivo. In the initial studies, curcumin feeding (1, 20 or 40 mg/kg) up to five weeks showed no effect on the NK cell activity in rats but enhanced the antibody (Ab) responses in rats [43]. In another study, Yadav et al. showed that curcumin treatment can augment NK cell cytotoxicity in-vitro that can further be enhanced by IFN-γ treatment [44]. The generation of IL-2 induced non-specific cytotoxic LAK cells (similar to cytotoxic NK cells) in the presence of curcumin (at 10–20 μM/l) was evaluated by Gau et al. and the cytotoxic activity of LAK cells was determined against NK sensitive YAC-1 lymphoma cells. The results showed little effect on the generation of LAK cell-mediated cytotoxicity, whereas higher dose (30 μM/l) of curcumin inhibited the cytotoxic LAK cell generation [24]. Few serious concerns for the use of curcumin in the melanoma treatment were however raised, which were based on the facts that NK cells from healthy donors treated with curcumin (10 or 20 μM/l) and IL-12 (10 or 50 ng/ml) secreted less amount of IFN-γ. Moreover, curcumin-treated NK cells also showed reduced granzyme B to kill K562 and A375 melanoma cell lines and curcumin slightly reduced production of IFN-γ by NK cells in the presence of A375 melanoma and K562 target cell lines. Although this study found the direct effect of curcumin on tumor cells it could not provide appreciable reasons for the use of curcumin as the ‘modifiers of NK mediated immune responses’ in favour of its use in anti-tumor therapies [45]. Similarly, our previous study had shown that curcumin injections for prolonged duration had no effect on the NK cell activity in-vivo, during the progression of ascites tumor [25]. However, we observed larger solid tumor with curcumin in the transplanted subcutaneous AK-5 tumor that interestingly underwent rapid spontaneous regression. Also, an enhanced activation of NK cells was observed after curcumin treatment that correlated with the response of curcumin on the tumor in-vivo [46]. Such an effect clearly describes the effect of curcumin as ‘NK cells modifier’, however its in-vivo effects may be highly dependent on the specific pathology of the diseases. Recently, tumor derived exosomes attracted much attention as they can effectively modulate the anti-tumor immune responses. Zhang et al. showed that curcumin enhanced the proteasomal degradation of tumor derived exosomal proteins that inhibit IL2-induced NK cell activity against breast carcinoma, partially restoring the NK cell activity against tumor. Such an action of curcumin displayed that curcumin can also target the immune escape strategies that are critical for the immune responses [47]. 5. Immunomodulatory effect of curcumin on monocytes and macrophages (Mϕ) Monocyte recruitment at the inflammatory site plays a vital role in the inflammatory response. Curcumin inhibited the tumor necrosis factor α (TNF-α) induced adhesion of monocytes on human endothelial cells. The TNF-α induced upregulation of Inter-Cellular Adhesion Molecule 1 (ICAM-1), vascular cell adhesion molecule-1 R.M. Srivastava et al. / International Immunopharmacology 11 (2011) 331–341 (VCAM-1) and endothelial cell leukocyte adhesion molecule-1 (ELAM-1) on monocytes was completely inhibited by curcumin. The curcumin mediated blocking of these adhesion molecules was attributed to the inhibitory effect on NF-κB activation. These results showed the promising activities of curcumin in the local inflammatory responses like arthritis as well as in metastasis [48]. PMA or LPSinduced production of inflammatory cytokines viz. TNF-α, IL-8, macrophage inflammatory protein 1 alpha (MIP-1α), monocyte chemoattractant protein (MCP-1) and IL-1β in monocytes and alveolar Mϕs was significantly inhibited by curcumin in a dose dependent manner [49]. Lim et al. also showed that curcumin blocked the enhanced expression and secretion of PMA induced inflammatory cytokine MCP-1 in U937 monocytic cell line [50]. Although curcumin completely blocked LPS mediated NO production in RAW264.7 cell line, it enhanced the phagocytosis of fluorescent beads and the surface expression of CD14. The enhancement of pahgocytic activity and the surface expression of CD14 followed a similar pattern, however no direct role of curcumin mediated enhanced CD14 surface expression was described for the phagocytic capcity [51]. Previously, an independent study had also shown a significant increase in Mϕ phagocytic activity in curcumin-treated animals. These actions of curcumin on Mϕs also described the enhanced scavenging capacity under non-inflammatory conditions [52]. Prolonged alcohol treatment led to oxidative stress and the mononuclear cells obtained from alcoholic animals showed lesser capacity for collagen surface attachment; however alcohol in conjunction with curcumin showed normal adhesion potential of mononuclear cells. This study showed the reduction in the toxic effect of alcohol by curcumin in the prolonged duration [53]. Pretreatment with curcumin inhibited the LPS-induced TLR-2 mRNA and NF-κB level in RAW264.7 cells [54]. Treatment with bisdemethoxycurcumin inhibited the LPS-induced NO production in RAW264.7 cells, which was abrogated by blocking the activity or the expression level of heme oxygenase-1. It was also shown that anti-inflammatory effects mediated by bisdemethoxycurcumin signaling to heme oxygenase-1 involve **Ca2+/calmodulinCaMKII-ERK1/2-Nrf2 cascade in RAW264.7 Mϕ cells [55]. Sumanont et al. have furthermore reported that curcumin manganese complex (CpCpx) and diacetylcurcumin manganese complex (AcylCpCpx) have greater NO radical scavenging activity than their parent compounds, curcumin and acetylcurcumin, respectively [56]. In diabetic condition, a massive increase in the inflammatory cytokines has been reported. To evaluate the anti-inflammatory effect of curcumin under high glucose mediated inflammatory responses, Jain et al. studied the effect of curcumin and placebo supplementation on plasma level of TNF-α, IL-6, MCP-1, glucose and oxidative stress in streptozotocin-treated diabetic rats [57]. Curcumin treatment significantly reduced the high glucose mediated upregulation of inflammatory cytokines along with increasing the lipid peroxidation. However curcumin had no effect on the reduced insulin level under diabetic conditions. In this study, the anti-inflammatory action of curcumin on the high glucose induced IL-8, TNF-α, IL-6, MCP-1 was also shown by using human promonocytic U937 cell line. Androgen receptor is a nuclear receptor that translocates to the nucleus following ligand binding and modulates the function of various genes. In the healing skin androgen receptor were detected in infilitrating Mϕs, keratinocytes and in dermal fibroblasts that indicated its possible function in the healing process. Androgen receptor activity in the presence of 5α-dihydrotestosterone induced TNF-α promoter activity in Mϕs. The curcumin derivative ASC-J9 disrupts the androgen receptor and its co-regulator interaction resulting in the increased androgen receptor degradation and the decreased androgen receptor transactivation. The topical application of ASC-J9 cream in mouse resulted in quick wound healing and also decreased local TNF-α expression. This study concluded that the curcumin derivative ASC-J9, which acts by inhibiting androgen receptor activity, could be utilized in wound healing as an anti- 335 inflammatory agent [58]. Alzheimer's disease is a complex disorder mainly characterized by deposition of large amount of amyloid-β (Aβ) peptide and subsequent massive inflammatory response. Heavy infilitration of monocytes and Mϕ has been observed in the affected tissue with Aβ deposition. Giri et al. demonstrated that both Aβ1–40 and fibrilar Aβ1–42 peptide are abundantly present in the plasma of patients along with the increase in the level of cytokines TNF-α and IL-1β and chemokines MCP-1, IL-8 and MIP-1β. Activation of transcription factors AP-1 and EGR-1 regulates the level of cytokines and chemokines in THP-1 monocytic cells and in peripheral blood monocytes [59]. Based on the ability of curcumin to block inflammation as well as to modulate the activities of β-secretase and acetylcholinesterase, in-vitro and in-vivo studies with curcumin led to suppressed Aβ deposition and aggregation in experimental animals [60]. In the evaluation study on the role of curcumin in Alzheimer disorder Giri et al. furthermore showed that curcumin could block the Aβ1–40-induced expression of TNF-α, IL-1β, MCP-1, IL-8, MIP-1β and CCR5. Also, it was reported that curcumin could inhibit Aβ-induced Egr-1 DNA-binding activity. These results provided the mechanism of the anti-inflammatory action of curcumin in this disease [61]. 6. Immunomodulatory effect of Curcumin on B cells Decoté-Ricardo et al. evaluated the effects of curcumin on murine spelnic B cells. LPS-induced IgM secretion as well as CpG and TLR4induced proliferation of B cells was inhibited following curcumin treatment. However curcumin failed to exert anti-proliferative effect when the B cell prolifearion was induced by the T-independent type 2 stimuli anti-delta-dextran or by the anti-IgM Ab. Moreover curcumin (10 μM) had no effect on the calcium mobilization induced by antiIgM (10 μg/ml) Ab. Interestingly, however, curcumin inhibited the TLR ligand and anti-IgM induced phosphorylation of ERK, Iκ-B and p38 kinase along with inhibiting NF-κB activation. These observations indicated the anti-inflammatory effects of curcumin in the B cell response [62]. Another study described that the mitogen-LPS-induced proliferation of B cells can be dose dependently inhibited by curcumin (1–20 μM); additionally the LPS-induced secretion of IgG1 and IgG2a was inhibited by curcumin. However, the curcumin mediated inhibition of IgG1 secretion was more pronounced than the inhibition of IgG2a secretion [63]. An independent study also described that curcumin (10 μM) can also inhibits the production of IgE from rat splenocytes [64]. Epstein barr virus (EBV) can immortalize human B lymphocytes in-vitro and immortalization is promoted by the oxidative stress induced by potent immunosuppressive drug cyclosporine A and with hydrogen peroxide. Curcumin (20 μM) aborted the EBV induced B cell immortalization process. This effect of curcumin may be exploited to prevent post-transplant lymphoproliferative disorders in patient receiving cyclosporine A, which otherwise may promote EBV induced B cell immortalization [65]. Later on, it was found that the curcumin modulates this immortalization process by enhanced apoptosis in the virus infected B cells [66]. In animals with spontaneous polyps in the intestinal mucosa, curcumin treatment resulted in 40% increase in B cell numbers in the intestinal mucosa, suggesting the therapeutic responses to curcumin [26]. B cell receptor (BCR) signaling regulates the induction of apoptosis in chronic lymphocytic lymphoma. The central mediator of BCRsignaling is the spleen tyrosine kinases, that govern the function and survival of B cells, and a high level of phosphorylated spleen tyrosine kinase was found in lymphoma cells in comparison to healthy B cells [67]. Curcumin differentially modulated the cytotoxicity of primary chronic lymphocytic lymphoma in comparison to healthy B cells [67]. Rats that received 1, 20 or 40 mg/kg curcumin for 5 weeks showed significantly enhanced IgG only at 40 mg/kg levels whereas animals receiving lower dietary concentrations (1 or 20 mg/kg) of curcumin had same IgG level as that of control with no dietary curcumin. These observations suggest that a threshold level of bioavailable curcumin is 336 R.M. Srivastava et al. / International Immunopharmacology 11 (2011) 331–341 also needed to modulate the IgG mediated responses [43]. A recent study showed that curcumin (6 to 50 μM) could suppress the expression of division dependent upregulation of activation-induced cytosine deaminase, which plays pivotal role in the Ig class switch recombination and somatic hyper-mutation and participates in tumorigenesis. Also the decrease in the recovery of IgG + classswitched B cells within the divided population was observed. These observations suggest the potential of curcumin in the treatment of B cell autoimmune disease [68]. 7. Immunomodulatory effect of curcumin on neutrophils and eosinophils and mast cells and its anti-oxidant properties Several independent studies have provided the evidence that curcumin can act on various aspects of neutrophil function, in a stimulus specific manner and may thus dampen the neutrophil mediated inflammatory response [69]. Chemotactic peptide Nformyl-methionyl-leucyl-phenylalanine (FMLP) and zymosan activated plasma induced aggregation of the monkey neutrophils could be inhibited by the curcumin (1 mM). FMLP peptide, zymosan and arachidonic acid induced production of oxygen radical was attenuated by the curcumin treatment. Calcium ionophore A23187 could nullify the curcumin effect by interfering with the effect of curcumin in neutrophils [69]. Neutrophils play significant role in the damage of joint tissue in the rheumatoid arthritis. A recent study demonstrated the reduced level of oxygen radical generation by neutrohphils upon treatment with curcumin both in-vitro and in-vivo. Adjuvant induced arthritis enhanced the neutrophil frequency in the blood that remain unaltered by curcumin. The stimulation of neutrophils by PMA led to increased level of PKC isozymes, α and β II, which was abrogated by curcumin treatment without interfering with neutrophils vital functions [70]. Similarly, the crystal induced neutrophil activation that served as a model of induced arthritis or rheumatoid arthritis condition was inhibited by curcumin [71]. Oral administration of curcumin (40–60 mg/kg body weight) increased survival of mice by 70% in response to heavy dose of LPS (40 mg/kg body weight). Moreover curcumin suppressed the LPS mediated neutrophil infiltration in liver that was the primary cause of liver damage. However, the reduction of infiltration was limited to the liver only, because whereas hepatic venules had same frequency of neutrophils as that of without curcumin. The reduction of LPS-induced infiltration of neutrophils was also correlated with the reduced levels of ICAM-1 and VCAM-1 in the liver tissue that influence neutrophil adhesion [72]. Without affecting the viability, curcumin (100 μM) significantly reduced the IL8 induced chemotactic activity of neutrophils in dose dependent manner and curcumin modulated this chemotaxis by dampening the IL-8 induced Ca++ ion mobilization. Surface CXCR1 and CXCR2 were internalized upon IL-8 treatment and curcumin treatment enhanced the intracellular level of CXCR1 and CXCR2 in conjunction with IL-8, which indicated that the effect of curcumin on the reduced migration of neutrophils might be attributed to the reduced IL-8 receptors. However, curcumin itself downregulated surface IL-8 receptor CXCR1 and CXCR2 and also blocked the recycling of these receptors on neutrophils. The Rab GTPase family (Ras superfamily of monomeric G proteins) plays pivotal role in the cellular transport mechanism. Interestingly, both CXCR1 and CXCR2 showed enhanced binding with Rab11 upon curcumin treatment, which could potentially block the recovery of IL-8 to cell surface. This study revealed the intricate mechanism that curcumin triggers to achieve anti-inflammatory responses meadiated by neutrophils [73]. LPS induced lung damage and reduction in lung and bronchoalveolar lavage fluid protein content, which was accompanied by enhanced numbers of neutrophils and elevated myeloperoxidase activity in cell-free lavage. Elevation in the cytokine-induced neutrophil chemoattractant-I protein level was seen in response to LPS in the lung tissue, which was significantly reduced by the pretreatment with curcumin. This shows an important protective response of curcumin by dampening neutrophil function in lung injury [74]. In a murine model of asthama, which was induced by OVA-Ag and which had airway hyper-responsiveness to allergens, curcumin (i.p, 10 or 20 mg/kg body weight) decreased the frequency of eosinophils and the inflammatory cells, inhibited iNOS (inducible nitric oxide synthase) expression in lungs and also suppressed the level of IL-4 and IL-5 in bronchoalveolar lavage fluid [75]. An interesting study involving the action of curcumin on mast cells indicated that curcumin reversibly inhibits the degranulation of mast cells along with inhibiting secretion of IL-4 and TNF-α. The evaluation of the antiallergic affect of curcumin was performed by utilizing passive cutaneous anaphylaxis in the mouse ear model. Oral administration of curcumin (50 mg/kg) suppressed the mast cell dependent IgE and Ag-induced local passive cuataneous anaphylaxis [76,77]. Effect of curcumin during myocardial ischemia/reperfusion injury with cardioplegia was also investigated [78]. The postoperative increase in the IL-8, IL-10, TNF-α levels in the plasma was decreased by curcumin. Also curcumin inhibited the activation of neutrophils in myocardium that was estimated by the myloperoxidase activity assay [78]. 8. Curcumin in health and disease Due to the fact that curcumin has been shown to be associated with a number of physiological processes and that it has a wide variety of cellular targets, its therapeutic role has been studied in several inflammatory and non-inflammatory disorders. In this section, we discuss most recent findings related to its direct application in health and disease. 8.1. Role of curcumin in the neoplastic diseases Curcumin has received maximum attention owing to its antitumor properties. Several hundred reports in the last two decades have shown its ability to selectively kill transformed cells across almost all types of tumors. Curcumin can exert its anti-tumor effects at two levels, (i) at the level of tumorigenesis or (ii) in selectively inducing apoptosis in tumor cells. Huang et al. have discussed the anti-carcinogenic effects of curcumin in duodenal and colon cancer in mice. In this study, dietary curcumin could significantly reduce tumor load during both pre initiation and post initiation of chemical induced carcinogenesis [79]. Similarly, curcumin application inhibited the induction of epidermal DNA synthesis and the tumor promotion in skin following 12-0-tetradecanoyl phorbol-13-acetate (TPA) treatment [80] as well as benzopyrene induced DNA adducts and skin tumors and DMBA induced skin tumors [79]. Rao et al. have shown that curcumin (200 ppm) in the diet could significantly suppress azoxymethane-induced colonic aberrant crypt foci formation, which are early preneoplastic lesions, and colon tumor incidence and tumor multiplicity [81]. These effects of curcumin in inhibiting tumorigenesis involve inhibition of arachidonic acid metabolism; decrease in TPA induced ornithine decarboxylase activity and inhibition of DNA synthesis. It was thought that metabolites of arachidonic acid such as HPETEs, HETEs, leukotrienes and prostaglandins play an important role in TPA induced inflammation and tumor promotion [82,83]. Similarly ornithine decarboxylase (ODC) is a rate limiting enzyme in polyamine synthesis [81] and its overexpression has been linked with cell transformation and carcinogenesis in skin, breast and colon. Thus, it is logical to speculate that inhibiting arachidonic acid metabolism and/or ODC activity shall result in an inhibition of tumorigenesis. In addition, curcumin has also been shown to cleave β-catenin, which impairs Wnt signaling and cell–cell adhesion pathways, which are critical in the development and promotion of many types of tumors including colorectal cancer [84]. Curcumin also induced downregulation of cyclin D1 expression and CDK-4 activity in breast and squamous cell carcinoma cell lines [85]. The suppression of cyclin R.M. Srivastava et al. / International Immunopharmacology 11 (2011) 331–341 D1 by curcumin led to inhibition of CDK-4 mediated phosphorylation of retinoblastoma, which is a crucial step for the cell to pass through the G1 phase of cell cycle and become transformed [86]. In addition to its inhibitory effects on neoplastic transformation, curcumin has been shown to induce apoptosis in tumor cells by various mechanisms, which include impairment of the ubiquitin proteasome pathway, upregulation of proto-oncoprotein Bax, activation of caspases and induction of Fas receptor aggregation in a Fas ligand dependent manner and the generation of free radicals [87–90]. One of the several possible mechanisms of apoptosis induction by curcumin involves the inhibition of proteasome complex. In mouse neuro 2a cells, exposure to curcumin revealed a dose dependent decrease in the proteasome activity and an increase in the ubiquitinated proteins. Curcumin also decreased the turnover of the destabilized enhanced green fluorescent protein suggesting an inhibition of the cellular proteasome machinery [90]. Another mode of apoptosis induction by curcumin involves the upregulation of p53 in tumor cells. In human basal cell carcinoma, apoptosis induction by curcumin resulted in induction of p53 and its downstream targets, p21 waf1/cip1 and GADD45, which are known to regulate apoptosis under stress conditions [91]. Work in our laboratory has shown that curcumin induced apoptosis involves the production of reactive oxygen intermediates (ROIs) and involves activation of caspase-3 [87]. A large number of reports confirm that curcumin induced typical apoptotic mode of cell death in a wide variety of tumors complete with mitochondrial depolarization and caspase-3 activation. However, some studies suggest that apoptosis induced by curcumin is independent of caspase-3 [92,93]. Curcumin has also been reported to induce an apoptosis like pathway, which is independent of not only caspases but mitochondria as well [93]. These effects of curcumin in Jurkat T cells were accompanied by DNA fragmentation into high but not low molecular weight fragments and the frequency of opening of the mitochondrial permeability transition pores in curcumin-treated cells was decreased compared to the control untreated cells. However, one of the most commonly shown effects of curcumin on tumors is its ability to induce the opening of mitochondrial permeability transition pore, which in turn induces the collapse of the mitochondrial membrane potential, respiration impairment ultimately leading to cell death [94,95]. This observed difference in curcumin's action with respect to the opening of permeability transition pore could be attributed to the large difference in its concentrations that were used during these studies. 8.2. Curcumin in cardiovascular disease The therapeutic effects of curcumin in the development and progression of cardiovascular disease have been studied to some depth in the last decade. Owing to its ability to regulate oxidant stress, curcumin has been shown to be effective against cardiac hypertrophy, cardiomyocyte apoptosis following myocardial infarction and ischemia/reperfusion injury [96–98]. Cardiac hypertrophy is the remodeling of the left ventricle following pressure or volume overload that results in ventricular wall thickening and an increase in overall cardiac dimensions. It begins as a compensatory process that becomes maladaptive over time and leads to heart failure [99,100]. Development of hypertrophy involves activation of the calcium and redox sensitive transcription factor NF-AT that brings about the metabolic and biochemical changes within the cardiomyocyte [101]. Transcriptional activation associated with hypertrophy has been recently shown to be regulated by acetylation and deacetylation events at histone lysine tails [102]. Acetylation and deacetylation of histones is carried out by enzymes called histone acetyl transferases (HATs) and histone deacetylases (HDACs), respectively. Various HDACs have been implicated in the pathogenesis of cardiac hypertrophy. For example, loss of class 2 HDAC results in development of hypertrophy while loss of class-1 HDAC confers resistance to hypertrophic growth [102,103]. 337 Morimoto et al. studied the effects of curcumin on HAT and progression of hypertrophy and subsequent decompensated heart failure. They have shown that exposure of isolated neonatal rat cardiomyocytes (NRCMs) to 5 or 10 μM curcumin completely suppressed the induction of hypertrophic response following phenylephrine treatment, a known inducer of cardiac hypertrophy [97]. In an in-vivo setting also, administration of curcumin prevented the development of hypertension induced heart failure in salt sensitive Dahl rat model of hypertension [97]. These data strongly suggest that curcumin possesses anti-hypertrophic properties both in-vitro and invivo. It has been proposed that curcumin may inhibit hypertrophic remodeling by two mechanisms (i) by inhibition of histone acetylation through inhibition of HATs and (ii) by disrupting p300/ GATA4 transcriptional complex through a completely independent mechanism. Curcumin has also been shown to inhibit p300 mediated acetylation of p53, both in-vitro as well as in-vivo [104]. Similarly, inhibition of NF-κB by curcumin could also be involved in the antihypertrophic effects of curcumin since NF-κB signaling is involved in cardiomyocyte hypertrophy [105]. Oxidative stress is a major outcome determinant in myocardial infarction and ischemia/reperfusion and curcumin's anti-oxidant property has been shown to prevent isoproterenol induced myocardial necrosis in rats [106]. In models of experimentally induced myocardial infarctions such as isoproterenol treatment, decrease in lysosomal stability leading to increase in lysosomal autolytic enzymes has been reported [107,108]. Curcumin has been shown to stabilize membranes and thereby suppress the infarct induced increase in myocardial lysosomal enzymes [109,110]. Besides, the generation of free radicals following ischemia/reperfusion can also be controlled by curcumin due to its strong anti-oxidant properties. The cardiotoxicity associated with doxorubicin, a potent drug for treatment of a broad array of cancers, is a major concern for cancer patients [111]. Animal studies have shown that doxorubicin treatment induces free radical generation and p53 activation, decreases glutathione and increases serum peroxidase and catalase [96,112]. Curcumin treatment significantly attenuated the cardiotoxic effects of doxorubicin [113]. The beneficial effect of curcumin in blockade of doxorubicin cardiotoxicity can be linked to modulation of intracellular redox status by curcumin. In a study by Feng et al., curcumin completely abrogated the induction of glucose induced hypertrophy in cardiomyocytes [114]. Glucose induced cardiomyocyte hypertrophy is mediated by p300 upregulation and subsequent activation of p300 dependent transcription factors. Since, curcumin can inhibit p300, exposure to curcumin prevented the induction of p300 mediated hypertrophic response in cardiomyocytes [114]. In human patients curcumin could markedly reduce the generation of glucose induced reactive oxygen species (ROS) in red blood cells. Myocardial tissue from diabetic rats exhibited higher levels of eNOS and iNOS mRNA and curcumin treatment considerably inhibited the upregulation in both eNOS and iNOS transcript levels [115]. Collectively, these studies have established the usefulness of curcumin in the treatment of various cardiovascular ailments. However, caution need to be exercised while reproducing curcumin's anti-tumor effects in the cardiovascular system due to the extremely different metabolic and biochemical nature of the two cell types. The molecular pathways targeted by curcumin in tumor cells may not be targeted at all in the cardiomyocytes or may be modulated in a markedly different manner so as to drastically change the physiological outcome. One of most important difference between the two cell types is the metabolic signature; tumor cells have a higher dependence on glycolysis while cardiomyocytes mainly depend on lipid oxidation for their metabolic needs. Similarly, the proteome and the transcriptome within cardiomyocytes are regulated quite differently than in a transformed cell type. This calls for a careful examination and analysis of curcumin dosage along with mechanistic details of its physiological effects within different cell types before establishing curcumin in any independent or combinatorial drug regimen. 338 R.M. Srivastava et al. / International Immunopharmacology 11 (2011) 331–341 8.3. Curcumin in neurodegenerative disease Brain is perhaps the most susceptible organ to oxidative damage due to the highly oxidative intracellular environment of the neurons and glial cells [116]. Oxidative stress has been shown to increase both with normal brain ageing as well as with brain injury [117,118]. ROS generation in the brain can enhance the production of nitric oxide by activating neuronal nNOS and iNOS. Nitric oxide is a known mediator of glutamatergic transmission and has been shown to be involved in ageing and age related neurodegenerative disorders [117]. Accumulation of redox active metals such as iron, copper and zinc, due to high levels of ascorbic acid in the brain that facilitates redox metal reactions, aggravates the oxidative load in the brain [119]. An elevation in the free radicals and oxidative stress in turn induces the activation of NF-κB and other inflammatory molecules such as IL1β and TNF-α [120,121]. The neuroprotective effects of curcumin have been described in a variety of stress models. In an oxidative damage induced neurodegeneration model, Guangwei et al. have shown the ability of curcumin to attenuate acrylonitrile induced oxidative damage in the brain [122]. In this study, curcumin dose of 100 mg/kg of body weight prevented lipid peroxidation and glutathione depletion in response to acrylonitrile exposure. In another study, curcumin could increase the cholinergic activity of neurons and free radical scavenging in streptozotocin induced dementia in rats [123]. The ability of curcumin to increase cholinergic activity in the brain is mediated by an increase in the acetylcholinesterase enzyme. It has been previously reported that curcumin attenuated diabetic encephalopathy by a similar free radical scavenging effect and increase in acetylcholinesterase activity [124]. Acute traumatic brain injury results in a widespread ‘secondary brain damage’ following the primary mechanical damage. One of the most critical mediators of the rather chronic secondary brain injury is the oxygen derived free radical species. Kontos and Povlishock have shown upregulation of the superoxide radical (O2−) in the brain microvasculature immediately following acute injury [125,126]. The superoxide radical can be generated from various enzymatic reactions such as arachidonic acid cascade, oxidation of amine neurotransmitters, mitochondrial leakage and xanthine oxidase activity [127]. The ability of curcumin to sacavenge oxygen derived free radicals has been implicated in its potential as a neuroprotective agent. Dietary curcumin supplementation has been shown to maintain energy homeostasis after brain trauma [128]. Cerebral edema, a cause of increased intracranial pressure and poor clinical outcome after acute brain injury, was significantly controlled by pretreatment (75– 150 mg/kg body weight) as well as post treatment (300 mg/kg body weight) with curcumin [129]. The protective effects of curcumin were associated with inhibition of IL-1β expression and inhibition of aquaporin-4 induction. Wakade et al. have shown that curcumin can attenuate vascular inflammation following subarachnoid hemorrhage while another study by Zhao et al. has described neuroprotection conferred by curcumin after cerebral ischemia [130,131]. These findings support the notion that intervention with curcumin treatment at any point during the brain injury can change the clinical outcome. Effects of curcumin on the pathophysiology of Alzheimer's disease have been studied somewhat extensively and several groups have shown its ability to inhibit Aβ-plaque formation [132,133]. In a mouse model of Alzheimer's disease, low doses of curcumin (160 ppm) decreased the plaque burden and reduced the soluble as well as insoluble forms of Aβ by about 50% [134]. Yang et al. have described the ability of curcumin to inhibit the formation of Aβ-oligomers. They have also shown that curcumin can bind to the amyloid plaques and significantly reduce in-vivo plaque formation [135]. They have earlier shown the efficacy of curcumin in reducing CNS lipid peroxidation and iNOS [136], which in turn can lower the oxidative stress. One of the possible mechanisms suggested in curcumin's ability to inhibit plaque formation is the high affinity with which it binds redox reactive metals such as copper and iron and therefore may act as a potent anti-oxidant by chelating redox reactive metals [137]. Amyloid plaque burden has been associated with depolarizing of the neuronal membrane and enhanced glutamate-mediated excitotoxicity [138,139] that results in impaired electrical firing of the neurons. Curcumin administration has been shown to prevent misfiring of neurons following Aβ burden in embryonic hippocampal neurons [140]. In nitrosourea induced neurotoxicity, curcumin administration prevented increase in the activity of glucose metabolic pathway enzymes including hexokinase, LDH and SDH [141]. Similarly, in mercury induced neurotoxicity, pretreatment with curcumin abrogated the increase in metallothinine mRNA and suppressed the toxic and oxidative stress load following mercury exposure in rats [142]. Cumulatively, these studies suggest that curcumin can help in maintaining the oxidative intraneuronal environment and thereby protect brain from a variety of oxidative, toxic and mechanical injuries. 8.4. Immunomodulatory action of curcumin in the prevention of inflammatory diseases Curcumin administration has been shown to be associated with a positive outcome in a large number of chronic inflammatory diseases due to its ability to inhibit NF-κB activation and subsequent inflammatory pathways. Starting with its use in biliary disease in 1937, curcumin has now been shown to ameliorate almost all kinds of liver toxicity and disease. Curcumin can inhibit the increase in serum ALT and AST enzymes following iron induced liver toxicity [143]. The biochemical and histopathological changes induced by ethanol toxicity were abrogated by curcumin administration [144]. Curcumin could also protect against thiodoacetamide induced hepatitis and cirrhosis in rats. It also protected against carbon tetrachloride induced livertoxicity and reversed carbon tetrachloride induced cirrhosis [145]. The underlying mechanism for the effects of curcumin on liver involves its ability to act as an oxidant and inhibit NFκB activation thereby inhibiting the inflammatory signaling cascade. Curcumin could protect against dinitrobenzene sulfonic acid induced model of murine colitis by suppressing p38 kinase and IL-1β activation [146]. In a similar murine model of inflammatory bowel disease, intragastric administration of curcumin inhibted the increase in intestinal neutrophil infiltration and serine protease activity, suggesting its promising therapeutic potential in the treatment of inflammatory bowel disease [147]. Rheumatoid arthritis is another chronic pro-inflammatory disease that has been shown to be targeted by curcumin. Several studies have reported the physiologically beneficial effects of curcumin in the management of rheumatoid arthritis [148,149]. These studies have shown the ability of curcumin to inhibit the increase in serum acidic glycoproteins and matrix metalloproteinase expression that is generally associated with the progression of disease in rheumatoid arthritis patients. Decreased apoptosis of synovial fibroblasts is one of causes for joint inflammation and stiffness in rheumatoid arthritis patients and Park et al. have shown that curcumin could induce apoptosis in synovial fibroblast by upregulation of proapoptotic genes including bax and a simultaneous downregulation of anti-apoptotic genes including bcl-2 and XIAP [150]. Chronic inflammatory bowel disease is a life threatening disease that affects children and adults. Elevated level of pp38 kinase was seen in the intestinal mucosa of ulcerative colitis and Crohn's disease biopsies, which was inhibited by curcumin (5–20 μM) ex-vivo. Curcumin suppressed the production of pro-inflammatory cytokine IL-1β and enhanced the production of IL-10 in the ex-vivo cultured mucosal biopsies. However, it had modest yet consistent effect on the reduction of IL-1β level [151]. Due to its strong inhibitory effects on cyclooxygenases1 and cyclooxygeanse-2, lipoxygenase, TNF-α, IFN-γ, iNOS and NF-κB, curcumin (360 mg/dose; 3 or 4 times/day for three months) showed promising response in patients and could reduce R.M. Srivastava et al. / International Immunopharmacology 11 (2011) 331–341 clinical relapse in patients with quiescent inflammatory bowel disease [152]. TNF-α plays instrumental role in the pathogenesis of inflammatory skin disorder psoriasis and since curcumin is a strong antiinflammatory agent, its action in the HaCa T keratinocytes was investiagted. Curcumin (20 μM) aborted the TNF-α induced expression of IL-1β, IL-6, IL-8 and TNF-α in keratinocytes. Curcumin also blocked the activation of NF-κBp65, pJNK, pp38 kinase activation and downregulated Cyclin E level. Without modulating the TNF receptor I and II expression, TNF-α induced activation of NF-κB in human umbilical vein endothelial cells was blocked by curcumin. Also, curcumin inhibited the pJNK level, pP38 kinase level and STAT-3 activation along with lowering the intracellular ROS level. The expression of ICAM-1, MCP-1, and IL8 was attenuated by curcumin at both mRNA and protein level. These studies indicate the protective effect of curcumin in the treatment of various pro-inflammatory diseases [153]. 9. Concluding remarks and future perspectives Immunomodulatory properties of curcumin are mostly immunosuppressive, but in some cases immunostimulative effects have been reported. 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