Impact of Curcumin/Piperine on Obese Mice
It has been well-accepted that the high fat/high caloric content present in the typical Western diet adversely impacts health and is a common cause of obesity. In turn, studies have provided evidence linking obesity to the high risk of developing type 2 diabetes, cardiovascular disease, and metabolic syndrome. It is believed that a possible mechanism underlying this link is a chronic, low-grade inflammatory state developed as a consequence of increased adiposity, which has been supported by numerous studies in the past decade. While the obesity-associated inflammation has been extensively studied, the impact of obesity on the innate and adaptive immune response, in particular that of T cell-mediated function, has not been well-clarified thus far, though available evidence suggests that obesity may compromise the immune-surveillance pathways leading to impaired immune response against pathogens. Further, the effect of weight reduction by CR and its combination with spice components on immune and inflammatory responses of obese mice has not been well-defined. Here we report that CR results in a reduction in body weight, fat mass, and inflammatory response, as well as an enhancement in T cell-mediated function compared to the mice fed the same high fat diet ad libitum. We also report that the addition of spice components curcumin and piperine does not further enhance CR's benefit, but rather it blunts the effect of CR on T cell-mediated function.
In this study, we found that CR applied to the obese mice provided metabolic benefits as indicated by the reduced weight gain and fat mass and by lowered levels of fast blood glucose and insulin. These results agree with previous reports on CR in both humans and animals. Contrary to what we anticipated, concurrent consumption of curcumin and/or piperine had no further effect on these CR-induced changes. However, this is consistent with the similar lack of additional effects of these spice compounds on inflammation profile given the currently recognized association between adiposity, inflammatory state, and glucose metabolism.
Previous studies have reported that CR administered to lean or obese rodents increases total T cell population and both CD4 and CD8 subpopulations. Consistent with these previous results, in the current study, which focused on CD4 T cells and their subpopulations, we found that spleens from CR mice had higher percentages of CD4 T cells. This increase in the proportion of CD4 T cells may partly explain the higher levels of splenocyte proliferation in CR mice because only T cells in the spleen cell mixture should proliferate after being stimulated by T cell mitogens or TCR antibodies, and a larger number of T cells to begin with would be expected to result in a larger number of proliferating cells in the end. However, we cannot rule out that CR might have enhanced the T cell division and thus the proliferation rate.
Also consistent with the previous studies showing enhanced T cell proliferation and IL-2 production in CR mice and humans, we found that obese mice subjected to CR had higher levels of splenocyte proliferation and IL-2 production in response to both T cell mitogen and anti-CD3/CD28 stimulation. Of note, CR mice also had lower PGE2 production compared to the control mice, which confirmed our previous findings in mice and humans. Previously, we showed that 21% CR for 13 mo in Emory mice reduced PGE2 production in spleen; in a recent human study, we found that 30% CR for 6 mo decreased PGE2 production in LPS-stimulated whole blood culture. Given that PGE2 is a suppressive factor for T cell proliferation and IL-2 production, a reduction in PGE2 production by CR represents another mechanism through which CR might have improved T cell proliferation and IL-2 production in obese mice. We did not evaluate the mechanism of CR-induced decrease in PGE2 production in the current study. However, a study by Kim et al. provided insight into the underlying mechanism. They reported that in kidneys from CR rats, the activity of cyclooxygenase (COX), a rate-limiting enzyme for eicosanoid synthesis, was lower compared to the control rats, and this change was accompanied by a lower expression of COX-2 at both protein and mRNA levels. Although their findings for CR-induced changes were from different tissues (kidney vs. spleen in the current study), we speculate that the down-regulated COX-2 abundance/activity may represent a common mechanism shared across different tissues for the observed effect of CR on PGE2 production. CR's inhibitory effect on PGE2 production also provided another piece of evidence to support the suggested anti-inflammatory property of CR given the fact that PGE2 is a very potent lipid inflammatory mediator and it is involved in the pathogenesis of many inflammatory diseases.
In this study, we found that CR's enhancing effect on T cell proliferation and IL-2 production was lessened by supplementation of CR diets with spice compounds to such a level that CR's effect was no longer significant in any of the CR groups that were supplemented with curcumin and/or piperine. This may reflect a suppressive effect of curcumin and piperine on T cell-mediated functions as reported in several previous studies. For example, it has been shown that dietary curcumin in mice suppresses CD4 T cell proliferation and IL-2 production while in vitro curcumin supplementation inhibits T cell activation, human peripheral blood mononuclear cell proliferation and IL-2 production, and IL-2 synthesis as well as IL-2 signaling in mouse CD4+ T cells. Although little information is available for piperine in this regard, some studies have indeed suggested that piperine may inhibit T cell-related functions.
Both CR and curcumin have been shown to have an anti-inflammatory effect, respectively. Therefore, in this study, we investigated whether inclusion of curcumin in a CR diet would further potentiate CR's beneficial effects. While our results clearly showed an anti-inflammatory effect of CR, which was indicated by a significant reduction of IL-1β and TNF-α as well as a strong trend toward some reduction of IL-6, curcumin and piperine demonstrated no additional benefits. These results lend further support to the suggested beneficial effects of CR toward mitigating inflammation caused by an HFD/obesity condition. Of note, it was a study limitation that all spice compound-supplemented mice were on CR treatment; therefore, it is presently unknown whether these bioactive compounds would have produced an anti-inflammatory effect if administered to HFD control mice without CR regimen.
An increased presence of immune cells in adipose tissue is the hallmark of obesity-induced inflammation. Macrophages were first found in the adipose tissue of obese animals and humans nearly a decade ago. Emerging evidence now suggests that during obesity, T cells precede macrophages in entering adipose tissue and produce pro-inflammatory cytokines to activate macrophages. It is still unresolved as to which of the CD4 and CD8 T cells are the predominant T cells present in the adipose tissue of obese humans and animals and how obesity affects the balance within CD4 T cells and among their different subpopulations, i.e., Th1, Th2, Th17, Treg, etc. The qualitative and quantitative changes in the immune cells present in adipose tissue are proposed to be key factors in the obesity-inflammation-metabolic disorder pathway. In particular, different CD4 T cell subsets have recently attracted increasing attention due to the fact that specific types of CD4 T cells are known to play different roles in maintaining homeostasis as well as regulating immune and inflammatory responses. In contrast to our study's hypothesis, we determined that CR and spice compounds, alone or in combination, have no effect on CD4 subpopulations in peripheral lymphoid tissue (spleen). This indicates that neither CR nor the spice compounds affect CD4 T cell differentiation under HFD/obesity. However, it remains possible that CR may favorably affect CD4 T cell differentiation under certain other conditions. In a recent study, CR in fact attenuated experimental autoimmune encephalomyelitis (EAE, a rodent model for multiple sclerosis) in mice, an autoimmune disease driven by antigen-specific Th1 and Th17 response and mitigated by Treg cells; however, CD4 T cell subsets were not examined in that study. Information regarding the impact of spices on CD4 subpopulation is limited. Dietary curcumin was shown to ameliorate EAE in rats through inhibition of Th17 response and in mice through inhibition of Th1/Th17 responses and enhancement of Th2/Treg responses. Taken together, these results indicate a potential area for future studies to explore: whether the beneficial effect of CR on autoimmune diseases is mediated through its effect on CD4 T cell differentiation and further, whether CR interacts with spice compound supplementation to mutually potentiate their respective effectiveness.
Discussion
It has been well-accepted that the high fat/high caloric content present in the typical Western diet adversely impacts health and is a common cause of obesity. In turn, studies have provided evidence linking obesity to the high risk of developing type 2 diabetes, cardiovascular disease, and metabolic syndrome. It is believed that a possible mechanism underlying this link is a chronic, low-grade inflammatory state developed as a consequence of increased adiposity, which has been supported by numerous studies in the past decade. While the obesity-associated inflammation has been extensively studied, the impact of obesity on the innate and adaptive immune response, in particular that of T cell-mediated function, has not been well-clarified thus far, though available evidence suggests that obesity may compromise the immune-surveillance pathways leading to impaired immune response against pathogens. Further, the effect of weight reduction by CR and its combination with spice components on immune and inflammatory responses of obese mice has not been well-defined. Here we report that CR results in a reduction in body weight, fat mass, and inflammatory response, as well as an enhancement in T cell-mediated function compared to the mice fed the same high fat diet ad libitum. We also report that the addition of spice components curcumin and piperine does not further enhance CR's benefit, but rather it blunts the effect of CR on T cell-mediated function.
In this study, we found that CR applied to the obese mice provided metabolic benefits as indicated by the reduced weight gain and fat mass and by lowered levels of fast blood glucose and insulin. These results agree with previous reports on CR in both humans and animals. Contrary to what we anticipated, concurrent consumption of curcumin and/or piperine had no further effect on these CR-induced changes. However, this is consistent with the similar lack of additional effects of these spice compounds on inflammation profile given the currently recognized association between adiposity, inflammatory state, and glucose metabolism.
Previous studies have reported that CR administered to lean or obese rodents increases total T cell population and both CD4 and CD8 subpopulations. Consistent with these previous results, in the current study, which focused on CD4 T cells and their subpopulations, we found that spleens from CR mice had higher percentages of CD4 T cells. This increase in the proportion of CD4 T cells may partly explain the higher levels of splenocyte proliferation in CR mice because only T cells in the spleen cell mixture should proliferate after being stimulated by T cell mitogens or TCR antibodies, and a larger number of T cells to begin with would be expected to result in a larger number of proliferating cells in the end. However, we cannot rule out that CR might have enhanced the T cell division and thus the proliferation rate.
Also consistent with the previous studies showing enhanced T cell proliferation and IL-2 production in CR mice and humans, we found that obese mice subjected to CR had higher levels of splenocyte proliferation and IL-2 production in response to both T cell mitogen and anti-CD3/CD28 stimulation. Of note, CR mice also had lower PGE2 production compared to the control mice, which confirmed our previous findings in mice and humans. Previously, we showed that 21% CR for 13 mo in Emory mice reduced PGE2 production in spleen; in a recent human study, we found that 30% CR for 6 mo decreased PGE2 production in LPS-stimulated whole blood culture. Given that PGE2 is a suppressive factor for T cell proliferation and IL-2 production, a reduction in PGE2 production by CR represents another mechanism through which CR might have improved T cell proliferation and IL-2 production in obese mice. We did not evaluate the mechanism of CR-induced decrease in PGE2 production in the current study. However, a study by Kim et al. provided insight into the underlying mechanism. They reported that in kidneys from CR rats, the activity of cyclooxygenase (COX), a rate-limiting enzyme for eicosanoid synthesis, was lower compared to the control rats, and this change was accompanied by a lower expression of COX-2 at both protein and mRNA levels. Although their findings for CR-induced changes were from different tissues (kidney vs. spleen in the current study), we speculate that the down-regulated COX-2 abundance/activity may represent a common mechanism shared across different tissues for the observed effect of CR on PGE2 production. CR's inhibitory effect on PGE2 production also provided another piece of evidence to support the suggested anti-inflammatory property of CR given the fact that PGE2 is a very potent lipid inflammatory mediator and it is involved in the pathogenesis of many inflammatory diseases.
In this study, we found that CR's enhancing effect on T cell proliferation and IL-2 production was lessened by supplementation of CR diets with spice compounds to such a level that CR's effect was no longer significant in any of the CR groups that were supplemented with curcumin and/or piperine. This may reflect a suppressive effect of curcumin and piperine on T cell-mediated functions as reported in several previous studies. For example, it has been shown that dietary curcumin in mice suppresses CD4 T cell proliferation and IL-2 production while in vitro curcumin supplementation inhibits T cell activation, human peripheral blood mononuclear cell proliferation and IL-2 production, and IL-2 synthesis as well as IL-2 signaling in mouse CD4+ T cells. Although little information is available for piperine in this regard, some studies have indeed suggested that piperine may inhibit T cell-related functions.
Both CR and curcumin have been shown to have an anti-inflammatory effect, respectively. Therefore, in this study, we investigated whether inclusion of curcumin in a CR diet would further potentiate CR's beneficial effects. While our results clearly showed an anti-inflammatory effect of CR, which was indicated by a significant reduction of IL-1β and TNF-α as well as a strong trend toward some reduction of IL-6, curcumin and piperine demonstrated no additional benefits. These results lend further support to the suggested beneficial effects of CR toward mitigating inflammation caused by an HFD/obesity condition. Of note, it was a study limitation that all spice compound-supplemented mice were on CR treatment; therefore, it is presently unknown whether these bioactive compounds would have produced an anti-inflammatory effect if administered to HFD control mice without CR regimen.
An increased presence of immune cells in adipose tissue is the hallmark of obesity-induced inflammation. Macrophages were first found in the adipose tissue of obese animals and humans nearly a decade ago. Emerging evidence now suggests that during obesity, T cells precede macrophages in entering adipose tissue and produce pro-inflammatory cytokines to activate macrophages. It is still unresolved as to which of the CD4 and CD8 T cells are the predominant T cells present in the adipose tissue of obese humans and animals and how obesity affects the balance within CD4 T cells and among their different subpopulations, i.e., Th1, Th2, Th17, Treg, etc. The qualitative and quantitative changes in the immune cells present in adipose tissue are proposed to be key factors in the obesity-inflammation-metabolic disorder pathway. In particular, different CD4 T cell subsets have recently attracted increasing attention due to the fact that specific types of CD4 T cells are known to play different roles in maintaining homeostasis as well as regulating immune and inflammatory responses. In contrast to our study's hypothesis, we determined that CR and spice compounds, alone or in combination, have no effect on CD4 subpopulations in peripheral lymphoid tissue (spleen). This indicates that neither CR nor the spice compounds affect CD4 T cell differentiation under HFD/obesity. However, it remains possible that CR may favorably affect CD4 T cell differentiation under certain other conditions. In a recent study, CR in fact attenuated experimental autoimmune encephalomyelitis (EAE, a rodent model for multiple sclerosis) in mice, an autoimmune disease driven by antigen-specific Th1 and Th17 response and mitigated by Treg cells; however, CD4 T cell subsets were not examined in that study. Information regarding the impact of spices on CD4 subpopulation is limited. Dietary curcumin was shown to ameliorate EAE in rats through inhibition of Th17 response and in mice through inhibition of Th1/Th17 responses and enhancement of Th2/Treg responses. Taken together, these results indicate a potential area for future studies to explore: whether the beneficial effect of CR on autoimmune diseases is mediated through its effect on CD4 T cell differentiation and further, whether CR interacts with spice compound supplementation to mutually potentiate their respective effectiveness.
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