Curcumin limits weight gain, adipose tissue growth, and glucose intolerance following the cessation of exercise and caloric restriction in rats

Weight regain after dieting and exercise is a common phenomenon plaguing many individuals. The biological mechanisms underlying weight regain are incompletely understood and are likely multifactorial. In this paper, we examined the metabolic implications of curcumin, a compound known for its anti-inflammatory properties and inhibitory action on the enzyme 11β-HSD1, in a rodent model of adiposity rebound after the cessation of diet and exercise.

Abstract

Weight regain, adipose tissue growth, and insulin resistance can occur within days after the cessation of regular dieting and exercise. This phenomenon has been attributed, in part, to the actions of stress hormones as well as local and systemic inflammation. We investigated the effect of curcumin, a naturally occurring polyphenol known for its anti-inflammatory properties and inhibitory action on 11β-HSD1 activity, on preserving metabolic health and limiting adipose tissue growth following the cessation of daily exercise and caloric restriction (CR). Sprague-Dawley rats (6–7 wk old) underwent a “training” protocol of 24-h voluntary running wheel access and CR (15–20 g/day; ~50–65% of ad libitum intake) for 3 wk (“All Trained”) or were sedentary and fed ad libitum (“Sed”). After 3 wk, All Trained were randomly divided into one group which was terminated immediately (“Trained”), and two detrained groups which had their wheels locked and were reintroduced to ad libitum feeding for 1 wk. The wheel locked groups received either a daily gavage of a placebo (“Detrained + Placebo”) or curcumin (200 mg/kg) (“Detrained + Curcumin”). Cessation of daily CR and exercise caused an increase in body mass, as well as a 9- to 14-fold increase in epididymal, perirenal, and inguinal adipose tissue mass, all of which were attenuated by curcumin (P < 0.05). Insulin area under the curve (AUC) during an oral glucose tolerance test, HOMA-IR, and C-reactive protein (CRP) were elevated 6-, 9-, and 2-fold, respectively, in the Detrained + Placebo group vs. the Trained group (all P < 0.05). Curcumin reduced insulin AUC, HOMA-IR, and CRP vs. the placebo group (all P < 0.05). Our results indicate that curcumin has a protective effect against weight regain and impaired metabolic control following a successful period of weight loss through diet and exercise, perhaps via inhibition of glucocorticoid action and inflammation.

NEW & NOTEWORTHY Weight regain after dieting and exercise is a common phenomenon plaguing many individuals. The biological mechanisms underlying weight regain are incompletely understood and are likely multifactorial. In this paper, we examined the metabolic implications of curcumin, a compound known for its anti-inflammatory properties and inhibitory action on the enzyme 11β-HSD1, in a rodent model of adiposity rebound after the cessation of diet and exercise.

physical activity and caloric restriction (CR) are cornerstone lifestyle modifications to induce weight loss and combat chronic disease. However, many individuals often experience difficulty in sustaining successful weight loss (54). The biological mechanisms underlying weight regain remain incompletely understood but have been attributed to altered metabolic characteristics in the skeletal muscle, liver, adipose tissue, brain, gut, and neuroendocrine system. Furthermore, these biological changes can occur rapidly once higher levels of daily physical activity and CR are discontinued. For example, reducing daily step count in humans (23, 32, 36) and cessation of wheel running in rodents (24, 25, 28) induces skeletal muscle insulin resistance, glucose intolerance, and a substantial increase in adipose mass within days. Importantly, if these deleterious effects could be counteracted with further modifications to lifestyle or with pharmacological strategies, then the biological drive for weight regain, even after cessation of daily physical activity, may be attenuated.

A growing field of research indicates that some easily accessible dietary compounds, such as the turmeric compound curcumin, have therapeutic value in preventing or treating obesity and metabolic disorders (1, 2). Curcumin is a naturally occurring polyphenolic compound that has potent anti-inflammatory (20), antioxidative (35), and anticarcinogenic (49) effects. Curcumin also plays a role in inhibiting lipid synthesis (56), adipogenesis (12), and it acts as a selective inhibitor of 11β-HSD1 activity in humans and rats (19). Interestingly, previous studies have identified a link between low-grade inflammation, insulin resistance, and elevated adipose tissue levels of 11β-HSD1 expression (16, 21, 48), which is an intracellular enzyme that converts glucocorticoids (GCs) from their inactive to active form. In humans and in rodents, elevations in 11β-HSD1 and GCs are associated with central obesity and insulin resistance (13, 17, 30). We previously observed that voluntary exercise plus food restriction in rodents increases 11β-HSD1 expression in adipose tissue (47) which we speculate may set the stage for adiposity “rebound” when the exercise training stops. Moreover, mifepristone, a GC receptor antagonist, prevents the development of insulin resistance and attenuates weight regain if it is introduced in the posttraining period (47). These studies suggest that changes in stress hormone action and inflammation drive some of the metabolic impairments associated with the cessation of exercise and CR. Therefore, targeting these pathways may have a therapeutic benefit toward limiting weight regain and impaired metabolic control following cessation of training.

In the present study, we have used a modified rodent “wheel lock” model, as first described by Rhodes et al. (42) and further developed by Booth and colleagues (24–26, 28), to examine the effect of curcumin intake on metabolism following the cessation of exercise training and CR. Our rationale for using the combination of training and caloric restriction was twofold. First, exercise and reduced caloric intake are well known for their individual and additive ability to improve glucose tolerance, insulin sensitivity, blood lipid profiles, as well as to induce a reduction in body weight and body fat (8, 41, 46, 52). Based on this, and our previous work using the same experimental model (47), we know that this dual approach will result in marked improvements in fitness following the training period (lean body weight, low body fat mass, highly insulin sensitive and glucose tolerant). Second, cessation of regular exercise (23, 36) and cessation of reduced caloric intake (9, 10, 34) have been shown to induce insulin resistance, glucose intolerance, dyslipidemia, as well as body fat and body mass regain. Thus an advantage of this dual approach is that it allows for the study of a more considerable transition in energy balance than cessation of exercise or CR alone. This robust stimulus highlights the extensive and concerning alterations in physical and metabolic fitness that can occur when transitioning from being regularly active with restricted caloric intake to becoming sedentary and eating in excess.

Using this training-detraining model, the overall purpose of this study was to further characterize the early mechanisms that lead to initial weight regain, insulin resistance, and adipose tissue growth. Based on the variety of curcumin’s biological properties which culminate in reduced inflammation, inhibition of adipose tissue growth signals, and reduced active GC production, we hypothesized that curcumin administration during the detraining period would attenuate weight regain and the metabolic dysregulation associated with detraining (i.e., insulin resistance, glucose intolerance).

MATERIALS AND METHODS

Ethics statement.

This study was approved by the York University Animal Care Committee in accordance with the Canadian Council for Animal Care guidelines.

Rodent treatment and experimental design.

The voluntary wheel running model used in this study involved providing 24 h access to running wheels plus daily CR for 3 wk followed by locking the wheels and the reintroduction to ad libitum feeding with or without daily curcumin gavage. For this, 35 male Sprague-Dawley rats (Charles River Laboratories, 50–75 g upon arrival) were individually housed in a humidity- and temperature-controlled environment (50% relative humidity, 22–23°C) with a 12:12-h light-dark cycle (0800–2000). An overview of the experimental design is depicted in Fig. 1. Briefly, rats were randomly divided into two groups, a sedentary group fed ad libitum (Sed) (n = 9) and a voluntary running group (All Trained, n = 26) fed a CR diet of 15–20 g/day (~50–65% of ad libitum food intake) of standard rodent chow (Purina Labdiet, no. 5012, St. Louis, MO). For practical purposes, pellets were weighed in a weigh boat until the gross mass was 15–20 g/day for each calorically restricted rat. After 3 wk, the All Trained group was further subdivided into the following groups: “Trained”, “Detrained + Placebo”, and “Detrained + Curcumin” (n = 8–10 per group, see Fig. 1). The Trained group was euthanized at the end of the 3rd wk and used as a pre-wheel lock comparison. The detrained groups had their running wheels locked, were reintroduced to ad libitum feeding, and were given a daily gavage of either a placebo (Detrained + Placebo) or curcumin (200 mg/kg) (Detrained + Curcumin) for 1 wk. All groups had an oral glucose tolerance test (OGTT) administered on the day of harvest.

Fig. 1.

Experimental timeline. Sprague-Dawley rats were initially divided into 2 groups: Sed and All Trained. The All Trained group had access to voluntary running wheels 24 h/day for 3 wk and was placed on a CR diet of 15–20 g/day (~50–65% of ad libitum intake) of standard rodent chow. The Sed group did not have wheel access and were fed ad libitum throughout the study. After 3 wk, the All Trained group was subdivided into the following groups: Trained, Detrained + Placebo, and Detrained + Curcumin. On day 21, the placebo and curcumin groups had their wheels locked, were reintroduced to ad libitum feeding, and were given a daily oral gavage of either curcumin or a placebo. The Trained group was harvested on day 21 and used as pre-wheel lock comparisons. The placebo and curcumin groups were harvested on day 28, 1 wk following cessation of daily exercise and CR. The Sed group was also harvested on day 28. All groups had an oral glucose tolerance test (OGTT) administered on harvest days.

Curcumin administration.

Curcumin (no. C1386, Sigma, Oakville, ON, Canada) was dissolved in dimethyl sulfoxide (DMSO, 100% wt/vol) at a concentration of 200 mg/ml, and an oral gavage was administered at a 1 ml/kg dose for a final dose of 200 mg/kg daily. The placebo group received only DMSO at a 1 ml/kg dose. This dose was selected based on a study in which this dose attenuated weight gain and improved blood lipid profile in high-fat-fed obese rats (19). Importantly, several studies have reported no treatment-related toxicity with curcumin feeding in both humans (27) and rodents (40), even at very high doses (3,000–12,000 mg/kg). Accordingly, we had a high level of confidence that the dose in this study would be well-tolerated. On the other hand, this dose would equal 14 g daily for a 70 kg adult and may not be a feasible long-term supplementation strategy. Of note, curcumin is known to have poor bioavailability and recent strategies to overcome this include the use of nanoparticles, liposomes, phospholipid complexes, and micelles (39) which may reduce the dose of curcumin necessary to observe clinical effects.

Oral glucose tolerance test.

An oral glucose tolerance test (OGTT) was administered (1.5 g/kg body mass, 50% dextrose solution) to overnight-fasted animals via oral gavage, as previously described (47), at the end of the 3rd wk for the trained group or at the end of the 4th wk for the Sed group and for both detrained groups (see Fig. 1 for a timeline). Blood glucose concentrations were measured via a handheld glucometer (Bayer, Contour, NY). Plasma was also isolated from saphenous blood collection throughout the OGTT for subsequent analysis of insulin concentrations (see below). Glucose area under the curve (AUC) was calculated relative to the fasting glucose levels of each individual rat. All rats were euthanized immediately following the completion of the OGTT.

Plasma analysis.

Approximately 125 µl of whole blood was collected from each animal via saphenous vein puncture to measure plasma insulin (no. 90060, Crystal Chem, Downer’s Grove, IL) and corticosterone (no. 07120102, MP Biomedicals, Solon, OH) levels. For this, a small area of hair on the lateral upper hindlimb was shaved with an electric razor and wiped with a thin layer of petroleum jelly. Once the saphenous vein was located, a sterile 25-gauge needle was used to puncture the vein, and whole blood was collected in lithium-heparin coated microvette capillary tubes (no. 16.443.100, Sarstedt, Canada) and centrifuged at 14,500 g (Eppendorf Mini-Spin Plus, Brinkman Instruments) for 5 min, transferred into polyethylene tubes, and stored at −80°C until further analysis. Nonesterified fatty acids (NEFAs) (nos. 999–34691, 995–34791, 991–34891, 993–35191, Wako Diagnostics, Richmond, VA) and CRP (no. RAB0097, Sigma-Aldrich) were measured from blood collected during decapitation following the OGTT.

Measures of insulin resistance and β-cell function.

Homeostasis model assessment (HOMA) has been described in detail previously (31). Briefly, insulin resistance (HOMA-IR) was calculated as follows: [fasted glucose (mM) × fasted insulin (mU/l)/22.5]. Homeostasis model assessment for β-cells (HOMA-β) was calculated as follows: [20 × fasted insulin (mU/l]/[fasted glucose (mM) − 3.5].

Western blotting.

Tissues were homogenized in lysis buffer (135 mM NaCl, 1 mM MgCl₂, 2.7 mM KCl, 20 mM Tris-base, 1% Triton, 10% glycerol) containing a protease inhibitor cocktail (no. P8340, Sigma-Aldrich) and phosphatase inhibitor cocktail (no. P0044, Sigma-Aldrich) and were quantified for protein via the Bradford method. Thirty to fifty micrograms of protein lysate from the plantaris was separated on 10% (11β-HSD1, GLUT4, total Akt, pSer⁴⁷³ and pTh³⁰⁸) or 6% (AS160) SDS-page gel and transferred to a PVDF membrane (no. 1620177, Bio-Rad). Membranes were blocked in 5% powdered milk and Tris-buffered saline with 0.1% Tween-20 (TBST) at room temperature for 1 h. Membranes were then incubated overnight in their respective primary antibody at 4°C at a concentration of 1:1,000 (11β-HDS1, no. 10004303, Cayman Chemical; GLUT4, no. ab65267, Abcam, Toronto, Canada; Akt, no. ab8805, Abcam; pSer⁴⁷³, no. ab28821, Abcam; pTh³⁰⁸, no. ab5626, Abcam; AS160, no. C69A7, Cell Signaling). The following morning, the membranes were washed with TBST and incubated with anti-mouse (1:10,000, no. ab6789, Abcam) or anti-rabbit (1:10,000, no. ab6721, Abcam) secondary antibodies for 1 h at room temperature. Membranes were then washed and imaged. Images were detected on a Kodak In vivo FX Pro imager, and Carestream Image MI SE (version S.0.2.3.0, Rochester, NY) was used to quantify protein content. Alpha-tubulin (1:40,000, no. ab7291, Abcam) and GAPDH (1:10,000, no. ab9484, Abcam) were used as loading controls.

Statistical analysis.

All data was analyzed using a repeated-measures or one-way ANOVA with a criterion of P < 0.05. Significant differences for repeated-measures ANOVAs were evaluated with a Bonferroni post hoc test. One-way ANOVAs were evaluated using a Tukey HSD post hoc test (GraphPad Prism version 5). In each bar graph, a significant difference is present if bars do not share a letter. All data are means ± SE.

RESULTS

Curcumin reduces body weight regain following exercise training and CR.

The Sed group had a significantly higher body mass compared with the trained rats from day 9 onward. In addition, sedentary rats had a higher body mass than both placebo and curcumin groups post wheel lock (both P < 0.05, Fig. 2A). Also during the post-wheel lock period, the curcumin group had a significantly lower body mass on days 27 and 28 vs. the placebo group (P < 0.05, Fig. 2A). In response to 3 wk of daily running and CR, the final body mass of the Trained group (198 ± 4.2 g) was significantly lower than all other groups (P < 0.05, Fig. 2B), perhaps in part due to their slightly younger age at the time of euthanization (i.e., study end point was 1 wk earlier). Ad libitum food intake and wheel lock for 1 wk, following the 3 wk of daily exercise and CR, caused a significant increase in body mass in both the placebo (318 ± 4.8 g) and curcumin (288 ± 4.4 g) groups vs. the Trained group, although the curcumin group gained back significantly less weight than the placebo group (P < 0.05, Fig. 2B). However, the curcumin group ate significantly less food (in absolute g and kcal) during the detraining period vs. the placebo group (P < 0.05, Fig. 2, D and E). Thus we cannot conclude that curcumin had a direct effect on preventing weight regain. Rather, the lower average body mass in the curcumin group may have been due to an indirect effect of suppressing appetite. As expected, the Sed ad libitum group had significantly greater food intake compared with the rats undergoing exercise training and CR (P < 0.05, Fig. 2C). The amount of wheel running slowly increased over time up to a steady state of ~10–12 km/night in the final 10 days of having access to voluntary running wheels (Fig. 2F).

Fig. 2.

Body mass, food intake, and running data. A: body mass over time in all groups. B: final body mass on harvest date. C: food intake during the CR (1st to 3rd wk) and ad libitum (4th wk) periods. The Trained group was harvested at the end of the 3rd wk. The Sed, placebo, and curcumin groups were harvested at the end of the 4th wk. D and E: total food intake in g and kcal, respectively, during the detraining period (final week). F: daily running distance throughout the protocol in all 3 running groups (“All Trained”). *Sed vs. All Trained (P < 0.05). #Sed vs. “Detrained + Placebo” and “Detrained + Curcumin” (P < 0.05). ψ “Detrained + Placebo” vs “Detrained + Curcumin” (P < 0.05). A significant difference is present when bars do not share a letter (P < 0.05). All data are means ± SE; n = 8–10. A and C were analyzed using a repeated-measures ANOVA with a Bonferroni post hoc test. B was analyzed using a one-way ANOVA and a Tukey post hoc test. D and E were analyzed using an unpaired t-test. Note: In C, the dashed lines indicate the range of daily food intake for the All Trained group (food pellets were weighed daily at ~15–20 g).

Curcumin reduces adipose tissue and liver growth following exercise training and CR.

Compared with the Sed group, the Trained group which underwent daily wheel running and CR for 3 wk had dramatically reduced absolute and relative adipose tissue mass in all adipose tissue depots (P < 0.05, Fig. 3, A–F). Following 1 wk of ad libitum food intake and wheel lock, the placebo group had a significant increase in absolute and relative adipose tissue mass in the epididymal, perirenal, and inguinal depots compared with the Trained group (P < 0.05, Fig. 3, A–F). The curcumin-treated group also had a significant increase in adipose tissue mass relative to the Trained group; however, they gained back significantly less adipose tissue vs. the placebo group (P < 0.05, Fig. 3, A–F). The placebo group had a twofold increase in absolute liver mass vs. the Trained group, and this growth was significantly reduced in the curcumin-treated rats (Table 1). A summary of relative and absolute tissue weights for skeletal muscle and the liver is shown in Table 1.

Fig. 3.

Adipose tissue mass. Epididymal (A, B), perirenal (C, D), and inguinal fat pads (E, F) were harvested at the end of the 3rd (Trained) or 4th wk (Sed, Detrained + Placebo, Detrained + Curcumin). A significant difference is present if bars do not share a letter (P < 0.05). All data are means ± SE, n = 8–10, and were analyzed using a one-way ANOVA and a Tukey post hoc test.

Table 1.

Tissue masses

	Sed	Trained	Detrained + Placebo	Detrained + Curcumin
Tibialis anterior	1.77 ± 0.05a	1.92 ± 0.06a	1.76 ± 0.03a	1.86 ± 0.04a
	0.64 ± 0.02a	0.39 ± 0.02b	0.54 ± 0.01c	0.54 ± 0.01c
Plantaris	0.95 ± 0.03a	0.87 ± 0.03a	0.86 ± 0.03a	0.95 ± 0.02a
	0.34 ± 0.01a	0.17 ± 0.01b	0.27 ± 0.01c	0.27 ± 0.01c
Soleus	0.40 ± 0.01a	0.54 ± 0.03b	0.41 ± 0.01ac	0.46 ± 0.01c
	0.14 ± 0.01a	0.10 ± 0.01b	0.13 ± 0.01a	0.13 ± 0.01a
Gastrocnemius	4.79 ± 0.23a	4.89 ± 0.10a	4.54 ± 0.08a	4.87 ± 0.10a
	1.71 ± 0.04a	0.98 ± 0.04b	1.44 ± 0.02c	1.40 ± 0.03c
Liver	33.9 ± 1.50ab	30.0 ± 1.39b	38.4 ± 0.83c	34.8 ± 0.49ac
	12.2 ± 0.58a	6.03 ± 0.27b	12.23 ± 0.29a	10.03 ± 0.24c

All data are means ± SE; n = 8–10. Relative (g/kg body mass) and absolute (g) tissue masses, respectively, for skeletal muscle and liver. A significant difference is present when groups do not share a letter (P < 0.05). All data were analyzed by a one-way ANOVA and Tukey post hoc test.

Curcumin improves glucose tolerance and insulin sensitivity following exercise training and CR.

As expected, daily wheel running and CR for 3 wk in the Trained group significantly improved oral glucose tolerance and insulin sensitivity vs. sedentary rats fed ad libitum (Fig. 4). In the placebo group, 1 wk of wheel lock and unrestricted food intake deteriorated both glucose tolerance and insulin sensitivity to values similar to, or higher than, what was observed in the sedentary ad libitum group (Fig. 4). More specifically, the placebo group had a two- and sixfold increase in glucose and insulin AUC, respectively, during an OGTT, vs. the Trained group (P < 0.05, Fig. 4, B and E). Following wheel lock and CR, the placebo group also had a ninefold and threefold increase in HOMA-IR and HOMA-β, respectively, vs. the Trained group (P < 0.05, Fig. 4, C and F). Compared with the placebo group, the curcumin group maintained blood glucose and plasma insulin AUC to levels observed in the Trained group (Fig. 4). Moreover, HOMA-IR in the curcumin group, although still higher than in the Trained group, was lower than that observed in the Placebo group, suggesting a role of curcumin in preserving improved glucose tolerance and insulin sensitivity.

Fig. 4.

Glucose tolerance and insulin sensitivity. All groups had an oral glucose tolerance test (OGTT) administered (1.5 g/kg body mass) at either the end of the 3rd (Trained) or 4th wk (Sed, Detrained + Placebo, and Detrained + Curcumin). A: glucose concentrations (mM) in the fasted state (t = 0) and at t = 5, 15, 30, 60, and 120 min post-oral glucose load. B: area under the curve (AUC) for glucose during the OGTT was calculated relative to the t = 0 glucose value for each individual rat. C: HOMA-IR was calculated as follows: [fasting insulin (mU/l) × fasting glucose (mM)]/22.5]. D: insulin concentrations (ng/ml) in the fasted state (t = 0) and at t = 15, 30, 60, and 120 min. E: AUC for insulin during the OGTT was calculated relative to the t = 0 insulin value for each individual rat. F: relative HOMA-β was calculated as follows: [20 × fasting insulin (mU/l)]/[fasting glucose (mM) – 3.5]. A significant difference is present if bars do not share a letter (P < 0.05). All data are means ± SE; n = 8–10. All data were analyzed using a one-way ANOVA and Tukey post hoc test.

Curcumin lowers circulating corticosterone levels and reduces inflammation following exercise training and CR.

Circulating corticosterone concentrations were measured during the pre-wheel lock (days 10 and 20) and post-wheel lock (day 25) periods at 0800 (i.e., “basal”). During the pre-wheel lock period, there were significantly greater circulating levels of corticosterone in the All Trained group (daily exercise + CR) vs. the Sed group (sedentary + ad libitum food intake) (P < 0.05, Fig. 5A). In the post-wheel lock period, the placebo group retained this tendency for hypercorticosteronemia, whereas the curcumin group experienced a significant reduction in corticosterone levels compared with either the placebo group or the All Trained group (both P < 0.05, Fig. 5A). Daily wheel running and CR caused a significant reduction in circulating CRP and NEFAs in the Trained group vs. the Sed group (P < 0.05, Fig. 5, B and C). Cessation of wheel running and CR in the placebo group resulted in a twofold increase in CRP and NEFAs vs. the Trained group (both P < 0.05). Curcumin supplementation resulted in a significant attenuation in the rise in CRP compared with the placebo group (P < 0.05) but circulating NEFA levels were not significantly different vs. the placebo group (Fig. 5, B and C).

Fig. 5.

Plasma analytes. A: AM (0800) corticosterone concentrations were collected on days 10, 20, and 25 from the saphenous vein. Days 10 and 20 represent the pre-wheel lock values, and day 25 represents the post-wheel lock value. B: C-reactive protein. C: NEFAs. A significant difference is present if bars do not share a letter (P < 0.05). All data are means ± SE; n = 6–9 for Sed, Detrained + Placebo, and Detrained + Curcumin, and n = 18 for All Trained. Data were analyzed using a one-way ANOVA and Tukey post hoc test.

Assessment of insulin signaling markers and 11β-HSD1 content in skeletal muscle.

To examine some of the potential mediators of the changes in basal insulin signaling and glucose tolerance with wheel running and CR, with or without curcumin in administration, select markers of insulin signaling in skeletal muscle (plantaris) were measured. No significant differences were observed in 11β-HSD1, AktT308, AktS473, Total Akt, AS160, or GLUT4, between the placebo and curcumin post-WL groups in the plantaris (Fig. 6, A–F). There was a tendency in the placebo group for elevated 11β-HSD1 content vs. the curcumin group, but this difference failed to reach statistical significance (P = 0.11). This elevated 11β-HSD1 content in the placebo group did reach significance vs. the Sed group (P < 0.05, Fig. 6A).

Fig. 6.

11β-HSD1 and insulin signaling protein content in skeletal muscle. Relative 11β-HSD1 (A), Akt^T308 (B), Akt^S473 (C), total Akt (D), AS160 (E), and GLUT4 (F) protein content in plantaris skeletal muscle. All data are means ± SE; n = 5–8. A significant difference is present if bars do not share a letter (P < 0.05). All data were analyzed using a one-way ANOVA and Tukey post hoc test.

DISCUSSION

Weight maintenance after weight loss is a significant challenge for most individuals. Evidence has been growing in support of a biological drive toward weight regain that counters weight loss efforts. Using a previously published rodent “wheel lock” model (47), we observed in the current study that 3 wk of CR and daily wheel running, followed by 1 wk of ad libitum food intake and physical inactivity induced by wheel lock, causes a rapid increase in epididymal, perirenal, and inguinal (subcutaneous) adipose tissue mass as well as liver mass. These previously physically active rats rapidly developed insulin resistance, glucose intolerance, had elevated circulating CRP, and regained much of the adipose tissue mass that was lost due to daily running and CR. Our study also demonstrates, for the first time, that curcumin supplementation after exercise training and CR is ceased leads to a significant improvement in insulin sensitivity, glucose tolerance, body fat content, and lowers circulating corticosterone and CRP levels.

These findings, although novel, are in line with previous data supporting the potent anti-inflammatory actions of curcumin (20) and its selective inhibition of 11β-HSD1 activity (19). Taken together, and as reviewed recently by Goel et al. (14), these data suggest that curcumin supplementation may be of therapeutic benefit to limit weight regain in individuals who have achieved weight loss from dieting and exercise.

We recently found evidence of stress-induced mechanisms that promote adipose tissue regain and metabolic dysfunction after dieting and exercise is ceased (47). A major regulator of GC exposure within the body is the enzyme 11β-HSD1 which converts GCs from their inactive to active form intracellularly. We and others have found that exercise causes an increase in 11β-HSD1 systemically (4) and within white adipose tissue (6, 7), and we hypothesized that this could be a mechanism that contributes to adipose tissue regain and insulin resistance following cessation of daily exercise and CR. Whorwood et al. (53) found that there is a positive association between the expression of 11β-HSD1, the GC receptor, and the extent of insulin resistance in human skeletal muscle myoblasts. In addition, previous studies have observed that 11β-HSD1-deficient mice are resistant to dietary-induced insulin resistance (22, 33). Thus 11β-HSD1 appears to be positively linked with body adiposity and may play a key role in regulating metabolic health. However, the mechanism by which 11β-HSD1 and glucocorticoids affect glucose metabolism and insulin sensitivity in skeletal muscle is not completely understood.

GCs are known to affect muscle insulin signaling at several levels along the insulin-signaling cascade. For example, rats injected with dexamethasone for 11 days showed decreased expression of Akt/PKB and insulin-stimulated Ser(473) and Thr(308) phosphorylation (5, 44). GCs in excess are also known to induce skeletal muscle atrophy, particularly in glycolytic fibers (11), and given the inverse correlation between muscle mass and whole body insulin resistance (45) we decided to evaluate a largely glycolytic muscle like the plantaris to evaluate changes in insulin signaling. To determine if tissue-specific regulation of 11β-HSD1 was associated with impaired skeletal muscle insulin action we evaluated total 11β-HSD1 protein content as well as key insulin signaling proteins in the plantaris muscle. We hypothesized that a curcumin-induced reduction of 11β-HSD1 activity or content, and thus less exposure to active corticosterone, might prevent impairments in insulin signaling and sensitivity. To our surprise, we found no difference in pAkt Thr(308), Ser(473), total Akt/PKB, and GLUT4 between all four treatment groups. There was a trend for AS160 to be elevated in the trained, placebo, and curcumin groups, although there was no significant difference between any of the groups. Importantly, we recognize that the lack of differences seen between our groups may be attributed to the fact that these are markers of basal, non-insulin-stimulated insulin signaling and therefore any conclusions drawn from these data must be made in light of this limitation. Future studies should consider measuring insulin signaling markers directly in skeletal muscle in an insulin-stimulated state and perhaps measuring whole body substrate metabolism using in vivo calorimetry.

We observed a significant increase in plantaris 11β-HSD1 protein content in the placebo group vs. the Sed group. However, there was no significant difference in 11β-HSD1 content observed between the placebo and curcumin groups despite a trend for a reduction with curcumin treatment. Although curcumin has been found to inhibit 11β-HSD1 activity (19), our results indicate that this compound does not alter the adipose tissue total protein content of 11β-HSD1. Interestingly, the curcumin group had a reduction in circulating corticosterone during the post-wheel lock period, and this may be related to a reduction in whole body activity of 11β-HSD1. Thus, relative to the curcumin treated rats, the placebo group was hypercorticosteronemic, which is a physiological stimulus that is well known to impair insulin signaling in a number of tissues (55). Why this elevation in GCs in the placebo-treated group during the detraining period did not result in a reduction in some of the molecular markers of insulin signaling in skeletal muscle currently remains unclear.

Systemic and local low-grade inflammation and release of pro-inflammatory cytokines is associated with obesity and numerous inflammatory diseases, and this is often implicated in the development and progression of insulin resistance (15, 29). The proinflammatory state of obesity includes elevated circulating CRP, elevated macrophage infiltration of adipose tissue, and increased adipose tissue content of nuclear factor κB, c-Jun NH₂-terminal kinase, monocyte chemoattractant protein 1, TNFα, inducible nitric oxide synthase, and plasminogen activator inhibitor-1 (18, 37, 51). In line with this, it was recently reported that there is an upregulation in gene transcripts related to macrophage infiltration, immunity, proinflammation, and extracellular matrix remodeling following cessation of daily wheel running (43). This suggests that inflammation may be playing an important role in the rapid metabolic impairments that are observed after suddenly reducing daily physical activity. We thus sought to explore if curcumin, by acting as a potent anti-inflammatory agent, in addition to acting as an inhibitor of 11β-HSD1 activity, could potentially prevent insulin resistance observed in the wheel lock model. Curcumin treatment reduced both glucose and insulin AUC during an OGTT and attenuated the increase in HOMA-IR and HOMA-β vs. rats that received a placebo. We observed a significant increase in CRP in the placebo group vs. the Trained group, indicative of a more proinflammatory environment that is induced by suddenly discontinuing daily exercise and “dieting.” Daily gavage of curcumin for 1 wk significantly reduced circulating CRP vs. the placebo group. In support of our findings, curcumin has been shown to ameliorate inflammation in white adipose tissue and the liver, improve glycemic control and insulin sensitivity, and reduce body weight gain in mouse models of Type 2 diabetes (50). Further research is required to determine how curcumin may be affecting the proinflammatory milieu within various tissues and how this is linked with the observations from this study and others of reducing body fat, improving insulin sensitivity, and inhibiting the activity of 11β-HSD1.

It is interesting to note that the Placebo group, despite all the exercise that had been performed over the span of 3 wk, had a similar level of adiposity as the Sed AL group once the wheels had been locked for 1 wk. Despite the similar fat mass, and lower body mass, the Placebo group had worse insulin sensitivity than the Sed AL group. This highlights the complex relationship between body adiposity, metabolic health, and physical activity levels and suggests that metabolic protection of regular exercise is soon lost because of rapid adiposity rebound (3). In line with this, we observed that curcumin had a specific capacity to limit adipose tissue growth, perhaps by limiting the dramatic rise in food intake after CR stopped.

In summary, curcumin intake, following a period of successful weight loss through daily exercise and CR, is effective at attenuating adipose tissue growth and the development of insulin resistance caused by 1 wk of “detraining” (physical inactivity and ad libitum food intake). The beneficial effects of curcumin observed in this study were linked with reduced systemic inflammation and lower circulating corticosterone levels relative to placebo-treated rats. However, we observed no significant difference in 11β-HSD1 protein content in skeletal muscle in response to curcumin treatment during the detraining period. Based on our findings, and other emerging reports of the physiological benefits of curcumin in rodents and humans (38), further investigation is warranted to determine if this is a compound that can improve weight management in humans following a period of weight loss.

GRANTS

This work was funded by the Natural Science and Engineering Research Council of Canada Discovery Grant (Grant No. 261306) to M. C. Riddell. T. Teich is the recipient of the York University Muscle Health Research Centre Graduate Student Fellowship. D. P. Porras is a recipient of the Ontario Graduate Scholarship. E. C. Dunford is a recipient of the Natural Science and Engineering Research Council of Canada Doctoral Scholarship.

DISCLOSURES

No conflicts of interest, financial or otherwise, are declared by the authors.

AUTHOR CONTRIBUTIONS

T.T., J.A.P., D.P.P., and M.C.R. conceived and designed research; T.T., J.A.P., D.P.P., and E.C.D. performed experiments; T.T., J.A.P., D.P.P., and E.C.D. analyzed data; T.T., J.A.P., D.P.P., E.C.D., and M.C.R. interpreted results of experiments; T.T., J.A.P., D.P.P., E.C.D., and M.C.R. prepared figures; T.T., J.A.P., D.P.P., and E.C.D. drafted manuscript; T.T., J.A.P., E.C.D., and M.C.R. edited and revised manuscript; T.T., J.A.P., D.P.P., E.C.D., and M.C.R. approved final version of manuscript.