Nicotinamide overload may play a role in the development of type 2 diabetes

Abstract

Aim: To investigate whether nicotinamide overload plays a role in type 2 diabetes.

Methods: Nicotinamide metabolic patterns of 14 diabetic and 14 non-diabetic subjects were compared using HPLC. Cumulative effects of nicotinamide and N(1)-methylnicotinamide on glucose metabolism, plasma H(2)O(2) levels and tissue nicotinamide adenine dinucleotide (NAD) contents of adult Sprague-Dawley rats were observed. The role of human sweat glands and rat skin in nicotinamide metabolism was investigated using sauna and burn injury, respectively.

Results: Diabetic subjects had significantly higher plasma N(1)-methylnicotinamide levels 5 h after a 100-mg nicotinamide load than the non-diabetic subjects (0.89 +/- 0.13 micromol/L vs 0.6 +/- 0.13 micromol/L, P < 0.001). Cumulative doses of nicotinamide (2 g/kg) significantly increased rat plasma N(1)-methylnicotinamide concentrations associated with severe insulin resistance, which was mimicked by N(1)-methylnicotinamide. Moreover, cumulative exposure to N(1)-methylnicotinamide (2 g/kg) markedly reduced rat muscle and liver NAD contents and erythrocyte NAD/NADH ratio, and increased plasma H(2)O(2) levels. Decrease in NAD/NADH ratio and increase in H(2)O(2) generation were also observed in human erythrocytes after exposure to N(1)-methylnicotinamide in vitro. Sweating eliminated excessive nicotinamide (5.3-fold increase in sweat nicotinamide concentration 1 h after a 100-mg nicotinamide load). Skin damage or aldehyde oxidase inhibition with tamoxifen or olanzapine, both being notorious for impairing glucose tolerance, delayed N(1)-methylnicotinamide clearance.

Conclusion: These findings suggest that nicotinamide overload, which induced an increase in plasma N(1)-methylnicotinamide, associated with oxidative stress and insulin resistance, plays a role in type 2 diabetes.

Figure 1

Urinary excretion patterns of 2Py and N¹-methylnicotinamide in diabetic and non-diabetic subjects. A and B: Representative HPLC chromatograms of urinary excretions of 2Py (indicated by arrow) and N¹-methylnicotinamide (NMN) of a non-diabetic (Aa and Ba) and a diabetic (Ab and Bb) subject, before and after 100 mg nicotinamide loading at 7:00 am Urine samples taken at given time were normalized to equal volumes before HPLC analysis; C and D: Summaries of the results from the measurements shown in A and B, respectively. Bar graphs indicate mean ± SD.

Figure 2

Plasma N¹-methylnicotinamide levels of diabetic and non-diabetic subjects after nicotinamide loading. A: Representative HPLC chromatograms of plasma N¹-methylnicotinamide levels from a non-diabetic (a, b) and a diabetic (c, d) subject before (a, c) and 5 h after (b, d) 100 mg nicotinamide loading. 1: N¹-methylnicotinamide; 2: Internal standard: N¹-methylnicotinamide; B: Summary of the results from the measurements shown in A. Bar graph indicates mean ± SD.

Figure 3

Effects of nicotinamide and N¹-methylnicotinamide on glucose metabolism of rats. A: Changes in blood glucose, muscle glycogen, plasma insulin and plasma N¹-methylnicotinamide in rats treated with or without cumulative nicotinamide (0.5 or 2 g/kg) after glucose loading; B: Comparable effects of cumulative N¹-methylnicotinamide (0.5 or 2 g/kg). NM: Nicotinamide; NMN: N¹-methylnicotinamide. ^aP < 0.05 vs control, ^bP < 0.01 vs control, ^dP < 0.001 vs control. Bar graphs show mean ± SD.

Figure 4

Effects of N¹-methylnicotinamide on H₂O₂ generation and NAD levels. A, B: Cumulative effects of nicotinamide (NM, 0.5 or 2 g/kg) and N¹-methylnicotinamide (NMN, 0.5 or 2 g/kg) on rat plasma H₂O₂ levels; C: H₂O₂ concentrations in the supernatant of cultured human erythrocytes with or without 3 h exposure to different concentrations of NMN. For each concentration, n = 4; D, E: NAD (NAD⁺ and NADH) contents in muscle (D) and liver (E) of rats treated with saline (control) or a cumulative dose of 2 g/kg NMN; F: NAD and NADH contents and NAD/NADH ratio in the erythrocytes (RBCs) of rats with or without cumulative exposure to NMN; G: NAD and NADH contents, and NAD/NADH ratio in human RBCs with (n = 4) or without (control, n = 4) 4 h exposure to 10 μmol/L NMN in vitro. ^aP < 0.05 vs control, ^bP < 0.01 vs control. Bar graphs indicate mean ± SD.

Figure 5

Effects of aldehyde oxidase (AOX) inhibitors on plasma N¹-methylnicotinamide levels and glucose metabolism in rats. A and B: Plasma N¹-methylnicotinamide (NMN) levels 5 h after nicotinamide load (100 mg/kg, ip) in rats treated with or without AOX inhibitors tamoxifen (Tam, A) or olanzapine (Olan, B) (each group, n = 6). ^aP < 0.05 vs control, ^bP < 0.01 vs control; C: Liver AOX expression in rats with or without 7 wk tamoxifen treatment. The blot is representative of four independent experiments; D: Responses to a glucose tolerance test in rats after 9 wk treatment with tamoxifen with or without NMN (100 mg/kg per day) treatment in the last 2 wk. FBG: Fasting blood glucose; GTT (1 h): Blood glucose measured 1 h after glucose tolerance test. ^bP < 0.01 vs control. Bar graph indicates mean ± SD.

Figure 6

Role of skin in nicotinamide metabolism and insulin resistance. A: Representative HPLC chromatograms showing changes of sweat nicotinamide (NM) and N¹-methylnicotinamide (NMN) concentrations in a subject before and 1, 2 and 3 h after 100 mg nicotinamide loading. 1 and 2 in Aa are NM and internal standard N¹-ethylnicotinamide, respectively; 1 and 2 in Ab are NMN and internal standard N¹-ethylnicotinamide, respectively; B: Summary of the measurements shown in A. ^bP < 0.0001 vs control; C: Comparison of plasma NMN and insulin levels, muscle and liver glycogen contents, and blood glucose between sham-burn (n = 7) and burn (n = 11) rats after glucose load. FBG: Fasting blood glucose; GTT: Blood glucose 1 h after glucose injection. Bar graphs show mean ± SD. ^cP < 0.05, ^dP < 0.01 vs control.

Figure 7

Proposed model of the role of nicotinamide overload in the development of type 2 diabetes. Normally, if nicotinamide intake is slightly more than the body needs, excess nicotinamide will be detoxified rapidly and eliminated mainly via the N¹-methylnicotinamide to 2Py pathway, which involves liver and skin functions (left). Frequent excess nicotinamide intake, low N¹-methylnicotinamide detoxification, or sweat gland inactivity induces a substantial rise in plasma N¹-methylnicotinamide concentrations and residence time after each meal, and consequently induces oxidative stress and insulin resistance (right). The change trends are indicated by red arrows or line thickness.