The Effect of a Hydrogen Sulphide (H2S)-Releasing Non-Steroidal Anti-Inflammatory Drug (NSAID) on Sex-Differences in Gastric and Renal Damage in Goto-Kakizaki Diabetic Rats

MV Chan, RW Blackler, GW McKnight, JL Wallace. McMaster University, Hamilton, Ontario, Canada

Introduction

Non-steroidal anti-inflammatory drugs (NSAIDs) have been shown to induce gastrointestinal (GI) and renal damage, which are exacerbated in people with co-morbidities such as diabetes. Hydrogen sulphide (H2S) has been shown to counteract these adverse GI effects. In addition, there is a greater severity in diabetes in females than males. We investigated the GI and renal effects of naproxen and an H2S-releasing NSAID, 2-(6-methoxy-napthalen-2-yl)-propionic acid 4-thiocarbamoyl-phenyl ester (ATB-346), in a non-obese rat model of type II diabetes.

Methods

10-month old male (414.8±5.1 g, n=14) and female (236.0±2.7 g, n=14) Goto-Kakizaki (GK) rats (Charles River, Montreal, Canada) and age-matched male (697.0±49.4 g, n = 4) and female (414.8±5.1 g, n=4) Wistar rats were treated twice daily by oral gavage for 4.5 days with vehicle (dimethylsulfoxide/1% carboxymethylcellulose; 5:95), naproxen acid (10 mg/kg) or ATB-346 (14.5 mg/kg). Rats were euthanised by overdose of sodium pentobarbital (65 mg/kg, i.p.) 3 hours after the final dose. Stomach and small intestine (SI) were removed for blind scoring of haemorrhagic damage by summing the lengths and areas of lesions (mm2), respectively. As markers of renal dysfunction, blood was taken after NSAID dosing, allowed to clot, and serum obtained for blood urea nitrogen (BUN) analysis, while urine was obtained by cystocentesis for protein and creatinine analysis by IDEXX (North Grafton, MA, USA). Blood was also taken, diclofenac (1 mM) added and serum analysed for thromboxane (TX) B2 by ELISA. Comparisons were made by one-way ANOVA and data are expressed as mean±S.E.M.

Results

In male GK rats, naproxen (71.2±12.0 mm2, p<0.001) caused significantly more SI damage than those treated with vehicle or ATB-346, as well as Wistar controls. Similarly, in females, significant SI damage was observed with naproxen (33.0±10.4 mm2, p<0.05) but not in other groups. No gastric damage was seen in any group. TxB2 levels, a stable metabolite of TxA2, were significantly greater (p<0.05) in both male and female GK rats compared to Wistar controls (2400±0.6 and 2400±0.6, respectively. Treatment of GK rats with either naproxen or ATB-346 significantly (p<0.05) attenuated these levels. BUN was greater (p<0.05) in Wistar controls compared to GK rats of both sexes. Urine protein:creatinine ratios were lower in females than males but, after dosing with either naproxen or ATB-346, there was no difference when compared to vehicle in either sex. Conversely, bile acid levels were greater in females than males and lower in GK rats. Treatment with ATB-346 restored bile acid levels to those of control.

Conclusions

Naproxen caused SI, but not gastric, damage in diabetic rats of both sexes, though the damage was more severe in males. ATB-346 treatment did not induce SI damage, consistent with the protective effect of H2S released from ATB-346. Males had greater renal dysfunction than females. Interestingly, in females, bile acid flow was decreased in GK rats, but these levels were restored to those of control Wistar rats with ATB-346 treatment. Taken together, these data suggest that females fair better to naproxen-induced SI damage, possibly due to an enhancement in levels of bile.