Analagesic and Anti-Inflammatory Potentials of a Less Ulcerogenic Thiadiazinethione Derivative in Animal Models: Biochemical and Histochemical Correlates

Abstract

Purpose: Gastric ulcer induced by NSAIDs is the major medical concern and researchers are utilizing several approaches to combat this medical issue. In the current study, we investigated the efficacy of thiadiazinethione derivative (2,2'(2-thioxo-1,3,5-thiadiazinane-3,5-diyl) diacetic acid, as new less ulcerogenic compound.

Methods: 2,2'(2-thioxo-1,3,5-thiadiazinane-3,5-diyl) diacetic acid was evaluated using standard animal models including hot plate, writhing test and formalin induced nociceptive models. Anti-inflammatory activity was assessed via carrageenan-induced paw oedema model. Involvement of opioidergic nociceptive mechanism was confirmed via naloxone administration in hot plat assay. The gastro-ulcerogenic potential of test and standard compounds were evaluated via NSAID-induced pyloric ligation model followed by standard histopathological and biochemical analysis.

Results: In acetic acid-induced writhing test, our compound significantly reduced abdominal constrictions at the tested doses of 15 (p < 0.05), 30 (p < 0.01) and 45 mg kg^-1 (p < 0.001) as compared to control (p < 0.001). In hot plate test, after 30 min of administration, our test compound showed significant anti-nociceptive potential (p < 0.05 at 15 and 30 mg kg^-1 and p < 0.01 at 45 mg kg^-1) and tramadol (p ˂ 0.001) at 30 mg kg^-1 dose. After 60 min tramadol (30 kg^-1) and test sample (30, 45 mg kg^-1) exhibited significant anti-nociceptive activity p < 0.001. In Formalin-induced nociceptive response, a significant decline (p ˂ 0.001) was observed for aspirin and test compound during acute and chronic phases. Decline in the anti-nociceptive potential of tramadol and test sample via administration of naloxone indicate the involvement of opioidergic mechanism. Our compound exhibited significant anti-inflammatory activity in second phase of carrageenan induced paw oedema model. Histological and biochemical parameters exhibited less ulcerogenic potential as compared to aspirin.

Conclusion: Our findings suggests that our test compound has desirable anti-nociceptive and anti-inflammatory potentials with less propensity to cause gastric ulcer.

Keywords: NSAIDs; gastric ulcer; hot plate; inflammation; nociception.

Figure 1

Chemical structure of 2,2’(2-thioxo-1,3,5-thiadiazinane-3,5-diyl) diacetic acid.

Figure 2

Acetic acid induced writhing test results. Exhibits the aspirin and test compound anti-nociceptive effect in the acetic acid induced writhing test. Statistical analysis by post-hoc Dunnett’s test after one way ANOVA reveals that aspirin and test compound at all the three doses shows a significant anti-nociceptive activity. Each bar shows mean number of writhing ± SEM (n=6). * p ˂ 0.05, ** p ˂ 0.01, ***p ˂ 0.001 as compared with control saline group.

Figure 3

Results of Hot plate test: Shows the result of tramadol anti-nociceptive effect at 30 mg/kg dose and test compound anti-nociceptive effect at 15, 30 and 45 mg/kg doses in hot plate induced nociceptive test. Statistical analysis by post hoc Dunnet’s test after one way ANOVA shows that tramadol and test compound has significant anti-nociceptive activity. Each bar shows percentage protection mean value ± SEM (n = 6). * p ˂ 0.05, ** p ˂ 0.01, *** p ˂ 0.001 as compared with control saline group.

Figure 4

Results of formalin induced nociception study. Represents anti-nociceptive activity of aspirin and test compound at 15, 30 and 45 mg/kg body weight doses in formalin induced nociceptive model. Statistical analysis by post hoc Dunnet’s test after one way ANOVA shows that aspirin and test compound has significant anti-nociceptive activity. Each bar represents mean number of nociceptive response ± SEM (n = 6). ***p˂0.001 as compared to control saline group.

Figure 5

Naloxone antagonism results shows the effect of Naloxone (1 mg/kg) on the tramadol and test compound anti-nociceptive effect. Statistical analysis by post hoc Tukey’s test after one way ANOVA shows naloxone antagonism in the hot plate test. *** p ˂ 0.001, *p ˂ 0.05.

Figure 6

Results of carrageenan-induced paw oedema model. Shows the anti-inflammatory effect of aspirin 50 mg/kg and test compound at 15, 30 and 45 mg/kg doses in the carrageenan induced paw edema model. Statistical analysis of data by post hoc Dunnet’s test after one way ANOVA shows that aspirin at 50 mg/kg dose and test compound at all the three doses shows significant anti-inflammatory effect after 3, 4 and 5 hour of the study. Each bar represents mean value of paw volume in mL ± SEM (n=6). ***p˂0.001 as compared to the control saline group.

Figure 7

Efficacy of test sample and aspirin in terms of ulcer sore. Figure 7 shows ulcer score of normal saline, aspirin at 150 mg/kg dose, test compound at 15, 30 and 45 mg/kg doses treated groups. Statistical analysis of data was performed by post hoc Dunnet’s test after one way ANOVA. Each bar represents mean value of ulcer score ± SEM (n=6). ***p˂0.001, ** p ˂0.01 as compared to the control saline group.

Figure 8

Effect of aspirin, test compound 15, 30 and 45 mg/kg on gastric juice volume (8A)., pH (8B), free acidity (8C), total acidity (8D) and pepsin concentration (8E). Data was analyzed by post hoc Dunnett’s test after applying one way ANOVA. Data presented as mean ± S.E.M. (n = six) compared to normal saline control group *** p ˂0.001, ** p ˂0.01.

Figure 9

Histopathological assessment of control, aspirin, test compound at 15, 30 and 45 mg/kg dose (H and E staining; 4X original magnification: (A) Middle part of stomach; Saline control group showing normal appearance of mucosa (green arrow) and submucosa (blue arrow), (B) Lower part of stomach; aspirin treated group showing disruption of mucosal epithelial lining (green arrow), normal histological appearance of mucosa (green arrow) and sub mucosa (blue arrow) were found in test compound treated groups (C) Lower part of stomach; test compound 15 mg /kg treated group, (D) Lower part of tissue; test compound 30 mg/kg treated group, (E) Middle part of stomach; test compound 45 mg/kg treated group.