Study of salivary arecoline in areca nut chewers

Abstract

Aims: Arecoline, a predominant alkaloid present in arecanut, has been implicated in the pathogenesis of several oral diseases because of its mutagenic and carcinogenic potential. The response of cultured cells to arecoline is highly dependent on its concentration; arecoline stimulates cultured cells above 0.1 μg/ml and is cytotoxic above 10 μg/ ml. Although this alkaloid seems important for areca nut induced oral diseases and carcinogenesis, little is known of the levels achieved before, during and after chewing. Also, it is prudent to understand its effects in arecanut chewers for a comprehensive understanding of its pathogenesis. Accordingly, the present study quantified the salivary arecoline levels in arecanut chewers. Materials and Methods: The study participants were divided into Study Group A & B and Control Group C; unstimulated whole saliva was collected by spitting method for a period of 5 min. Then, participants in Group A and C chewed 0.5 g of areca nut without any other additives while in Group B were asked to chew 0.5 g of inert rubber base impression material. Stimulated whole saliva from all three groups was collected into graduated tubes during chewing at time intervals of 1, 3, 5, 10, 15, 20 and 25 min. Then, all participants were asked to remove nut particles or inert rubber base material from the mouth, and saliva samples were collected further up to 20 min, changing tubes at 5 min interval. Salivary arecoline was quantitated by HPLC-MS. The tabulation and descriptive statistics of the study were carried out. Results: In the present study, baseline levels of arecoline were zero in all three groups, whereas mean salivary arecoline levels during chewing were 76.93 ng/ml, 129.83 ng/ml and 64.83 ng/ml and after chewing were 196.17 ng/ml, 321.12 ng/ml and 43.75 ng/ml in Groups A, B and Control respectively, which were significantly higher than reported threshold levels. Conclusions: The data from this study reveals that a significant amount of arecoline would be trapped in oral cavity, or being re-circulated between blood and saliva might have resulted in surprisingly high levels of arecoline even 10 mins after chewing in both groups after which the levels started declining. The higher levels of salivary arecoline achieved during and after chewing are enough to cause cytotoxic and genotoxic effects on oral tissues over a period of time in chronic chewers. The great differences in salivary arecoline levels achieved during chewing, may contribute to the variable response to areca nut seen in communities where this habit is widespread. Areca nut users have persistent background salivary arecoline levels long after chewing, whereas concentrations achieved are highly variable and consistent with a role in oral pre-malignancy and malignancy..

Egr-1 mediates low dose Arecoline induced human oral mucosa fibroblasts proliferation by transaction the expression of Wnt5a

“…IEGs are genes which are activated transiently and rapidly in response to a wide variety of cellular stimuli. These characteristics of Egr-1 are consistent with the wavy change of Arecoline concentration in areca nut chewer's saliva [20]. And in this study, we also produced an Arecoline concentration uctuation by replacing half of the cell culture medium every 24 hours.…”

“…However, it is true that the areca nut chewing increases the risk of oral cancer and oral submucous brosis. In fact, the salivary Arecoline level during chewing were about 0.1 µg/ml, and increased to about 0.3 µg/ml after chewing [20], which is much lower than the concentration used in most experiments.…”

“…Chang et al found that Arecoline was cytotoxic to human oral broblasts at a concentration higher than 50 µg/ml by depleting intracellular thiols and inhibiting mitochondrial activity [8,30]; Jeng et al found Arecoline inhibited the migration, attachment, spreading, growth and collagen synthesis of human oral broblasts at concentrations of 0.4 mM (62 µg/ml) and 1 mM (155 µg/ml) [7]. In fact, Venkatesh et al found that the salivary Arecoline level were about 0.1 µg/ml to about 0.3 µg/ml after chewing commercially available areca nut [20], which is much lower than the concentration used in most experiments. In this study, we found that low dose Arecoline could promote the proliferation of human oral broblasts.…”

Egr-1 mediates low-dose arecoline induced human oral mucosa fibroblast proliferation via transactivation of Wnt5a expression

“…Chewing areca nuts is known to increase the risk of oral cancer and OSF. The level of arecoline present in the saliva during areca nut chewing has been found to be around 0.1 μg/ml, and has been found to increase to about 0.3 μg/ml after chewing [ 24 ]; these concentrations are much lower than the concentration of arecoline used in most experiments.…”

“…Chang et al found that arecoline was cytotoxic to human oral fibroblasts at concentrations greater 50 μg/ml due to depletion of intracellular thiols and inhibition of mitochondrial activity [ 12 , 30 ]; Jeng et al found that arecoline treatment inhibited the migration, attachment, spreading, growth, and collagen synthesis of human oral fibroblasts at concentrations of 0.4 mM (62 μg/ml) and 1 mM (155 μg/ml) [ 11 ]. Venkatesh et al found that arecoline levels in the saliva were about 0.1 μg/ml to about 0.3 μg/ml after chewing commercially available areca nuts [ 24 ], which is much lower than the concentration of arecoline used in most experiments. In this study, we found that low doses of arecoline could promote the proliferation of human oral fibroblasts.…”