Capsaicin, the pungent ingredient of chili peppers, displays potent anti-neoplastic activity in a wide array of human cancer cells. The present manuscript examines the signaling pathways underlying the apoptotic activity of capsaicin in human small cell lung cancer (SCLC) in vitro and in vivo. Studies in neuronal cells show that capsaicin exerts its biological activity via the transient receptor potential vanilloid (TRPV) superfamily of cation-channel receptors. The TRPV family is comprised of six members (TRPV1-6). Capsaicin is an agonist of the TRPV1 receptor. We observed that capsaicin-induced apoptosis in human SCLC cells was mediated via the TRPV receptor family; however it was independent of TRPV1. Surprisingly, the apoptotic activity of capsaicin required the TRPV6 receptor. Depletion of TRPV6 receptor by siRNA methodology abolished the apoptotic activity of capsaicin in SCLC cells. Immunostaining and ELISA showed that TRPV6 receptor was robustly expressed on human SCLC tissues (from patients) and SCLC cell lines but almost absent in normal lung tissues. This correlates with our results that capsaicin induced very little apoptosis in normal lung epithelial cells. The proapoptotic activity of was mediated by the intracellular calcium and calpain pathway. The treatment of human SCLC cells with capsaicin induced increased the activity of calpain 1 and 2 by three-fold relative to untreated SCLC cells. Such calpain activation, in response to capsaicin, was downstream of the TRPV6 receptor. Taken together, our data provide insights into the mechanism underlying the apoptotic activity of capsaicin in human SCLCs.

In the clinical setting, antidotes are generally administered after the occurrence of a drug overdose. Therefore, the most pertinent evaluation of any new agent should model human exposure. This study tested whether acetaminophen (APAP) hepatotoxicity was reversed when S-adenosyl-L-methionine (SAMe) was administered after APAP exposure, similar to what occurs in clinical situations. Comparisons were made for potency between SAMe and N-acetylcysteine (NAC), the current treatment for APAP toxicity. Male C57BL/6 mice were fasted overnight and divided into groups: control (VEH), SAMe treated (SAMe), APAP treated (APAP), N-acetylcysteine treated (NAC), SAMe or NAC administered 1h after APAP (SAMe+APAP) and (NAC+APAP), respectively. Mice were injected intraperitoneal (i.p.) with water (VEH) or 250 mg/kg APAP (15 ml/kg). One hour later, mice were injected (i.p.) with 1.25 mmol/kg SAMe (SAMe+APAP) or NAC (NAC+APAP). Hepatotoxicity was evaluated 4h after APAP or VEH treatment. APAP induced centrilobular necrosis, increased liver weight and alanine transaminase (ALT) levels, depressed total hepatic glutathione (GSH), increased protein carbonyls and 4-hydroxynonenal (4-HNE) adducted proteins. Treatment with SAMe 1h after APAP overdose (SAMe+APAP) was hepatoprotective and was comparable to NAC+APAP. Treatment with SAMe or NAC 1h after APAP was sufficient to return total hepatic glutathione (GSH) to levels comparable to the VEH group. Western blot showed reversal of APAP mediated effects in the SAMe+APAP and NAC+APAP groups. In summary, SAMe was protective when given 1h after APAP and was comparable to NAC.

Acetaminophen (APAP) overdose leads to severe hepatotoxicity, increased oxidative stress and mitochondrial dysfunction. S-adenosyl-L-methionine (SAMe) protects against APAP toxicity at a mmol/kg equivalent dose to N-acetylcysteine (NAC). SAMe acts as a principle biological methyl donor and participates in polyamine synthesis which increase cell growth and has a role in mitochondrial protection. The purpose of the current study tested the hypothesis that SAMe protects against APAP toxicity by maintaining critical antioxidant enzymes and markers of oxidative stress. Male C57Bl/6 mice were treated with vehicle (Veh; water 15 ml/kg, ip), SAMe (1.25 mmol/kg, ip), APAP (250 mg/kg, ip), and SAMe + APAP (SAMe given 1 h following APAP). Liver was collected 2 and 4 h following APAP administration; mitochondrial swelling as well as hepatic catalase, glutathione peroxidase (GPx), glutathione reductase, and both Mn- and Cu/Zn-superoxide dismutase (SOD) enzyme activity were evaluated. Mitochondrial protein carbonyl, 3-nitrotyrosine cytochrome c leakage were analyzed by Western blot. SAMe significantly increased SOD, GPx, and glutathione reductase activity at 4 h following APAP overdose. SAMe greatly reduced markers of oxidative stress and cytochrome C leakage following APAP overdose. Our studies also demonstrate that a 1.25 mmol/kg dose of SAMe does not inhibit CYP 2E1 enzyme activity. The current study identifies a plausible mechanism for the decreased oxidative stress observed when SAMe is given following APAP.

Acetaminophen (APAP) is the leading cause of drug induced liver failure in the United States. Previous studies in our laboratory have shown that S-adenosyl methionine (SAMe) is protective for APAP hepatic toxicity. SAMe is critical for glutathione synthesis and transmethylation of nucleic acids, proteins and phospholipids which would facilitate recovery from APAP toxicity. SAMe is synthesized in cells through the action of methionine adenosyltransferase (MAT). This study tested the hypothesis that total hepatic and subcellular SAMe levels are decreased by APAP toxicity. Studies further examined MAT expression and activity in response to APAP toxicity. Male C57BL/6 mice (16-22 grams) were treated with vehicle (Veh; water 15ml/kg ip injections). 250 mg/kg APAP (15 ml/kg, ip), SAMe (1.25 mmol/kg) or SAMe administered one h after APAP injection (SAMe and SAMe+APAP). Hepatic tissue was collected 2, 4, and 6 h after APAP administration. Levels of SAMe and its metabolite S-adenosylhomocysteine (SAH) were determined by HPLC analysis. MAT expression was examined by Western blot. MAT activity was determined by fluorescence assay. Total liver SAMe levels were depressed at 4 h by APAP overdose, but not at 2 or 6 h. APAP depressed mitochondrial SAMe levels at 4 and 6 h relative to the Veh group. In the nucleus, levels of SAMe were depressed below detectable limits 4 h following APAP administration. SAMe administration following APAP (SAMe+APAP) prevented APAP associated decline in mitochondrial and nuclear SAMe levels. In conclusion, the maintenance of SAMe may provide benefit in preventing damage associated with APAP toxicity.

Solid chemical inhibitors placed in the formation during hydraulic fracturing have provided inhibition protection for up to five years. Deepwater operators, especially, desire longer inhibition periods. This paper provides a summary on treatment results over five years and over 1500 wells. It covers inhibition for scale, paraffin and asphaltene either as a single application or as a multiple product application. These applications have been in various formations including deepwater, tight gas and coal bed methane formations. The longest documented treatment has been in the ground for over five years.Placing a solid chemical inhibitor into the formation via the fracturing process requires a product that is compatible with the fracturing fluid, does not adversely affect conductivity and provides long term inhibition through the controlled release of the inhibitor into the produced fluid. The solid inhibitor is added to the fracturing proppant. Primarily this is a mass-balance process by which a finite amount of inhibitor desorbs over time. The goal is to maximize the inhibitor loading, minimize the chemical release rate without negatively impacting the stimulation.In the treated wells inorganic and organic deposition has been arrested for extended periods when compared to alternative liquid addition applications. Solid scale inhibitors have been the most common treatment and are monitored through residual analysis. The paraffin and asphaltene inhibitors are monitored through comparative testing. The paper details both the time and the cumulative production that has flowed through the respective proppant packs.Wells that are scheduled for hydraulic fracturing and that display tendencies for organic and/or inorganic deposition are candidates for solid chemical placement during the fracture. The results from over 1500 wells indicate that this method for deposition inhibition reduces intervention costs and lowers lifting costs.