Fisiopatología del canal iónico trpa1 en el agrandamiento gingival y en la neuropatía inducida por oxaliplatino

Resumo

TRPA1 channel as a potential mediator of drug-induced gingival overgrowth Gingival enlargement is a common side effect observed in patients treated with antiepileptic (phenytoin and carbamazepine), immunosuppressant (cyclosporine A) and some antihypertensive (nifedipine) drugs. However, the molecular mechanisms behind gingival overgrowth by these agents are still unknown. In a recent study, it was demonstrated that 1,4 dihydropyridines, including nifedipine, can activate TRPA1 channel, a cationic channel expressed in nociceptors, which is activated by many irritant compounds. Hence, we hypothesized that TRPA1 could be the molecular target involved in drug-induced gingival overgrowth. In our studies, phenytoin, carbamazepine and nifedipine increased the intracellular calcium levels in CHO cells expressing mouse TRPA1, and in HEK cells expressing human TRPA1. Interestingly, these responses were not observed in cells lacking TRPA1 or expressing TRPM8 or TRPV1. No such effect was observed with cyclosporine A. By RT-PCR, we demonstrated that TRPA1, TRPV1 and its capsaicin-insensitive isoform TRPV1b are expressed in cultured human gingival fibroblasts (HGF). Calcium imaging showed that HGF cells responded to TRPA1 agonists (e.g. mustard oil) and drugs producing gingival enlargement. Moreover, activation of TRPA1 by phenytoin was blocked by HC030031, a specific blocker for this channel. Similarly, the use of shRNAs against hTRPA1 in HGF cells reduced TRPA1 expression and activation by phenytoin. Finally, this activation was markedly reduced in the presence of ascorbic acid, folic acid and ¿-tocopherol, antioxidants that are reduced in blood samples from patients with gingival enlargement. Gingival enlargement may be caused by fibroblast proliferation or higher synthesis and secretion of collagen by fibroblasts. By performing MTT cell proliferation assay, we showed that none of these drugs induced proliferation of HGFs. These results indicate that phenytoin, nifedipine and carbamazepine can activate TRPA1 in native gingival fibroblasts increasing intracellular calcium, this increase in calcium could induce overexpression of cytokines and connective tissue growth factor (CTGF) leading to gingival enlargement by increasing collagen deposits. ¿ TRPA1 channel is a mediator of oxaliplatin-induced neuropathic pain: Oxaliplatin is a chemotherapeutic drug used in the treatment of colorectal cancer. This treatment can induce acute side effects, such as cold hypersensitivity and paraesthesia, in 80% of the patients, and can evolve to a long-term neuropathy in a fraction of the affected patients (around 15-20%). The mechanism responsible for the peripheral neuropathy induced by oxaliplatin is poorly understood. Oxaliplatin affects different ionic channels expressed in peripheral sensory neurons, including reductions in sodium and potassium currents leading to a decrease in the action potential amplitude. We found that oxaliplatin reduces the response to elevated potassium chloride in Dorsal Root Ganglia neurons and induces calcium channel Cav1.2 overexpression. These changes in voltage-gated channel expression could modify the sensory neuron biophysics perpetuating long-term neuropathy symptoms. As these symptoms, such as cold hypersensitivity, are triggered by cold exposure, we hypothesized that cold-sensing TRP channels TRPA1 and/or TRPM8 could be mediating cold hypersensivity in oxaliplatin-induced neuropathy. In the present work, we demonstrated that oxaliplatin activates TRPA1 channels expressed in heterologous systems (HEK 293 and CHO¿TRPA1). This activation was blocked by the specific TRPA1 antagonist HC030031, and also by the reactive oxygen species chelator GSH. These results suggest that TRPA1 activation by oxaliplatin depends on oxidative stress. Moreover, we described how oxaliplatin increased cold response and temperature threshold in TRPA1, and reduced cold and menthol responses in TRPM8. At the molecular level, oxaliplatin treatment induces overexpression of TRPA1, but not TRPM8. Taking all this into account, we demonstrated that cold hypersensitivity in oxaliplatin-induced neuropathy could be mediated by TRPA1 cold activation at warm temperatures, but also by TRPM8 cold response inhibition. These results suggest that at the chronic phase, TRPA1 overexpression would extend cold hypersensitivity in oxaliplatin treated patients.