Novel strategies for the prevention of cisplatin-induced ototoxicity

Cisplatin is a commonly used chemotherapeutic agent. Unfortunately, serious side effects limit its clinical use such as ototoxicity, which presents as bilateral and progressive sensorineural hearing loss. Regrettably, there is currently no treatment for cisplatin-induced ototoxicity. The pathophysiology remains unclear, however, it is believed that inflammation and oxidative stress are the main mechanisms leading to cell death. In the present thesis, various aspects of cisplatin-induced ototoxicity and potential treatment strategies are evaluated. We begin with a review of the literature in what concerns the entrance and egress of cisplatin from cochlear cells. Cisplatin has a predilection for the inner ear tissues and the reason for such an occurrence is unknown. We describe the receptors that may play a role in cisplatin-induced ototoxicity and that are present in cochlear cells. Understanding the circulating pathways of cisplatin within the inner ear can provide some insight into the mechanisms of cisplatin-induced ototoxicity. Once inside the cell, cisplatin can elicit an inflammatory response. For this reason, we decided to evaluate the potential of dexamethasone as a protective agent against cisplatin's toxic effects in vivo. It was observed that a central regulator of inflammation was decreased as a result of the therapy; however, the hearing was not preserved. An anti-inflammatory did not provide sufficient protection to preserve hearing following the cisplatin treatment. ROS have also been implicated in cisplatin-induced cytotoxicity. It appears that cisplatin can lead to an increased expression of ROS that can overwhelm the natural antioxidant response of the cochlea. Thus, the potential of an exogenous antioxidant as a protective agent was evaluated in vivo. Erdosteine, a derivative of methionine, provided protection against cisplatin-induced ototoxicity at high frequencies of hearing as well as partially prevented OHC loss. Because these two compounds provided only but partial benefits, we decided to evaluate a more specific and targeted approach, gene therapy for cisplatin-induced ototoxicity. We performed a systematic review of the literature in order to evaluate the potential of genetic manipulation in experimental animal and in vitro studies. Interestingly, a variety of genes have been evaluated as potential targets for inhibiting cisplatin-induced cytotoxicity such as apoptotic suppressors, copper transporters, regulators of the antioxidant response and neural growth factors. Consequences of genetic manipulation in the inner ear tissues remain to be assessed in order for gene therapy to become a conventional therapeutic option. Because the cochlea is embedded in bone, is fluid filled and contains various cell types, it has been a challenge to detect the expression of manipulated genes in a particular cell type of interest. While homogenization of a whole cochlea and posterior RNA extraction can provide us with the general expression levels of a certain gene, it does not allow for the determination in a cellular subpopulation of the cochlea. One possibility is the use of laser capture microdissection of cells of interest from a histological section. With this approach, the cells of interest are obtained and RNA can then be extracted and gene expression levels determined. However, the process of obtaining histological sections from cochlear samples requires fixation and decalcification steps which are known to cause RNA degradation. Hence, we decided to evaluate combinations of fixatives and decalcifying agents in order to determine which protocol would yield the greatest quantity of RNA from the cochlea and also preserve the morphology. The resulting protocol with methacarn fixation and decalcification in Morse's solution can therefore be used in future studies that aim to determine genetic expression, a regularly performed experiment, in a specific cellular subtype of interest in the cochlea.

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