Alsbury, S.J.; (2004) Neuroprotection and ageing in sensory neurons. Doctoral thesis , University of London.

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Abstract

Cardiotrophin-l (CT-1) and Urocortin (UCN) are peptides which have previously been shown to have protective effects in cardiac myocytes and which induce heat shock protein expression. CT-1 is a member of the interleukin-6 family of cytokines. UCN is a peptide expressed in the brain that binds to the corticotrophin releasing hormone receptors. In this investigation the ability of CT-1 and UCN to protect cells against a lethal stress was tested in neonatal and adult sensory neurons. CT-1 has a protective effect against hypoxic ischaemia in neonatal sensory neurons; in contrast although CT-1 induces the expression of HSP 70 in adult sensory neurons it does not have a protective effect. Results from studies using the kinase inhibitors PD 98059, SB 203580 and LY 294002 suggest that CT-1 is activating the P42/44 MAPK pathway in neonatal sensory neurons. The western blots also reveal a higher level of phosphorylated P42 MAPK in the adult sensory neurons than in the neonatal sensory neurons suggesting that CT-1 does not have a protective effect in the adult neurons because this pathway is already activated. Surprisingly, UCN does not have a protective effect in sensory neurons. In the neonatal cells this could be explained by a lack of receptor expression since RT-PCR revealed an absence of mRNA for both receptors 1 and 2, however; in the adult neurons mRNA was present for both receptors. In conclusion CT-1 but not UCN has an age dependent protective effect in sensory neurons. CT-1 is not the only treatment known to have a protective effect that is lost with age, for instance ischaemic preconditioning is protective in young hearts but not in senescent hearts. The induction of heat shock proteins by mild stress is protective against further stress in young animals but the induction of HSPs is impaired with age. Investigations into the possible therapeutic benefit of increasing heat shock protein expression have previously been performed but this work was carried out using young animals and cells from young animals. This investigation therefore progressed to explore whether increasing heat shock protein expression in neurons from aged animals could protect them from lethal stresses. An HSV-1 based viral vector was used to individually overexpress three members of the heat shock family, HSP 27, HSP 70 and HSP 56, in sensory neurons of aged and neonatal Sprague Dawley rats in vitro. Transcription of the HSP genes in this vector is controlled by a CMV promoter, producing high levels of protein expression. The protective effect of the three proteins against heat shock and hypoxic ischaemia was tested in sensory neurons from aged and neonatal rats. As has been previously shown, in neonatal sensory neurons HSP 27 and HSP 70 protect against cell death due to heat shock. Encouragingly, both HSP 27 and HSP 70 protect the sensory neurons of aged animals against heat shock, although only HSP 27 gave a significant level of protection against hypoxic ischaemia in aged sensory neurons. In conclusion, it is possible to protect neuronal cells of aged animals against stress if the levels of heat shock proteins can be restored.

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