Silencing of Hsp70 Intensifies 6-OHDA-Induced Apoptosis and Hsp90 Upregulation in PC12 Cells

Parkinson's disease (PD) is a neurodegenerative disorder characterized by a preferential loss of the dopaminergic neurons of the substantia nigra pars compacta. Although the etiology of PD is unknown, major biochemical processes such as oxidative stress and mitochondrial inhibition are largely described. However, despite these findings, the actual therapeutics are essentially symptomatical and are not able to block the degenerative process. Recent histological studies performed on brains from PD patients suggest that nigral cell death could be apoptotic. However, since post-mortem studies do not allow precise determination of the sequence of events leading to this apoptotic cell death, the molecular pathways involved in this process have been essentially studied on experimental models reproducing the human disease. These latter are created by using neurotoxic compounds such as 6-hydroxydopamine (6-OHDA), 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) or dopamine (DA). Extensive study of these models have shown that they mimick, in vitro and in vivo, the histological and/or the biochemical characteristics of PD and thus help to define important cellular actors of cell death presumably critical for the nigral degeneration. This review reports recent data concerning the biochemical and molecular apoptotic mechanisms underlying the experimental models of PD and correlates them to the phenomena occurring in human disease.

Human idiopathic Parkinson's disease (PD) is a progressive neurodegenerative disorder that is primarily characterized by degeneration of the dopaminergic neurons of the nigrostriatal pathway. Different 6-OHDA rat models of PD have been developed in which this toxin has been injected into different parts of the nigrostriatal pathway: (a) the medial forebrain bundle which leads to extensive dopamine (DA) depletion; (b) the substantia nigra pars compacta, which leads to more specific and moderate DA depletions; and (c) subregions of the caudate-putamen complex (CPu), which also leads to specific DA depletions. In this article we review the dopaminergic depletion and behavioral consequences of 6-OHDA lesions in the rat. It was examined whether the relation between DA depletion and behavioral deficits mimic idiopathic PD. In addition, it was evaluated which model most closely approximates the human situation, especially in relation to the stage of this progressive disease. It was concluded that with respect to the site of the lesion, rats with partial lesions of the ventrolateral CPu are the most appropriate models to study early and late stages of PD. The choice of the behavioral parameters determines the use of unilateral or bilateral lesions, although it is obvious that the bilateral model mimics the human situation more closely. (c) 2002 Elsevier Science (USA).

Resistance to stress-induced apoptosis was examined in cells in which the expression of hsp70 was either constitutively elevated or inducible by a tetracycline-regulated transactivator. Heat-induced apoptosis was blocked in hsp70-expressing cells, and this was associated with reduced cleavage of the common death substrate protein poly(ADP-ribose) polymerase (PARP). Heat-induced cell death was correlated with the activation of the stress-activated protein kinase SAPK/JNK (c-Jun N-terminal kinase). Activation of SAPK/JNK was strongly inhibited in cells in which hsp70 was induced to a high level, indicating that hsp70 is able to block apoptosis by inhibiting signaling events upstream of SAPK/JNK activation. In contrast, SAPK/JNK activation was not inhibited by heat shock in cells with constitutively elevated levels of hsp70. Cells that constitutively overexpress hsp70 resist apoptosis induced by ceramide, a lipid signaling molecule that is generated by apoptosis-inducing treatments and is linked to SAPK/JNK activation. Similar to heat stress, resistance to ceramide-induced apoptosis occurs in spite of strong SAPK/JNK activation. Therefore, hsp70 is also able to inhibit apoptosis at some point downstream of SAPK/JNK activation. Since PARP cleavage is prevented in both cell lines, these results suggest that hsp70 is able to prevent the effector steps of apoptotic cell death. Processing of the CED-3-related protease caspase-3 (CPP32/Yama/apopain) is inhibited in hsp70-expressing cells; however, the activity of the mature enzyme is not affected by hsp70 in vitro. Caspase processing may represent a critical heat-sensitive target leading to cell death that is inhibited by the chaperoning function of hsp70. The inhibition of SAPK/JNK signaling and apoptotic protease effector steps by hsp70 likely contributes to the resistance to stress-induced apoptosis seen in transiently induced thermotolerance.