A cybrid cell model for the assessment of the link between mitochondrial deficits and sporadic Parkinson's disease

Abstract

Parkinson's disease (PD) is a multifactorial and clinically complex age-related movement disorder. The cause of its most common form (sporadic PD, sPD) is unknown, but one prominent causal factor is mitochondrial dysfunction. Although several genetic- and toxin-based models have been developed along the last decades to mimic the pathological cascade of PD, cellular models that reliably recapitulate the pathological features of the neurons that degenerate in PD are scarce.We describe here the generation of cytoplasmic hybrid cells (or cybrids) as a cellular model of sPD. This approach consists on the fusion of platelets harboring mtDNA from sPD patients with cells in which the endogenous mtDNA has been depleted (Rho0 cells).The sPD cybrid model has been successful in recapitulating most of the hallmarks of sPD, constituting now a validated model for addressing the link between mitochondrial dysfunction and sPD pathology.

Fig. 1

Generation of cybrid cell lines. Tumor or immortalized cell lines are grown in the presence of ethidium bromide, which efficiently eliminates functional mtDNA resulting in a Rho0 cell line. Rho0 cells are then fused with patient platelets, which contain mitochondria but not nuclei. This creates cytoplasmic hybrid (cybrid) cells that can be isolated and expanded. The expanded cybrid cell cultures are biochemically analyzed. Differences in function between cell lines mostly likely arise through differences in their mtDNA

Fig. 2

Schematic representation of the end result after centrifugation of anticoagulated venous blood layered onto Histopaque. During centrifugation, erythrocytes and granulocytes are aggregated by polysucrose and rapidly sediment, whereas lymphocytes and other mononuclear cells such as platelets remain at the plasma–Histopaque interface