The results of MSc Niina Piippo’s doctoral thesis indicate that impaired cellular degradation activates the NLRP3 inflammasome in retinal pigment epithelial cells via increased oxidative stress. Retinal pigment epithelial cells play a major role in the pathogenesis of age-related macular degeneration.
Age-related macular degeneration (AMD) is one of the leading causes of blindness throughout the world. Its prevalence is expected to increase significantly especially as the citizens of the Western countries grow older. AMD causes severe visual impairment and thus it is associated with social problems for the individual and a financial burden to society. The degeneration of retinal pigment epithelium (RPE) is fundamental in the progression of AMD. Many factors affect RPE degeneration e.g. the accumulation of cellular waste, elevated oxidative stress, and increased inflammation. While initially a beneficial response to cellular damage, inflammation subsequently evokes tissue damage in situations where the original trigger cannot be resolved and inflammation gets prolonged. Tissue damage in the RPE is followed by the death of photoreceptors causing a loss of vision. The innate immune receptor Nucleotide-binding domain and Leucine-rich repeat Receptor containing a Pyrin domain 3 (NLRP3) has recently been shown to participate in inflammation in RPE cells.
In this study, the intracellular degradation in RPE cells was inhibited by blocking proteasomes with MG-132 and preventing autophagy with bafilomycin A (BafA). The increased expression of NLRP3, increased activation of caspase-1 enzyme and elevated levels of secreted interleukin (IL)-1β cytokine suggest that MG-132 and BafA treatments had induced NLRP3 inflammasome activation in the RPE cells. Preliminary experiments were conducted using the ARPE-19 cell line; later the functionality of the protocol was confirmed using primary human RPE cells and human embryonal stem cell (hESC)-derived RPE cells. The role of oxidative stress in the NLRP3 activation was revealed by measuring intracellular reactive oxygen species (ROS) and 4-hydroxy-2-nonenal (HNE) adducts as well as by inhibiting intracellular ROS with several scavengers of intracellular ROS. It was demonstrated that oxidative stress plays the principal role in the activation of NLRP3 in this model. The NLRP3 activity induced by dysfunctional intracellular clearance could be prevented by geldanamycin, a well-known Hsp90 inhibitor. 8
The results of this thesis indicate that impaired cellular degradation activates the NLRP3 inflammasome in RPE cells via increased oxidative stress. RPE cells play a major role in the pathogenesis of AMD. Furthermore, NLRP3 signaling could be prevented by inhibiting the actions of the heat shock protein, Hsp90.
The doctoral dissertation of Niina Piippo, Master of Science, entitled Inflammasome signaling and its regulation in human retinal pigment epithelial cells with dysfunctional intracellular clearance will be examined at the Faculty of Health Sciences on 19 November 2019. The Opponent in the public examination will be Professor Harri Alenius of Karolinska Institutet and the University of Helsinki, and the Custos will be Professor Anu Kauppinen of the University of Eastern Finland.