Abstract
Purpose :
Protein aggregation has been one of the leading triggers of various disease conditions, such as Alzheimer's, Parkinson's and other amyloidosis. TGFBI-associated corneal dystrophies are protein aggregation disorders in which the mutant TGFBIp aggregates and accumulates in the cornea, leading to a reduction in visual acuity and blindness in severe cases. Currently, the only therapy available is invasive and there is a known recurrence after surgery. In this study, we tested the inhibitory and amyloid dissociation properties of four osmolytes in an in-vitro TGFBI peptide aggregation model. We also tested the toxicity of the disaggregated fibrils on human corneal fibroblasts.
Methods :
The 23-amino acid long peptide (TGFBIp 611-633 with the mutation c.623 G>R) from the 4th FAS-1 domain of TGFBIp that rapidly forms amyloid fibrils was used in the study. Several biophysical methods like Thioflavin T (ThT) fluorescence, Circular Dichroism (CD), fluorescence microscopy, and Transmission electron microscopy (TEM) were used to study the inhibitory and amyloid disaggregation properties of four osmolytes (Betaine, Raffinose, Sarcosine, and Taurine). The toxicity of the disaggregated amyloid fibrils on corneal fibroblasts was studied using xCELLigence and Incucyte assays.
Results :
The osmolytes were effective in both inhibition and disaggregation amyloid fibrils derived from TGFBIp 611-633 c.623 G>R peptide. The osmolytes did not have an adverse toxic effect on cultured human corneal fibroblast cells and could potentially be a useful therapeutic strategy for patients with TGFBIp corneal dystrophies. The disaggregated amyloid fibrils after treatment with osmolytes did not have any toxic effect on the human corneal fibroblasts.
Conclusions :
The osmolytes were not toxic to the human corneal fibroblast cells. The osmolytes were effective to disaggregate the in-vitro amyloid fibrils. The osmolytes were also effective to delay or inhibit amyloid fibril formation in these peptides with G623R mutation. The disaggregated amyloid fibrils after osmolyte treatment added to the human corneal fibroblast cell did not show any toxicity.
This is a 2021 ARVO Annual Meeting abstract.