Lecithin: cholesterol acyltransferase (LCAT) is a soluble enzyme that converts cholesterol to cholesteryl esters on the surface of HDL for their subsequent transport to the liver. Mutations in the human LCAT gene result either in fish eye disease (FED), where patients have a partial deficiency of LCAT activity or in familial LCAT deficiency (FLD). FLD patients have a complete deficiency of LCAT activity and clinically present with corneal opacities, normochromic anemia, and renal disease (proteinuria). The LCAT-KO mice are characterized by lipid, hematologic, and renal abnormalities similar to familial LCAT deficiency patients.
In this study, we aim to: 1) investigate if the Cretan patients with FLD, who have a novel mutation in the LCAT gene, carry any of seven other known polymorphisms, 2) develop a refined three - dimensional (3D) model of LCAT and locate the 7 mutations examined in Cretan patients as well as the new one, 3) measure the LCAT activity in these patients, 4) examine the expression pattern of activin A, a member of the TGF superfamily, of activin A receptors as well as TGF-β, a key mediator of renal fibrosis in mice exhibiting nephropathy.
Our results show that apart from the new mutation, no other mutations were found in the Cretan patients and that the novel mutation seems to be Cretan-specific. Concerning the LCAT activity assay, no activity of the enzyme was detected neither in heterozygotes nor in homozygotes. Using the method of homology modeling, which deals with the prediction of the tertiary structure of an unknown protein based on the known structure of a homologous one, we constructed a temporary 3D-model, based on 2PVS (human pancreatic lipase related protein). This model revealed that the novel mutation is located at the beginning of a β-strand and may influence the secondary structure of the protein. We then analyzed the expression of activin A, its receptors and TGF-β in 1, 3, 6 and 9 months old NZBW mice, which were used as a surrogate for LCAT KO mice, since they exhibit the same renal lesions at the final stages of the disease. We found that activin’s A expression increases with the age of mice, whereas we cannot conclude safely for the pattern of expression of its receptors and TGF-β.