Autosomal recessive hypercholesterolaemia: Discrimination of ARH protein and LDLR function in the homozygous FH phenotype Aron B. Abera a , A. David Marais a , Frederick J. Raal b , Felicity Leisegang c , Sheena Jones a , Peter George d , Howard E. Henderson c, ⁎ a Department of Medicine, University of Cape Town, UCT/NHLS, Cape Town, South Africa b Department of Medicine University of Witwatersrand, Johannesburg, South Africa c Division of Chemical Pathology, UCT/NHLS, Cape Town, South Africa d Canterbury Health Laboratories, Christchurch, New Zealand Received 22 September 2006; received in revised form 4 October 2006; accepted 4 October 2006 Available online 13 October 2006 Abstract Background: Phenocopies of homozygous familial hypercholesterolemia (hoFH) having autosomal recessive inheritance, were recently found to arise from defects in the LDL receptor (LDLR) adapter protein, called ARH, which facilitates the clearance of circulating LDL. Discrimination between the two causes of the phenotype at a clinical level may not be possible when parents display moderate hypercholesterolaemia. An effective strategy is thus required to identify the appropriate mechanism for the disorder. Methods: Fibroblast LDL uptake studies were coupled with Western blotting for ARH protein in cell extracts, to identify the defective gene before DNA studies were initiated. Two subjects with the hoFH phenotype, but with indeterminate dyslipidaemia in their parents, were fully worked up. Results: Defective LDL metabolism was established in both patients by functional and protein studies and further confirmed by detecting deleterious mutations, in the LDLR and ARH genes. The ARH patient is the first subject of Negroid identity to be described and records a specific mutation in this racial grouping. Conclusion: This study highlights the occasional complexity and uncertainty of a clinical diagnosis of hoFH and presents Western blotting of leucocyte extracts for ARH protein, as a rapid strategy for the detection of ARH before sequencing the gene for mutation(s). This strategy may be particularly useful in populations where founder mutations for ARH and LDLR defects are rare or co-exist. © 2006 Elsevier B.V. All rights reserved. Keywords: Autosomal recessive hypercholesterolaemia; ARH deficiency; LDLR function; Western blotting; Gene mutations 1. Introduction Patients presenting with premature coronary artery disease (CAD), tendon and cutaneous xanthomata, absence of hyper- triglyceridaemia and plasma LDL cholesterol levels in excess of 5 mmol/l are overwhelmingly heterozygous or homozygous for mutations in the LDL receptor (LDLR) gene and are diagnosed as having familial hypercholesterolaemia (FH; OMIM 143890) [1]. However, three additional genes involved in LDL metabo- lism can give rise to phenocopies of FH. These include familial defective apolipoprotein-B (FDB; OMIM 144010; [2], autoso- mal dominant hypercholesterolaemia type 3 (HCHOLA3; OMIM 603776) where mutations in the proprotein convertase subtilisin/kexin type 9 (PCSK9; OMIM 607786) are causative [3], and autosomal recessive hypercholesterolaemia (ARH; OMIM 603813) where patients have deficiencies in the LDL receptor adapter protein, ARH [4,5]. The heterozygous FH phenotype (heFH) is diagnosed by the concurrence of plasma LDL cholesterol, in the range of 5– 12 mmol/l, tendon xanthoma and premature CAD whereas homozygous FH (hoFH) presents with cutaneous and tendon xanthoma, LDL cholesterol, usually N 12 mmol/l and very pre- mature CAD. The South African experience with genetic hypercholesterolaemia to date (unpublished data) indicates that Clinica Chimica Acta 378 (2007) 33 – 37 www.elsevier.com/locate/clinchim ⁎ Corresponding author. Division of Chemical Pathology, University of Cape Town, Observatory 7925, Cape Town, South Africa. Tel.: +27 21 4066217; fax: +27 21 4488150. E-mail address: hendersn@chempath.uct.ac.za (H.E. Henderson). 0009-8981/$ - see front matter © 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.cca.2006.10.005