A human neuronal tissue culture model for Lesch‐Nyhan disease

Abstract

Mutations in the gene encoding the purine salvage enzyme, hypoxanthine-guanine phosphoribosyltransferase (HPRT) cause Lesch-Nyhan disease, a neurodevelopmental disorder characterized by cognitive, neurological, and behavioral abnormalities. Despite detailed knowledge of the enzyme's function, the key pathophysiological changes that accompany loss of purine recycling are unclear. To facilitate delineating the consequences of HPRT deficiency, four independent HPRT-deficient sublines of the human dopaminergic neuroblastoma, SK-N-BE(2) M17, were isolated by targeted mutagenesis with triple helix-forming oligonucleotides. As a group, these HPRT-deficient cells showed several significant abnormalities: (i) impaired purine recycling with accumulation of hypoxanthine, guanine, and xanthine, (ii) reduced guanylate energy charge and GTP : GDP ratio, but normal adenylate energy charge and no changes in any adenine nucleotide ratios, (iii) increased levels of UTP and NADP + , (iv) reduced DOPA decarboxylase, but normal monoamines, and (v) reduction in cell soma size. These cells combine the analytical power of multiple lines and a human, neuronal origin to provide an important tool to investigate the pathophysiology of HPRT deficiency.

Lesch-Nyhan disease: from mechanism to model and back again

“…HPRT-deficient dopamine neuronlike lines show a loss of dopamine content that is analogous to that reported for the LND brain (Bitler and Howard, 1986;Yeh et al, 1998;Lewers et al, 2008). These models also reveal changes in neuronal microstructure in the form of abnormal numbers or morphology of neurites, which are comparable to dendrites or axons in vivo (Stacey et al, 1999;Connolly et al, 2001;Shirley et al, 2007).…”

“…An unexpected finding has been the lack of an obvious purine deficiency, which most investigators expected would result from the failure of purine recycling. Some studies reveal small decreases or increases in one or another purine, but results are inconsistent and many studies reveal normal levels of purines (Shirley et al, 2007).…”

“…A final limitation is that findings might be idiosyncratic to the cell model being evaluated (Shirley et al, 2007;Lewers et al, 2008). Because LND is so rare, models derived from blood samples or fibroblast biopsies often come from a single patient, or very small numbers of patients.…”

“…Several investigators have sought, more specifically, to model the processes responsible for neural dysfunction by studying HPRT-deficient neuronal or glial cells in culture (Shirley et al, 2007). Such cells cannot be obtained readily from affected human patients, so most were developed by creating HPRT-deficient sublines of established neuroblastoma or glioma cell lines.…”

Lesch-Nyhan Syndrome: Models, Theories, and Therapies

“…To circumvent this cell type problem, researchers developed HPRTdeficient subclones of glioma and neuroblastoma cell lines [Shirley et al, 2007]. These neural models have allowed researchers to assess specific deficits in neurotransmitters [Bitler and Howard, 1986;Yeh et al, 1998;Lewers et al, 2008] and neuronal structure abnormalities [Connolly et al, 2001;Shirley et al, 2007] that occur when HPRT activity is absent. However, results obtained from these models may lose relevance, as the cells were obtained from nonpatient neuronal tumors.…”