Hypoxanthine impairs morphogenesis and enhances proliferation of a neuroblastoma model of Lesch Nyhan syndrome

Abstract

Extracellular purines have essential roles in neuronal development; hence, disruptions in their metabolism as reported in Lesch Nyhan syndrome (LNS) could result in developmental abnormalities. The deficiency of hypoxanthine-guanine phosphoribosyl transferase (HGPRT) in LNS leads to increased hypoxanthine and uric acid production. We have reported that HGPRT-deficient B103-4C neuroblastoma, a neuronal model of LNS, proliferated less and differentiated more than their HGPRT-positive B103 counterparts. Here, we sought to determine whether differences in proliferation and differentiation would occur when these cells were cultured in the presence of hypoxanthine or in a hypoxanthine-/serum-free chemically defined media (NBMN2). In media with 1% serum, hypoxanthine (50 microM) significantly increased the proliferation of both cell lines with a greater effect on B103-4C cells. In 1% serum media, hypoxanthine increased differentiation of B103 but decreased B103-4C differentiation. In NBMN2, B103 proliferated far more than B103-4C, but both cell types differentiated to the same extent. These results are interpreted to suggest that elevated levels of central nervous system (CNS) hypoxanthine as reported in LNS may affect neuronal development, and to implicate hypoxanthine and abnormal neuronal development as causative factors in the etiology of LNS.

Impairment of adenylyl cyclase 2 function and expression in hypoxanthine phosphoribosyltransferase-deficient rat B103 neuroblastoma cells as model for Lesch–Nyhan disease: BODIPY–forskolin as pharmacological tool

“…Implications of our study for understanding LND HPRT deficiency has been repeatedly associated with defects in neuronal differentiation (Ma et al 2001;Connolly 2001;Jinnah 2009;Erdorf et al 2011a;Kang et al 2011;Guibinga et al 2010Guibinga et al , 2012Mastrangelo et al 2012). We could confirm this differentiation defect since HPRT − cells underwent much less striking morphological changes upon exposure to FS and BODIPY-FS than control cells (Fig.…”

“…BODIPY control dye itself had no effect on the morphology on HPRT − cells. Impaired differentiation of B103 cells had already been observed in previous studies (Connolly 2001;Ma et al 2001). As was the case in B103 control cells, FS, BODIPY-FS, and BODIPY (1 μM each) had no effect on cell viability of HPRT − cells as assessed by ethidium bromide/acridine orange staining (data not shown).…”

“…5). The differentiation defect of HPRT − cells is seen under different experimental conditions and involves multiple signal transduction pathways including purinoceptors, microRNAs and the canonical Wnt pathway (Ma et al 2001;Connolly 2001;Jinnah 2009;Erdorf et al 2011a;Kang et al 2011;Guibinga et al 2010Guibinga et al , 2012Mastrangelo et al 2012).…”

“…In HPRT-deficient cells, 6-thioguanine cannot be converted to the cytotoxic 6-thioguanosine 5′-triphosphate and hence, cells survive (Zoref-Shani et al 1993;Hacke et al 2012). HPRTdeficient B103 cells are an established cell culture model for LND and have already been used for two decades to study various aspects of the pathophysiology of LND including alterations in neuronal differentiation, nucleotide metabolism, purinoceptor function, nucleotidase activity, nucleotidase gene expression, and adenylyl cyclase (AC) regulation (Zoref-Shani et al 1993;Ma et al 2001;Connolly 2001;Pinto et al 2005;Pinto and Seifert 2006;Lorenz et al 2007;Erdorf et al 2011a).…”

Intrastriatal hypoxanthine administration affects Na+,K+‐ATPase, acetylcholinesterase and catalase activities in striatum, hippocampus and cerebral cortex of rats

“…A common factor in Lesch–Nyhan is that patients exhibit a characteristically raised level of hypoxanthine in urine (Harkness et al, 1988), plasma and cerebrospinal fluid (Rosenbloom et al, 1967; Harkness et al, 1988; Puig and Mateos, 1993). Although the underlying mechanisms of brain dysfunction in Lesch–Nyhan are poorly understood, the accumulation of oxypurines such as hypoxanthine has been proposed to contribute to the neurological dysfunction present in this disease (Dasheiff, 1980; Kisch et al, 1985; Visser et al, 2000; Ma et al, 2001).…”