tretinoin and Lesch-Nyhan-Syndrome

Neurological manifestations in Lesch-Nyhan disease (LND) are attributed to the effect of hypoxanthine-guanine phosphoribosyltransferase (HPRT) deficiency on the nervous system development. HPRT deficiency causes the excretion of increased amounts of hypoxanthine into the extracellular medium and we hypothesized that HPRT deficiency related to hypoxanthine excess may then lead, directly or indirectly, to transcriptional aberrations in a variety of genes essential for the function and development of striatal progenitor cells. We have examined the effect of hypoxanthine excess on the differentiation of neurons in the well-established human NTERA-2 cl.D1 (NT2/D1) embryonic carcinoma neurogenesis model. NT2/D1 cells differentiate along neuroectodermal lineages after exposure to retinoic acid (RA). Hypoxanthine effects on RA-differentiation were examined by the changes on the expression of various transcription factor genes essential to neuronal differentiation and by the changes in tyrosine hydroxylase (TH), dopamine, adenosine and serotonin receptors (DRD, ADORA, HTR). We report that hypoxanthine excess deregulate WNT4, from Wnt/β-catenin pathway, and engrailed homeobox 1 gene and increased TH and dopamine DRD1, adenosine ADORA2A and serotonin HTR7 receptors, whose over expression characterize early neuro-developmental processes.

Topics: Adenosine; Cell Culture Techniques; Cell Differentiation; Cell Line, Tumor; Homeodomain Proteins; Humans; Hypoxanthine Phosphoribosyltransferase; Lesch-Nyhan Syndrome; Neurons; Receptors, Dopamine D1; Tretinoin; Tyrosine 3-Monooxygenase; Wnt Signaling Pathway; Wnt4 Protein

Lesch-Nyhan disease (LND), caused by complete deficiency of hypoxanthine guanine phosphoribosyltransferase (HPRT), is characterized by a neurological deficit, the etiology of which is unknown. Evidence has accumulated indicating that it might be related to dysfunction of the basal ganglia with a prominent loss of striatal dopamine fibers. Guanine nucleotide depletion has been shown to occur in cells from Lesch-Nyhan patients. In this study we demonstrate that chronic guanine nucleotide depletion induced by inhibition of inosine monophosphate dehydrogenase with low levels (50 nM) of mycophenolic acid (MPA) lead human neuroblastoma cell lines to differentiate toward the neuronal phenotype. The MPA-induced morphological changes were more evident in the dopaminergic line LAN5, than in the cholinergic line IMR32. MPA-induced differentiation, unlike that induced by retinoic acid, caused a less extensive neurite outgrowth and branching (similar to that observed in cultured HPRT-deficient dopaminergic neurons) and involved up-regulation of p53, p21 and bax, and bcl-2 down-regulation without p27 protein accumulation. These results suggest that guanine nucleotide depletion following HPRT deficiency, might lead to earlier and abnormal brain development mainly affecting the basal ganglia, displaying the highest HPRT activity, and could be responsible for the specific neurobehavioral features of LND.

Topics: Basal Ganglia; Basal Ganglia Diseases; Cell Cycle Proteins; Cell Differentiation; Cell Enlargement; Cell Line; Enzyme Inhibitors; Guanine Nucleotides; Humans; Hypoxanthine Phosphoribosyltransferase; IMP Dehydrogenase; Lesch-Nyhan Syndrome; Models, Neurological; Mycophenolic Acid; Neurites; Neuroblastoma; Neurofilament Proteins; Tretinoin; Up-Regulation

5'-Aminoimidazole-4-carboxamide riboside (AICA riboside) has been previously shown to be toxic to two neuronal cell models [Neuroreport 11 (2000) 1827]. In this paper we demonstrate that AICA riboside promotes apoptosis in undifferentiated human neuroblastoma cells (SH-SY5Y), inducing a raise in caspase-3 activity. In order to exert its effect on viability, AICA riboside must enter the cells and be phosphorylated to the ribotide, since both a nucleoside transport inhibitor, and an inhibitor of adenosine kinase produce an enhancement of the viability of AICA riboside-treated cells. Short-term incubations (2 h) with AICA riboside result in five-fold increase in the activity of AMP-dependent protein kinase (AMPK). However, the activity of AMPK is not significantly affected at prolonged incubations (48 h), when the apoptotic effect of AICA riboside is evident. The results demonstrate that when the cell line is induced to differentiate both toward a cholinergic phenotype (with retinoic acid) or a noradrenergic phenotype (with phorbol esters), the toxic effect is significantly reduced, and in the case of the noradrenergic phenotype differentiation, the riboside is completely ineffective in promoting apoptosis. This reduction of effect correlates with an overexpression of Bcl-2 during differentiation. AICA riboside, derived from the hydrolysis of the ribotide, an intermediate of purine de novo synthesis, is absent in normal healthy cells; however it may accumulate in those individuals in which an inborn error of purine metabolism causes an increase in the rate of de novo synthesis and/or an overexpression of cytosolic 5'-nucleotidase, that appears to be the enzyme responsible for AICA ribotide hydrolysis. In fact, 5'-nucleotidase activity has been shown to increase in patients affected by Lesch-Nyhan syndrome in which both acceleration of de novo synthesis and accumulation of AICA ribotide has been described, and also in other neurological disorders of unknown etiology. Our results raise the intriguing clue that the neurotoxic effect of AICA riboside on the developing brain might contribute to the neurological manifestations of syndromes related to purine dismetabolisms.

Topics: 5'-Nucleotidase; Acetylcholine; Aminoimidazole Carboxamide; Apoptosis; Brain; Caspase 3; Caspases; Cell Differentiation; Cells, Cultured; Dipyridamole; Enzyme Inhibitors; Humans; Lesch-Nyhan Syndrome; Neuroblastoma; Neurons; Norepinephrine; Phorbol Esters; Proto-Oncogene Proteins c-bcl-2; Purines; Ribonucleosides; Ribose-Phosphate Pyrophosphokinase; Tretinoin