Title

 

Authors

S. De Iudicibus¹, E. Cuzzoni², F. Bartoli¹, A. Ventura¹, G. Decorti<sup>2

1</sup>Department of Reproductive and Developmental Sciences and IRCCS, Istituto per l'Infanzia Burlo Garofolo, Clinica Pediatrica, Trieste, Italy; ²Department of Life Sciences, University of Trieste, Trieste, Italy

 

Abstract

Glucocorticoids (GCs) are used to induce remission in patients with chronic inflammation diseases, however a considerable variability in response to these drugs is evident in paediatric subjects. GCs exert their biological effects through binding to the GC receptor, which regulates either positively or negatively the expression of target genes. Polymorphisms in the GC receptor gene (NR3C1) have been describedwithin the normal population.An association between an intronic polymorphism in this gene, and GC response in paediatric patients with Inflammatory Bowel Diseases (IBD) has been described (De Iudicibus et al, 2011).  BclI single nucleotide polymorphism (SNP) at position 647 in intron 2, was significantly more frequent in paediatric patients who exhibited a good response to GC treatment, than in patients that did not respond to these agents. The aim of this study was to evaluate the relationship between individual variations in the anti-proliferative potency of methylprednisolone and  BclI genotype on lymphocytes obtained from healthy subjects. The study was conduct on 42 buffy coats of healthy donors (35 males and 7 females; mean age 42.0  years, range 18-64 years). The effect of methyl-prednisolone on the proliferation of peripheral blood mononuclear cells was determined by [methyl-h3] thymidine incorporation into the proliferating lymphocytes. The drug concentration that would give 50% of lymphocyte inhibition (IC50) was determined from the sigmoidal dose response curve. All subjects were genotyped for BclI polymorphism using TaqMan^® genotyping technologies. From the dose response curves, the methyl-prednisolone IC50 was determined for each subjects: (median 1.4x10^-7 M; mean 1.2x10^-6 M; range 7.4x10^-10 M - 2.9x10^-4 M). The frequency of the mutated BclI genotype in our study group was comparable with literature data (11.9%). Subjects with the mutated BclI genotype displayed significantly increased methyl-prednisolone responsiveness (median IC50 2.4x10^-9 M) compared to nonmutated carriers (median IC50 2.8 X10^-7 M, Wilcoxon test p value=0.003). In conclusion, we have developed a simple and reproducible in vitro assay to study GC effect on lymphocyte proliferation. This in vitro test could be easily applied to predict the interindividual GCs sensitivity in chronic inflammatory diseases before starting the clinical treatment. Moreover, we have confirmed also with the in vitro assay a relation between BclI mutated genotype and increased GC sensitivity. These results suggested that BclI polymorphism associated with a lymphocytes proliferation assay could therefore represent a useful diagnostic tool to predict the  clinical response to steroid treatment in patients with chronic inflammation diseases.
De Iudicibus et al. (2011), J Clin Gastroenterol.45(1): e1-7