2 ANALYSIS OF METHYLATION PATTERN IN FEMALES WITH LOW FACTOR VIII: C LEVEL

Abstract

The analysis of X chromosome activation through the metilation pattern could be useful in the study of affected females for X-linked recessive diseases, such as Hemophilia A (HA). Low FVIII;C level in female can be due to: acquired inhibitors to FVIII, X chromosome abnormalities, homozygosity at the hemophilia locus, extreme lyonization in an heterozygote. It is possible to reveal a non random inactivation of X chromosomes by hybridization of genomic DNA with M27ß probe (locus DXS 255). We analyzed with this probe 3 females with low FVIII:C levels (range 2%-5%). G.V. is a 33 year old female (FVIIhC 5%, FVIIIvW:Ag 80%) born from a HA carrier and a normal man. Karyotype was 46,XX and no inhibitors against FVIII were detected. The study with M27ß probe showed that only the paternal X chromosome was inactive. HA in this female is probably due to a non random X-inactivation. P.M. is a 48 year old female (FVIII:C 12%; FVIIIvW:Ag 100%) born from first cousins parents. Her father and brother are hemophilics (FVIII:C 11%) and her mother was a carrier. Cytogenetic studies revealed no abnormalities. M27ß analysis showed a balanced lyonization. An homozygosity for the HA defective gene, was demonstrated by the aplotype analysis with RFLPs. P.T. is a 19 year old female (FVIII:C 2.2%; FVIIIvW:Ag 73%) with Turner Syndrome (46,X,-idic(X)) born from healthy unrelated parents without history of bleeding disorders. The RFLP studies showed the maternal origin of idic(X). The methylation pattern obtained by M27ß probe confirmed the whole inactivation of the abnormal X therefore we hypothesized a mutation in FVIII gene in the normal X coming from her healthy father. In conclusion, the possibility to demonstrate the degree of X-chromosome activation is a very important and useful tool in understanding the pattern of HA carriers.