Prenatal diagnosis of X-linked chronic granulomatous disease by percutaneous umbilical blood sampling

In this study, we investigated how to prenatally diagnose X-linked chronic granulomatous disease (X-CGD) effectively and accurately. Percutaneous umbilical blood sampling was conducted in the 22nd week of pregnancy. NADPH oxidase activity and gp91(phox) protein expression of neutrophils were analysed using flow cytometry. Direct sequencing was used to detect CYBB gene mutations. Umbilical blood was obtained from seven foetuses whose mothers were X-CGD carriers. Six foetuses, whose mothers needed prenatal diagnosis because of other diseases, were used as control. The neutrophils in all 13 foetuses showed lower hydrogen peroxide generation (stimulation index < 100) and gp91(phox) protein expression. Among the seven foetuses whose mothers were X-CGD carriers, four foetuses (Family 2, 4, 5 and 7) had CYBB gene mutations and showed very low hydrogen peroxide generation (stimulation index < 10) and no gp91(phox) expression. The other three foetuses (Family 1, 3 and 6) had no CYBB gene mutations and showed stimulation index of 20-50 and partial or normal gp91(phox) protein expression (no difference with controls). Two of the three mothers (Family 1 and 3) have delivered healthy infants with normal hydrogen peroxide generation and expression of gp91(phox) in neutrophils. Combined with direct sequencing, dihydrorhodamine oxidation analysis and gp91(phox) protein detection is an effective and accurate method for prenatal screening for X-CGD.