PMC8295588_01
(A) The pedigree of the patient's family. III.2: female carrier; III.3, III.4, and III.5: affected male; IV.2: female proband. (B) Sanger sequencing of the WAS gene of c.173C>T. The proband and her mother were heterozygous of the mutated gene, and her older brother and her father were normal. (C) WASp expression levels in the PBMCs by flow cytometry assay. The proportion of M1 area was 98.5% of normal control, 50.6% of her mother and 20.6% of the proband, respectively. (D) WASp expression levels were analyzed by western blotting. The gray mean value of the proband's mother is 10.8% of the control level and the gray mean value of proband is 5.4% of the level. (E) X-chromosome inactivation analysis of the patient and her parents. The experiments were performed in triplicate.
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F1
Upon investigating her family history, her father and elder brother showed no symptoms (Figure 1A), while her mother showed decreased IgM and IgA levels, with normal PLT. Sanger sequencing revealed a heterozygous missense mutation in exon 2 of the WAS gene, leading to a change from proline to leucine at position 58 in the patient and her mother (Figure 1B). Meanwhile, flow cytometry showed the level of WASp from the proband and in the cells from her mother at 20.6 and 50.4%, respectively (Figure 1C). WASp was reduced by ~5-fold in mononuclear cells from the proband compared to the normal control (98.5%). Western blot analysis revealed significantly decreased WASp levels in the patient and her mother compared to normal control (Figure 1D). The results showed that the proband had only one paternal allele amplified after HpaII digestion (Figure 1E). In contrast, her mother showed two bands in the presence of HpaII; however, the rate of random inactivation indicated a mildly skewed pattern of XCI in mature blood cells (Figure 1E).
Platelet fluctuations of the patient within one and a half years (13-Feb-2019-26-Aug-2020) after admission.
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F2
Her platelet count fluctuated between 30x 109/L and 60x 109/L within one and a half years after admission (Figure 2).