Paroxysmal nocturnal hemoglobinuria is a clonal disease caused by PIG-A mutation of hematopoietic stem cells. At present, there is no suitable PNH animal model for basic research, therefore, it is urgent to establish a stable animal model. We constructed a Pig-a conditional knock-out mice model by ES targeting technique and Vav-iCre. The expressions of GPI and GPI-AP were almost completely absent in CKO homozygote mice, and the proportion of the deficiency remained stable from birth. In CKO heterozygote mice, the proportion of the deficiency of GPI and GPI-AP was partially absent and decreased gradually from birth until it reached a stable level at 3 months after birth and remained there for life. Compared with normal C57BL/6N mice and Flox mice, pancytopenia was found in CKO homozygous mice, and leukopenia and anemia were found in CKO heterozygotes mice. Meanwhile, in CKO mice, the serum LDH, TBIL, IBIL, complement C5b-9 levels were increased, and the concentration of plasma FHb was increased. Hemosiderin granulosa cells can be seen more easily in the spleens of CKO mice. What’s more, CKO mice had stable transcription characteristics. In conclusion, our mouse model has stable GPI-deficient and mild hemolysis, which may be an ideal in vivo experimental model for PNH.

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Paroxysmal nocturnal hemoglobinuria (PNH) is a disease involving hematopoietic stem cell membrane defects caused by acquired phosphatidylinositol glycan anchor biosynthesis class A (PIGA) mutations. In this study, 97 target genes were selected as a target gene panel and screened in 23 PNH patients via the sequencing of specific DNA target regions. Through functional analysis, we identified that suppressor-of-Zeste 12 (SUZ12) may be involved in the proliferation of PNH clones. mRNA and protein expression levels of SUZ12 and the trimethylation level of histone H3 at lysine 27 (H3K27) in CD59peripheral blood leukocytes from PNH patients were higher than those in CD59 + cells from PNH patients and peripheral blood leukocytes from healthy controls.In addition, the relative expression of SUZ12 in PNH patients was positively correlated with Ret% and the proportion of PNH clones. When we knocked down SUZ12 expression in a PIGA knockdown THP-1 cell line (THP-1 KD cells), the trimethylation of histone H3K27(H3K27me3) and cell proliferation decreased, apoptosis increased, and cell cycle arrest occurred in G0/G1 phase. In conclusion, SUZ12 participates in the proliferation of PNH clones by regulating histone H3K27me3 levels. Our results may provide new therapeutic targets and possibilities for PNH patients.

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