Chimaeric mice with disruption of the gene coding for phosphatidylinositol glycan class A (Pig-a) were defective in embryogenesis and spermatogenesis.

Abstract

Mutations in the gene encoding PIG-A (phosphatidylinositol glycan class A) are found in patients with paroxysmal nocturnal haemoglobinuria (PNH), an acquired haematopoietic stem cell disorder. Individuals with hereditary PIG-A mutations have never been identified, which is also manifested by the difficulties in generating Pig-a knockout (KO) mice. This study investigated the effect of Pig-a mutations on the development of visceral and genital organs in addition to the haematopoietic system by the generation of Pig-a KO chimaeric mice. Of a total of 54 live births out of 1684 blastocysts injected, chimaerism for Pig-a knockout was detected in 29 mice, suggesting the importance of Pig-a in embryogenesis and in live birth. Quantification of the degree of chimaerism in different organs of the surviving chimaeric mice revealed extremely low levels of Pig-a KO cells in the liver and spleen. In contrast, high levels of KO signals were usually detected in the brain, heart, lung and kidney. Haematopoiesis proceeded normally in these chimaeric mice (as measured by 'complete blood cell counting') and the Pig-a KO cells were present at low levels in red blood cells and B lymphocytes but at high levels in T lymphocytes, although these KO cells did not gain any growth advantage. The effect of Pig-a knockout was also prominent in the reproductive system, another organ with high mitotic activity. Breeding the male chimaeras revealed a high rate of infertility and abnormality in the male genital organs, including abnormally shaped testes, epididymis and seminal vesicles. Even in the absence of gross abnormalities of the genital organs, low counts of motile sperm were also discernible. Pig-a KO sperm was detected in these organs; however, no transmission of the KO allele was observed. The results suggest a possible mechanism underlying the non-transmission of the Pig-a KO gene in germlines.