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The objectives of this study were to determine in humans the relationship between sex chromosome loss and increased micronucleus frequencies with age, the establishment of sex chromosome loss frequencies unbiased by cellular survival factors, the involvement of kinetochore proteins in sex chromosome loss and the effect of smoking. Additionally, this study was to establish the involvement of the active and inactive X chromosomes in micronucleus formation. Blood samples were obtained from 38 adult females, 8 newborn females, 35 adult males, 18 newborn males, 2 females with stable X;9 translocations and 7 Turner syndrome individuals. Lymphocytes were separated, cultured and blocked with cytochalasin B. A modified micronucleus assay was used to determine the kinetochore status of each micronucleus. Fluorescent in situ hybridization (FISH) was then utilized to determine if micronuclei contained either the X or Y chromosomes, or the active or inactive X chromosome. Kinetochore labeling showed 59.0% of the micronuclei to be kinetochore-positive for females, whereas only 23.8% of the male micronuclei were kinetochore-positive. FISH showed 72.2% of the female micronuclei contained at least one X chromosome. Male micronuclei contained the Y chromosome 13.0% of the time. For both males and females there was a significant increase in sex chromosome-positive micronuclei with age. Further analysis showed 64.9% that kinetochore-negative micronuclei in females contained an X chromosome while 87.8% of the Y chromosome-positive micronuclei were previously scored as kinetochore-negative. The active X chromosome was observed 6.7% to 33.3% of the time in those micronuclei scored specificly for the active X chromosome. Further, the inactive X chromosome was incorporated into 83.3% of the X chromosome-positive micronuclei observed from t(X;9) females. The relationship between sex chromosome-positive micronuclei and kinetochore-negative micronuclei suggests that a dysfunctional kinetochore or centromere is the likely cause of chromosome lagging which leads to the production of micronuclei. The low frequency of active X chromosome-positive micronuclei suggests that there is preferential involvement of the inactive X chromosome in micronucleus formation.