Kaposi's sarcoma-associated herpesvirus induction of chromosome instability in primary human endothelial cells

Abstract

Chromosome instability contributes to the multistep oncogenesis of cancer cells. Kaposi's sarcoma (KS), an angiogenic vascular spindle cancer of endothelial cells, displays stage advancement with lesions at early stage being hyperproliferative, whereas lesions at late stage are clonal or multiclonal and can exhibit a neoplastic nature and chromosome instability. Although infection with KS-associated herpesvirus (KSHV) has been associated with the initiation and promotion of KS, the mechanism of KS neoplastic transformation remains unclear. We show that KSHV infection of primary human umbilical vein endothelial cells induces abnormal mitotic spindles and centrosome duplication. As a result, KSHV-infected cells manifest chromosome instability, including chromosomal misalignments and laggings, mitotic bridges, and formation of micronuclei and multinucleation. Our results indicate that KSHV infection could predispose cells to malignant transformation through induction of genomic instability and contributes to the development of KS.

Figure 1

Induction of abnormal mitotic spindles and centrosome numbers and chromosomal misalignments during Kaposi’s sarcoma-associated herpesvirus (KSHV) infection of human umbilical vein endothelial cells (HUVECs). A, 4′,6-diamidino-2-phenylindole (DAPI) and α-tubulin staining shows normal mitotic spindles and chromosomal alignments in mock-infected HUVECs (a) and abnormal mitotic spindles and chromosomal misalignments in three different representative fields of KSHV-infected HUVECs (b– d). B, DAPI and γ-tubulin staining shows abnormal centrosome numbers and chromosomal misalignments in three different representative fields of KSHV-infected HUVECs. C, quantification of abnormal mitotic spindles in mock- or KSHV-infected mitotic cells.

Fig. 2

Induction of chromosomal laggings and mitotic bridges during Kaposi’s sarcoma-associated herpesvirus (KSHV) infection of human umbilical vein endothelial cells (HUVECs). A, 4′,6-diamidino-2-phenylindole (DAPI) and α-tubulin staining shows chromosomal laggings (b and c) and mitotic bridges (c–f) in KSHV-infected cells. Some cells have chromosomal laggings and mitotic bridges (c). Mock-infected cells have neither chromosomal laggings nor mitotic bridges (a). B, quantification of chromosomal laggings in mock- or KSHV-infected mitotic cells. C, quantification of mitotic bridges in mock- or KSHV-infected mitotic cells.

Fig. 3

Induction of micronuclei during Kaposi’s sarcoma-associated herpesvirus (KSHV) infection of human umbilical vein endothelial cells (HUVECs). A, micronuclei detected by 4′,6-diamidino-2-phenylindole staining of mock-infected HUVECs (a) or HUVECs infected by KSHV (b). B, quantification of micronuclei in mock- or KSHV-infected cells.

Fig. 4

Induction of multinucleation during Kaposi’s sarcoma-associated herpesvirus (KSHV) infection of human umbilical vein endothelial cells (HUVECs). A, 4′,6-diamidino-2-phenylindole (DAPI) and α-tubulin staining shows multinucleation in KSHV-infected cells (b) but not in mock-infected cells (a). B, quantification of multinucleation in mock- or KSHV-infected cells. C, DAPI and γ-tubulin staining shows abnormal centrosome numbers in cells with multinucleation (a–d represent four different fields).

Fig. 5

Multistep oncogenesis model for Kaposi’s sarcoma-associated herpesvirus (KSHV) induction of Kaposi’s sarcoma.