Proliferation status defines functional properties of endothelial cells

Abstract

Homeostasis of solid tissue is characterized by a low proliferative activity of differentiated cells while special conditions like tissue damage induce regeneration and proliferation. For some cell types it has been shown that various tissue-specific functions are missing in the proliferating state, raising the possibility that their proliferation is not compatible with a fully differentiated state. While endothelial cells are important players in regenerating tissue as well as in the vascularization of tumors, the impact of proliferation on their features remains elusive. To examine cell features in dependence of proliferation, we established human endothelial cell lines in which proliferation is tightly controlled by a doxycycline-dependent, synthetic regulatory unit. We observed that uptake of macromolecules and establishment of cell-cell contacts was more pronounced in the growth-arrested state. Tube-like structures were formed in vitro in both proliferating and non-proliferating conditions. However, functional vessel formation upon transplantation into immune-compromised mice was restricted to the proliferative state. Kaposi's sarcoma-associated herpes virus (KSHV) infection resulted in reduced expression of endothelial markers. Upon transplantation of infected cells, drastic differences were observed: proliferation arrested cells acquired a high migratory activity while the proliferating counterparts established a tumor-like phenotype, similar to Kaposi Sarcoma lesions. The study gives evidence that proliferation governs endothelial functions. This suggests that several endothelial functions are differentially expressed during angiogenesis. Moreover, since proliferation defines the functional properties of cells upon infection with KSHV, this process crucially affects the fate of virus-infected cells.

An endothelial cell line infected by Kaposi’s sarcoma–associated herpes virus (KSHV) allows the investigation of Kaposi’s sarcoma and the validation of novel viral inhibitors in vitro and in vivo

“…After infection with KSHV the cells become spindle-like, lose endothelial properties and undergo transcriptional changes corresponding to EndMT, thereby reflecting the features as described for patients. Upon transplantation into immunocompromised mice the infected cells form lesions, histologically mimicking the lesions seen in Kaposi's sarcoma patients [18]. These properties suggest that this culture system may be suitable not only for the investigation of KSHV infection mechanisms but also for the validation of novel compounds.…”

“…The cells form 3D spheroids that can be maintained in presence of doxycycline without increase of cell number [17]. rKSHV-HuARLT cells [18] were derived upon infection of HuARLTs with recombinant KSHV.219 carrying the constitutively expressed GFP and puromycin selection genes as well as RFP under control of a viral lytic promoter [20].…”

“…We recently described conditionally immortalized human endothelial cells (HuARLT) that are permissive for KSHV infection [17,18]. The cell line preserves the important properties of primary endothelial cells in vitro and gives rise to functional blood vessels when transplanted into mice.…”

One Step Ahead: Herpesviruses Light the Way to Understanding Interferon-Stimulated Genes (ISGs)

“…In accordance with previous observations regarding the nuclease activity of UL41, this viral protein was shown to degrade ZAP mRNA during HSV-1 infection. Correspondingly, growth of a mutant virus *These studies used iSLK or HuARLT2 cells that were latently infected with recombinant rKSHV.219 (Vieira and O'Hearn, 2004;Myoung and Ganem, 2011;Lipps et al, 2017). HFF, human foreskin fibroblasts; HF, human fibroblasts.…”

Function or Expansion: How to Investigate Cells of Human Blood Vessels

“…Bottom: to see how much the cells proliferated, the cells in each dish were counted on days 1, 4, 6, 8, 11, and 13. In the presence of doxycycline (green line), the number of cells on the dish were increased over time; in contrast, in the absence of doxycycline (orange line), cells did not divide and the cell number stayed the same [1].…”