Analysis of the potential of cancer cell lines to release tissue factor-containing microvesicles: correlation with tissue factor and PAR2 expression

Abstract

BackgroundDespite the association of cancer-derived circulating tissue factor (TF)-containing microvesicles and hypercoagulable state, correlations with the incidence of thrombosis remain unclear.MethodsIn this study the upregulation of TF release upon activation of various cancer cell lines, and the correlation with TF and PAR2 expression and/or activity was examined. Microvesicle release was induced by PAR2 activation in seventeen cell lines and released microvesicle density, microvesicle-associated TF activity, and phoshpatidylserine-mediated activity were measured. The time-course for TF release was monitored over 90 min in each cell line. In addition, TF mRNA expression, cellular TF protein and cell-surface TF activities were quantified. Moreover, the relative expression of PAR2 mRNA and cellular protein were analysed. Any correlations between the above parameters were examined by determining the Pearson’s correlation coefficients.ResultsTF release as microvesicles peaked between 30–60 min post-activation in the majority of cell lines tested. The magnitude of the maximal TF release positively correlated with TF mRNA (c = 0.717; p < 0.001) and PAR2 mRNA (c = 0.770; p < 0.001) expressions while the percentage increase correlated with PAR2 mRNA (c = 0.601; p = 0.011) and protein (c = 0.714; p < 0.001). There was only a weak correlation between resting TF release, and microvesicle release. However, TF release in resting cells did not significantly correlate with any of the parameters examined. Furthermore, TF mRNA expression correlated with PAR2 mRNA expression (c = 0.745; p < 0.001).Discussion and ConclusionsIn conclusion, our data suggest that TF and PAR2 mRNA, and PAR2 protein are better indicators of the ability of cancer cells to release TF and may constitute more accurate predictors of risk of thrombosis.

Microvesicles and Cancer Associated Thrombosis

“…Cultured human tumor cell lines of different origins express variable levels of TF and release cancer MVs with various procoagulant potentials in their supernatant. 31,32 As illustrated for breast cancer, according to the cell line, mechanisms involve TF-independent pathways which promote platelet activation and aggregation and/or TF-dependent thrombin generation. 33 Evidence of MVs involvement in thrombus formation in vivo is supported by different studies using endogeneously formed or exogeneously injected MVs and mice mouse models of thrombosis.…”

The role of tissue factor in metastasising, neoangiogenesis and hemostasis in cancer

“…тканевый фактор и нарушения системы гемостаза. Культуры клеточных линий рака молочной железы MDA-MB-231 и MCF-10A экспрессируют TF, и он является прокоагулянтно активным [85,86]. При этом метастазирующая клеточная линия MDA-MB-231 активирует свертывание лучше, чем неметастазирующая MCF-10A [86].…”

“…Концентрация везикул коррелирует с клинической стадией опухоли: она выше для III и IV стадий по сравнению с I и II [56]. TF-положительные микровезикулы в плазме пациентов имеют моноцитарную и тромбоцитарную природу [59], а также могут быть испущены опухолевыми клетками [85]. Несмотря на то что в норме тромбоциты не имеют детектируемого TF на поверхности [95], исследователи утверждают, что TF на тромбоцитарных везикулах может быть синтезирован другими клетками, а впоследствии перенесен на тромбоциты [59].…”

“…заболеваниями источником TF на тромбоцитарных везикулах могут быть опухолевые клетки [59], поскольку эти клетки также способны испускать TF-положительные микровезикулы [85,97].…”

“…тканевый фактор и нарушения системы гемостаза. Для аденокарциномы толстой кишки на клеточной линии Caco-2 было показано, что данные клетки экспрессируют прокоагулянтно активный TF, а также способны испускать TF-положительные прокоагулянтные везикулы в ответ на стимуляцию рецептора PAR2 пептидом SLIGRL [85].…”