apyrase and Fibrosarcoma

Tumour cells activate and aggregate platelets [tumour cell-induced platelet aggregation (TCIPA)] and this process plays an important role in the successful metastasis of cancer cells. To date, most studies on TCIPA have been conducted under no-flow conditions. In this study, we have investigated TCIPA in real time under flow conditions, using an ultrasound standing wave trap that allows formation and levitation of cancer cell clusters in suspension, thus mimicking the conditions generated by flowing blood.. Using 59M adenocarcinoma and HT1080 fibrosarcoma cells and human platelets, cancer cell cluster-platelet aggregates were imaged in real time using epi-fluorescence microscopy (F-actin) and investigated in detail using confocal microscopy (matrix metalloproteinase-2-GPIIb/IIIa co-localization) and scanning electron and helium-ion microscopy (<1 nm resolution). The release of gelatinases from aggregates was studied using zymography.. We found that platelet activation and aggregation takes place on the surface of cancer cells (TCIPA), leading to time-dependent disruption of cancer cell clusters. Pharmacological modulation of TCIPA revealed that EDTA, prostacyclin, o-phenanthroline and apyrase significantly down-regulated TCIPA and, in turn, delayed cell cluster disruption, However, EGTA and aspirin were ineffective. Pharmacological inhibition of TCIPA correlated with the down-regulation of platelet activation as shown by flow-cytometry assay of platelet P-selectin.. Our results show for the first time, that during TCIPA, platelet activation disrupts cancer cell clusters and this can contribute to metastasis. Thus, selective targeting of platelet aggregate-cancer cell clusters may be an important strategy to control metastasis.

Topics: Actins; Adenocarcinoma; Apyrase; Blood Platelets; Cell Communication; Cell Line, Tumor; Down-Regulation; Edetic Acid; Epoprostenol; Female; Fibrosarcoma; Flow Cytometry; Humans; Matrix Metalloproteinase 2; Microscopy, Confocal; Microscopy, Fluorescence; Ovarian Neoplasms; P-Selectin; Phenanthrolines; Platelet Activation; Platelet Aggregation; Platelet Aggregation Inhibitors; Platelet Glycoprotein GPIIb-IIIa Complex; Tumor Cells, Cultured; Ultrasonics; Ultrasonography

We established five clones (ML-01, ML-02, MH-01, MH-02, MH-03) from murine 3-methylcholanthrene-induced fibrosarcoma A (Meth A), and investigated their experimental metastatic potentials in relation to their platelet-aggregating activities. A clone with a high metastatic potential (MH-02) showed a characteristic biphasic pattern of platelet aggregation, of which the first peak was not present in the aggregation patterns of the clone with low metastatic potential (ML-01). The first peak was eliminated by treatment of the cells with apyrase, indicating that adenosine diphosphate (ADP) was the causative substance of this particular peak. The metastatic potential of clones correlated well with the ADP concentration of the culture media. These results suggest that the increased ADP production and consequential enhancement of platelet-aggregating activity are closely related to the increment of pulmonary metastatic potential of MH-02 clone.

Topics: Animals; Apyrase; Fibrosarcoma; Hot Temperature; Lung Neoplasms; Male; Mice; Mice, Inbred BALB C; Microbial Collagenase; NADP; Neoplasm Metastasis; Neuraminidase; Platelet Aggregation; Trypsin; Tumor Cells, Cultured; Ultracentrifugation

In a previous study we found a correlation between metastatic potential and platelet aggregating activity in sublines of a benzopyrene-induced murine fibrosarcoma ( mFS6 ); the purpose of the present work was to elucidate the role of thromboxane biosynthesis by platelets and/or by neoplastic cells in the activation of platelets in this system. The cells of the more malignant subline induced higher aggregation and TxB2 production than those of the non metastasizing one. The supernatants of aggregating cell suspensions contained very few TxB2; furthermore, preincubation of platelets with ASA or Apyrase resulted in inhibition of aggregation and TxB2 production, while preincubation of the cells was ineffective; these results suggest the platelet origin of the measured TxB2 and indicate that platelet-derived ADP plays an important role in their activation, while the production of ADP by the cells does not seem to be relevant in this model. The involvement of platelet prostaglandin biosynthesis pathway in neoplastic cell induced platelet activation could play an important role in the development of platelet-dependent tumour metastasis.

Topics: Animals; Apyrase; Aspirin; Blood Platelets; Cell Line; Cell Transformation, Neoplastic; Fibrosarcoma; Humans; Mice; Platelet Aggregation; Thromboxane B2; Thromboxanes