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Anagrelide Modulates Proplatelet Formation Resulting in Decreased Number and Increased Size of Platelets.

ABSTRACT: We retrospectively evaluated 48 essential thrombocythemia (ET) patients who were treated in our institute (male/female, 14/34, median age, 61.5 years). In 14 patients treated with anagrelide (ANA), the degree of platelet count reduction (median, -56.6%) was strongly correlated with increase of mean platelet volume (MPV) (median, +11.7%) (R = 0.777). This correlation was not observed in ET patients treated with hydroxycarbamide alone (R = 0.245). The change in size of platelets strongly suggested that ANA affected the final process of platelet production. Thus, we hypothesized that ANA modifies the process by which platelets are released from proplatelets. To verify the association in an in vitro setting, we compared MEG-01 cells treated with PMA ± ANA. The number of platelet-like particles (PLPs) was decreased (P < 0.05) and the size of PLPs estimated by using flow cytometry was significantly increased when MEG-01 cells were treated with PMA + ANA (P < 0.05 vs PMA alone), recapitulating the clinical findings. The cytoplasmic protrusions extending from MEG-01 cells were shorter and thicker and the number of proplatelets was decreased when MEG-01 cells were treated with PMA + ANA (P < 0.01 vs PMA alone). Western blotting analysis showed that ANA treatment resulted in increased phosphorylation of MLC2 and reduced phosphorylation of focal adhesion kinase (FAK). The morphological change of proplatelets were reversed by blebbistatin, a specific inhibitor of myosin II. These findings indicated that ANA modulates the FAK-RhoA-ROCK-MLC2-myosine IIA pathway and suppresses proplatelet maturation, leading to a decrease in platelet count and increase in MPV.

SUBMITTER: Miyashita N

PROVIDER: S-EPMC6745917 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundInsertion force has been shown to significantly reduce chondrocyte viability during osteochondral allograft transplantation. How graft size influences the required insertion force and chondrocyte viability has yet to be determined. Hypothesis/Purpose: The purpose was to characterize how graft size influences insertion force requirements and chondrocyte viability during osteochondral transplantation. The hypothesis was that larger grafts would require greater force and reduce chondrocyte viability.Study designControlled laboratory study.MethodsFour graft sizes-15 × 5 mm, 15 × 10 mm, 25 × 5 mm, and 25 × 10 mm (diameter × depth)-were harvested from 13 thawed fresh-frozen human cadaveric distal femurs. Average, maximum, and cumulative force and number of impacts were recorded for 44 grafts by a surgical mallet embedded with a calibrated force sensor. In a separate experiment, fresh osteochondral tissues were subjected to mechanical loading. To capture a range of clinically important forces, categories were selected to correspond to impaction force data. Chondrocyte viability was assessed with confocal laser microscopy and live/dead staining.ResultsTotal force for all grafts averaged 4576 N. Median number of impacts for all grafts was 20 (range, 7-116). The mean number of impacts for 5-mm-deep grafts was 14.2 (95% CI, 10.8-18.6), as compared with 26.3 (95% CI, 19.9-34.4) for 10-mm-deep grafts ( P < .001). The mean cumulative force for 5-mm-deep grafts was 2128 N (95% CI, 1467-3087), as opposed to 4689 N (95% CI, 3232-6803) for 10-mm-deep grafts ( P = .001). For every 1 mm in graft depth, an average of 13.1% (95% CI, 6.2%-20.3%) more impacts are required when controlling for diameter and density ( P < .001). For every 1 mm in graft depth, the force required increases on average by 17.1% (95% CI, 7.7%-27.4%) when controlling for diameter and density ( P = .001). There was a significant reduction in chondrocyte viability for the forces required for graft thickness values >10 mm. Only forces associated with graft thickness <10 mm had chondrocyte viabilities consistently >70%.ConclusionInsertion force increases significantly with increasing graft depth. Controlling for diameter and bone density, a 1-mm increase in graft depth is associated with 13.1% more impacts and 17.1% more force. Chondrocyte viability was significantly reduced to <70% at average forces associated with grafts thicker than 10 mm.Clinical relevanceBased on the current data, graft depth is an important consideration for surgeons when sizing osteochondral allograft transplant for chondral lesions of the knee.

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Anagrelide Modulates Proplatelet Formation Resulting in Decreased Number and Increased Size of Platelets.

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