Platelets are classified as terminally differentiated cells that are incapable of cellular division. However, we observe that anucleate human platelets, either maintained in suspension culture or captured in microdrops, give rise to new cell bodies packed with respiring mitochondria and ␣-granules. Platelet progeny formation also occurs in whole blood cultures. Newly formed platelets are structurally indistinguishable from normal platelets, are able to adhere and spread on extracellular matrix, and display normal signaldependent expression of surface Pselectin and annexin V. Platelet progeny formation is accompanied by increases in biomass, cellular protein levels, and protein synthesis in expanding populations. Platelet numbers also increase during ex vivo storage. These observations indicate that platelets have a previously unrecognized capacity for producing functional progeny, which involves a form of cell division that does not require a nucleus. Because this new function of platelets occurs outside of the bone marrow milieu, it raises the possibility that thrombopoiesis continues in the bloodstream. (Blood. 2010;115(18):3801-3809) IntroductionAfter they are shed from the cytoplasm of megakaryocytes, 1 platelets circulate in the bloodstream for 9 to 11 days. There is no evidence that these anucleate cytoplasts undergo cellular division, but recent studies by our group and others have identified unexpected cellular functions of platelets, 2-4 including the capacity to process pre-mRNA 2-5 and translate mRNA into protein. [6][7][8][9][10] Platelets also continue to synthesize protein for several days when they are stored ex vivo. 11 These findings indicate that, despite their terminally differentiated state, platelets are biosynthetically more sophisticated than previously thought. 12 They also suggest that platelets may be able to adjust their phenotypic composition in response to environmental cues.Here we show that platelets give rise to new cells that are structurally and functionally similar to their parent counterparts. The formation of platelet progeny is associated with increases in platelet biomass, protein synthetic events, and total intracellular protein. This heretofore undescribed proliferative capacity of platelets may have significant consequences for normal and pathologic thrombopoiesis in humans in addition to having clinical implications for transfusion medicine. Methods Platelet isolation and cultureAll studies were approved by the University of Utah Institutional Review Board committee (no. 392). Leukocyte-depleted platelets were isolated as previously described. 2,5 Washed platelets were resuspended at 100 000/L in serum-free M199 medium, placed in round-bottom polypropylene tubes (BD Biosciences), and cultured in a 37°C humidified incubator under gentle rotation (MacsMix, slow, 45°angle; Miltenyi). The same suspension culture conditions were also used for the whole blood studies shown in Figure 2A.Stored platelets were obtained from the ARUP Blood Transfusion Services at the University o...

Megakaryocytes transfer a diverse and functional transcriptome to platelets during the final stages of thrombopoiesis. In platelets, these transcripts reflect the expression of their corresponding proteins and, in some cases, serve as a template for translation. It is not known, however, if megakaryocytes differentially sort mRNAs into platelets. Given their critical role in vascular remodeling and inflammation, we determined whether megakaryocytes selectively dispense transcripts for matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) into platelets. Next-generation sequencing (RNA-Seq) revealed that megakaryocytes express mRNA for 10 of the 24 human MMP family members. mRNA for all of these MMPs are present in platelets with the exception of MMP-2, 14, and 15. Megakaryocytes and platelets also express mRNA for TIMPs 1-3, but not TIMP-4. mRNA expression patterns predicted the presence and, in most cases, the abundance of each corresponding protein. Nonetheless, exceptions were observed: MMP-2 protein is present in platelets but not its transcript. In contrast, quiescent platelets express TIMP-2 mRNA but only traces of TIMP-2 protein. In response to activating signals, however, platelets synthesize significant amounts of TIMP-2 protein. These results demonstrate that megakaryocytes differentially express mRNAs for MMPs and TIMPs and selectively transfer a subset of these into platelets. Among the platelet messages, TIMP-2 serves as a template for signal-dependent translation. (Blood. 2011;118(7): 1903-1911) IntroductionThe biogenesis of platelets from megakaryocytes is a complex and intricate process that is incompletely understood. What is generally agreed on, however, is that platelets are assembled along intermediate pseudopodial-like extensions that derive from the cytoplasm of megakaryocytes. 1 These extensions are referred to as proplatelets. 1 Proplatelet formation has been modeled in vitro 2 and more recently observed in vivo. 3 Mechanisms that control proplatelet formation and platelet release are emerging, including the critical role of microtubules in driving the elongation of proplatelets. 4 Microtubules are also thought to be involved in sorting mitochondria as well as ␣ and dense granules into individual platelet buds. 5,6 Granules readily track along microtubular-rich shafts of proplatelets and distinct granule subpopulations are partitioned into platelets. 6 This suggests that platelet biogenesis is under discrete control.Protein synthesis also ramps up as proplatelets form. Adhesion molecules, secretome constituents, and other proteins are synthesized and packaged into discrete locations, including membranes and granules, so that platelets are loaded with requisite components before they are released into the bloodstream. 7 In addition to proteins, megakaryocytes send thousands of mRNAs and miRNAs into platelets. [8][9][10] The mRNAs are capped and polyadenylated on their 5Ј-and 3Ј-untranslated region (UTR), respectively, and code for protein when they are placed in in vitro tra...

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Android obesity is associated with a concomitant reduction of IL-10 and adiponectin levels. However, the antiinflammatory status of obesity might require prolonged periods of energy-restricted diets to revert to normal.

These data suggest that metformin could improve oxidative stress, preserve antioxidant function and restrain platelet activation in type 2 diabetes.

Glycoprotein (GP) IIbIIIa inhibitors are used in the treatment of acute coronary syndromes. Transient immune-mediated acute thrombocytopenia is a recognized side effect of GPIIbIIIa inhibitors. We provide evidence that GPIIbIIIa inhibitor-induced antibodies can affect megakaryo-cytes in the presence of eptifibatide. In a patient with acute coronary syndrome, acute thrombocytopenia occurred after a second exposure to eptifibatide 20 days after the initial treatment. Despite the short half-life of eptifibatide (t 1/2 2 hours), thrombocytopenia less than 5 10 9 /L and gastrointestinal and skin hemorrhage persisted for 4 days. Glycoprotein-specific enzyme-linked im-munosorbent assay showed eptifibatide-dependent, GPIIbIIIa-specific antibodies. Bone marrow examination showed predominance of early megakaryocyte stages, and platelet transfusion resulted in an abrupt platelet count increase. Viability of cultured cord blood-derived megakaryocytes was reduced in the presence of eptifibatide and patient IgG fraction. These findings can be explained by impaired megakaryocytopoiesis complicating anti-GPIIbIIIa antibody-mediated immune thrombocytopenia. This mechanism may also apply to some patients with autoimmune thrombocytopenia. (Blood. 2009;114:1250-1253) Introduction Glycoprotein (GP) IIbIIIa antagonists are a heterogeneous group of integrin inhibitors used for treatment in acute coronary syndromes. A relatively frequent adverse effect of GPIIbIIIa inhibitors is immune-mediated thrombocytopenia. The risk of thrombocytopenia from immune and nonimmune causes differs between the drugs: abciximab carries a risk of 0.3%-1.6%; tirofiban, 0.2%-0.4% 1 ; and eptifibatide, 0%-0.2%. 2,3 GPIIbIIIa inhibitor-immune thrombocytopenia typically manifests within the first 24 hours of treatment, which indicates that these patients already have circulating antibodies in their plasma. 4 The risk for thrombocytopenia is higher during second exposure compared with first-time treatment. 5 In addition, severe thrombocytopenia (20 10 9 /L) was more frequent after a second exposure with abciximab. 6 Usually, platelet counts decrease to values less than 50 10 9 /L, often less than 20 10 9 /L. Patients have an increased risk for bleeding, but, especially with tirofiban and eptifibatide, platelet counts normalize after cessation of the GPIIbIIIa inhibitor within 2 to 3 days. However, some patients have an unusually prolonged thrombo-cytopenia, 7 which is inconsistent with the relatively short half-lives of tirofiban and eptifibatide. Methods Case report A 67-year-old male patient was treated with eptifibatide (20 mg intravenous bolus) during coronary stent implantation (Figure 1) and reexposed (20 mg intravenously) during a second coronary intervention 20 days later. Within 1 hour, diffuse bleeding occurred and the platelet count decreased to less than 5 10 9 /L (Figure 1). Despite transfusion of 2 platelet concentrates, pericardial hemorrhage occurred, platelet counts remained less than 5 10 9 /L for 4 days, and gastrointestinal bleeding, trachea...

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Increased levels of sCD40L (soluble CD40 ligand) have been associated with enhanced in vivo platelet activation, and may represent a molecular link between inflammation and a prothrombotic state. The aim of the present study was to analyse the relationship between platelet activation, endothelial dysfunction, low-grade inflammation and sCD40L in patients with hypertension with or without MA (microalbuminuria). A cross-sectional comparison of sCD40L levels was performed in 25 patients with MH (essential hypertension with MA) pair-matched for gender and age with 25 patients with EH (essential hypertension) and 25 HS (healthy subjects with normotension). Circulating levels of CRP (C-reactive protein), a marker of inflammation, sP-selectin (soluble P-selectin), a marker of in vivo platelet activation, and ADMA (asymmetric dimethylarginine) and vWF (von Willebrand factor), markers of endothelial dysfunction, were analysed in each subject. sCD40L levels were increased in patients with MH compared with either patients with EH (P<0.001) or HS (P<0.0001). A highly significant correlation between plasma sCD40L and sP-selectin (P<0.0001), vWF (P<0.001) or CRP levels (P<0.05) was observed in patients with MH. Multivariate regression analysis showed that sP-selectin was the strongest independent predictor of sCD40L levels (P<0.0001) in patients with MH. Patients with hypertension with both vWF and CRP levels above the median had the highest sCD40L levels (P<0.0001). Factorial ANOVA of all of the patients with hypertension confirmed that only patients with MH with low-grade inflammation had elevated levels of sCD40L. In conclusion, sCD40L levels appear to discriminate a subset of patients characterized by MA and low-grade inflammation, suggesting that inhibition of the CD40/CD40L system may represent a potential therapeutic target in subjects with hypertension at a high risk of cardiovascular events.