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Background Platelets release platelet-derived extracellular vesicles (PDEVs) upon activation – in a process that is regulated by generation of reactive oxygen species (ROS). Platelet NADPH oxidase-1 (Nox-1) contributes to ROS generation and thrombus formation downstream of the collagen receptor GPVI. Objectives We aimed to investigate whether PDEVs contain Nox-1 and whether this is relevant for PDEV-induced platelet activation. Methods PDEVs were isolated through serial centrifugation after platelet activation with thrombin receptor agonist TRAP-6 (activated PDEVs) or in the absence of agonist (resting PDEVs). The physical properties of PDEVs were analyzed through nanoparticle tracking analysis. Nox-1 levels, fibrinogen binding and P-selectin exposure were measured using flow cytometry, and protein levels quantified by immunoblot analysis. ROS were quantified using DCF fluorescence and electron paramagnetic resonance. Results Nox-1 was found to be increased on the platelet outer membrane upon activation and was present in PDEVs. PDEVs induced platelet activation, while co-addition of GPVI agonist collagen-related peptide (CRP) did not potentiate this response. PDEVs were shown to be able to generate superoxide in a process at least partially mediated by Nox-1, while Nox-1 inhibition with ML171 (also known as 2-APT) did not influence PDEV production. Finally, inhibition of Nox-1 abrogated PDEV-mediated platelet activation. Conclusions PDEVs are able to generate superoxide, bind to and activate platelets in a process mediated by Nox-1. These data provide novel mechanisms by which Nox-1 potentiates platelet responses, thus proposing Nox-1 inhibition as a feasible strategy to treat and prevent thrombotic diseases., Graphical abstract Image 1, Highlights • Activated platelets have increased NADPH oxidase-1 (Nox-1) exposure on the outer membrane. • Platelet-derived extracellular vesicles (PDEVs) express Nox-1 and generate superoxide in a process mediated by Nox-1. • PDEVs bind and activate platelets in a Nox-1-dependent manner.

Folliculitis decalvans (FD) is a rare, chronic primary scarring alopecia that classically presents with perifollicular pustules, scaling, crusting, and tufted hair in the affected areas of the scalp, accompanied by pruritus or pain.1 The cause of FD remains unclear; however, on histopathology, it presents as a predominantly neutrophilic inflammatory process targeting hair follicles, resulting in irreversible damage and permanent hair loss.1 Currently, there is no cure for FD. Most treatment regimens are directed at controlling the scalp-associated microbes with antibiotics and antiseptics, managing the inflammation, and preventing further hair loss with immunomodulators.2 Recent case reports have suggested that autologous platelet-rich plasma (PRP) can be a promising adjunct therapy for the treatment of lymphocytic scarring alopecia.3,4 However, there are no data on the efficacy of PRP in neutrophilic scarring alopecias, such as FD. Here, we present 2 patients with long-standing, refractory FD who experienced stability in hair loss and significant symptomatic improvement in inflammation with concurrent PRP treatments. The severity of scalp inflammation was graded (Table I) by a single dermatologist (NAM) during each clinical visit. Table I Grading scale of disease severity

Objective The magnitude of the therapeutic effects of intra-articular injection of platelet-rich plasma (PRP) on osteoarthritis (OA) is still under debate. The goal of this study that was a systematic review of randomised controlled trials ​of PRP injections for the treatment of OA was to elucidate the therapeutic efficacy of PRP. Methods Electronic databases of PubMed, CENTRAL, EMBASE, EBSCO, ClinicalTrials.gov, and International Clinical Trials Registry Platform ​were searched from inception to June 2018 for RCTs that compared PRP injections to controls in patients with OA. A random-effects approach was used to compile data and subgroups according to trial size (large trials versus small trials), patient profile (age and gender), and PRP preparation method was performed. Results Thirty trials met the inclusion criteria and were analysed. All results had unexplained statistical heterogeneity. Patients treated with PRP compared with control showed statistically relevant pain relief and function improvement at short term (standardised mean difference [SMD] ​= ​−0.62, 95% confidence interval [CI]: −0.98 to −0.27, P ​= ​0.0006, SMD ​= ​−0.74, 95% CI: −1.11 to 0.36, P ​= ​0.0001, respectively), medium term (SMD ​= ​−0.53, 95% CI: −0.83 to −0.23, P ​= ​0.0006, SMD ​= ​−0.50, 95% CI: −0.75 to −0.25, P ​= ​0.0006), and long term (SMD ​= ​−0.69, 95% CI: −1.08 to −0.30, P ​= ​0.0006, SMD ​= ​−0.68, 95% CI: −0.1.09 to −0.27, P ​= ​0.001, respectively). A subgroup analysis of the data from large trials and from trials composed of less than 50% female patients revealed that therapeutic effects of the treatment are insignificant. Conclusions According to the currently available data, PRP injections are beneficial for pain relief and function improvement in patients with OA. This meta-analysis, however, demonstrated that the efficacy of PRP is related to sample size and gender composition. Thus, more randomised controlled trials of high quality and larger patient size, also including gender aspects, are required to understand this phenomenon. The translational potential of this article The translation potential of this meta-analysis is that provided another perspective to analyse the treatment effect of PRP for OA. In future research, phenotypes subpopulation and gender difference of OA patient should be considered for PRP treatment.

BACKGROUND Dyslipidemia induces platelet hyperactivation and hyper-aggregation, which are linked to thrombosis. Anthocyanins could inhibit platelet function in vitro and in mice fed high-fat diets with their effects on platelet function in subjects with dyslipidemia remained unknown. This study aimed to investigate the effects of different doses of anthocyanins on platelet function in individuals with dyslipidemia. METHODS A double-blind, randomized, controlled trial was conducted. Ninety-three individuals who were initially diagnosed with dyslipidemia were randomly assigned to placebo or 40, 80, 160 or 320 mg/day anthocyanin groups. The supplementations were anthocyanin capsules (Medox, Norway). Platelet aggregation by light aggregometry of platelet-rich plasma, P-selectin, activated GPⅡbⅢa, reactive oxygen species (ROS), and mitochondrial membrane potential were tested at baseline, 6 weeks and 12 weeks. FINDINGS Compared to placebo group, anthocyanins at 80 mg/day for 12 weeks reduced collagen-induced platelet aggregation (-3.39±2.36%) and activated GPⅡbⅢa (-8.25±2.45%) (P

Introduction Current non-invasive treatments for erectile dysfunction (ED) include oral medications, intracavernosal injections, and vacuum-assisted devices. Though these therapies work well for many, a subset of patients have contraindications or are unsatisfied with these options. Restorative therapies for ED are a new frontier of treatments focused on regenerating diseased tissue and providing a potential “cure” for ED. Aim The aim of this position statement is to examine existing clinical trial data for restorative therapies and identify elements that require further research before widespread adoption. Methods A literature review was performed to identify all clinical trials performed with regenerative therapy for ED. This includes treatments such as stem cell therapy (SCT), platelet rich plasma (PRP), and restorative related technologies like low-intensity shockwave therapy (LiSWT). Main Outcome Measures Most clinical trials in restorative therapies were assessed for safety, feasibility, or efficacy. This included recording adverse events, changes in sexual function and erectile function questionnaires, and diagnostics measures. Results To date there is an absence of robust clinical data supporting the efficacy of restorative therapies regarding ED, though technologies such as LiSWT have established relative safety. Conclusions Restorative therapies are a promising technology that represents a new frontier of treatment geared towards reversing disease pathology rather than just treating symptoms. However, current published clinical studies are limited. Future work needs to be adequately powered, multi-center, randomized, sham/placebo-controlled trials in well-characterized patient populations to ensure safety and demonstrate efficacy. Until these studies are done, restorative therapies should be reserved for clinical trials and not offered in routine clinical practice.

Freeze-drying methods not only enable the delivery of growth factors using platelets, but also extend the shelf-life of platelet concentrates. The present study shows the clinical results of treating knee osteoarthritis with freeze-dried platelet-derived factor concentrate (PFC). While it improved pain, activities of daily living, sports and recreational activities, and knee-related quality of life, it did not significantly improve symptoms other than pain, such as restricted range of motion and mechanical symptoms. As such, the treatment effect may be attributed to anti-inflammatory action rather than actual cartilage regeneration., Highlights • Intra-articular injection of freeze-dried PFC generally improved knee KOOS scores. • Pain, activities of daily living, sports and recreational activities, and knee-related quality of life improved. • However, symptoms other than pain, such as restricted range of motion and mechanical symptoms, did not improve.

Introduction Platelet-rich plasma (PRP) has been shown to be effective in treating partial tears of the ulnar collateral ligament (UCL) of the elbow in overhead throwing athletes, but it is still unknown whether it has a role in complete tears. The aim of this study was to assess the effectiveness of PRP in treating complete as well as partial UCL tears. We hypothesized that trephination of the injured UCL followed by injection with PRP can promote healing of both partial and complete tears. Methods Thirty-four baseball players with partial or complete UCL tears confirmed by magnetic resonance imaging (MRI) were included in the study. They were all recalcitrant to more than two months of rest and physical therapy. Under ultrasound guidance, trephination of the UCL was performed using an 18-gauge needle, followed by PRP injection. Visual analog scale (VAS) scores, Disabilities of the Arm, Shoulder, and Hand (DASH) sports module scores, and sonographic ulnohumeral joint space measurements with valgus stress were all obtained prior to the procedure and six months after. Results Twenty-six of 30 athletes were able to return to sport with pre-injury level of play within six months after the procedure, at an average time of 12.4 weeks (range: 10–18). Four subjects needed surgical treatment for persistent UCL insufficiency. The average follow-up was 54.2 weeks (range: 26–148). The average VAS and DASH scores improved from 53.5 to 17.2 (p, Highlights • Platelet-rich plasma can be used to treat tears of the ulnar collateral ligament. • Subjective measures of arm function significantly improved with treatment. • Ulnohumeral joint space size on ultrasound significantly decreases with treatment. • Return to sport, even for complete ruptures, is faster than with surgical therapy. • The treatment is more effective for proximal tears of the ulnar collateral ligament.

Autologous biologics, defined as platelet-rich plasma (PRP) and bone marrow aspirate concentrate (BMC), are cell-based therapy treatment options in regenerative medicine practices, and have been increasingly used in orthopedics, sports medicine, and spinal disorders. These biological products are produced at point-of-care; thereby, avoiding expensive and cumbersome culturing and expansion techniques. Numerous commercial PRP and BMC systems are available but reports and knowledge of bio-cellular formulations produced by these systems are limited. This limited information hinders evaluating clinical and research outcomes and thus making conclusions about their biological effectiveness. Some of their important cellular and protein properties have not been characterized, which is critical for understanding the mechanisms of actions involved in tissue regenerative processes. The presence and role of red blood cells (RBCs) in any biologic has not been addressed extensively. Furthermore, some of the pathophysiological effects and phenomena related to RBCs have not been studied. A lack of a complete understanding of all of the biological components and their functional consequences hampers the development of clinical standards for any biological preparation. This paper aims to review the clinical implications and pathophysiological effects of RBCs in PRP and BMC; emphasizes hemolysis, eryptosis, and the release of macrophage inhibitory factor; and explains several effects on the microenvironment, such as inflammation, oxidative stress, vasoconstriction, and impaired cell metabolism., Highlights • Different biological formulations optimize disease specific regenerative treatment protocols. • Disintegrated RBC's release harmful components to regenerative therapy treatment vials. • The effectiveness of MSC injection depends on the quality of the bone marrow aspiration procedure. • PRP and BMC should contain minimal to no erythrocytes.

Graphical abstract, Platelet aggregometry assays are generally used for the analysis of platelet function but can also be adapted for further research and therapy focused applications. This method describes the procedures for the preparation of human platelet-rich plasma (PRP) and platelet-poor plasma (PPP) for the assessment of human platelet aggregation induced by agonists such as platelet-activating factor (PAF), thrombin, collagen, adenosine diphosphate (ADP), arachidonic acid, etc. • This method can be applied in vitro to evaluate the aggregatory effects of these agonists and to assess the antiaggregatory effects of several bioactive antiplatelet agents (compounds of natural or pharmacological origin) in human PRP. • This versatile method can be used in both basic and clinical research for the assessment of platelet aggregation (a major cardiovascular risk factor), platelet agonists, and inhibitors, in physiological or pathological conditions. • This method can be adapted to assess platelet activity in postprandial and intervention studies ex vivo.

Lysophosphatidic acid (LPA) is a potent signaling lipid, and state-dependent alterations in plasma LPA make it a promising diagnostic marker for various diseases. However, plasma LPA concentrations vary widely among reports, even under normal conditions. These variations can be attributed, at least in part, to the artificial metabolism of LPA after blood collection. Here, we aimed to develop an optimized plasma preparation method that reflects the concentration of LPA in the circulating blood. The main features of the devised method were suppression of both LPA production and degradation after blood collection by keeping whole blood samples at low temperature followed by the addition of an autotaxin inhibitor to plasma samples. Using this devised method, the LPA level did not change for 30 min after blood collection. Also, human and mouse LPA levels were found to be much lower than those previously reported, ranging from 40 to 50 nM with minimal variation across the individual. Finally, the increased accuracy made it possible to detect circadian rhythms in the levels of certain LPA species in mouse plasma. These results demonstrate the usefulness of the devised plasma preparation method to determine accurate plasma LPA concentrations.

Introduction and importance Atraumatic splenic rupture(ASR) is a rare event with challenging management, due to absence of clinical history of trauma and delayed diagnosis. Current clinical report could provide detailed information regarding clinical presentation and management to physicians. Case presentation A 61 years-old woman underwent percutaneous coronary intervention(PTCA) after ST elevation myocardial infarction(STEMI). In the first day after PTCA epigastric abdominal disconfort was reported, and new PTCA excluded early complication. During hospitalization, due to anemization and hypotension CT scan was performed which revealed ASR with large hemoperitoneum. Emergency surgical splenectomy was performed. Postoperative course was uneventful and patient started 90 mg Ticageclor twice daily in the first post-operative day(POD) plus low molecular weight Heparin and restarted dual antiplatelet therapy(DAPT) the seventh POD. During follow up, patient underwent to assessment of platelet function showing normal level of DAPT inhibition. Due to the lack of pathological aggregation activity, DAPT was maintained. Clinical discussion ASR is mainly linked to oncological, malformative, inflammatory and thromboembolic conditions. Despite anticoagulant and anti-aggregating drug-related ASR has been already described, we report the first case of drug-related ASR as immediate complication of PTCA due to DAPT. After surgery, careful anti-aggregating management was required to balance in stent restenosis and hemorragic risk. Assessment of platelet activity was performed to design a tailored anti-aggregating therapy. Conclusion Drug-related ASR is dangerous complication due to the high mortality rate and misleading symptoms. After major bleeding events, such as drug-related ASR, evaluation of platelet function could provide a tailored DAPT., Highlights • Atraumatic splenic rupture (ASR) is a rare event with misleading symptoms. • Our report describes the first case of ASR as an early complication after stenting. • Aggregation test could support a tailored therapy to avoid further bleeding events.

Redox-related plasma proteins are candidate reporters of protein signatures associated with endothelial structure/function. Thiol-proteins from protein disulfide isomerase (PDI) family are unexplored in this context. Here, we investigate the occurrence and physiological significance of a circulating pool of PDI in healthy humans. We validated an assay for detecting PDI in plasma of healthy individuals. Our results indicate high inter-individual (median = 330 pg/mL) but low intra-individual variability over time and repeated measurements. Remarkably, plasma PDI levels could discriminate between distinct plasma proteome signatures, with PDI-rich (>median) plasma differentially expressing proteins related to cell differentiation, protein processing, housekeeping functions and others, while PDI-poor plasma differentially displayed proteins associated with coagulation, inflammatory responses and immunoactivation. Platelet function was similar among individuals with PDI-rich vs. PDI-poor plasma. Remarkably, such protein signatures closely correlated with endothelial function and phenotype, since cultured endothelial cells incubated with PDI-poor or PDI-rich plasma recapitulated gene expression and secretome patterns in line with their corresponding plasma signatures. Furthermore, such signatures translated into functional responses, with PDI-poor plasma promoting impairment of endothelial adhesion to fibronectin and a disturbed pattern of wound-associated migration and recovery area. Patients with cardiovascular events had lower PDI levels vs. healthy individuals. This is the first study describing PDI levels as reporters of specific plasma proteome signatures directly promoting contrasting endothelial phenotypes and functional responses., Graphical abstract Image 1, Highlights • A newly-validated plasma PDI assay shows high inter- but low intra-individual variability. • PDI-poor plasma carries protein signatures related to coagulation/immunoinflammation • PDI-rich plasma protein signatures relate to cell adhesion/housekeeping functions. • Endothelial exposure to such plasmas triggers corresponding gene and secretome patterns. • PDI-poor plasma impairs endothelial adhesion and migration vs. PDI-rich plasma.

BACKGROUND: Thrombocytopenia is a serious issue for all patients with classical Wiskott-Aldrich syndrome (WAS) and X-linked thrombocytopenia (XLT) because it causes severe and life-threatening bleeding. Lentiviral gene therapy (GT) for WAS has shown promising results in terms of immune reconstitution. However, despite the reduced severity and frequency of bleeding events, platelet counts remain low in GT-treated patients. OBJECTIVE: We carefully investigated platelet defects in terms of phenotype and function in untreated patients with WAS and assessed the effect of GT treatment on platelet dysfunction. METHODS: We analyzed a cohort of 20 patients with WAS/XLT, 15 of them receiving GT. Platelet phenotype and function were analyzed by using electron microscopy, flow cytometry, and an aggregation assay. Platelet protein composition was assessed before and after GT by means of proteomic profile analysis. RESULTS: We show that platelets from untreated patients with WAS have reduced size, abnormal ultrastructure, and a hyperactivated phenotype at steady state, whereas activation and aggregation responses to agonists are decreased. GT restores platelet size and function early after treatment and reduces the hyperactivated phenotype proportionally to WAS protein expression and length of follow-up. CONCLUSIONS: Our study highlights the coexistence of morphologic and multiple functional defects in platelets lacking WAS protein and demonstrates that GT normalizes the platelet proteomic profile with consequent restoration of platelet ultrastructure and phenotype, which might explain the observed reduction of bleeding episodes after GT. These results are instrumental also from the perspective of a future clinical trial in patients with XLT only presenting with microthrombocytopenia.

Platelets promote tumor metastasis by inducing promalignant phenotypes in cancer cells and directly contributing to cancer-related thrombotic complications. Platelet-derived extracellular vesicles (EVs) can promote epithelial-mesenchymal transition (EMT) in cancer cells, which confers high-grade malignancy. 12S-hydroxyeicosatetraenoic acid (12-HETE) generated by platelet-type 12-lipoxygenase (12-LOX) is considered a key modulator of cancer metastasis through unknown mechanisms. In platelets, 12-HETE can be esterified into plasma membrane phospholipids (PLs), which drive thrombosis. Using cocultures of human platelets and human colon adenocarcinoma cells (line HT29) and LC-MS/MS, we investigated the impact of platelets on cancer cell biosynthesis of 12S-HETE and its esterification into PLs and whether platelet ability to transfer its molecular cargo might play a role. To this aim, we performed coculture experiments with CFSE[5-(and-6)-carboxyfluorescein diacetate, succinimidyl ester]-loaded platelets. HT29 cells did not generate 12S-HETE or express 12-LOX. However, they acquired the capacity to produce 12S-HETE mainly esterified in plasmalogen phospholipid forms following the uptake of platelet-derived medium-sized EVs (mEVs) expressing 12-LOX. 12-LOX was detected in plasma mEV of patients with adenomas/adenocarcinomas, implying their potential to deliver the protein to cancer cells in vivo. In cancer cells exposed to platelets, endogenous but not exogenous 12S-HETE contributed to changes in EMT gene expression, mitigated by three structurally unrelated 12-LOX inhibitors. In conclusion, we showed that platelets induce the generation of primarily esterified 12-HETE in colon cancer cells following mEV-mediated delivery of 12-LOX. The modification of cancer cell phospholipids by 12-HETE may functionally impact cancer cell biology and represent a novel target for anticancer agent development.

Visual Abstract, Highlights • As expected, ticagrelor reduced ex-vivo ADP-induced aggregation in patients with pneumonia compared with placebo. • Ticagrelor reduced platelet–leukocyte interactions as well as plasma interleukin-6 within 24 h in patients with pneumonia compared with placebo. • Ticagrelor acutely altered NETosis biomarkers, whereas placebo had no effect. • Ticagrelor improved lung function and reduced need for supplemental oxygen in patients with pneumonia compared with placebo., Summary Despite treatment advances for sepsis and pneumonia, significant improvements in outcome have not been realized. Antiplatelet therapy may improve outcome in pneumonia and sepsis. In this study, the authors show that ticagrelor reduced leukocytes with attached platelets as well as the inflammatory biomarker interleukin (IL)-6. Pneumonia patients receiving ticagrelor required less supplemental oxygen and lung function tests trended toward improvement. Disruption of the P2Y12 receptor in a murine model protected against inflammatory response, lung permeability, and mortality. Results indicate a mechanistic link between platelets, leukocytes, and lung injury in settings of pneumonia and sepsis, and suggest possible therapeutic approaches to reduce complications.(Targeting Platelet-Leukocyte Aggregates in Pneumonia With Ticagrelor [XANTHIPPE]; NCT01883869)

Platelets store and secrete the chemokine stromal cell-derived factor (SDF)-1α upon platelet activation, but the ability of platelet-derived SDF-1α to signal in an autocrine/paracrine manner mediating functional platelet responses relevant to thrombosis and haemostasis is unknown. We sought to explore the role of platelet-derived SDF-1α and its receptors, CXCR4 and CXCR7 in facilitating platelet activation and determine the mechanism facilitating SDF-1α-mediated regulation of platelet function. Using human washed platelets, CXCR4 inhibition, but not CXCR7 blockade significantly abrogated collagen-mediated platelet aggregation, dense granule secretion and thromboxane (Tx) A2 production. Time-dependent release of SDF-1α from collagen-activated platelets supports a functional role for SDF-1α in this regard. Using an in vitro whole blood perfusion assay, collagen-induced thrombus formation was substantially reduced with CXCR4 inhibition. In washed platelets, recombinant SDF-1α in the range of 20–100 ng/mL− 1 could significantly enhance platelet aggregation responses to a threshold concentration of collagen. These enhancements were completely dependent on CXCR4, but not CXCR7, which triggered TxA2 production and dense granule secretion. Rises in cAMP were significantly blunted by SDF-1α, which could also enhance collagen-mediated Ca(2 +) mobilisation, both of which were mediated by CXCR4. This potentiating effect of SDF-1α primarily required TxA2 signalling acting upstream of dense granule secretion, whereas blockade of ADP signalling could only partially attenuate SDF-1α-induced platelet activation. Therefore, this study supports a potentially novel autocrine/paracrine role for platelet-derived SDF-1α during thrombosis and haemostasis, through a predominantly TxA2-dependent and ADP-independent pathway., Highlights • Collagen-induced platelet aggregation, TxA2 production and dense granule secretion require CXCR4 signalling. • CXCR4 regulates platelet thrombus formation. • SDF-1α-induced changes in cAMP and Ca(2 +) signalling require CXCR4. • SDF-1α, via CXCR4, enhances platelet activation responses to collagen, primarily through a TxA2-dependent and ADP-independent pathway.

Highlights • New treatment options for challenging procedures in hernia surgery are necessary. • Possibility of improving prosthetic compatibility and reducing future recurrences. • Tissue engineering offers new strategies to improve fascial healing. • Case of a surgeon – challenging abdominal incisional hernia. • Treatment provided was PRP and BM-MSCs on a biological mesh., Introduction The necessity to develop new treatment options for challenging procedures in hernia surgery is becoming even more evident and tissue engineering and biological technologies offer even newer strategies to improve fascial healing. The present case reports a patient-tailored surgical technique performed to repair a grade IV abdominal incisional hernia, with a combined use of platelet-rich plasma and bone marrow-derived mesenchymal stromal cells, implanted on a biological mesh. Presentation of the case A 71 year-old female patient complained of an abdominal incisional hernia, complicated by enterocutaneous fistula, four-months following laparostomy. Contrast enhanced computed tomography showed an incisional hernia defect of 15.5 × 20 cm, with a subcutaneous abscess and an intestinal loop adherent to the anterior abdominal wall, with a concomitant enterocutaneous fistula. Surgery involved abdominal wall standardized technique closure, with in addition platelet-rich plasma and bone marrow-derived mesenchymal stromal cells implanted on a biological mesh. Two years follow up showed no recurrences of incisional hernia. Discussion Coating surgical meshes with patient’s own cells may improve biocompatibility, by reducing inflammation and adhesion formation. Moreover, platelet-rich plasma is a good source of growth factors for wound healing, as well as a good medium for bone marrow multinucleate cells introduction into fascial repair. Conclusion This approach is likely to improve abdominal wall repair in high grade (IV) incisional hernia, with the real possibility of improving prosthetic compatibility and reducing future recurrences. The authors agree with the necessity of further studies and trials to assure the safety profile and superiority of this procedure.

The integrin alpha(IIb)beta(3) plays a critical role in mediating clot retraction by platelets which is important in vivo in consolidating thrombus formation. Actin-myosin interaction is essential for clot retraction. In the present study, we demonstrate that the structurally distinct Src kinase inhibitors, PP2 and PD173952, significantly reduced the rate of clot retraction, but did not prevent it reaching completion. This effect was accompanied by abolition of alpha(IIb)beta(3)-dependent protein tyrosine phosphorylation, including PLCgamma2. A role for PLCgamma2 in mediating clot retraction was demonstrated using PLCgamma2-deficient murine platelets. Furthermore, platelet adhesion to fibrinogen leads to MLC phosphorylation through a pathway that is inhibited by PP2 and by the PLC inhibitor, U73122. These results demonstrate a partial role for Src kinase-dependent activation of PLCgamma2 and MLC phosphorylation in mediating clot retraction downstream of integrin alpha(IIb)beta(3).

Ingestion of vegetables rich in inorganic nitrate has emerged as an effective method, via the formation of a nitrite intermediate, for acutely elevating vascular NO levels. As such a number of beneficial effects of dietary nitrate ingestion have been demonstrated including the suggestion that platelet reactivity is reduced. In this study we investigated whether inorganic nitrate supplementation might also reduce platelet reactivity in healthy volunteers and have determined the mechanisms involved in the effects seen. We conducted two randomised crossover studies each in 24 (12 of each sex) healthy subjects assessing the acute effects of dietary nitrate (250 ml beetroot juice) or potassium nitrate capsules (KNO3, 8 mmol) vs placebo control on platelet reactivity. Inorganic nitrate ingested either from a dietary source or via supplementation raised circulating nitrate and nitrite levels in both sexes and attenuated ex vivo platelet aggregation responses to ADP and, albeit to a lesser extent, collagen but not epinephrine in male but not female volunteers. These inhibitory effects were associated with a reduced platelet P-selectin expression and elevated platelet cGMP levels. In addition, we show that nitrite reduction to NO occurs at the level of the erythrocyte and not the platelet. In summary, our results demonstrate that inorganic nitrate ingestion, whether via the diet or through supplementation, causes a modest decrease in platelet reactivity in healthy males but not females. Our studies provide strong support for further clinical trials investigating the potential of dietary nitrate as an adjunct to current antiplatelet therapies to prevent atherothrombotic complications. Moreover, our observations highlight a previously unknown sexual dimorphism in platelet reactivity to NO and intimate a greater dependence of males on the NO-soluble guanylate cyclase pathway in limiting thrombotic potential., Highlights • Dietary nitrate or nitrate salt supplementation attenuates platelet activation in healthy male but not female volunteers. • These effects of nitrate relate to its bioactivation via the enterosalivary circuit to nitrite and then NO. • Nitrite attenuates platelet activation following reduction to NO at least in part at the level of the erythrocyte. • The antiplatelet effects of inorganic nitrate or nitrite in males are due to elevation of cGMP: an effect not evident in females. • Our findings intimate moderate antiplatelet effects of inorganic nitrate that might prove useful in therapeutics.