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Immune thrombocytopenia (ITP) is an autoimmune disorder characterized by low platelet count which can cause fatal hemorrhage. ITP patients with antiplatelet glycoprotein (GP) Ib-IX autoantibodies appear refractory to conventional treatments, and the mechanism remains elusive. Here we show that the platelets undergo apoptosis in ITP patients with anti-GPIbα autoantibodies. Consistent with these findings, the anti-GPIbα monoclonal antibodies AN51 and SZ2 induce platelet apoptosis in vitro. We demonstrate that anti-GPIbα antibody binding activates Akt, which elicits platelet apoptosis through activation of phosphodiesterase (PDE3A) and PDE3A-mediated PKA inhibition. Genetic ablation or chemical inhibition of Akt or blocking of Akt signaling abolishes anti-GPIbα antibody-induced platelet apoptosis. We further demonstrate that the antibody-bound platelets are removed in vivo through an apoptosis-dependent manner. Phosphatidylserine (PS) exposure on apoptotic platelets results in phagocytosis of platelets by macrophages in the liver. Notably, inhibition or genetic ablation of Akt or Akt-regulated apoptotic signaling or blockage of PS exposure protects the platelets from clearance. Therefore, our findings reveal pathogenic mechanisms of ITP with anti-GPIbα autoantibodies and, more importantly, suggest therapeutic strategies for thrombocytopenia caused by autoantibodies or other pathogenic factors.

Background: Fat deposition is associated with peripheral arterial disease. Adipose tissue has recently been implicated in vascular remodeling and angiogenic activity. We hypothesized that the transplantation of adipose tissues from normal mice improves blood flow perfusion and neovascularization in high-fat diet fed mice. Methods: After 14 weeks of high-fat diet (HFD)-fed mice, unilateral hind limb ischemia was performed. Subcutaneous white adipose tissue (WAT) and brown adipose tissue (BAT) fat pads were harvested from normal EGFP mice, and subcutaneously transplanted over the region of the adductor muscles of HFD mice. Blood flow was measured using Laser Doppler Scanner. Vascular density, macrophages infiltration, and macrophage polarization were examined by RT-qPCR, and immunohistochemistry. Results: We found that the transplantation of WAT derived from normal mice improved functional blood flow in HFD-fed mice compared to mice transplanted with BAT and sham-treated mice. WAT transplantation increased the recruitment of pericytes associated with nascent blood vessels, but did not affect capillary formation. Furthermore, transplantation of WAT ameliorated HFD-induced insulin resistance, M2 macrophage predominance and the release of arteriogenic factors in ischemic muscles. Mice receiving WAT also displayed a marked reduction in several proinflammatory cytokines. In contrast, mice transplanted with BAT were glucose intolerant and demonstrated increased IL-6 levels in ischemic muscles. Conclusion: These results indicate that transplantation of adipose tissue elicits improvements in blood perfusion and beneficial effects on systemic glucose homeostasis and could be a promising therapeutic option for the treatment of diabetic peripheral arterial disease.

Background: Fat deposition is associated with peripheral arterial disease. Adipose tissue has recently been implicated in vascular remodeling and angiogenic activity. We hypothesized that the transplantation of adipose tissues from normal mice improves blood flow perfusion and neovascularization in high-fat diet fed mice. Methods: After 14 weeks of high-fat diet (HFD)-fed mice, unilateral hind limb ischemia was performed. Subcutaneous white adipose tissue (WAT) and brown adipose tissue (BAT) fat pads were harvested from normal EGFP mice, and subcutaneously transplanted over the region of the adductor muscles of HFD mice. Blood flow was measured using Laser Doppler Scanner. Vascular density, macrophages infiltration, and macrophage polarization were examined by RT-qPCR, and immunohistochemistry. Results: We found that the transplantation of WAT derived from normal mice improved functional blood flow in HFD-fed mice compared to mice transplanted with BAT and sham-treated mice. WAT transplantation increased the recruitment of pericytes associated with nascent blood vessels, but did not affect capillary formation. Furthermore, transplantation of WAT ameliorated HFD-induced insulin resistance, M2 macrophage predominance and the release of arteriogenic factors in ischemic muscles. Mice receiving WAT also displayed a marked reduction in several proinflammatory cytokines. In contrast, mice transplanted with BAT were glucose intolerant and demonstrated increased IL-6 levels in ischemic muscles. Conclusion: These results indicate that transplantation of adipose tissue elicits improvements in blood perfusion and beneficial effects on systemic glucose homeostasis and could be a promising therapeutic option for the treatment of diabetic peripheral arterial disease.

Background Coagulation factor V ( FV ) plays a key role in hemostasis, is present in plasma and platelets, and has both pro‐ and anticoagulant properties; however, the contribution of platelet‐derived FV to arterial thrombosis remains undetermined. Methods and Results Using transgenic mice with various levels of FV gene expression that was restricted to the plasma or platelets, the roles of platelet FV were evaluated in the regulation of arterial thrombosis and platelet activation. Mice with higher levels of platelet FV exhibited faster thrombotic occlusion of the carotid artery after injury compared with mice with lower platelet FV levels. Infusion of platelets with higher levels of FV into transgenic mice with undetectable levels of platelet FV reduced the time to carotid artery occlusion. In contrast, infusion of purified recombinant plasma FV into mice with undetectable platelet FV levels failed to reduce the carotid occlusion times following injury. Evaluation of isolated platelets revealed that platelet‐derived FV was critical for the regulation of platelet activation. These effects were associated with an increased level of expression of P‐selectin and increased cGMP in platelets. Conclusions We established that platelet‐derived FV is a critical mediator of arterial thrombosis that involves platelet activation.

Methylglyoxal (MGO), an active metabolite of glucose, has been reported to induce vascular cell apoptosis in diabetic complication. Polydatin (PD), a small natural compound from Polygonum cuspidatum, has a number of biological functions, such as antioxidative, anti-inflammatory, and nephroprotective properties. However, the protective effects of PD on MGO-induced apoptosis in endothelial cells remain to be elucidated. In this study, human umbilical vein endothelial cells (HUVECs) were used to explore the effects of PD on MGO-induced cell apoptosis and the possible mechanism involved. HUVECs were pretreated with PD for 2 h, followed by stimulation with MGO. Then cell apoptosis, reactive oxygen species (ROS) generation, mitochondrial membrane potential (MMP) impairment, mitochondrial morphology alterations, and Akt phosphorylation were assessed. The results demonstrated that PD significantly prevented MGO-induced HUVEC apoptosis. PD pretreatment also significantly inhibited MGO-induced ROS production, MMP impairment, mitochondrial morphology changes, and Akt dephosphorylation. These results and the experiments involving N-acetyl cysteine (antioxidant), Cyclosporin A (mitochondrial protector), and LY294002 (Akt inhibitor) suggest that PD prevents MGO-induced HUVEC apoptosis, at least in part, through inhibiting oxidative stress, maintaining mitochondrial function, and activating Akt pathway. All of these data indicate the potential application of PD for the treatment of diabetic vascular complication.

Background: The flowers of Carthamus tinctorius L. are widely used in traditional Chinese medicine to treat cerebrovascular and cardiovascular diseases. Hydroxysafflor yellow A (HSYA), the main constituent of C. tinctorius L. flowers, is known for its multiple biological activities. The present study investigated the effects of HSYA on angiogenesis in vitro and in a mouse hindlimb ischemia model. Methods: Using human umbilical vein endothelial cells (HUVEC) in vitro and a mouse hindlimb ischemia model in vivo, the angiogenic role of HSYA was evaluated. Results: HSYA significantly increased the capillary-like tube formation and migration of HUVEC. HSYA not only induced a rise in the expression of angiopoietin 1 and Tie-2 but it also increased phosphorylation of Tie-2, Akt, and extracellular signal-regulated kinase 1/2. Furthermore, an anti-Tie-2 neutralizing antibody significantly inhibited HSYA-induced HUVEC tube formation and migration. In vivo, the recovery of perfusion of ischemic hindlimb tissue after femoral artery interruption was significantly increased in HSYA-treated mice compared to vehicle controls. Consistent with these results, the arteriole and capillary densities in ischemic gastrocnemius muscles were significantly increased in HSYA-treated mice. Conclusions: These results indicate the potential utility of HSYA for the treatment of ischemic diseases.