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Platelets have emerged as key inflammatory cells implicated in the pathology of sepsis, but their contributions to rapid clinical deterioration and dysregulated inflammation have not been defined. Here, we show that the incidence of thrombocytopathy and inflammatory cytokine release was significantly increased in patients with severe sepsis. Platelet proteomic analysis revealed significant upregulation of gasdermin D (GSDMD). Using platelet-specific Gsdmd -deficient mice, we demonstrated a requirement for GSDMD in triggering platelet pyroptosis in cecal ligation and puncture (CLP)-induced sepsis. GSDMD-dependent platelet pyroptosis was induced by high levels of S100A8/A9 targeting toll-like receptor 4 (TLR4). Pyroptotic platelet-derived oxidized mitochondrial DNA (ox-mtDNA) potentially promoted neutrophil extracellular trap (NET) formation, which contributed to platelet pyroptosis by releasing S100A8/A9, forming a positive feedback loop that led to the excessive release of inflammatory cytokines. Both pharmacological inhibition using Paquinimod and genetic ablation of the S100A8/A9–TLR4 signaling axis improved survival in mice with CLP-induced sepsis by suppressing platelet pyroptosis.

Rationale: Kawasaki disease (KD) is an acute vasculitis of early childhood that can result in per-manent coronary artery structural damage. The etiology for this arterial vulnerability in up to 15% of KD patients is unknown. Vascular smooth muscle cell (VSMC) dedifferentiation play a key role in the pathophysiology of medial damage and aneurysm formation, recognized arterial pathology in KD. Platelet hyperreactivity is also a hallmark of KD. We recently demonstrated that uptake of platelets and platelet-derived miRNAs influences VSMC phenotype in vivo. Objective: We set out to explore whether platelet/vascular smooth muscle cell interactions contrib-ute to coronary pathology in KD. Methods and Results: We prospectively recruited and studied 242 KD patients, 75 of whom had doc-umented coronary artery pathology. Genome-wide miRNA sequencing and droplet digital PCR (ddPCR) demonstrated that KD patient platelets have significant induction of miR-223 compared to healthy controls. Platelet-derived miR-223 has recently been shown to promote vascular smooth muscle quiescence and resolution of wound healing after vessel injury. Paradoxically, KD patients with the most severe coronary pathology (giant coronary artery aneurysms) exhibited a lack of miR-223 induction. Hyperactive platelets isolated from KD patients are readily taken up by VSMCs, delivering functional miR-223 into the VSMCs promoting VSMC differentiation via downregulation of platelet-derived growth factor receptor β (PDGFRβ). The lack of miR-223 induction in patients with severe coronary pathology leads to persistent VSMC dedifferentiation. In a mouse model of KD (Lactobacillus casei cell wall extract (LCWE) injection), miR-223 knockout (miR-223 KO) mice exhibited increased medial thickening, loss of contractile VSMCs in the media, and fragmenta-tion of medial elastic fibers compared to WT mice, which demonstrated significant miR-223 induc-tion upon LCWE challenge. The excessive arterial damage in the miR-223 knockout could be res-cued by adoptive transfer of platelet, administration of miR-223 mimics, or the PDGFRβ inhibitor imatinib mesylate. Interestingly, miR-223 levels progressively increase with age, with the lowest levels found in less than five-year-old. This provides a basis for coronary pathology susceptibility in this very young cohort. Conclusions: Platelet-derived miR-223 (through PDGFRβ inhibition) promotes VSMC differentia-tion and resolution of KD induced vascular injury. Lack of miR-223 induction leads to severe coro-nary pathology characterized by VSMC dedifferentiation and medial damage. Detection of platelet-derived miR-223 in KD patients (at the time of diagnosis) may identify patients at greatest risk of coronary artery pathology. Moreover, targeting platelet miR-223 or VSMC PDGFRβ represents po-tential therapeutic strategies to alleviate coronary pathology in KD.

Arnebin-1, a naphthoquinone derivative, plays a crucial role in the wound healing properties of Zicao (a traditional wound healing herbal medicine). It has been noted that Arnebin-1, in conjunction with vascular endothelial growth factor (VEGF), exerts a synergistic pro-angiogenic effect on human umbilical vein endothelial cells (HUVECs) and accelerates the healing process of diabetic wounds. However, the mechanisms responsible for the pro-angiogenic effect of arnebin-1 on HUVECs and its healing effect on diabetic wounds have not yet been fully elucidated. In this study, in an aim to elucidate these mechanisms of action of arnebin-1, we investigated the effects of arnebin-1 on the VEGF receptor 2 (VEGFR2) and the phosphoinositide 3-kinase (PI3K)-dependent signaling pathways in HUVECs treated with VEGF by western blot analysis. The pro-angiogenic effects of arnebin-1 on HUVECs, including its effects on proliferation and migration, were evaluated by MTT assay, Transwell assay and tube formation assay in vitro. The expression levels of hypoxia-inducible factor (HIF)-1α, endothelial nitric oxide synthase (eNOS) and VEGF were determined by western blot analysis in the HUVECs and wound tissues obtained from non-diabetic and diabetic rats. CD31 expression in the rat wounds was evaluated by immunofluorescence staining. We found that the activation of the VEGFR2 signaling pathway induced by VEGF was enhanced by arnebin-1. Arnebin-1 promoted endothelial cell proliferation, migration and tube formation through the PI3K-dependent pathway. Moreover, Arnebin-1 significantly increased the eNOS, VEGF and HIF-1α expression levels in the HUVECs and accelerated the healing of diabetic wounds through the PI3K-dependent signaling pathway. CD31 expression was markedly enhanced in the wounds of diabetic rats treated with arnebin-1 compared to the wounds of untreated diabetic rats. Therefore, the findings of the present study indicate that arnebin-1 promotes the wound healing process in diabetic rats by eliciting a pro-angiogenic response.

As a BCS II drug, the atypical antipsychotic agent lurasidone hydrochloride (LH) has low oral bioavailability mainly because of its poor aqueous solubility/dissolution. Unexpectedly, amorphous LH exhibited a much lower dissolution than that of its stable crystalline form arising from its gelation during the dissolution process. In the current study, a supramolecular coamorphous system of LH with l-cysteine hydrochloride (CYS) was prepared and characterized by powder X-ray diffraction and differential scanning calorimetry. Surprisingly, in comparison to crystalline and amorphous LH, such a coamorphous system dramatically enhanced solubility (at least ∼50-fold in the physiological pH range) and dissolution (∼1200-fold) of LH, and exhibited superior physical stability under long-term storage condition. More importantly, the coamorphous system was able to eliminate gelation of amorphous LH during dissolution. In order to further explore the mechanism of such improvement, the internal interactions of the coamorphous system in the solid state and in aqueous solution were investigated. Fourier transform infrared spectroscopy, Raman spectroscopy, and solid-state 13C NMR suggested that intermolecular hydrogen bonds formed between the nitrogen atom in the benzisothiazole ring of LH and the NH3 + group of CYS after coamorphization. A fluorescence quenching test with a Stern–Volmer plot and density functional theory modeling, phase-solubility study, and NMR test in D2O indicated that ground-state complexation occurred between LH and CYS in aqueous solution, which contributed to the solubility and dissolution enhancement of LH. The current study offers a promising strategy to overcome poor solubility/dissolution and be able to eliminate gelation of amorphous materials by coamorphization and complexation.

The coamorphous tadalafil–repaglinide (molar ratio, 1 : 1) prepared by solvent-evaporation method significantly improve the physicochemical properties of tadalafil and repaglinide.

Zicao (Lithospermum erythrorhizon) has been used in clinics as a traditional Chinese medicine for thousands of years. Acetylshikonin (AS) is the main ingredient of Zicao, Xinjiang, China. The objective of this study was to investigate the anti-obesity and anti-nonalcoholic fatty liver disease (NAFLD) efficacy of AS in a model of spontaneous obese db/db mice. Mice were divided into Wild Type (WT) groups and db/db groups, which received no treatment or treatment with 100 mg/kg/day clenbuterol (CL) hydrochloride or 540 mg/kg/day AS by oral gavage for eight weeks. The results provided the evidence that AS prevented obesity and NAFLD including reduction in body weight, food efficiency ratio, serum triglyceride (TG) and free fatty acid (FFA) levels in db/db mice. Administration of AS markedly suppressed the levels of hepatic alanine aminotransferase (ALT), aspartate aminotransferase (AST) and pro-inflammatory cytokines in treated groups when compared with that of db/db groups. Further investigation of the lipid synthesis-related protein using Western blotting revealed that hepatic protein expression of sterol regulatory element-binding protein-1 (SREBP-1), fatty acid synthetase (FAS) and 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR) were significantly downregulated by AS treatment. These findings suggest that AS exerts anti-obesity and anti-NAFLD effects through the regulation of lipid metabolism and anti-inflammatory effects.