A pre‐clinical functional assessment of an acellular scaffold intended for the treatment of hard‐to‐heal wounds

The majority of the population experience successful wound‐healing outcomes; however, 1–3% of those aged over 65 years experience delayed wound healing and wound perpetuation. These hard‐to‐heal wounds contain degraded and dysfunctional extracellular matrix (ECM); yet, the integrity of this structure is critical in the processes of normal wound healing. Here, we evaluated a novel synthetic matrix protein for its ability to act as an acellular scaffold that could replace dysfunctional ECM. In this regard, the synthetic protein was subjected to adsorption and diffusion assays using collagen and human dermal tissues; evaluated for its ability to influence keratinocyte and fibroblast attachment, migration and proliferation and assessed for its ability to influence in vivo wound healing in a porcine model. Critically, these experiments demonstrate that the matrix protein adsorbed to collagen and human dermal tissue but did not diffuse through human dermal tissue within a 24‐hour observation period, and facilitated cell attachment, migration and proliferation. In a porcine wound‐healing model, significantly smaller wound areas were observed in the test group compared with the control group following the third treatment. These data provide evidence that the synthetic matrix protein has the ability to function as an acellular scaffold for wound‐healing purposes.     

Tanshinone IIA attenuates interleukin-17A-induced systemic sclerosis patient-derived dermal vascular smooth muscle cell activation via inhibition of the extracellular signal-regulated kinase signaling pathway

OBJECTIVE:Salvia miltiorrhiza has long been used to treat systemic sclerosis. Tanshinone IIA, one of the phytochemicals derived from the roots of Salvia miltiorrhiza, exhibits multiple biological activities. The present study aimed to investigate whether tanshinone IIA has an effect on the interleukin-17A-induced functional activation of systemic sclerosis patient-derived dermal vascular smooth muscle cells.METHODS:Systemic sclerosis patient-derived dermal vascular smooth muscle cells were incubated with various dosages of tanshinone IIA in the presence of interleukin-17A or the serum of systemic sclerosis patients. Cell proliferation was assessed using Cell Counting Kit-8. The expression of collagen 1 and 3 in cells was evaluated by immunofluorescence. Cell migration was measured using a transwell assay. The expression of phospho-extracellular signal-regulated kinase was detected by Western blotting.RESULTS:Our data demonstrate that tanshinone IIA exerts an inhibitory effect on interleukin-17A-induced systemic sclerosis patient-derived dermal vascular smooth muscle cell proliferation, collagen synthesis and migration.CONCLUSION:These findings suggest that tanshinone IIA might serve as a promising therapeutic agent for the treatment of systemic sclerosis.     

神经病理痛模型大鼠前扣带皮层不同水平磷酸化细胞外信号调节激酶表达差异比较

目的探讨痛相关情绪模型大鼠前扣带皮层(ACC)磷酸化细胞外信号调节激酶(p-ERK)的分布特点。方法将12只雄性SD大鼠随机分为对照组和模型组。模型组大鼠进行右侧腰5脊神经结扎制做模型。采用右后足跖机械痛阈检测观察行为变化,旷场实验和高架O迷宫实验检测痛相关情绪变化,免疫荧光技术检测同侧前扣带皮层前囟前3.2、2.7及2.2mm 3个水平p-ERK表达。结果大鼠经神经病理痛模型制做成功后机械痛阈显著下降,焦虑样行为产生。前扣带皮层前囟前3.2、2.7及2.2mm水平p-ERK阳性细胞表达量分别为11.89±2.57、32±4.67和17.56±2.04。对照组相应的p-ERK阳性细胞表达量分别为12.44±2.16、10±0.87和10.11±1.36。除前囟前3.2mm水平对照组与模型组相比没有显著性差异(P0.05)之外,其他两个水平p-ERK阳性细胞表达量模型组显著高于对照组(P0.01)。结论神经病理性疼痛能诱发大鼠焦虑情绪的产生及ACC脑区pERK的表达增高,这种变化可能主要与ACC脑区前囟前2.7及2.2mm水平p-ERK的变化相关,而与前囟前3.2mm水平无关。     

细胞外囊泡的检测及其在非酒精性脂肪性肝病诊断中的价值

细胞外囊泡是双层脂质膜小囊泡,可以由大多数类型的细胞释放,并在大多数体液中可检测到,细胞外囊泡可将其生物活性成分转移到受体细胞或激活靶细胞中的信号转导,发挥细胞间通讯的关键功能,从而参与多种疾病包括肝脏疾病的发生和发展。近年来非酒精性脂肪性肝病患病率上升,除有创的肝活组织检查外,目前还没有可靠的肝脏炎症诊断或纤维化分期方法,寻求相应的无创循环标志物的研究持续活跃,细胞外囊泡是其中较受关注者之一。为此综述了目前关于细胞外囊泡的物理特征、生物成分和分离方法的知识,并介绍了利用循环细胞来源囊泡作为非酒精性脂肪性肝病诊断新标志物的概念。     

转化生长因子β1/细胞外信号调节激酶/基质金属蛋白酶通路对腺样囊性癌侵袭和迁移的影响

目的 探讨转化生长因子β1/细胞外信号调节激酶/基质金属蛋白酶2(TGF-β1/ERKl/2/MMP-2)通路在腺样囊性癌(ACC)侵袭和迁移过程中的作用及机制.方法 以腺样囊性癌ACC-2细胞株为研究对象.用转化生长因子β1(TGF-β1)以及ERK通路抑制剂UO126处理ACC-2细胞.MTT检测ACC-2细胞的增殖情况,Transwell实验检测细胞迁移、侵袭能力,Westem blot蛋白印迹检测ACC-2细胞中ERKI/2的活化及MMP-2的表达情况,实时荧光定量聚合酶链反应检测ACC-2细胞中MMP-2的mRNA的表达情况.结果 TGF-β1及UO126干预后,ACC-2细胞增殖能力无明显变化;TGF-β1刺激可增强ACC-2细胞迁移、侵袭能力,增加ACC-2细胞p-ERKI/2和MMP-2蛋白以及MMP-2 mRNA的表达.而UO126阻断ERK磷酸化后,抑制了TGF-β1刺激的增强作用,ACC-2细胞的迁移、侵袭能力降低,MMP-2蛋白和mRNA的表达均下降.结论 TGF-β1/ERKl/2/MMP-2通路参与了人唾液腺ACC侵袭和迁移能力的调节.TGF-β1可通过上调ERK1/2,继而上调MMP-2,促进人唾液腺ACC细胞的侵袭和迁移能力,ERKl/2可能成为人唾液腺ACC侵袭防治的新靶点.     

Specific VEGF sequestering to biomaterials: Influence of serum stability

Vascular endothelial growth factor (VEGF) was originally discovered as a tumor-derived factor that is able to induce endothelial cell behavior associated with angiogenesis. It has been implicated during wound healing for the induction of endothelial cell proliferation, tube formation and blood vessel remodeling. However, previous investigations into the biological effect of VEGF concluded that a particular range of growth factor concentrations are required for healthy vasculature to form, motivating recent studies to regulate VEGF activity via molecular sequestering to biomaterials. Numerous VEGF sequestering strategies have been developed, and they have typically relied on extracellular matrix mimicking moieties that are not specific for VEGF and can affect many growth factors simultaneously. We describe here a strategy for efficient, specific VEGF sequestering with poly(ethylene glycol) (PEG) microspheres, using peptides designed to mimic VEGF receptor type 2 (VEGFR2). By immobilizing two distinct peptides with different serum stabilities, we examined the effect of serum on the specific interaction between peptide-containing PEG microspheres and VEGF. We addressed the hypothesis that VEGF sequestering in serum-containing solutions would be influenced by the serum stability of the VEGF-binding peptide. We further hypothesized that soluble VEGF could be sequestered in serum-containing cell culture media, resulting in decreased VEGF-dependent proliferation of human umbilical vein endothelial cells. We show that soluble VEGF concentration can be effectively regulated in serum-containing environments via specific molecular sequestering, which suggests potential clinical applications.     

In vivo response to dynamic hyaluronic acid hydrogels

Tissue-specific elasticity arises in part from developmental changes in extracellular matrix over time, e.g. ～10-fold myocardial stiffening in the chicken embryo. When this time-dependent stiffening has been mimicked in vitro with thiolated hyaluronic acid (HA-SH) hydrogels, improved cardiomyocyte maturation has been observed. However, host interactions, matrix polymerization, and the stiffening kinetics remain uncertain in vivo, and each plays a critical role in therapeutic applications using HA-SH. Hematological and histological analysis of subcutaneously injected HA-SH hydrogels showed minimal systemic immune response and host cell infiltration. Most importantly, subcutaneously injected HA-SH hydrogels exhibited time-dependent porosity and stiffness changes at a rate similar to hydrogels polymerized in vitro. When injected intramyocardially host cells begin to actively degrade HA-SH hydrogels within 1 week post-injection, continuing this process while producing matrix to nearly replace the hydrogel within 1 month post-injection. While non-thiolated HA did not degrade after injection into the myocardium, it also did not elicit an immune response, unlike HA-SH, where visible granulomas and macrophage infiltration were present 1 month post-injection, likely due to reactive thiol groups. Altogether these data suggest that the HA-SH hydrogel responds appropriately in a less vascularized niche and stiffens as had been demonstrated in vitro, but in more vascularized tissues, in vivo applicability appears limited.     

Regulator of G-protein signaling 2 inhibits acid-induced mucin5AC hypersecretion in human airway epithelial cells

Mucus hypersecretion is a common pathological feature of inflammatory airway diseases. Previous studies have shown that acidic microenvironment of inflamed airways may provoke the pathophysiology of inflammatory airway diseases. However, the acidic-sensing and negative regulatory mechanisms that mediate mucus hypersecretion in inflamed airway remain elusive. Thus, we sought to explore the role of ovarian cancer G-protein-coupled receptor 1 (OGR1) in acid-induced mucin5AC (MUC5AC) hypersecretion in human airway epithelium and the inhibitory effect of regulator of G-protein signaling 2 (RGS2) in this process. We found that airway acidification increased [Ca²⁺]_i, which was required for MUC5AC secretion. Knocking-down OGR1 and G_q with siRNAs and pretreating cells with phospholipase C inhibitor effectively attenuated acid-induced cellular responses. Moreover, the overexpression of wild-type RGS2 attenuated acid-induced cellular responses. In contrast, reducing RGS2 with siRNA enhanced the increases in acid-induced cellular responses. These data suggest that airway acidification can induce MUC5AC hypersecretion through an OGR1-mediated mechanism and RGS2 can inhibit acid-induced MUC5AC hypersecretion at OGR1 receptor level.