血清TM、P-selectin、HDL-C以及凝血功能指标与创伤性骨折患者术后深静脉血栓形成的关系研究

摘要 目的：探讨血清血栓调节蛋白（TM）、P-选择素（P-selectin）、高密度脂蛋白（HDL-C）以及凝血功能指标与创伤性骨折患者术后深静脉血栓形成（DVT）的关系。方法：选择2018年1月至2020年1月我院收治的术后发生DVT的创伤性骨折患者100例作为DVT组,同期术后未发生DVT的创伤性骨折患者100例作为无DVT组,比较两组血清TM、P-selectin、HDL-C、血浆凝血功能指标[凝血酶时间（TT）、凝血酶原时间（PT）、活化部分凝血酶时间（APTT）、纤维蛋白原（FIB）、D-二聚体（D-D）],应用Pearson相关性分析血清TM、P-selectin、HDL-C与凝血功能指标的相关性,应用多因素Logistic回归分析创伤性骨折患者术后DVT的影响因素。结果：DVT组血清TM、P-selectin、血浆D-D、FIB水平、年龄≥60岁比例高于无DVT组,血清HDL-C水平低于无DVT组（P＜0.05）。DVT组血清TM和P-selectin与血浆D-D、FIB呈正相关（P＜0.05）,血清HDL-C与血浆D-D、FIB呈负相关（P＜0.05）。多因素Logistic回归分析显示,年龄≥60岁、血清TM≥9.50 IU/mL、P-selectin ≥70.00 ng/mL、HDL-C<1.00 mmol/L、血浆D-D≥700.00 μg/L、FIB≥4.00 g/L是创伤性骨折患者术后DVT的危险因素（P＜0.05）。结论：创伤性骨折患者术后发生DVT患者血清TM、P-selectin较无DVT患者升高,HDL-C较无DVT患者降低,联合检测血清TM、P-selectin、HDL-C和凝血功能指标可能有助于降低DVT的发生风险。     

P-selectin glycoprotein ligand 1 promotes T cell lymphoma development and dissemination

P-selectin glycoprotein ligand-1 (PSGL-1) is a membrane-bound glycoprotein expressed in lymphoid and myeloid cells. It is a ligand of P-, E- and L-selectin and is involved in T cell trafficking and homing to lymphoid tissues, among other functions. PSGL-1 expression has been implicated in different lymphoid malignancies, so here we aimed to evaluate the involvement of PSGL-1 in T cell lymphomagenesis and dissemination. PSGL-1 was highly expressed at the surface of human and mouse T cell leukemia and lymphoma cell lines. To assess its impact on T cell malignancies, we stably expressed human PSGL-1 (hPSGL-1) in a mouse thymic lymphoma cell line, which expresses low levels of endogenous PSGL-1 at the cell surface. hPSGL-1-expressing lymphoma cells developed subcutaneous tumors in athymic nude mice recipients faster than control empty vector or parental cells. Moreover, the kidneys, lungs and liver of tumor-bearing mice were infiltrated by hPSGL-1-expressing malignant T cells. To evaluate the role of PSGL-1 in lymphoma cell dissemination, we injected intravenously control and hPSGL-1-expressing lymphoma cells in athymic mice. Strikingly, PSGL-1 expression facilitated disease infiltration of the kidneys, as determined by histological analysis and anti-CD3 immunohistochemistry. Together, these results indicate that PSGL-1 expression promotes T cell lymphoma development and dissemination to different organs.     

Differential localization of P-selectin and von Willebrand factor during megakaryocyte maturation

Abstract

An important step in megakaryocyte maturation is the appropriate assembly of at least two distinct subsets of α-granules. The mechanism that sorts the α-granule components into distinct structures and mediates their release in response to specific stimuli is now emerging. P-selectin and von Willebrand factor are two proteins present in the α-granules that recognize P-selectin glycoprotein ligand on neutrophils and collagen in the subendothelial matrix. These proteins may play an important role in determining the differential release of the α-granule contents in response to external stimuli. If P-selectin and von Willebrand factor are localized in the same or different α-granules is not known. To clarify this question, we analyzed by immunoelectron microscopy the localization of von Willebrand factor and P-selectin during the maturation of wild-type and Gata1^low megakaryocytes induced in vivo by treating animals with thrombopoietin. Gata1^low is a hypomorphic mutation that blocks megakaryocyte maturation, reduces the levels of von Willebrand factor expression and displaces P-selectin on the demarcation membrane system. The maturation block induced by this mutation is partially rescued by treatment in vivo with thrombopoietin. In immature megakaryocytes, both wild-type and Gata1^low, the two receptors were co-localized in the same cytoplasmic structures. By contrast, the two proteins were segregated to separate α-granule subsets as the megakaryocytes matured. These observations support the hypothesis that P-selectin and von Willebrand factor may ensure differential release of the α-granule content in response to external stimuli.     

T lymphocytes migrate upstream after completing the leukocyte adhesion cascade

The leukocyte adhesion cascade is of critical importance for both the maintenance of immune homeostasis and the ability of immune cells to perform effector functions. Here, we present data showing CD4⁺ T cells migrate upstream (against the direction of flow) after completing the leukocyte adhesion cascade on surfaces displaying either ICAM-1 or ICAM-1 and VCAM-1, but migrate downstream on surfaces displaying only VCAM-1. Cells completing the cascade on HUVECs initially migrate upstream before reverting to more random migration, partly caused by transmigration of cells migrating against the flow. Furthermore, cells migrating upstream transmigrate faster than cells migrating downstream. On HUVECs, blocking interactions between LFA-1 and ICAM-1 resulted in downstream migration and slower transmigration. These results further suggest a possible physiological role for upstream migration in vivo.     

AP-3 adaptor functions in targeting P-selectin to secretory granules in endothelial cells

P-selectin, a cell adhesion protein participating in the early stages of inflammation, contains multiple sorting signals that regulate its cell surface expression. Targeting to secretory granules regulates delivery of P-selectin to the cell surface. Internalization followed by sorting from early to late endosomes mediates rapid removal of P-selectin from the surface. We show here that the P-selectin cytoplasmic domain bound AP-2 and AP-3 adaptor complexes in vitro . The amino acid substitution L768A, which abolishes endosomal sorting and impairs granule targeting of P-selectin, reduced binding of AP-3 adaptors but not AP-2 adaptors. Turnover of P-selectin was 2.4-fold faster than turnover of transferrin receptor in AP-3-deficient mocha fibroblasts, similar to turnover of these two proteins in AP-3-competent cells, demonstrating that AP-3 function is not required for endosomal sorting. However, sorting P-selectin to secretory granules was defective in endothelial cells from AP-3-deficient pearl mice, demonstrating a role for AP-3 adaptors in granule assembly in endothelial cells. P-selectin sorting to platelet α-granules was normal in pearl mice, consistent with earlier evidence that granule targeting of P-selectin is mechanistically distinct in endothelial cells and platelets. These observations establish that AP-3 adaptor functions in assembly of conventional secretory granules, in addition to lysosomes and the 'lysosome-like' secretory granules of platelets and melanocytes.     

Weibel-Palade body membrane proteins exhibit differential trafficking after exocytosis in endothelial cells

Weibel-Palade bodies, the secretory granules of endothelial cells, possess two different membrane proteins. However, P-selectin is seen only in Weibel-Palade bodies in HUVECs, whereas CD63 is also seen in late endosomes/lysosomes. Since P-selectin is targeted to lysosomes in heterologous expression studies, we have determined whether a lysosomal targeting signal also operates within HUVECs. We have also examined the trafficking of CD63 to its two different intracellular locations. By following antibodies bound at the plasma membrane during stimulation, we have discovered that while half of the P-selectin recycles to the WPBs, 50% is rapidly delivered to a lamp-1-positive compartment. Thus, the lysosomal targeting signal of this protein also operates in HUVECs. CD63 is found constitutively at the cell surface of HUVECs and most of it is delivered to the late endosomes/lysosomes after internalisation. However, stimulation causes both a rise in the CD63 plasma membrane level and in the amount that recycles to the WPBs. Our data strongly suggest that the CD63 that originates in the WPB preferentially recycles to the granule rather than being delivered to the late endosome/lysosome, and that there are, therefore, two separate pools of this protein within HUVECs. Our findings indicate that although P-selectin and CD63 are both targeted to the same compartments from the PM, the kinetics and the ratio of their targeting to Weibel-Palade bodies versus lysosomes are very different.     

急性ST段抬高型心肌梗死患者血清NT-proBNP、P-selectin联合IMA预测PCI术后心电图ST段回落的临床价值研究

摘要 目的：探讨急性ST段抬高型心肌梗死（ASTEMI）患者血清N末端B型利钠肽前体（NT-proBNP）、P-选择素（P-selectin）联合缺血修饰白蛋白（IMA）预测经皮冠状动脉介入治疗（PCI）术后心电图ST段回落（STR）不良的临床价值。方法：选取2020年1月～2022年7月南京医科大学第二附属医院急诊科收治的100例ASTEMI患者，根据PCI术后心电图STR分为STR不良组和STR良好组，另选取同期50名体检健康志愿者为对照组。采用酶联免疫吸附法检测血清NT-proBNP、P-selectin和IMA水平。采用多因素Logistic回归分析ASTEMI患者PCI术后心电图STR不良的影响因素，采用受试者工作特征（ROC）曲线分析血清NT-proBNP、P-selectin、IMA水平对ASTEMI患者PCI术后心电图STR不良的预测价值。结果：与对照组比较，ASTEMI组PCI术前血清NT-proBNP、P-selectin和IMA水平升高（P＜0.05）。根据心电图STR将ASTEMI患者分为STR不良组35例和STR良好组65例。STR不良组与STR良好组PCI术后血清NT-proBNP、P-selectin和IMA水平低于PCI术前（P＜0.05）；STR不良组PCI术前和PCI术后血清NT-proBNP、P-selectin和IMA水平高于STR良好组（P＜0.05）。STR不良组Killip分级≥2级比例和肌钙蛋白I高于STR良好组，ST段偏差总和低于STR良好组（P＜0.05）。多因素Logistic回归分析显示，Killip分级≥2级和NT-proBNP、P-selectin、IMA升高为ASTEMI患者PCI术后心电图STR不良的独立危险因素（P＜0.05）。ROC曲线分析显示，血清NT-proBNP、P-selectin联合IMA预测ASTEMI患者PCI术后心电图STR不良的曲线下面积（AUC）大于NT-proBNP、P-selectin和IMA单独预测。结论：血清NT-proBNP、P-selectin和IMA水平升高与ASTEMI患者PCI术后心电图STR不良独立相关，三者联合预测ASTEMI患者PCI术后心电图STR不良的价值较高。     

P-selectin and CD63 use different mechanisms for delivery to Weibel-Palade bodies

The biogenesis of endothelial-specific Weibel-Palade bodies (WPB) is poorly understood, despite their key role in both haemostasis and inflammation. Biogenesis of specialized organelles of haemopoietic cells is often adaptor protein complex 3-dependent (AP-3-dependent), and AP-3 has previously been shown to play a role in the trafficking of both WPB membrane proteins, P-selectin and CD63. However, WPB are thought to form at the trans Golgi network (TGN), which is inconsistent with a role for AP-3, which operates in post-Golgi trafficking. We have therefore investigated in detail the mechanisms of delivery of these two membrane proteins to WPB. We find that P-selectin is recruited to forming WPB in the trans-Golgi by AP-3-independent mechanisms that use sorting information within both the cytoplasmic tail and the lumenal domain of the receptor. In contrast, CD63 is recruited to already-budded WPB by an AP-3-dependent route. These different mechanisms of recruitment lead to the presence of distinct immature and mature populations of WPB in human umbilical vein endothelial cells (HUVEC).     

A high affinity human antibody antagonist of P-selectin mediated rolling

We have characterized the IgG form of a previously isolated and engineered single-chain Fv (scFv), named RR2r3s4-1, that binds to human PSGL-1. This fully human IgG was determined to have a Kd of 1.8+/-0.7 nM by fluorescence quenching titration. It better inhibits P-selectin-PSGL-1 interactions than a commercially available murine monoclonal antibody KPL1 and better inhibits neutrophil rolling than KPL1. Thus, RR2r3s4-1 is the most effective antibody at inhibiting P-selectin-PSGL-1 interactions known. Specificity analysis reveals that RR2r3s4-1 does not cross react with murine PSGL-1 and thus requires more than tyrosine sulfate for binding to human PSGL-1. This evidence demonstrates the therapeutic potential of this antibody as a potent anti-inflammatory therapeutic.     

The utility of inflammation and platelet biomarkers in patients with acute coronary syndromes

Introduction

Thrombotic and inflammatory mechanisms are involved in the pathophysiology of acute coronary syndrome (ACS). The aim of the study was the evaluation of inflammation (white blood cells count/WBC, C-reactive protein/CRP, interleukin-6/IL-6) and platelet (platelet count/PLT, mean platelet volume/MPV, large platelet/LPLT, beta-thromboglobulin/β-TG) biomarkers in the groups of ACS patients depending on the severity of signs and symptoms and compared to controls without coronary artery disease.