Expression and activity of thrombomodulin in human gingival epithelium: in vivo and in vitro studies

Epidermal keratinocytes thrombomodulin (TM) has been shown to regulate thrombin at sites of cutaneous injury in addition to a role for epidermal differentiation. TM, a major anticoagulant proteoglycan of the endothelial cell membrane, is a thrombin receptor that acts as a co‐factor for protein C activation. Thrombin has pro‐inflammatory effects for periodontitis. However, little is known about TM in gingival tissue with periodontitis. We used immunohistochemistry to examine expression of TM in gingival epithelium from patients with periodontitis. In vitro , we observed TM expression at varying Ca²⁺ concentrations by confocal laser scanning microscopy, examined the expression of TM mRNA and tested TM co‐factor activity. Furthermore, we measured TM concentration in gingival crevicular fluid (GCF) from 11 severe adult cases of periodontitis using enzyme‐linked immunosorbent assay. Immunoreactive TM was present in gingival epithelium and junctional epithelium, and was reduced in inflamed gingival epithelium compared to healthy gingival epithelium. Ultrastructurally, TM, including microvilli, was observed on the cell membrane. TM localization in cells cultured in 0.09 m m Ca²⁺ differed from that in cells exposed to 1.2 m m Ca²⁺. Northern analysis demonstrated TM mRNA in gingival keratinocytes more than in human umbilical vein endothelial cells (HUVEC). Gingival keratinocytes also facilitated protein C activation by thrombin, although less strongly than HUVEC. TM in GCF at sites with bleeding on probing in patients was significantly elevated ( p <0.001, Student's t ‐test). TM in gingival epithelium may regulate thrombin activity at sites of coagulation and inflammation with periodontal disease, although inflammation may impair this regulation of thrombin.     

血栓调节蛋白抑制慢性肾脏病微炎症的机制研究进展*

慢性肾脏病患者体内普遍存在一种微炎症状态,这种持续存在的炎症状态会进一步加重肾脏损 伤。如何改善慢性肾脏病患者的微炎症状态是当今国内外研究关注的热点之一。而与肾脏血管内皮受损密切 相关的血栓调节蛋白是调控体内多种生理反应的关键位点,具有抗凝、抗炎、保护内皮细胞及维持血管稳态 的功能,可为慢性肾脏病抗炎治疗提供新的思路。因此,该文就血栓调节蛋白抑制慢性肾脏病微炎症的机制 展开综述。     

Increased thrombin generation in splanchnic vein thrombosis is related to the presence of liver cirrhosis and not to the thrombotic event

Introduction

In recent years there have been increasing evidence associating liver disease with hypercoagulability, rather than bleeding. The aim of the study was to evaluate the haemostatic potential in patients with liver disease.

Patients and methods

We measured thrombin generation in the presence and absence of thrombomodulin in patients with portal vein thrombosis (PVT, n = 47), Budd-Chiari syndrome (BCS, n = 15) and cirrhosis (n = 24) and compared the results to those obtained from healthy controls (n = 21). Fifteen patients with PVT and 10 patients with BCS were treated with warfarin and were compared to an equal number of patients with atrial fibrillation matched for prothrombin time-international normalized ratio. We assessed resistance to thrombomodulin by using ratios [marker measured in the presence/absence of thrombomodulin].

Results

There were no differences in thrombin generation between patients on warfarin treatment and their controls. Cirrhotic patients generated more thrombin in the presence of thrombomodulin and exhibited thrombomodulin resistance compared to controls [p = 0.006 for endogenous thrombin potential (ETP) and p < 0.001 for peak thrombin and both ratios ETP and peak] and patients with non-cirrhotic PVT (p = 0.001, p = 0.006, p < 0.001, p < 0.001 for ETP, peak, ratio ETP, ratio peak, respectively). The patients with cirrhotic PVT exhibited higher ETP (p = 0.044) and peak (p = 0.02) in the presence of thrombomodulin than controls, as well as thrombomodulin resistance (ETP and peak ratios: p = 0.001).

Conclusions

Hypercoagulability and thrombomodulin resistance in patients with cirrhosis were independent of the presence of splanchnic vein thrombosis. The hypercoagulability in patients with cirrhotic PVT could have implications for considering longer or more intensive treatment with anticoagulants in this group.     

2型糖尿病患者血浆P-选择素和血栓调节蛋白水平变化及其临床意义

目的：通过检测2型糖尿病患者血浆P-选择素和血栓调节蛋白水平变化，探讨检测患者血浆P-选择素和血栓调节蛋白的临床意义。方法：采用ELISA法检测了95例2型糖尿病患者和50例健康对照组血浆P-选择素和血栓调节蛋白水平。结果：2型糖尿病患者血浆P-选择素和血栓调节蛋白水平分别依次升高，且相差显著，与对照组相比有显著性差异（P〈0．01）；2型糖尿病患者血浆P-选择素与血栓调节蛋白水平呈显著的正相关。结论：检测血浆P-选择素和血栓调节蛋白水平，可作为2型糖尿病并发血管病变、病情判定以及治疗效果观察的指标。     

可溶性P-选择素及血栓调节蛋白在多器官功能障碍综合征早期诊断中的意义

目的 :观察多器官功能障碍综合征 (MODS)患者的内皮细胞功能改变 ,寻求早期诊断方法。方法 :采用免疫荧光技术连续监测 30例 ICU中严重感染、创伤患者外周血可溶性 P选择素 (s P选择素 )和血栓调节蛋白 (TM)浓度的动态变化 ,并计算其诊断 MODS的灵敏度、特异性和正确诊断指数。结果 :发生 MODS患者的 s P选择素、TM水平比未发生 MODS患者明显升高 (P均 <0 .0 5 )。 s P选择素对 MODS的诊断灵敏度高 ,但特异度不理想 ,诊断能力不高 ;提高诊断的截断值其特异度和正确诊断能力提高。TM对 MODS诊断的灵敏度高而特异度不足 ;诊断能力低 ;提升截断值后灵敏度下降 ,特异度提高 ,诊断能力提高 ,但 TM仍不足以作为独立的 MODS诊断指标。两个指标联合诊断意义更大。结论 :s P 选择素联合 TM有助于 MODS早期诊断     

Recombinant thrombomodulin ameliorates experimental autoimmune encephalomyelitis by suppressing high mobility group box 1 and inflammatory cytokines

Recombinant thrombomodulin (rTM) has pleiotrophic properties, including anti‐coagulation and anti‐inflammation; however, its effectiveness as a treatment for multiple sclerosis (MS) has not been evaluated fully. High mobility group box 1 (HMGB1) and proinflammatory cytokines, working as inflammatory mediators, are reportedly involved in the inflammatory pathogenesis of MS. The aim of this study was to determine whether rTM can be a potential therapeutic agent for experimental autoimmune encephalomyelitis (EAE). EAE mice received rTM treatment (1 mg or 0·1 mg/kg/day) from days 11 to 15 after immunization. The clinical variables, plasma levels of inflammatory cytokines and HMGB1 and pathological findings in EAE were evaluated. rTM administration ameliorated the clinical and pathological severity of EAE. An immunohistochemical study of the spinal cord showed weaker cytoplasmic HMGB1 staining in the rTM‐treated EAE mice than in the untreated EAE mice. Plasma levels of inflammatory cytokines and HMGB1 were suppressed by rTM treatment. In conclusion, rTM down‐regulated inflammatory mediators in the peripheral circulation and prevented HMGB1 release from nuclei in the central nervous system, suppressing EAE‐related inflammation. rTM could have a novel therapeutic potential for patients with MS.     

血必净注射液对脓毒症大鼠血栓调节蛋白及内皮蛋白C受体基因表达的影响

目的探讨血必净注射液对脓毒症大鼠血栓调节蛋白（TM）及内皮蛋白C受体（EPCR）基因表达的影响。方法采用盲肠结扎穿孔术（CLP）制备脓毒症模型。将96只健康Wistar大鼠按随机数字表法分为正常对照组、假手术组、模型组和血必净治疗组,后两组又按处死时间分为术后2、8、24、48和72h亚组,每组8只。留取肝、肺组织,分别检查各组动物组织TM和EPCR的mRNA表达。结果正常对照组和假手术组肝、肺组织TM和EPCR的mRNA有一定表达。CLP后2h肝、肺组织TM和EPCR的mRNA表达无明显变化（P均〉0.05）,8～48h肝、肺组织TM和EPCR的mRNA表达均有不同程度的增强（P均〈0.01）,至伤后72h基本恢复到术前水平（P均〉0.05）;与模型组比较,血必净治疗组CLP后8h和24h组织TM和EPCR的mRNA表达均有不同程度下降,48h和72h的基因表达有不同程度提高。结论血必净注射液可以从基因水平影响脓毒症动物组织TM及EPCR的基因表达。     

Loxosceles spider venom induces the release of thrombomodulin and endothelial protein C receptor: implications for the pathogenesis of intravascular coagulation as observed in loxoscelism

BACKGROUND: The venom of the spider Loxosceles can cause both local and systemic effects including disseminated intravascular coagulation. AIM: The aim of this study was to investigate the effects of the venom of Loxosceles intermedia (L. intermedia) and the purified Sphingomyelinase D (SMaseD) toxin upon the Protein C (PC) natural anticoagulant pathway. RESULTS: Both the venom and e purified SMaseD reduced the cell surface expression of thrombomodulin (TM) and Endothelial PC Receptor on endothelial cells in culture. The reduction of cell surface expression was caused by cleavage from the cell surface mediated by activation of an endogenous metalloproteinase. Reduction of TM and Endothelial PC Receptor on the surface of these cells resulted in an impaired ability of the cells to assist in the thrombin-induced activation of PC. CONCLUSION: This novel observation gives further insight into the mechanisms of the pathology induced by venom from Loxosceles spiders and may aid the development of a suitable therapy.