Imaging of muscle activity-induced morphometric changes in fibril network of myofascia by two-photon microscopy

Myofascia, deep fascia enveloping skeletal muscles, consists of abundant collagen and elastin fibres that play a key role in the transmission of muscular forces. However, understanding of biomechanical dynamics in myofascia remains very limited due to less quantitative and relevant approaches for in vivo examination. The purpose of this study was to evaluate the myofascial fibril structure by means of a quantitative approach using two-photon microscopy (TPM) imaging in combination with intravital staining of Evans blue dye (EBD), a far-red fluorescence dye, which potentially labels elastin. With focus on myofascia of the tibial anterior (TA) muscle, the fibril structure intravitally stained with EBD was observed at the depth level of collagen fibrous membrane above the muscle belly. The EBD-labelled fibril structure and orientation in myofascia indicated biomechanical responses to muscle activity and ageing. The orientation histograms of EBD-labelled fibrils were significantly modified depending upon the intensity of muscle activity and ageing. Moreover, the density of EBD-labelled fibrils in myofascia decreased with habitual exercise but increased with muscle immobilization or ageing. In particular, the diameter of EBD-labelled fibrils in aged mice was significantly higher. The orientation histograms of EBD-labelled fibrils after habitual exercise, muscle immobilization and ageing showed significant differences compared to control. Indeed, the histograms in bilateral TA myofascia of exercise mice made simple waveforms without multiple sharp peaks, whilst muscular immobilization or ageing significantly shifted a histogram with sustaining multiple sharp peaks. Therefore, the dynamics of fibre network with EBD fluorescence in response to the biomechanical environment possibly indicate functional tissue adaptation in myofascia. Furthermore, on the basis of the knowledge that neutrophil recruitment occurs locally in working muscles, we suggested the unique reconstruction mechanism involving neutrophilic elastase in the myofascial fibril structure. In addition to the elastolytic susceptibility of EBD-labelled fibrils, distinct immunoreactivities and activities of neutrophil elastase in the myofascia were observed after electric pulse stimulation-induced muscle contraction for 15 min. Our findings of EBD-labelled fibril dynamics in myofascia through quantitative approach using TPM imaging and intravital fluorescence labelling potentially brings new insights to examine muscle physiology and pathology.     

Neutrophil and platelet antibodies in autoimmune lymphoproliferative syndrome

BACKGROUND AND OBJECTIVES: Autoimmune lymphoproliferative syndrome (ALPS), is an inherited disorder characterized by defective lymphocyte apoptosis, lymphadenopathy, splenomegaly, accumulation of T-cell receptor (TCR)-alphabeta+ CD4- CD8- T cells (double-negative T cells) and autoimmunity. We investigated the incidence and nature of neutrophil and platelet antibodies in patients with ALPS. MATERIALS AND METHODS: Sera from 26 patients with ALPS were tested for neutrophil antibodies by granulocyte immunofluorescence, granulocyte agglutination and monoclonal antibody immobilization assays of granulocyte antigens, and for platelet antibodies using a solid-phase antibody-detection system. RESULTS: Neutrophil antibodies were detected in 46% of patients with ALPS. Antibody specificity could be defined in eight of the 12 patients with neutrophil antibodies. Among these eight patients, four had antibodies directed against more than one antigen. Overall, 14 antibodies directed to specific antigens were identified: three were directed to the HNA-1a antigen of FcgammaRIIIb; two to the HNA-1b antigen of Fcgamma-RIIIb; two to epitopes common to all FcgammaRIIIb molecules; four to the HNA-2a antigen of the NB1 glycoprotein; and three to neutrophil beta2 integrins. Platelet antibodies were detected in 35% of patients with ALPS. No antibody specificities were identified among the platelet antibodies. There was no association between the detection of neutrophil antibodies and a history of clinical neutropenia, or between the detection of platelet antibodies and a history of clinical thromobocytopenia. CONCLUSIONS: Neutrophil and platelet antibodies are important markers of ALPS, but do not always cause clinical cytopenias. The specificities of neutrophil antibody were similar to those found in children with autoimmune neutropenia but without ALPS.     

Azithromycin Enhances Phagocytic Killing of Aggregatibacter actinomycetemcomitans Y4 by Human Neutrophils

Background: Aggregatibacter actinomycetemcomitans resists killing by neutrophils and is inhibited by azithromycin (AZM) and amoxicillin (AMX). AZM actively concentrates inside host cells, whereas AMX enters by diffusion. The present study is conducted to determine whether AZM is more effective than AMX at enhancing phagocytic killing of A. actinomycetemcomitans by neutrophils. Methods: Killing assays were conducted in the presence of either 2 μg/mL AZM or 16 μg/mL AMX (equipotent against A. actinomycetemcomitans). Neutrophils were loaded by incubation with the appropriate antibiotic. Opsonized A. actinomycetemcomitans strain Y4 was incubated with the indicated antibiotic alone, with loaded neutrophils and antibiotic, or with control neutrophils (without antibiotic) at multiplicities of infection (MOIs) of 30 and 90 bacteria per neutrophil. Results: Neutrophil incubation with 2 μg/mL AZM yielded an intracellular concentration of 10 μg/mL. At an MOI of 30, neutrophils loaded with AZM failed to kill significantly more bacteria than control neutrophils during the 60‐ and 90‐minute assay periods. At an MOI of 90, neutrophils loaded with AZM killed significantly more bacteria than either AZM alone or control neutrophils during 60‐ and 90‐minute incubations (P <0.05), and killed significantly more bacteria after 90 minutes than the sum of the killing produced by AZM alone or neutrophils alone. Neutrophils incubated with AMX under identical conditions also killed significantly more bacteria than either AMX alone or control neutrophils, but there was no evidence of synergism between AMX and neutrophils. Conclusions: Neutrophils possess a concentrative transport system for AZM that may enhance killing of A. actinomycetemcomitans. Its effects are most pronounced when neutrophils are greatly outnumbered by bacteria.     

多指标联合检测在肝硬化合并感染诊断中的价值

目的 分析降钙素原(PCT)、白介素6(IL-6)、C反应蛋白(CRP)、中性粒细胞CD64、白细胞总数(WBC)、中性粒细胞百分比(NEU%)等多种指标诊断肝硬化患者早期感染的临床价值。方法 将肝硬化住院患者按照细菌培养结果和临床症状分为感染组和非感染组,血清PCT、IL-6采用化学发光分析仪进行检测,血清CRP采用生化分析仪进行检测,中性粒细胞CD64采用流式细胞分析仪进行检测,WBC和NEU%采用全血细胞分析仪进行检测。数据采用Logistic回归和ROC曲线进行分析。 结果 感染组各个指标均显著高于非感染组(P<0.01); Logistic回归分析结果表明PCT、IL-6和CD64具有早期预测肝硬化合并感染的能力,其OR值分别为7.199(95%CI,2.180-23.771),1.010(95%CI,1.002-1.017)和2.312(95%CI,1.485-3.600)。然而CRP、WBC和NEU%则不具有早期预测价值;ROC曲线分析结果显示,PCT、IL-6和CD64的曲线下面积AUC分别为0.791(95%CI,0.727-0.856),0.762(95%CI,0.693-0.832)和0.884(95%CI,0.835-0.933);三者联合检测的ROC曲线下面积AUC为0.932(95%CI,0.897-0.967),诊断准确率为86.9%。 结论 PCT、IL-6和CD64可以作为早期诊断肝硬化合并感染的指标,三者联合检测能够提高临床诊断效率。     

牙周致病菌免疫逃逸机制的研究进展

牙周致病菌可引起牙周组织慢性炎症,由其引发的宿主过度的免疫炎症反应会进一步加重牙周组织的损伤。本文就龈沟的天然免疫防御机制,微生物对龈沟防御机制的敏感性,中性粒细胞和牙龈上皮细胞对牙周致病菌的免疫反应进行系统阐述,旨在表明真正的牙周致病菌因不能很好地激发或者抑制宿主的免疫反应,从而逃避宿主的免疫防御,导致牙周炎的发生,为牙周病的治疗和早期预防提供参考依据。