Monomeric myosin V uses two binding regions for the assembly of stable translocation complexes

Myosin-motors are conserved from yeast to human and transport a great variety of cargoes. Most plus-end directed myosins, which constitute the vast majority of all myosin motors, form stable dimers and interact constitutively with their cargo complexes. To date, little is known about regulatory mechanisms for cargo-complex assembly. In this study, we show that the type V myosin Myo4p binds to its cargo via two distinct binding regions, the C-terminal tail and a coiled-coil domain-containing fragment. Furthermore, we find that Myo4p is strictly monomeric at physiologic concentrations. Because type V myosins are thought to require dimerization for processive movement, a mechanism must be in place to ensure that oligomeric Myo4p is incorporated into cargo-translocation complexes. Indeed, we find that artificial dimerization of the Myo4p C-terminal tail promotes stabilization of myosin-cargo complexes, suggesting that full-length Myo4p dimerizes in the cocomplex as well. We also combined the Myo4p C-terminal tail with the coiled-coil region, lever arm, and motor domain from a different myosin to form constitutively dimeric motor proteins. This heterologous motor successfully translocates its cargo in vivo, suggesting that wild-type Myo4p may also function as a dimer during cargo-complex transport.     

急性胸痛患者联合检测cTn-I、CK-MBmass和Myo的临床价值

目的：了解急性胸痛患者胸痛发作初期时血液中心肌钙蛋白－I（cTn-I）、肌酸激酶同工酶（CK-MBmass）和肌红蛋白（Myo）浓度的动态变化，探讨3项指标联合检测用于心肌梗死（AMI）早期诊断的临床价值。方法：于患者胸痛发生的1－2h、3－4h、5－6h，分别采血，用荧光免疫干片法同时定量检测cTn-I、CK-MBmass、Myo值。结果：42例急性胸痛患者最终确诊为AM125例，其3－4h、5－6hcTn-I、CK-MBmass和Myo的测定值较非AMI患者均明显升高（P＜0．05或0．01）；AMI患者3－4h、5－6h、cTn-I的阳性率较非AMI患者 显著升高（P＜0．01），1－2h 、3-4h、5-6h CK-MBmass和Myo的阳性率均较非AMI患者显著升高（P＜0．05或0．01）。结论：用荧光免疫干片法联合检测急性胸痛患者血中cTn-I、CK-MBmass、Myo浓度快速、简便、所得结果特异、敏感、稳定，有助于快速诊断AMI，为临床及时有效治疗AMI、降低死亡率创造条件。     

Tissue-associated self-antigens containing exosomes: Role in allograft rejection

Exosomes are extracellular vesicles that express self-antigens (SAgs) and donor human leukocyte antigens. Tissue-specific exosomes can be detected in the circulation following lung, heart, kidney and islet cell transplantations. We collected serum samples from patients who had undergone lung (n?=?30), heart (n?=?8), or kidney (n?=?15) transplantations to isolate circulating exosomes. Exosome purity was analyzed by Western blot, using CD9 exosome-specific markers. Tissue-associated lung SAgs, collagen V (Col-V) and K-alpha 1 tubulin (Kα1T), heart SAgs, myosin and vimentin, and kidney SAgs, fibronectin and collagen IV (Col-IV), were identified using western blot. Lung transplant recipients diagnosed with bronchiolitis obliterans syndrome had exosomes with higher expression of Col-V (4.2-fold) and Kα1T (37.1-fold) than stable. Exosomes isolated from heart transplant recipients diagnosed with coronary artery vasculopathy had a 3.9-fold increase in myosin and a 4.7-fold increase in vimentin compared with stable. Further, Kidney transplant recipients diagnosed with transplant glomerulopathy had circulating exosomes with a 2-fold increased expression of fibronectin and 2.5-fold increase in Col-IV compared with stable. We conclude that circulating exosomes with tissue associated SAgs have the potential to be a noninvasive biomarker for allograft rejection.     

A review on static splinting therapy to prevent burn scar contracture: Do clinical and experimental data warrant its clinical application?

Background

Static splinting therapy is widely considered an essential part in burn rehabilitation to prevent scar contractures in the early phase of wound healing. However, scar contractures are still a common complication. In this article we review the information concerning the incidence of scar contracture, the effectiveness of static splinting therapy in preventing scar contractures, and specifically focus on the – possible – working mechanism of static-splinting, i.e. mechanical load, at the cellular and molecular level of the healing burn wound.

Method

A literature search was done including Pubmed, Cochrane library, CINAHL and PEDRO.

Results

Incidence of scar contracture in patients with burns varied from 5% to 40%. No strong evidence for the effectiveness of static splinting therapy in preventing scar contracture was found, whereas in vitro and animal studies demonstrated that mechanical tension will stimulate the myofibroblast activity, resulting in the synthesis of new extracellular matrix and the maintenance of their contractile activity.

Conclusion

The effect of mechanical tension on the wound healing process suggests that static splinting therapy may counteract its own purpose. This review stresses the need for randomised controlled clinical trials to establish if static splinting to prevent contractures is a well-considered intervention or just wishful thinking.     

Transduced viral IL-10 is exocytosed from lacrimal acinar secretory vesicles in a myosin-dependent manner in response to carbachol

The purpose of this study was to determine the intracellular trafficking and release pathways for the therapeutic protein, viral IL-10 (vIL-10), from transduced acinar epithelial cells from rabbit lacrimal gland. Primary cultured rabbit lacrimal gland acinar cells (LGACs) were transduced with adenovirus serotype 5 containing viral interleukin-10 (AdvIL-10). The distribution of vIL-10 was assessed by confocal fluorescence microscopy. Carbachol (CCH)-stimulated release of vIL-10 was quantified by ELISA. vIL-10 localization and exocytosis was probed in response to treatments with agents modulating actin- and myosin-based transport. vIL-10 immunoreactivity was detected in large intracellular vesicles in transduced LGAC. vIL-10 was partially co-localized with biosynthetic but not endosomal compartment markers. vIL-10 release was sensitive to CCH, and the kinetics of release showed an initial burst phase that was similar but not identical to that of the secretory protein, β-hexosaminidase. Disassembly of actin filaments with latrunculin B significantly increased CCH-stimulated vIL-10 secretion, suggesting that vIL-10 was released from stores sequestered beneath the subapical actin barrier. That release required the activity of actin-dependent myosin motors previously implicated in secretory vesicle exocytosis was confirmed by findings that CCH-stimulated vIL-10 release was reduced by inhibition of non-muscle myosin 2 and myosin 5c function, using ML-7 and overexpression of dominant negative myosin 5c, respectively. These results suggest that the majority of vIL-10 transgene product is packaged into a subpopulation of secretory vesicles that utilize actin-dependent myosin motors for aspects of actin coat assembly, compound fusion and exocytosis at the apical plasma membrane in response to CCH stimulation.     

应用明胶海绵栓塞法建立猪的心肌梗死模型

目的经心导管用明胶海绵栓子栓塞猪的冠状动脉左前降支(LAD),建立心肌梗死的大动物模型。方法对7头猪用明胶海绵经导管栓塞LAD第二对角支远端,致前室壁梗死。四周后行心脏超声、冠状动脉造影及组织学检查。结果手术死亡率为1/7。与对照组相比,心肌梗死(MI)组左室舒张未内径(LVEDd)增大由37.0mm±3.4mm增至50.8mm±6.1mm(P<0.01),射血分数(EF)由62.3%±2.9%下降为36.6%±2.1%(P<0.001);复查冠脉造影提示LAD栓塞仍存在。MI组的左室被动容积为81.4ml±4.3ml,梗死区面积/左室心肌面积为18.4%±1.6%,梗死区厚3.5mm±0.8mm。心肌组织马森染色显示梗塞交界区有大块纤维形成,梗塞区有片状纤维形成。结论这种模型接近临床病理生理过程,稳定可靠,重复性好,可作为研究心肌梗死的技术平台。     

高敏肌钙蛋白T检测在心肌梗死中应用价值的研究

目的:观察高敏肌钙蛋白T(hs-cTnT)、肌酸激酶同工酶质量(CK-MBmass)、肌红蛋白(Myo)和肌钙蛋白T(cTnT)在疾病发生及发展过程中的变化特征,研究高敏肌钙蛋白T检测在诊断急性心肌梗死方面的应用价值。方法:选取100例急性心肌梗死患者,分别在0 h、4 h、12 h抽取静脉血,正常对照组为68例空腹抽取静脉血,分别测定4项指标的含量,比较它们在不同时间内对AMI的诊断特性。结果:AMI患者发病入院后,hs-TnT的敏感性为91%,特异性为83%,阳性预测值为89%,阴性预测值为86%。结论:高敏心肌肌钙蛋白具有最好的灵敏度和较好的组织特异性,适合作为AMI早期检测指标。     

Hereditary human myopathies in muscle culture

In this article I illustrated the use of regenerating human muscle cultures for studying the hereditary human myopathies. Although some of the data are still controversial, they do point up the great potential of this “in vitro system”. For hereditary myopathies due to developmentally regulated proteins that are expressed only at a more advanced stage of muscle differentiation, the use of highly differentiated nerve-muscle cocultures might contribute significantly to a better understanding of their developmental pathogenesis. More advanced techniques (permanent human muscle cell lines, heterokaryons, myoblast transfer, gene transfer, myogenic conversion of human non-muscle cells, cybrid clones) may provide a great deal of information at molecular level and may also have practical applications in the diagnosis or even in the treatment of hereditary human myopathies.

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Sommario L'Autore illustra l'utilità e i limiti delle colture di muscolo umano nello studio delle miopatie umane ereditarie. In particolare nelle miopatie dovute a difetti di enzimi-specifici muscolari, l'utilizzo di tecniche di cocoltura muscolo-nervo con avanzato grado di maturazione muscolare permette di delucidare i meccanismi molecolari patogenetici di tali malattie. L'uso di avanzate tecniche di biologia molecolare e cellulare, quali linee permanenti, trapianto di mioblasti, trasferimento genico e di mitocondri, oltre che fornire utili informazioni a livello molecolare potranno trovare applicazione, in futuro, persino nella terapia di molte miopatie ereditarie.