为探讨人乳头瘤病毒16(human papillomavirus 16,HPV16)E6癌蛋白对宫颈癌细胞的生物学行为及外泌体中β-联蛋白(β-catenin)和紧密连接蛋白(claudin-1)表达的影响,本研究利用RNA干扰技术建立HPV16 E6敲低细胞模型(shE6组),通过CCK8试剂盒、流式细胞仪、划痕实验和Transwell实验对细胞增殖、细胞周期、迁移和侵袭特征进行检测,发现shE6相对于对照组(NC组),细胞增殖速率减慢、细胞周期阻滞在G₀/G₁期向S期的过渡阶段,细胞迁移能力显著降低,侵袭能力下降。同时提取细胞上清液中外泌体,利用Western印迹对β-联蛋白和紧密连接蛋白-1表达量进行检测,发现相对于NC组,shE6组细胞内β-联蛋白表达量减少,但外泌体中β-联蛋白量增加,紧密连接蛋白-1在细胞内和外泌体中均增加。上述结果提示,HPV16 E6促进细胞恶性表型可能与E6蛋白能够抑制β-联蛋白以外泌体形式释放,从而增加其在细胞内的积累,以及抑制紧密连接蛋白-1在细胞内和外泌体中的积累有关。

In addition to intracellular organelles, eukaryotic cells also contain extracellular organelles that are released, or shed, into the microenvironment. These membranous extracellular organelles include exosomes, shedding microvesicles (SMVs) and apoptotic blebs (ABs), many of which exhibit pleiotropic biological functions. Because extracellular organelle terminology is often confounding, with many preparations reported in the literature being mixtures of extracellular vesicles, there is a growing need to clarify nomenclature and to improve purification strategies in order to discriminate the biochemical and functional activities of these moieties. Exosomes are formed by the inward budding of multivesicular bodies (MVBs) and are released from the cell into the microenvironment following the fusion of MVBs with the plasma membrane (PM). In this review we focus on various strategies for purifying exosomes and discuss their biophysical and biochemical properties. An update on proteomic analysis of exosomes from various cell types and body fluids is provided and host-cell specific proteomic signatures are also discussed. Because the ectodomain of ~42% of exosomal integral membrane proteins are also found in the secretome, these vesicles provide a potential source of serum-based membrane protein biomarkers that are reflective of the host cell. ExoCarta, an exosomal protein and RNA database (http://exocarta.ludwig.edu.au), is described.Copyright © 2010 Elsevier B.V. All rights reserved.

Exosomes are nanosized membrane vesicles secreted by wide variety of cells and found in abundance in biological fluids including semen. They contain cargo of lipids, proteins, microRNAs and mRNAs, and are known to play a major role in intracellular communication. Seminal exosomes mainly include epididymosomes and prostasomes. Most of the proteins associated with the epididymosomes are transferred to the sperm subcellular or membranous domains during their epididymal transit and are involved in the acquisition of fertilizing ability, modulation of motility and protection against oxidative stress. Proteins associated with prostasomes stimulate sperm motility and regulate the timing of capacitation to avoid premature induction of acrosome reaction. Furthermore, prostasomes protect the sperm from immune responses within the female reproductive tract. Overall, exosome-associated proteins play an indispensable role in maturation of spermatozoa and therefore, serve as an excellent biomarker in early diagnosis of male infertility.© 2020 Elsevier Inc. All rights reserved.

外泌体是一种广泛存在于多种细胞间质中的囊泡，多种细胞在正常情况下或在细胞外刺激等应激条件下都能产生外泌体，来自不同细胞类型的外泌体作用机制及功能有很大区别。外泌体可通过膜蛋白与靶细胞膜蛋白结合、膜蛋白碎片与细胞膜上受体结合、膜与靶细胞膜直接融合等方式传递生物活性分子，从而参与机体免疫、细胞分化等过程。由于外泌体具有来源天然、穿越屏障能力好、生物相容性强等特点，在许多研究领域均受到广泛的关注。作者介绍了外泌体的形态特点、形成过程、作用机制、分离纯化、生物学特性、功能及应用；总结了近年来外泌体在畜禽研究中的应用。外泌体不仅可应用于动物遗传育种中，还可作为工具或载体来研究动物体内各种生化途径。该综述可为外泌体的相关研究及应用提供参考。

外泌体（exosome）是通过细胞内多泡体与细胞膜融合产生的纳米级细胞外膜泡，广泛分布在血清、尿液、唾液和其他生物液体中。外泌体作为细胞间通信和遗传物质的重要转移载体，可通过受体介导的相互作用或通过各种生物活性分子（如细胞膜受体、蛋白质、mRNA和miRNA）的转移直接刺激靶细胞，从而发挥其生物学功能。文章主要综述了外泌体内含蛋白质和miRNA的功能及外泌体在临床中的应用。外泌体内蛋白质和miRNA的功能包括生理状态下诱导机体免疫、递呈抗原、参与细胞间信号传导；病理状态下改变肿瘤微环境、促进癌细胞增殖、侵袭、加快血管生成、促进癌症发展；以及某些病毒可将其组分包装、整合到外泌体中来实现细胞间传播，或劫持外泌体，实现免疫逃避。作者主要介绍了其作为肺癌、乳腺癌、胰腺癌及结直肠癌等多种癌症早期诊断的生物性标志物，以及作为稳定性高、转运效率高、靶向性强的药物载体在临床中的应用。

Exosomes, cell-derived vesicles of endosomal origin, are continuously released in the extracellular environment and play a key role in intercellular crosstalk. In this study, we have investigated whether transfer of integrins through exosomes between prostate cancer (PrCa) cells occurs and whether transferred integrins promote cell adhesion and migration. Among others, we have focused on the αvβ6 integrin, which is not detectable in normal human prostate but is highly expressed in human primary PrCa as well as murine PrCa in Pten(pc-/-) mice. After confirming the fidelity of the exosome preparations by electron microscopy, density gradient, and immunoblotting, we determined that the αvβ6 integrin is actively packaged into exosomes isolated from PC3 and RWPE PrCa cell lines. We also demonstrate that αvβ6 is efficiently transferred via exosomes from a donor cell to an αvβ6-negative recipient cell and localizes to the cell surface. De novo αvβ6 expression in an αvβ6-negative recipient cell is not a result of a change in mRNA levels but is a consequence of exosome-mediated transfer of this integrin between different PrCa cells. Recipient cells incubated with exosomes containing αvβ6 migrate on an αvβ6 specific substrate, latency-associated peptide-TGFβ, to a greater extent than cells treated with exosomes in which αvβ6 is stably or transiently down-regulated by shRNA or siRNA, respectively. Overall, this study shows that exosomes from PrCa cells may contribute to a horizontal propagation of integrin-associated phenotypes, which would promote cell migration, and consequently, metastasis in a paracrine fashion. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

In crustaceans, male differentiation and primary and secondary characteristics are regulated by the androgenic gland (AG). In gonochoristic crustaceans, the AG is also linked to intersexuality. Whereas the co-occurrence of various male and female characteristics has been demonstrated in intersex crustaceans, little is known regarding sexually dimorphic behavior patterns in such individuals. In the present study, we used an intersex crayfish model to investigate--for the first time in crustaceans--the agonistic and mating behavior of intersex individuals, and to explore the effects of AG ablation on behavior, morphology and physiology. As was the case for their morphological and physiological reproductive traits, intersex individuals--despite being genotypically females--generally resembled males in terms of behavior: they engaged in fighting with males and copulated with receptive females. However, fighting durations of intersex animals were intermediate between those of males and females, and the durations of the copulations were remarkably short. Adult intersex individuals that had been AG ablated at the juvenile stage were unlikely to engage in fighting with males (similar behavior to females) and did not exhibit any mating behavior with receptive females. AG ablation resulted in feminine morphological and physiological shifts in the treated intersex individuals and enabled vitellogenin gene transcription and the onset of secondary vitellogenesis. It thus appears that an as-yet-unknown AG-secreted factor(s) regulating maleness also seems to regulate the organization of male behavior in crustaceans.

Ginseng can help regulate brain excitability, promote learning and memory, and resist cerebral ischemia in the central nervous system. Ginsenosides are the major effective compounds of Ginseng, but their protein targets in the brain have not been determined.We screened proteins that interact with the main components of ginseng (ginsenosides) by affinity chromatography and identified the 14-3-3 ζ protein as a potential target of ginsenosides in brain tissues.Biolayer interferometry (BLI) analysis showed that 20(S)-protopanaxadiol (PPD), a ginseng saponin metabolite, exhibited the highest direct interaction to the 14-3-3 ζ protein. Subsequently, BLI kinetics analysis and isothermal titration calorimetry (ITC) assay showed that PPD specifically bound to the 14-3-3 ζ protein. The cocrystal structure of the 14-3-3 ζ protein-PPD complex showed that the main interactions occurred between the residues R56, R127, and Y128 of the 14-3-3 ζ protein and a portion of PPD. Moreover, mutating any of the above residues resulted in a significant decrease of affinity between PPD and the 14-3-3 ζ protein.Our results indicate the 14-3-3 ζ protein is the target of PPD, a ginsenoside metabolite. Crystallographic and mutagenesis studies suggest a direct interaction between PPD and the 14-3-3 ζ protein. This finding can help in the development of small-molecular compounds that bind to the 14-3-3 ζ protein on the basis of the structure of dammarane-type triterpenoid.© 2021 The Korean Society of Ginseng. Publishing services by Elsevier B.V.