Virus hijacking ESCRT system to promote self-replication: a review / 生物工程学报

Endosomal sorting complex required for transport (ESCRT) system drives various cellular processes, including endosome sorting, organelle biogenesis, vesicle transport, maintenance of plasma membrane integrity, membrane fission during cytokinesis, nuclear membrane reformation after mitosis, closure of autophagic vacuoles, and enveloped virus budding. Increasing evidence suggests that the ESCRT system can be hijacked by different family viruses for their proliferation. At different stages of the virus life cycle, viruses can interfere with or exploit ESCRT-mediated physiological processes in various ways to maximize their chance of infecting the host. In addition, many retroviral and RNA viral proteins possess "late domain" motifs, which can recruit host ESCRT subunit proteins to assist in virus endocytosis, transport, replicate, budding and efflux. Therefore, the "late domain" motifs of viruses and ESCRT subunit proteins could serve as promising drug targets in antiviral therapy. This review focuses on the composition and functions of the ESCRT system, the effects of ESCRT subunits and virus "late domain" motifs on viral replication, and the antiviral effects mediated by the ESCRT system, aiming to provide a reference for the development and utilization of antiviral drugs.

Papel de los exosomas en la angiogénesis, revascularización y respuesta inmune / Role of exosomes in angiogenesis, revascularization and immune response

Introducción: Los exosomas son vesículas extracelulares de tamaño nanométrico, que se generan cuando los endosomas multivesiculares se fusionan con la membrana plasmática y el contenido de las vesículas intraluminales se libera en el espacio extracelular. Son producidos por casi todos los tipos de células, en condiciones fisiológicas y patológicas. Transportan proteínas, lípidos y ácido ribonucleico (ARN) no codificante, desde la célula madre hasta la célula receptora, estos son considerados un punto clave en la regeneración de tejidos, lo que se ha demostrado en una serie de estudios, con diferentes tejidos corporales, como piel, cartílago, pancreático y tejidos cardiovasculares. Objetivo: Explicar los aspectos generales y posibles usos de los exosomas en el campo médico. Métodos: Se realizó una búsqueda de información mediante consulta en las bases de datos SciELO PubMed, Science Direct y Lilacs, en los idiomas español e inglés, con diferentes combinaciones de palabras claves y términos MESH como: exosomes, neovascularization, wound healing, immunity, micro RNA, immunology, therapy, classification. Se efectuó un análisis y resumen de la información revisada. Conclusiones: En la actualidad, los exosomas se han convertido en objeto de investigación para diversos tratamientos, medicamentos y uso como marcadores moleculares. Se destacan en terapias contra el cáncer, la inmunomodulación, la estimulación o supresión de la angiogénesis, regeneración cutánea, cicatrización y curación de heridas; por lo que de forma general resultan prometedores en el ámbito de las ciencias médicas(AU)

Introduction: Exosomes are nano-sized extracellular vesicles, which are generated when multivesicular endosomes fuse with the plasma membrane and the content of intraluminal vesicles released into the extracellular space. Are produced by almost all types of cells, under physiological and pathological conditions and they transport proteins, lipids and non-coding RNA (ribonucleic acid), from the stem cell to the recipient cell, these are considered a key point in tissue regeneration, which has been shown in a series of studies, with different body tissues, such as skin, cartilage, pancreatic and cardiovascular tissues. Objective: To explain the general aspects and possible uses of exosomes in the medical field. Methods: A search for information was carried out by consulting the Scielo, PubMed, ScienceDirect and Lilacs databases, in Spanish and English, with different combinations of keywords and MESH terms such as: exosomes, neovascularization, wound healing, immunity, microRNA, immunology, therapy, classification. Then, an analysis and summary of the reviewed information was carried out. Conclusions: Currently, exosomes have become the object of research for various treatments, drugs, and their use as molecular markers. They stand out in cancer therapies, immunomodulation, stimulation or suppression of angiogenesis, skin regeneration, and wound healing, which is why they are generally promising in the field of medical sciences(AU)

CD36 (SR-B2) as a Target to Treat Lipid Overload-Induced Cardiac Dysfunction

The heart faces the challenge of adjusting the rate of fatty acid uptake to match myocardial demand for energy provision at any given moment, avoiding both too low uptake rates, which could elicit an energy deficit, and too high uptake rates, which pose the risk of excess lipid accumulation and lipotoxicity. The transmembrane glycoprotein cluster of differentiation 36 (CD36), a scavenger receptor (B2), serves many functions in lipid metabolism and signaling. In the heart, CD36 is the main sarcolemmal lipid transporter involved in the rate-limiting kinetic step in cardiac lipid utilization. The cellular fatty acid uptake rate is determined by the presence of CD36 at the cell surface, which is regulated by subcellular vesicular recycling from endosomes to the sarcolemma. CD36 has been implicated in dysregulated fatty acid and lipid metabolism in pathophysiological conditions, particularly high-fat diet-induced insulin resistance and diabetic cardiomyopathy. Thus, in conditions of chronic lipid overload, high levels of CD36 are moved to the sarcolemma, setting the heart on a route towards increased lipid uptake, excessive lipid accumulation, insulin resistance, and eventually contractile dysfunction. Insight into the subcellular trafficking machinery of CD36 will provide novel targets to treat the lipid-overloaded heart. A screen for CD36-dedicated trafficking proteins found that vacuolar-type H⁺-ATPase and specific vesicle-associated membrane proteins, among others, were uniquely involved in CD36 recycling. Preliminary data suggest that these proteins may offer clues on how to manipulate myocardial lipid uptake, and thus could be promising targets for metabolic intervention therapy to treat the failing heart.

Progress in endosomal Na⁺,K⁺/H⁺ antiporter in Arabidopsis thaliana / 生物工程学报

Important progress has been made in the interpretation of subcellular location, ion transport characteristics and biological functions of endosomal Na⁺,K⁺/H⁺ antiporter in Arabidopsis thaliana. The endosomal Na⁺,K⁺/H⁺ antiporter contain two members, AtNHX5 and AtNHX6, whose amino acid sequence similarity is 78.7%. Studies have shown that AtNHX5 and AtNHX6 are functionally redundant, and they are all located in Golgi, trans-Golgi network (TGN), endoplasmic reticulum (ER) and prevacuolar compartment (PVC). AtNHX5 and AtNHX6 are critical for salt tolerance stress and the homeostasis of pH and K⁺. It has been reported that there are conservative acidic amino acid residues that can regulate their ion activity in the endosomal NHXs transmembrane domain, which plays a decisive role in their own functions. The results of the latest research indicate that endosomal NHXs affect vacuolar transport and protein storage, and participate in the growth of auxin-mediated development in A. thaliana. In this paper, the progress of subcellular localization, ion transport, function and application of endosomal NHXs in A. thaliana was summarized.

Advance in research on recycling antibody / 生物工程学报

Monoclonal antibodies have become the main type of antibody drug because of their high specificity and strong affinity to antigen. However, with the intensive study of the natural monoclonal antibody, many defects have faced, such as the limit times of binding to antigen, the unanticipated antibody clearance and antigen accumulation. Therefore, studies are no longer limited to the natural antibody screening, but rather to improve the efficiency of antibody drugs by engineering. In recent years, the bottlenecks in the development of conventional antibody have been solved effectively since the discovery of a novel recycling antibody. Recycling antibody binds to an antigen in plasma and dissociates from the antigen in endosome, thus maximizing the use of antibody and reducing antigen-mediated antibody clearance and antibody-mediated antigen accumulation. In addition, recycling antibodies can enhance the affinity with Fc receptors through further Fc modification. This paper reviews the research progress of circulating antibodies, including its characteristics, transformation methods and prospects.

Role of endocytosis in cell surface CXC chemokine receptor 4 expression of stem cells from apical papilla / 北京大学学报(医学版)

OBJECTIVE@#To evaluate the change of cell surface CXC chemokine receptor 4 (CXCR4) expression of stem cells from apical papilla (SCAP) after the inhibition of endocytotic pathway, thus to provide experimental basis for the mechanism of SCAP migration.@*METHODS@#The immunofluorescence analysis was conducted to examine the co-expression of CXCR4 and endocytotic compartments, including early endosomes, recycling endosomes and lysosomes in SCAP. Several Rab proteins were applied as markers of organelles in the endocytotic pathway, including Rab5 for early endosomes, Rab11A for recycling endosomes, and Lamp1 for lysosomes. The co-localization of CXCR4 with these endodontic compartments was further observed by proximity ligation assay (PLA). SCAP was treated with two kinds of endocytotic inhibitors, Blebbistatin and Dynasore, at a concentration of 80 μmol/L, respectively. The conditioning time was 1 hour. Flow cytometry was carried out to evaluate the proportion of SCAP that expressed CXCR4 on cell surface. The data were analysed by analysis of variance (ANOVA).@*RESULTS@#The red staining of CXCR4 on immunofluorescence confocal microscopy predominantly overlapped with the green staining of Rab5 and Rab11A, and partly overlapped with Lamp1. It indicated that most CXCR4 molecules were located in early endosomes and recycling endosomes, and some were located in lysosomes. The PLA results revealed that the co-localizaiton of CXCR4 with endocytotic compartments could be observed in early endosomes, recycling endosomes and lysosomes. According to the results of flow cytometry, the proportion of SCAP that expressed CXCR4 on cell surface was as low as 0.13%±0.10%. After the inhibition of endocytosis by pretreating the cells with the following two inhibitors, Blebbistatin and Dynasore, the percentage of SCAP that positively expressed CXCR4 on cell surface was significantly increased to 13.34%±1.31% in Blebbistatin group and 4.03%±0.92% in Dynasore group (F=16.721, P<0.001). Moreover, the number of SCAP that expressed CXCR4 on cell surface in Blebbistatin group was significantly higher than that in Dynasore group (P<0.001).@*CONCLUSION@#The inhibition of endocytotic pathway could increase the number of SCAP that expressed CXCR4 on cell surface, and provide potency for the migration of SCAP.

Justicidin A Reduces β-Amyloid via Inhibiting Endocytosis of β-Amyloid Precursor Protein

β-amyloid precursor protein (APP) can be cleaved by α-, and γ-secretase at plasma membrane producing soluble ectodomain fragment (sAPPα). Alternatively, following endocytosis, APP is cleaved by β-, and γ-secretase at early endosomes generating β-amyloid (Aβ), the main culprit in Alzheimer's disease (AD). Thus, APP endocytosis is critical for Aβ production. Recently, we reported that Monsonia angustifolia, the indigenous vegetables consumed in Tanzania, improved cognitive function and decreased Aβ production. In this study, we examined the underlying mechanism of justicidin A, the active compound of M. angustifolia, on Aβ production. We found that justicidin A reduced endocytosis of APP, increasing sAPPα level, while decreasing Aβ level in HeLa cells overexpressing human APP with the Swedish mutation. The effect of justicidin A on Aβ production was blocked by endocytosis inhibitors, indicating that the decreased APP endocytosis by justicidin A is the underlying mechanism. Thus, justicidin A, the active compound of M. angustifolia, may be a novel agent for AD treatment.

Cryo-EM structures of the mammalian endo-lysosomal TRPML1 channel elucidate the combined regulation mechanism

TRPML1 channel is a non-selective group-2 transient receptor potential (TRP) channel with Ca permeability. Located mainly in late endosome and lysosome of all mammalian cell types, TRPML1 is indispensable in the processes of endocytosis, membrane trafficking, and lysosome biogenesis. Mutations of TRPML1 cause a severe lysosomal storage disorder called mucolipidosis type IV (MLIV). In the present study, we determined the cryo-electron microscopy (cryo-EM) structures of Mus musculus TRPML1 (mTRPML1) in lipid nanodiscs and Amphipols. Two distinct states of mTRPML1 in Amphipols are added to the closed state, on which could represent two different confirmations upon activation and regulation. The polycystin-mucolipin domain (PMD) may sense the luminal/extracellular stimuli and undergo a "move upward" motion during endocytosis, thus triggering the overall conformational change in TRPML1. Based on the structural comparisons, we propose TRPML1 is regulated by pH, Ca, and phosphoinositides in a combined manner so as to accommodate the dynamic endocytosis process.

Expression of epithelial-mesenchymal transition markers at the invasive front of oral squamous cell carcinoma

Oral squamous cell carcinoma (OSCC) is one of the most common malignances. In epithelial-mesenchymal transition (EMT), epithelial cells switch to mesenchymal-like cells exhibiting high mobility. This migratory phenotype is significant during tumor invasion and metastasis. Objective : The aim of this study is to evaluate the expression of the EMT markers E-cadherin, N-cadherin and vimentin in OSCC. Material and Methods : Immunohistochemical detection of E-cadherin, N-cadherin and vimentin was performed on 20 OSCC samples. Differences in the expression of each protein at the invasive front (IF) and in the central/superficial areas (CSA) of the tumor were assessed. Differences in the expression of each protein at the IF of both histologically high- and low-invasive OSCCs were evaluated. Associations among expression of proteins at the IF were assessed. Correlations between the expression levels of each protein at the IF and the tumor stage and clinical nodal status were also evaluated. Results : Reduced expression of E-cadherin was detected in 15 samples (75%). E-cadherin expression was reduced at the IF when compared to the CSA and in high-invasive tumors when compared to low-invasive tumors. All samples were negative for N-cadherin, even though one sample showed an inconspicuous expression. Positive expression of vimentin was observed in 6 samples (30%). Nevertheless, there was no difference in vimentin expression between the IF and the CSA regions or between the low- and high-invasive tumors. Furthermore, no association was observed among protein expression levels at the IF. Finally, no correlations were observed between each protein’s expression levels and tumor stage or clinical nodal status. Conclusions : Reduced E-cadherin expression at the IF and its association with histological invasiveness suggest that this protein is a noteworthy EMT marker in OSCC. Although vimentin was also detected as an EMT marker, its expression was ...

Preparation and evaluation of doxorubicin hydrochloride liposomes modified by poly(2-ethyl-2-oxazoline)-cholesteryl methyl carbonate / 药学学报

In this study, the buffering capacity of amphiphilic pH-sensitivity copolymer poly(2-ethyl-2-oxazoline)-cholesteryl methyl carbonate (PEOZ-CHMC) was evaluated. The ammonium sulfate gradient method was used to prepare doxorubicin hydrochloride (DOX x HCl)-loaded liposomes (DOX-L), and then the post-insertion method was used to prepare PEOZ-CHMC and polyethylene glycol-distearoyl phosphatidyl ethanolamine (PEG-DSPE) modified DOX x HCl-loaded liposomes (PEOZ-DOX-L and PEG-DOX-L). The physico-chemical properties, in vitro drugs release behavior, cellular toxicity and intracellular delivery of liposomes were evaluated, separately. The results showed that PEOZ-CHMC has a satisfactory buffering capacity. The sephadex G-50 column centrifugation method and dynamic light scattering were used to determine the encapsulation efficiency (EE) and particle size of liposomes. The EE and particle size of DOX-L were (97.3 ± 1.4) % and 120 nm, respectively, and the addition of PEOZ-CHMC or PEG-DSPE had no influence on EE and particle size. The zeta potentials of three kinds of liposomes were negative. The release behavior of various DOX liposomes in vitro was investigated by dialysis method. In phosphate buffer solution (PBS) at pH 7.4, DOX x HCl was released from PEOZ-DOX-L in a sustained manner. While in PBS at pH 5.0, the release rate of DOX x HCl from PEOZ-DOX-L increased significantly, which suggested DOX x HCl was released from PEOZ-DOX-L in a pH-dependent manner. The intracellular delivery of liposomes was investigated by confocal laser scanning microscopy (CLSM). The CLSM images indicated that PEOZ-DOX-L showed efficient intracellular trafficking including endosomal escape and release DOX x HCl into nucleus, as well as the DOX-L and PEG-DOX-L had no this effect. The cytotoxicity of liposomes against MCF-7 cells was detected by using MTT assay. The results showed that antiproliferative effects of PEOZ-DOX-L enhanced with pH value decreased, whereas DOX-L and PEG-DOX-L did not have any significant difference in inhibitions at different pH conditions. Therefore, the problems of the inhibition of cellular uptake of liposomes and the failed endosomal escape of pH-sensitive liposomes by PEG chain can be overcome by the pH-sensitive liposomes constructed by PEOZ-CHMC.

ATP13A2/PARK9 Deficiency Neither Cause Lysosomal Impairment Nor Alter alpha-Synuclein Metabolism in SH-SY5Y Cells

Parkinson's disease is a multifactorial disorder with several genes linked to the familial types of the disease. ATP13A2 is one of those genes and encode for a transmembrane protein localized in lysosomes and late endosomes. Previous studies suggested the roles of this protein in lysosomal functions and cellular ion homeostasis. Here, we set out to investigate the role of ATP13A2 in lysosomal function and in metabolism of alpha-synuclein, another PD-linked protein whose accumulation is implicated in the pathogenesis. We generated non-sense mutations in both copies of ATP13A2 gene in SH-SY5Y human neuroblastoma cells. We examined lysosomal function of ATP13A2-/- cells by measuring the accumulation of lysosomal substrate proteins, such as p62 and polyubiquitinated proteins, induction of acidic compartments, and degradation of ectopically introduced dextran. None of these measures were altered by ATP13A2 deficiency. The steady-state levels of alpha-synuclein in cells or secretion of this protein were unaltered either in ATP13A2-/- compared to the normal cells. Therefore, the proposed roles of ATP13A2 in lysosomal functions may not be generalized and may depend on the cellular context. The ATP13A2-/- cells generated in the current study may provide a useful control for studies on the roles of PD genes in lysosomal functions.

The strategies of endosomal escape for intracellular gene delivery / 药学学报

The intracellular trafficking and subcellular distribution of exogenous gene is very important for gene delivery. A successful gene vehicle should overcome various barriers including endosomal membrane barriers to delivery gene to the target organelle. Traditional nonviral vehicle is unable to avoid endosomal pathway efficiently, so the efficiency of gene delivery is low and the application of gene drugs is limited. In order to achieve efficient nonviral gene delivery, a lot of researches based on endosomal escape have been carried out and some agents with the function of endsomal escape have been found. These agents facilitate the endsomal escape via various mechanisms, such as fusion into the lipid bilayer of endosomes, pore formation in the endosomal membrane, proton sponge effect and photochemical methods to rupture the endosomal membrane. In this review, various reported strategies for endsomal escape are described according to the escape mechanisms, and their applications in intracellular gene delivery are also discussed.

Progress in infection pathway and intracellular trafficking of adenovirus / 生物工程学报

The research on intracellular trafficking of adenovirus has been described mainly through observations of subgroup C adenoviruses in transformed cell lines. The basic elements of the trafficking pathway include binding to receptors at the cell surface, internalization by endocytosis, lysis of the endosomal membrane, escape to the cytosol, intracellular trafficking along microtubules, nuclear pore docking, and viral genome translocation into the nucleus. More than 80% of the adenovirus genome is delivered to the nucleus in a highly efficient manner in approximately 1 h. However, exceptions to this trafficking pattern have been noted, including: variations based on target cell type, cell physiology, and adenovirus serotype. This review summarizes mechanism of adenovirus infection pathway and intracellular trafficking, providinging a foundation for the development of clinical adenoviral vector.

Neuronal Functions of ESCRTs

The endosomal sorting complexes required for transport (ESCRTs) regulate protein trafficking from endosomes to lysosomes. Recent studies have shown that ESCRTs are involved in various cellular processes, including membrane scission, microRNA function, viral budding, and the autophagy pathway in many tissues, including the nervous system. Indeed, dysfunctional ESCRTs are associated with neurodegeneration. However, it remains largely elusive how ESCRTs act in post-mitotic neurons, a highly specialized cell type that requires dynamic changes in neuronal structures and signaling for proper function. This review focuses on our current understandings of the functions of ESCRTs in neuronal morphology, synaptic plasticity, and neurodegenerative diseases.

The hydrophobic amino acids involved in the interdomain association of phospholipase D1 regulate the shuttling of phospholipase D1 from vesicular organelles into the nucleus

Phospholipase D (PLD) catalyzes the hydrolysis of phosphatidylcholine to generate the lipid second messenger, phosphatidic acid. PLD is localized in most cellular organelles, where it is likely to play different roles in signal transduction. PLD1 is primarily localized in vesicular structures such as endosomes, lysosomes and autophagosomes. However, the factors defining its localization are less clear. In this study, we found that four hydrophobic residues present in the N-terminal HKD catalytic motif of PLD1, which is involved in intramolecular association, are responsible for vesicular localization. Site-directed mutagenesis of the residues dramatically disrupted vesicular localization of PLD1. Interestingly, the hydrophobic residues of PLD1 are also involved in the interruption of its nuclear localization. Mutation of the residues increased the association of PLD1 with importin-beta, which is known to mediate nuclear importation, and induced the localization of PLD1 from vesicles into the nucleus. Taken together, these data suggest that the hydrophobic amino acids involved in the interdomain association of PLD1 are required for vesicular localization and disturbance of its nuclear localization.

Hrs inhibits citron kinase-mediated HIV-1 budding via its FYVE domain

Hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs) is a key component of the endosomal sorting complexes required for transport and has been demonstrated to play a regulatory role in endocytosis/exocytosis and the accumulation of internal vesicles in multivesicular bodies. Citron kinase is a Ser/The kinase that we previously reported to enhance human immunodeficiency virus type 1 (HIV-1) virion production. However, the relationship between Hrs and citron kinase in HIV-1 production remains elusive. Here, we report that Hrs interacts with citron kinase via its FYVE domain. Overexpression of Hrs or the FYVE domain resulted in a significant decrease in HIV-1 virion production. Depletion of Hrs by RNA interference in HEK293T cells increased HIV-1 virion production and enhanced the activity of citron kinase. These data suggest that Hrs inhibits HIV-1 production by inhibiting citron kinase-mediated exocytosis.

Ribosomal Protein L19 and L22 Modulate TLR3 Signaling

BACKGROUND: Toll-like receptor 3 (TLR3) recognizes double-stranded RNA (dsRNA) and induces inflammation. In this study we attempted to ascertain if there are endogenous host molecules controlling the production of cytokines and chemokines. Two candidates, ribosomal protein L19 and L22, were analyzed to determine if they influence cytokine production followed by TLR3 activation. In this study we report that L19 acts upon production of IP-10 or IL-8 differently in glioblastoma cells. METHODS: L19 or L22 was transfected into HEK293-TLR3, A549 or A172 cells. After treatment with several inhibitors of NF-kB, PI3K, p38 or ERK, production of IL-8 or IP-10 was measured by ELISA. siRNA was introduced to suppress expression of L19. After Vesicular stomatitis virus infection, viral multiplication was measured by western blot. RESULTS: L19 increased ERK activation to produce IL-8. In A172 cells, in which TLR3 is expressed at endosomes, L19 inhibited interferon regulatory factor 3 (IRF3) activation and IP-10 production to facilitate viral multiplication, whereas L19 inhibited viral multiplication in A549 cells bearing TLR3 on their cell membrane. CONCLUSION: Our results suggest that L19 regulates TLR3 signaling, which is cell type specific and may be involved in pathogenesis of autoimmune diseases and chronic inflammatory diseases.

Alteraciones en el reclutamiento y activación de proteínas Rab durante la infección micobacteriana: [revisão] / Alterations in recruitment and activation of Rab proteins during mycobacterial infection: [review]

En el fagosoma, Mycobacterium spp. altera la activación y reclutamiento de diferentes proteínas “del gen Ras de cerebro de rata”, comúnmente conocidas como Rab. En este manuscrito se revisa una serie de reportes que han demostrado que los fagosomas que contienen micobacterias tienen una expresión mayor y sostenida de Rab5, Rab11, Rab14 y Rab22a, y menor o ninguna expresión de Rab7, Rab9 y Rab6. Esto se correlaciona con aumento de la fusión de estos fagosomas con endosomas tempranos y de reciclaje, lo que les permite mantener ciertas características de compartimentos tempranos, permite que las bacterias obtengan acceso a nutrientes y previene la activación de mecanismos contra la micobacteria. La expresión de mutantes constitutivamente activos de las Rab de endosomas tempranos impide la maduración de fagosomas que contienen esferas de látex o micobacterias inactivadas por calor. Mientras que su silenciamiento, mediante ARN de interferencia o mediante dominantes negativos, induce la maduración de fagosomas micobacterianos. Los mecanismos exactos por los que las micobacterias alteran la dinámica de expresión de estas GTPasas, afectando la maduración fagolisosómica, no se han establecido. El problema podría explicarse por defectos en el reclutamiento de las proteínas que interactúan con Rab, como la cinasa-3 del fosfatidilinositol y el antígeno endosómico temprano 1. La identificación de los mecanismos empleados por Mycobacterium spp. para interrumpir el ciclo de activación de las Rab, será esencial para comprender la fisiopatología de la infección micobacteriana y útil como posibles blancos farmacológicos.

At the phagosome level, Mycobacterium spp. alters activation and recruitment of several “Ras gene from rat brain” proteins, commonly known as Rab. Mycobacterial phagosomes have a greater and sustained expression of Rab5, Rab11, Rab14 and Rab22a, and lowered or no expression of Rab7, Rab9 and Rab6. This correlates with increased fusion of the phagosomes with early and recycling endosomes acquiring some features of early phogosomes, allowing the bacteria to gain access to nutrients and preventing the activation of anti-mycobacterial mechanisms. The expression of constitutively active mutants of Rab from the early stage endosomes prevents the maturation of phagosomes containing latex beads or heat-inactivated mycobacteria. Silencing of these mutants by interference RNA or dominant negative forms induces the maturation of mycobacterial phagosomes. The mechanisms have not been established by which mycobacteria alter the expression of these GTPases and thereby shift the phagolysosomal maturation. The problem can be explained by alterations in the recruitment of proteins that interact with Rab, such as phosphoinositide 3-kinases and early endosomal antigen 1. Identifying the mechanisms used by Mycobacterium spp. to disrupt the cycle of Rab activation will be essential to understand the pathophysiology of mycobacterial infections and usefully to potential drug targets.

Tumour-derived exosomes and their roles in cancer / 中南大学学报(医学版)

Exosomes are nanometer sized membrane vesicles, released in the extracellular milieu following the fusion of the external membrane of multivesicular body (MVB) with plasma membrane. They perform a certain function in immune regulation. Exosomes have been shown to be released by cells of hematopoietic and non-hematopoietic origin. Tumour-derived exosomes (TEX) exist in the supernatant of tumour cells, plasma and malignant effusions of tumour patients. They contain native candidate tumour associated antigen and are capable of transferring antigens to T lymphocytes, therefore efficiently promoting cytotoxic T lymphocyte (CTL) activation and producing antitumor immunity. However, recent evidence shows that tumor exosomes may induce immunologic tolerance and even activate immunosuppression which makes tumour escape from the immune surveillance of the host immune system. In addition, tumor exosomes may mediate a growth-promoting effect on tumor cells. These discrepancies are almost certainly due to differences in the phenotype of the exosomes.

The mechanism of HBV infection of human trophoblast cell / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To observe the changes of human trophoblast cells after infected with hepatitis B virus.</p><p><b>METHODS</b>HBV positive serum was used to infect human trophoblast cells in vitro. HBsAg in cell culture medium were detected by ELISA method and HBV DNA in cell culture medium and cells were detected by PCR method. HBV fluorescence polymerase chain reaction diagnose kit were used to detect the HBV DNA concentration. Ultra structure of trophoblast cells were observed with transmission electron microscopy (TEM).</p><p><b>RESULTS</b>HBsAg could be detected in infection group by ELISA. Infection group cell culture medium and infection group cells were HBV DNA positive. HBV DNA concentrations in HBV infection cell culture medium in 0, 12, 36, 60, 84 h after extensively PBS washed were < 10(3), 3 x 10(4), 6 x 10(5), 5 x 10(5), 3 x 10(5) copies/mL. HBV infected trophoblast cells were found many forms of endosomes, some of which contents virus like particle.</p><p><b>CONCLUSION</b>HBV might take advantage of clathrin-mediated endocytosis to enter trophoblast cell, which might lead to cell infection or across the cell bar by transcytosis.</p>