Endothelial tight junction proteins Essay Example for Free

Endothelial tight junction proteins Essay Endothelial tight junction proteins Introduction            The endothelium is situated at the inner side of all kinds of vessels and comprises of a monolayer of endothelial cells. Inter-endothelial junctions comprise junctional complexes, such as adherens junctions (AJ), tight junctions (TJ) and gap junctions (GJ) that play essential roles in tissue integrity, barrier function and intercellular communication respectively. These junctional complexes are related to those found at epithelial junctions with notable changes in terms of certain molecules and structure.            Endothelial junctional proteins play important roles in tissue integrity but also in vascular permeability, leukocyte extravasation and angiogenesis. Dormant endothelium may be exposed to stimuli provoking leukocyte extravasation at seditious sites and propagating angiogenesis. Both activities have an intense impact on endothelial cell-cell junctions.            Tight junctions aid the major functional objective of establishing a barrier inside the membrane, by controlling paracellular permeability and sustaining cell polarity. They achieve this by constricting apical or basolateral transmembrane diffusion of lipids and they have been suggested to contribute in regulating proliferation and differentiation of epithelial cells. However, the components that are involved and the signal routes concerned are unknown (Mitic Anderson 1998).            Tight junctions are made up of integral membrane proteins claudins, occludin, tricellulin, junctional adhesion molecules (JAMs), including many peripheral membrane proteins such as the scaffold PDZ- domain proteins. This review will however, focus on ZO-1 and ZONAB. Histology of endothelia junctions            The junctional structures situated at the endothelial intercellular fissure are related to those located at the epithelium; however, their formation is more inconsistent and in most vascular beds their topology is less constrained than in epithelial cells. Adherens junctions, tight junctions and gap junctions are in most cases intermingled and create a complex zonular system with disparities in depth and thickness of the sub-membrane plate associated with the junctional structure (Franke et al. 1988; Rhodin 1974). In contrast to epithelial cells, GJs are often found close to the luminal surface. Hence, the term â€Å"Apical junction† used to jointly describe epithelial TJ and AJ may not be applied to the endothelium. The endothelium forms the vascular barrier with controlled permeability properties between the blood and the underlying tissues.            Tight junctions exhibit considerable inconsistency among different segments of the vascular tree (Franke et al. 1988). This disparity composes a major evidence of vascular bed differentiation of endothelial cells and has a strong impact on vascular permeability and leukocyte extravasation. Variations concern the complexity degree of the occluding strands as well as tight junction composition.            Large Artery endothelial cells, which are exposed to high flow rates, display a well-developed system of tight junctions. Within the microvasculature, tight junctions are less complex in capillaries than in arterioles, and even less in venules. It is important to mention that, post-capillary venules are the primary site of leukocyte extravasation, and accordingly, they display a high content of permeability mediator receptors, such as those for histamine, serotonin and bradykinin. On the other hand, blood brain barrier (BBB) and the blood retinal barrier (BRB) are predominantly rich in Tight Junctions and endothelial tight junctions have been principally studied in these sites.            Endothelial intercellular realms differ from those of epithelial cells by the absence of desmosomes (Franke et al. 1988). The transitional filaments, comprised in the endothelium by vimentin molecules, are poorly connected to cell-cell contacts. However, contrary to the situation in epithelia, the vimentin filaments may be associated to endothelial adherens junctions in junctional structures similar to desmosomes, called complexus adherens.            It must be emphasized that interendothelial junctions are vibrant structures, subjected to multiple regulations. Moreover, leukocytes extravasate majorly in postcapillary venules either through transcellular or paracellular methods. Extravasation via the intercellular junction is a rapid and controlled process, through which the leukocyte is squeezed in the fissure (diapedesis), followed by rapid junction reformation.            ZO-1 is a protein located on the cytoplasmic membrane plate of intercellular tight junctions and is engaged in transducing signals at cell-to-cell junctions. ZO-1 links tight junction transmembrane proteins to a cytoplasmic plaque and the actin-based cytoskeleton (Aijaz et al. 2006; Tsukita et al. 2001). In epithelial cells, ZO-1 interrelates with the transcription factor ZONAB to regulate cells proliferation in a cell density related manner (Balda Matter 2000); however, the functions of ZO-1 and ZONAB in endothelial cells are still not clearly understood.            Unpublished work shows that downregulation of ZO-1 in endothelial cells stimulates redistribution of two transmembrane proteins; claudin-5 and JAM-A, and radical changes in the cytoskeleton affecting the localization of mechanosensor proteins and VE-cadherin role in the control of cell-cell tension.            These observations imply that one function of ZO-1 in endothelial cells is to coordinate components of the tight junction and associate them to the cortical cytoskeleton. However, it is unfamiliar whether the ZO-1 associated transcription factor ZONAB is linked to such ZO-1 effects.            Despite the fact that, ZO-1 explicitly associates with epithelial tight junctions (Stevenson et al. 1986), it has been observed that the protein appears in the nucleus in the process of proliferation (Gottardi et al. 1996). While the functional impact of the nuclear localization is currently not clear, studies reveal that these discrete subcellular distributions of ZO-1 are exquisitely sensitive to the state of cell-to-cell contact.            ZO-1 plays a major role of restraining ZONAB and regulates its accumulation in the nucleus through cytoplasmic sequestration. MDCK cells found in the epithelium exhibit two forms of this Y-box transcription factor (ZONAB) i.e. ZONAB -A and ZONAB -B which vary in a 68-amino acid supplement. Both categories of ZONAB bind to ZO-1 and link with intercellular junctions (Balda Matter 2000).            ZONAB was initially designated in canine kidney epithelial cells (MDCK) and is a Y-box transcription factor. Y-box transcription factors are multipurpose control mechanisms of gene expression and studies suggest that they play a common role in enhancing proliferation (Bargou et al. 1997). ZONAB is one of the tight junction-associated dual localization protein: it localizes to junctions where it attaches to the SH3 surface of the adaptor protein ZO-1, and to the nucleus where it regulates transcription.            The distribution of ZONAB is controlled by the cell density as it localizes to both junctions and nuclei in low density, proliferating cells, and becomes constrained to the cytoplasm in high density cells (Balda Matter, 2000). This distribution is also exhibited in its transcription activity, as ZONAB is transcriptionally vigorous in proliferating cells but inactive in non-proliferating cells. In the MDCK cells, ZONAB is necessary for normal rates of proliferation and controls G1/S phase transition (Balda et al. 2003).            ZONAB affects cell cycle development by two distinct processes: it controls the nuclear accumulation of CDK4 through a direct interaction and controls manifestation of genes encoding cell cycle regulators for example, PCNA and cyclin D1 (Balda et al. 2003; Sourisseau et al. 2006 ).            In 3D principles of MDCK cells, regular ZO-1 and ZONAB processes are necessary for epithelial cyst formation, implying that the Y-box transcription factor also controls epithelial differentiation (Sourisseau et al. 2006). Since ZO-1 and ZONAB can also relate with other types of intercellular junctions, for instance the gap junctions, in cells that lack tight junctions, it is possible that ZO-1 or ZONAB signaling is also of useful significance in other cell types other than epithelia (Ciolofan et al. 2006; Giepmans Moolenaar 1998). Aims of the study            The aim of the study is to understand the functional consequences of downregulation of ZONAB in endothelial cells, and whether and how ZONAB cross-talks with other junctional components to regulate endothelial cell migration, proliferation and angiogenesis. Currently, we are looking at similarities and differences between the phenotype of downregulation of ZO-1 or ZONAB by RNA interference. Changes in expression and localization of a given protein are analysed using specific antibodies for immunoblots and immunofluorescence. Preliminary Results            It is observed that downregulation of ZO-1 or ZONAB resulted in similar redistribution of actin and vinculin from cell-cell junctions to stress fibers and focal adhesions, respectively. However, the localization of transmembrane proteins such as Claudin-5 and JAM-A is affected by downregulation of ZO-1 rather than by downregulation of ZONAB. The localization of the polarity protein PAR-3 is changed in both conditions.            Additionally, downregulation of ZONAB causes changes in ZO-1 by immunofluorescence that needs to be tested for expression by immunoblots. Next, we will characterize other transmembrane proteins (e.g. MD3 and claudin-1), polarity proteins (PKCzeta), Rho regulators and mechanotransducers such as PAK2, Zyxin and YAP.            ZONAB is a DNA and RNA binding factor that it is involved in transcription (e.g. cyclin D1 and PCNA) in the nucleus and translation (e.g. cell cycle inhibitor p21) in the cytosol. Thus, we are also trying to identify new genes regulated. We have identified that expression of fibronectin is regulated by ZONAB. We are evaluating whether the changes in protein expression of fibronectin are due to ZONAB role on transcription or translation, using actinomicin D to inhibit transcription or cyclohexidimide to inhibit translation. Additionally, we are validating new genes identified by cDNA array analysis of endothelial cells with downregulation of ZONAB.            The tight junction localizing protein ZO-1 symptomatically forms a continuous band around the apices of well-differentiated, confluent, polarized epithelial cells in culture. However, under nonconfluent conditions, endogenous ZO-1 can localize to the nucleus in addition to the border of cell-cell contact.            ZONAB manifestation tends to be high in proliferating but low in growth-impeded MDCK cells, implying that high manifestation levels might be a necessity for cell proliferation (Balda Matter 2000).            ZONAB confines in the nucleus as well as tight junctions in proliferating cells, however, it is not noticeable in the nucleus of nonproliferating high density cells (Balda Matter 2000), proposing that accumulation of ZONAB in the nucleus might be necessary for efficient proliferation.            ZO-1 quantities are low in proliferating cells and they rise with cell density, and overexpression of ZO-1 hinders accumulation of ZONAB in the nucleus (Balda Matter 2000); hence, ZO-1 may control proliferation by inhibiting ZONAB from accumulating in the nucleus. Overexpression of ZO-1 in low density cells triggers a redistribution of ZONAB from the nucleus to the cytoplasm and reduced proliferation.            CDK4 is a major regulator of G1/s transition (Sherr 2000; Malumbres Barbacid 2001). Thus, ZONAB could control proliferation by regulating the process or the localization of CDK4. Since ZONAB binds CDK4, the nuclear pools of the two proteins may diminish in a parallel manner.            Symplekin is combined with ZONAB in the nucleus; hence, it could be argued that Symplekin modulates the transcription activity of ZONAB. Increased expression of Symplekin results in stimulation of the transcriptional suppressor ZONAB. However, it is also noted that Symplekin is absent in endothelial cells (Keon et al. 1996).            ZONAB controls cell cycle entry. ZO-1 overexpression results in a reduction in DNA synthesis, implying that entry into S-phase was distressed.            These experiments will allow understanding the role of ZO-1 and ZONAB in endothelial cells. Depending on the results, we plan to test how these two proteins are involved in endothelial stress conditions such as shear stress and high glucose. Conclusion            The collaboration of ZO-1 with tight junctions can only be significant for the stabilization of ZO-1, as opposed to attaching ZO-1 to the plasma membrane so as to constrain nuclear accumulation of related proteins. This is supported by the opinion that a truncated protein comprising only the HA-tagged SH3 domain accumulated in the Cytosol, but was adequate to decrease proliferation and nuclear accumulation of ZONAB (unpublished data).            ZONAB and ZO-1 control proliferation and the ultimate cell density of MDCK cells. Explanations that ZO-1 accumulates with increasing cell density, and overexpression of ZO-1 in transfected cells lowers the final density proposes a pattern in which ZO-1 serves as a measure for cell density whereby, on reaching the threshold level, provokes growth impediment by cytoplasmic sequestration of ZONAB and the related cell cycle kinase CDK4. It will be essential to control how the ZO-1 or ZONAB pathway associates with the other signaling methods that affect proliferation.            Vascular endothelial stress induces dysfunctions that have been implicated in many diseases such as diabetes and diabetic retinopathy. Therefore, characterization of the role of tight junction molecules in different endothelial cell behavior and functions will help us to understand the molecular mechanisms of disease pathogenesis and these findings may be implicated in prognosis and possibly to develop new treatment strategies. References Balda, MS and Matter, K 2000. The tight junction protein ZO-1 and an interacting transcription factor regulate ErbB-2 expression. EMBO J. 19, pp 2024-2033. Balda MS, Garrett MD and Matter K, 2003. The ZO-1 associated Y-box factor ZONAB regulates epithelial cell proliferation and cell density. J. Cell Biol. 160, pp 423-432. Bargou RC, K Jurchott, C Wagener, S Bergmann, S metzner, K Bommert, MY Mapara, KJ Winzer. M Dietel, B Dorken, and HD Royer, 1997. Nuclear localization and increased levels of transcription factor YB-1 in primary human breast cancers are associated with intrinsic MDR1 gene expression. Nat. Med. 3: pp 447-450. Ciolofan C, Li XB, Olson C, Kamasawa N, Gebhardt BR, Yasumura T, Morita M, Rash JE and Nagy JI, 2006. Association of connexin36 and Zonula occludens-1 with zonula occludens-2 and the transcription factor zonula occludens-1 associated nucleic acid-binding protein at neuronal gap junctions in rodent retina. Neuroscience 140: pp 433-451. Franke WW, P Cowin, C Grund, C Kuhn, HP Kapprell, 1998, The Endothelial Junction: the plaque and its component., in: N. Simionescu, M Simionescu (Eds.), Endothelial cell biology in health and diseases, Plenum publishing corporation, New York. pp 147-166. Giepmans BN and Moolenaar WH, 1998. The gap junction protein connexin43 interacts with the second PDZ domain of the zonal occludens-1 protein. Curr. Biol. 8. Pp 931-934. Gottardi CJ, M Arpin, AS Fanning and D Louvard, 1996. The junction-associated protein, zonular occludens-1, localizes to the nucleus before the maturation and during the remodeling of cell-cell contacts. Proc. Natl. Acad. Sci. USA. 93: pp 10779-10784. Keon BH, S Schafer, C Kuhn, C Grund, WW Franke, Symplekin, a novel type of tight junction plaque protein, J Cell Biol. 134 (1996) 1003-1018.Malumbres M and M Barbacid, 2001. To cycle or not to cycle: a critical decision in cancer. Nat. Rev. Cancer. 1: pp 222-231. Mitic LL and JM Anderson, 1998. Molecular architecture of tight junctions. Annu. Rev. Physiol. 60: pp 121-142. Rhodin, JAG 1974, Histology, Oxford University Press, New York. Sherr, CJ 2000. The Pezcoller lecture: cancer cell cycles revisited. Cancer res. 60: pp 3689-3695. Sourisseau T, Georgiadis A, Tsapara A, Ali RR, Pestell RG, Matter K and Balda MS, 2006. Regulation of PCNA and cyclin D1 expression and epithelial morphogenesis by the ZO-1 regulated transcription factor ZONAB/DbpA. Mol. Cell. Biol. 26, pp 2387-2398.Stevenson, BR, JD Siliciano, MS Mooseker, and DA Goodenough, 1986. Identification of ZO-1: a high molecular weight polypeptide associated with the tight junction (zonula occludens) in a variety of epithelia. J. Cell Biol. 103: pp 755-766. Source document

America Needs Some Gun Control Essay -- argumentative, persuasive, gun

Gun control is the effort to restrict or limit the possession and use of guns. The gun control debate may be one of the most important issues in our society. The U.S. Supreme Court ruled in 2008 that the 2nd Amendment restrains the government’s ability to ban handguns. Some politicians are passionate about gun control and make voting decisions based upon on this issue alone; hence, the people we elect into office directly affect our lives. A few republican politicians have a strong belief in the right to own guns. This battling contradiction can sometimes make it difficult to approach the issue reasonably. The arguments have different statistics and facts regarding how firearms can impact society. It is important to realize that there are strong and valid points to be made for both sides of gun control issues. With that in mind, is there a â€Å"right† or â€Å"wrong† side to this issue? Gun right advocates believe that the 2nd Amendment, â€Å"A well regulated militia, being necessary to the security of a free State, the right of the people to keep and bear arms, shall not be infringed† guarantees the right to own guns and that gun control laws are a violation of their constitutional rights.(constitution) Sarah Palin, Alaska’s former governor, believes that gun control is taking away the rights from the citizens in which the 2nd Amendment has granted them. She is in favor of the constitution and expresses that gun control laws are unconstitutional; â€Å"I support our constitutional right to bear arms†¦You start putting more and more laws on guns and you take away a second amendment right.† (Romano 2) The government is constantly proposing legislation for more and more gun control. However, they cannot be so naive to think criminal... ...ent, and More - Newsweek. 30 Jan. 2011. Web. 09 Feb. 2014. http://www.newsweek.com/2011/01/30/sarah-palin-s-gun-control-warnings-at-safari-club-international.html Romano, Andrew. "Sarah Palin's Gun Control Warnings at Safari Club International - Newsweek." Newsweek - National News, World News, Business, Health, Technology, Entertainment, and More - Newsweek. 30 Jan. 2011. Web. 17 Feb. 2014. http://www.newsweek.com/2011/01/30/sarah-palin-s-gun-control-warnings-at-safari-club-international.html Souter, David. "David Souter on Gun Control." OnTheIssues.org - Candidates on the Issues. 09 Feb. 2010. Web. 17 Feb. 2011. . United States. The U.S. Constitution - The 2nd Amendment. By James Madison, John Jay, and Alexander Hamilton. Web. 10 Feb. 2011. .

Speech on 14th August

Child labour and poverty are inevitably bound together and if you continue to use the labour of children as the treatment for the social disease of poverty, you will have both poverty and child labour to the end of time. (Grace Abbott) Today, there are millions of children who work as wage-earners. They are deprived of childhood, love, nutrition and social association. Child labour emerged during the industrial revolution and today it has become a very serious problem. It is a world-wide phenomenon. Extreme poverty large families, lack of free and compulsory education These children have no chance to attend school and have no choice except to work as unskilled labour. These children are compelled to live below poverty line all their lives. There are many laws against child labour in Pakistan and in other countries but these laws alone cannot control the exploitation of children. We must get the support of all the people of the society to control this menace. According to the law, no child below the age of fourteen can be employed in any hazardous job. Another law states that children should not be made to work beyond their capacity and they should be given opportunities and facilities to develop in a healthy manner. However, all these laws have failed to check the problem of child labour. Stringent laws should be enacted and exemplary punishment should be given to those who exploit children for their selfish end SPARC has conducted research that goes into producing its publications, including three major books on child labour, juvenile justice and child rights. Its annual report The State of Pakistan’s Children and a large number of brochures, SPARC has conducted a number of research studies. SPARC has continued to ask successive governments to upgrade their laws to set a legal age limit for employment in Pakistan, although they have not been successful in doing so.

The Prevalence Of Hiv Among Women - 1824 Words

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We intend recruiting 224~250 participants, based on the sample size calculation below (to accommodate potential losses to follow ups), nationwide in urban hospital clinics from t he largest antenatal centers in Gambia (Please send me the correct link [2]). An informed consent will be obtained and signed by participants. Afterwards, a detailed questionnaire about their demographic variables, physical examination findings, HIV staging, and mode of delivery, breastfeeding and laboratory tests will be performed (Lambert et al., 1997). The specimen will be collected and stored, and the women will be seen on subsequent antenatal or postnatal visits for follow up. The selectionShow MoreRelatedThe Prevalence Of Hiv Is Higher Among Women Essay2100 Words   |  9 PagesThe prevalence of HIV is higher among women (30%) than men (19%) (Ministry of Health [Lesotho] ICF International, 2014). For both men and women, HIV prevalence increases with age and then declines. 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