Unraveling the potential and pore-size dependent capacitance of slit-shaped graphitic carbon pores in aqueous electrolytes.

Understanding and leveraging physicochemical processes at the pore scale are believed to be essential to future performance improvements of supercapacitors and capacitive desalination (CD) cells. Here, we report on a combination of electrochemical experiments and fully atomistic simulations to study the effect of pore size and surface charge density on the capacitance of graphitic nanoporous carbon electrodes. Specifically, we used cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) to study the effect of potential and pore size on the capacitance of nanoporous carbon foams. Molecular dynamics simulations were performed to study the pore-size dependent accumulation of aqueous electrolytes in slit-shaped graphitic carbon pores of different widths (0.65 to 1.6 nm). Experimentally, we observe a pronounced increase of the capacitance of sub-nm pores as the applied potential window gets wider, from a few F g(-1) for narrow potential ranges (-0.3 to 0.3 V vs. Ag/AgCl) to ~40 F g(-1) for wider potential windows (-0.9 V to 0.9 V vs. Ag/AgCl). By contrast, the capacitance of wider pores does not depend significantly on the applied potential window. Molecular dynamics simulations confirm that the penetration of ions into pores becomes more difficult with decreasing pore width and increasing strength of the hydration shell. Consistent with our experimental results, we observe a pore- and ion-size dependent threshold-like charging behavior when the pore width becomes comparable to the size of the hydrated ion (0.65 nm pores for Na(+) and 0.79 nm pores for Cl(-) ions). The observed pore-size and potential dependent accumulation of ions in slit-shaped carbon pores can be explained by the hydration structure of the ions entering the charged pores. The results are discussed in view of their effect on energy-storage and desalination efficiency.

Dynamics of nonspherical capsules in shear flow.

Three-dimensional numerical simulations using a front-tracking method are presented on the dynamics of oblate shape capsules in linear shear flow by considering a broad range of viscosity contrast (ratio of internal-to-external fluid viscosity), shear rate (or capillary number), and aspect ratio. We focus specifically on the coupling between the shape deformation and orientation dynamics of capsules, and show how this coupling influences the transition from the tank-treading to tumbling motion. At low capillary numbers, three distinct modes of motion are identified: a swinging or oscillatory (OS) mode at a low viscosity contrast in which the inclination angle theta(t) oscillates but always remains positive; a vacillating-breathing (VB) mode at a moderate viscosity contrast in which theta(t) periodically becomes positive and negative, but a full tumbling does not occur; and a pure tumbling mode (TU) at a higher viscosity contrast. At higher capillary numbers, three types of transient motions occur, in addition to the OS and TU modes, during which the capsule switches from one mode to the other as (i) VB to OS, (ii) TU to VB to OS, and (iii) TU to VB. Phase diagrams showing various regimes of capsule dynamics are presented. For all modes of motion (OS, VB, and TU), a large-amplitude oscillation in capsule shape and a strong coupling between the shape deformation and orientation dynamics are observed. It is shown that the coupling between the shape deformation and orientation is the strongest in the VB mode, and hence at a moderate viscosity contrast, for which the amplitude of shape deformation reaches its maximum. The numerical results are compared with the theories of Keller and Skalak, and Skotheim and Secomb. Significant departures from the two theories are discussed and related to the strong coupling between the shape deformation, inclination, and transition dynamics.

The role of endothelial-to-mesenchymal transition in cancer progression.

Recent evidence has demonstrated that endothelial-to-mesenchymal transition (EndMT) may have a significant role in a number of diseases. Although EndMT has been previously studied as a critical process in heart development, it is now clear that EndMT can also occur postnatally in various pathologic settings, including cancer and cardiac fibrosis. During EndMT, resident endothelial cells delaminate from an organised cell layer and acquire a mesenchymal phenotype characterised by loss of cell-cell junctions, loss of endothelial markers, gain of mesenchymal markers, and acquisition of invasive and migratory properties. Endothelial-to-mesenchymal transition -derived cells are believed to function as fibroblasts in damaged tissue, and may therefore have an important role in tissue remodelling and fibrosis. In tumours, EndMT is an important source of cancer-associated fibroblasts (CAFs), which are known to facilitate tumour progression in several ways. These new findings suggest that targeting EndMT may be a novel therapeutic strategy, which is broadly applicable not only to cancer but also to various other disease states.

Susceptibility to anti-glomerular basement membrane disease and Goodpasture syndrome is linked to MHC class II genes and the emergence of T cell-mediated immunity in mice.

We developed a new mouse model of human anti-glomerular basement membrane (GBM) disease to better characterize the genetic determinants of cell-mediated injury. While all major histocompatibility complex (MHC) haplotypes (H-2a, k, s, b, and d) immunized with alpha3 NC1 domains of type IV collagen produce anti-alpha3(IV) NC1 antibodies that cross-react with human Goodpasture [anti-GBM/anti-alpha3(IV) NC1] autoantibodies, only a few strains developed nephritis and lung hemorrhage associated with Goodpasture syndrome. Crescentic glomerulonephritis and lung hemorrhage were MHC-restricted in haplotypes H-2s, b, and d (A beta/A alpha region in H-2s) and associated with the emergence of an IL-12/Th1-like T cell phenotype. Lymphocytes or anti-alpha3(IV) NC1 antibodies from nephritogenic strains transfer disease to syngeneic recipients. However, passive transfer of isogenic alpha3(IV) NC1 antibodies into -/- T cell receptor-deficient mice failed to produce nephritis. Finally, nephritis and its associated IL-12/Th1-like T cell response attenuate in disease-susceptible mice tolerized orally to alpha3(IV) collagen before immunization. Our findings suggest collectively, as a hypothesis, that anti-GBM antibodies in mice only facilitate disease in MHC haplotypes capable of generating nephritogenic lymphocytes with special T cell repertoires.

Isoform switching of type IV collagen is developmentally arrested in X-linked Alport syndrome leading to increased susceptibility of renal basement membranes to endoproteolysis.

Normal glomerular capillaries filter plasma through a basement membrane (GBM) rich in alpha3(IV), alpha4(IV), and alpha5(IV) chains of type IV collagen. We now show that these latter isoforms are absent biochemically from the glomeruli in patients with X-linked Alport syndrome (XAS). Their GBM instead retain a fetal distribution of alpha1(IV) and alpha2(IV) isoforms because they fail to developmentally switch their alpha-chain use. The anomalous persistence of these fetal isoforms of type IV collagen in the GBM in XAS also confers an unexpected increase in susceptibility to proteolytic attack by collagenases and cathepsins. The incorporation of cysteine-rich alpha3(IV), alpha4(IV), and alpha5(IV) chains into specialized basement membranes like the GBM may have normally evolved to protectively enhance their resistance to proteolytic degradation at the site of glomerular filtration. The relative absence of these potentially protective collagen IV isoforms in GBM from XAS may explain the progressive basement membrane splitting and increased damage as these kidneys deteriorate.

Comparative distribution of the alpha 1(IV), alpha 5(IV), and alpha 6(IV) collagen chains in normal human adult and fetal tissues and in kidneys from X-linked Alport syndrome patients.

We have shown previously that the 5' ends of the genes for the alpha 5(IV) and alpha 6(IV) collagen chains lie head-to-head on Xq22 and are deleted in patients with Alport syndrome (AS)-associated diffuse leiomyomatosis. In this study, we raised a rabbit anti-human alpha 6(IV)chain antibody, demonstrated its specificity by the analysis of recombinant NC1 domains af all six type IV chains, and studied the distribution of the alpha 6(IV) chain in relation to the alpha 1(IV) and alpha 5(IV) chains in human adult and fetal tissues involved in AS and diffuse leiomyomatosis. The alpha 6(IV) chain colocalizes with the alpha 5(IV) chain in basement membranes (BMs) of many tissues, but not in glomerular BM. These data exclude the alpha 6(IV) chain as a site for AS mutations. The head-to-head genomic pairing of the alpha 5(IV) and alpha 6 (IV) genes implies coordinate transcription of the two genes. Differential localization of the alpha 5(IV) and alpha 6(IV) chains shows that the two chains are not always coordinately regulated. The alpha 6(IV) chain, together with the alpha 3(IV)-alpha 5(IV) chains, was absent from all renal BMs in eight patients with X-linked AS while the alpha 1(IV) and alpha 2(IV) chains were increased. The data support the existence of two independent collagen networks, one for the alpha 3(IV)-alpha 6(IV) chains and one for the alpha 1(IV) and alpha 2(IV) chains.

A program of cell death and extracellular matrix degradation is activated in the amnion before the onset of labor.

Fetal membranes usually rupture during the process of labor. Premature fetal membrane rupture occurs not infrequently and is associated with significant fetal and maternal morbidity. The mechanisms of normal and pathologic fetal membrane rupture are not well understood. We have examined structural and biochemical changes in the rat amnion as labor approaches in order to characterize this process in normal pregnancy. Here we report that before the onset of active labor the amnion epithelial cells undergo apoptotic cell death which encompasses degradation of 28S ribosomal subunit RNA and associated P proteins and fragmentation of nuclear DNA. Concurrent with these cellular changes, the amnion type I collagen matrix is degraded with the accumulation of three-quarter length type I collagen fragments in extraembryonic fluid, characteristic of the cleavage of fibrillar collagen by interstitial collagenase. Western blot and immunohistochemical analyses confirmed that interstitial collagenase protein appears in association with the loss of amnion type I collagen. We conclude that amnion epithelial cells undergo a process of programmed cell death associated with orchestrated extracellular matrix degradation which begins before the onset of active labor. Thus, fetal membrane rupture is likely to be the result of biochemical changes as well as physical forces.

NM-3, an isocoumarin, increases the antitumor effects of radiotherapy without toxicity.

We examined the effects of a new antiangiogenic isocoumarin, NM-3, as a radiation modifier in vitro and in vivo. The present studies demonstrate that NM-3 is cytotoxic to human umbilical vein endothelial cells (HUVECs) but not to Lewis lung carcinoma (LLC) cells nor Seg-1, esophageal adenocarcinoma cells, in clonogenic survival assays. When HUVEC cultures are treated with NM-3 combined with ionizing radiation (IR), additive cytotoxicity is observed. In addition, the combination of NM-3 and IR inhibits HUVEC migration to a greater extent than either treatment alone. The effects of treatment with NM-3 and IR were also evaluated in tumor model systems. C57BL/6 female mice bearing LLC tumors were given injections for 4 consecutive days with NM-3 (25 mg/kg/day) and treated with IR (20 Gy) for 2 consecutive days. Combined treatment with NM-3 and IR significantly reduced mean tumor volume compared with either treatment alone. An increase in local tumor control was also observed in LLC tumors in mice receiving NM-3/IR therapy. When athymic nude mice bearing Seg-1 tumor xenografts were treated with NM-3 (100 mg/kg/day for 4 days) and 20 Gy (four 5 Gy fractions), significant tumor regression was observed after combined treatment (NM-3 and IR) compared with IR alone. Importantly, no increase in systemic or local tissue toxicity was observed after combined treatment (NM-3 and IR) when compared with IR alone. The bioavailability and nontoxic profile of NM-3 suggests that the efficacy of this agent should be tested in clinical radiotherapy.

Anti-angiogenic cues from vascular basement membrane collagen.

Vascular basement membrane is an important structural component of blood vessels and has been shown to interact with and modulate vascular endothelial behavior during angiogenesis. During the inductive phase of tumor angiogenesis, this membrane undergoes many degradative and structural changes and reorganizes to a native state around newly formed capillaries in the resolution phase. Such matrix changes are potentially associated with molecular modifications that include expression of matrix gene products coupled with conformational changes, which expose cryptic protein modules for interaction with the vascular endothelium. We speculate that these interactions provide important endogenous angiogenic and anti-angiogenic cues. In this report, we identify an important antiangiogenic vascular basement membrane-associated protein, the 26-kDa NC1 domain of the alpha1 chain of type IV collagen, termed arresten. Arresten was isolated from human placenta and produced as a recombinant molecule in Escherichia coli and 293 embryonic kidney cells. We demonstrate that arresten functions as an anti-angiogenic molecule by inhibiting endothelial cell proliferation, migration, tube formation, and Matrigel neovascularization. Arresten inhibits the growth of two human xenograft tumors in nude mice and the development of tumor metastases. Additionally, we show that the anti-angiogenic activity of arresten is potentially mediated via mechanisms involving cell surface proteoglycans and the alpha1beta1 integrin on endothelial cells. Collectively, our results suggest that arresten is a potent inhibitor of angiogenesis with a potential for therapeutic use.

Structure and composition of type IV collagen of bovine aorta.

To determine the chain composition of type IV collagen of bovine thoracic aorta, we analyzed collagenase-solubilized carboxyl-terminal noncollagenous (NC1)-domains by high-pressure liquid chromatography, two-dimensional electrophoresis, immunoblotting and enzyme-linked immunoassay. In addition to the classical alpha 1- and alpha 2-chains, we found small amounts of the recently discovered alpha 3-, alpha 4- and alpha 5-chains. The alpha 3- and alpha 4-chains were, collectively, 7-13% of the total, and the alpha 5-chain was present in a low amount. Seventy-nine percent of the NC1-domains were dimerized. Immunolocalization studies on sections of aorta showed that the alpha 3- and alpha 5-chains were present, along with alpha 1- and alpha 2-chains, in the subendothelium and media. In capillaries of the media, the alpha 3-chain was found at relatively high levels and was co-localized with alpha 1- and alpha 2-chains. Digestion of aorta with Pseudomonas aeruginosa elastase yielded soluble multimolecular assemblies of type IV collagen. Electron microscopy results provided a direct demonstration of the supramolecular structure, in which the collagen molecules were tetramerized at the amino-terminal end and dimerized at the carboxyl-terminal end.

Identification of alpha3, alpha4, and alpha5 chains of type IV collagen as alloantigens for Alport posttransplant anti-glomerular basement membrane antibodies.

BACKGROUND: Alport syndrome is a hereditary disorder of basement membranes especially affecting the kidneys, ears, and eyes. Some patients who undergo renal transplantation lose their kidneys as a result of posttransplant anti-glomerular basement membrane (anti-GBM) disease. METHODS: In the present study, we analyzed serum from 21 unselected Alport patients who underwent renal transplantation. Eleven samples were from patients without posttransplant anti-GBM nephritis, and 10 were from patients with this disease. RESULTS: Thirteen serum samples [10 alport posttransplant nephritis serum (APTN) and three Alport posttransplant serum (APT)] revealed linear binding to the GBM by indirect immunofluorescence. By using direct ELISA and immunoblotting with GBM constituents and type IV collagen NC1 domains from bovine, human, and recombinant sources, we detected anti-GBM antibodies in all Alport patients in varying titers. Five samples showed specific reactivity to the alpha3 chain, four to the alpha5 chain, six to both alpha3 and alpha5 chains, one to the alpha3 and alpha4 chains, and two to the alpha3, alpha4, and alpha5 chains of type IV collagen. The varied spectrum of reactivities was present equally in nephritic and non-nephritic sera. Ten control samples from non-Alport transplant patients did not exhibit specific binding to the GBM. CONCLUSIONS: These results suggest that the absence of alpha3, alpha4, and alpha5 chains of type IV collagen in the Alport kidney leads to alloantibodies in all Alport patients who receive transplants, irrespective of whether they develop nephritis or not. Although all Alport transplant patients develop this humoral response, only a select few develop anti-GBM disease. We suggest that this difference could be attributable to a genotypic effect on the ability of some individuals to launch a cell-mediated immune response.

Exploring the connection between chronic renal fibrosis and bone morphogenic protein-7.

Tubulointerstitial fibrosis is a hallmark feature of chronic renal injury. Specific therapies to control the progression of renal fibrosis towards end-stage renal failure are still limited. Transforming growth factor-beta1 (TGF-beta1) has been identified as a major mediator of renal fibrosis. Recent reports have suggested that Bone Morphogenic Protein-7 (BMP-7), another member of the TGF-beta superfamily, accelerates repair of acute renal injury and ameliorates progression of chronic renal fibrosis in a variety of animal models. Interestingly, BMP-7, an endogenous molecule which is present in the normal kidney, vastly decreases its expression during renal injury. Although, the mechanism of BMP-7 action in the kidney is not yet fully understood, the idea of an endogenous molecule with reno-protective function is intriguing.

The immunohistochemical composition of corneal basement membrane in keratoconus.

PURPOSE: Keratoconus is a gradually progressing disease of unknown cause, characterized by central thinning, increased curvature, and finally scarring of the cornea. This causes myopia and astigmatism and the ultimate treatment is keratoplasty. We studied the composition of basement membranes (BMs) in normal, scarred and keratoconus corneas to find out possible changes specific for keratoconus. METHODS: Frozen sections of normal, scarred and keratoconus corneas were immunostained with various antibodies against basement membrane (BM) proteins and integrin beta 4. RESULTS: In the keratoconus corneas, we found discontinuities or defects in Bowman's layer, sometimes distorted stroma beneath the defects, and also thinning of the stroma. The results show that within the defects in keratoconus corneas, there is an expression of proteins that are not normally present in the corneal BM, i.e. collagen alpha 1/2 (IV) chains, and on the contrary, absence of the expression of some proteins, i.e. collagen alpha 5-6 (IV) chains that normally are continuously expressed in the corneal epithelial BM. In addition, either increased or decreased expression of laminin-1 (alpha 1 beta 1 gamma 1), laminin-5 (alpha 3 beta 3 gamma 2) and collagen type VII, depending on the keratoconus defect, was seen and the expression of integrin beta 4 was decreased. These findings seem to be specific for keratoconus, as they were not found in scarred corneas. CONCLUSIONS: The results show that the defects in BM and changes in the BM composition are involved in the pathogenesis of keratoconus. Furthermore, it seems that scarring alone does not explain the breaks in Bowman's layer and immunohistochemical changes seen in keratoconus. Therefore, we suggest that a process similar to wound healing, which is initiated by breaks in Bowman's layer, would largely contribute to the differences seen in keratoconus corneas.

Tank-treading and tumbling frequencies of capsules and red blood cells.

This study is motivated in part by the discrepancy that exists in the literature with regard to the dependence of the tank-treading frequency of red blood cells on the shear rate and suspending medium viscosity. Here we consider three-dimensional numerical simulations of deformable capsules of initially spherical and oblate spheroidal shapes and biconcave discoid representing the red blood cell resting shape. By considering a much broader range of the viscosity ratio (ratio of capsule or cell interior to suspending fluid viscosity), shear rate, and aspect ratio (ratio of minor to major axes) than that considered in the previous experiments, we find several new characteristics of the tank-treading and tumbling frequencies that have not been reported earlier. These new characteristics are the result of the large shape deformation and the coupling between shape and angular oscillations of the capsules or cells. For the spherical and oblate spheroidal capsules, the tank-treading frequency shows a nonmonotonic trend that is characterized by an initial decrease leading to a minimum followed by an increase with increasing viscosity ratio. For red blood cells, we find two regimes of the viscosity dependence of the tank-treading frequency: an exponential regime in which the tank-treading frequency decreases at a slower rate with increasing viscosity ratio, and a logarithmic range in which it decreases at a much faster rate. While this trend agrees well with different theoretical models of shape-preserving capsules, it was not evident in previous experimental results. When the shear rate dependence is considered, the tank-treading frequency of red blood cells and capsules of highly elongated initial shapes exhibits a nonmonotonic trend that is characterized by an initial increase leading to a maximum followed by a sharp decrease with decreasing shear rate. This anomalous behavior of the tank-treading frequency is shown to be due to a breathing-like dynamics of the capsule or cell that is characterized by a repeated emergence and absence of deep, crater-like dimples, and a large swinging motion. We further observe that the tumbling frequency exhibits a decreasing trend with increasing viscosity ratio that is in contrast to the theoretical result for the shape-preserving capsules and is due to the periodic deformation and preferential alignment of the capsules in the extensional quadrant of the flow.

Rheology of a dilute suspension of liquid-filled elastic capsules.

Rheology of a dilute suspension of liquid-filled elastic capsules in linear shear flow is studied by three-dimensional numerical simulations using a front-tracking method. This study is motivated by a recent discovery that a suspension of viscous vesicles exhibits a shear viscosity minimum when the vesicles undergo an unsteady vacillating-breathing dynamics at the threshold of a transition between the tank-treading and tumbling motions. Here we consider capsules of spherical resting shape for which only a steady tank-treading motion is observed. A comprehensive analysis of the suspension rheology is presented over a broad range of viscosity ratio (ratio of internal-to-external fluid viscosity), shear rate (or, capillary number), and capsule surface-area dilatation. We find a result that the capsule suspension exhibits a shear viscosity minimum at moderate values of the viscosity ratio, and high capillary numbers, even when the capsules are in a steady tank-treading motion. It is further observed that the shear viscosity minimum exists for capsules with area-dilating membranes but not for those with nearly incompressible membranes. Nontrivial results are also observed for the normal stress differences which are shown to decrease with increasing capillary number at high viscosity ratios. Such nontrivial results neither can be predicted by the small-deformation theory nor can be explained by the capsule geometry alone. Physical mechanisms underlying these results are studied by decomposing the particle stress tensor into a contribution due to the elastic stresses in the capsule membrane and a contribution due to the viscosity differences between the internal and suspending fluids. It is shown that the elastic contribution is shear-thinning, but the viscous contribution is shear thickening. The coupling between the capsule geometry and the elastic and viscous contributions is analyzed to explain the observed trends in the bulk rheology.

Contribution of epithelial plasticity to renal transplantation-associated fibrosis.

Every year in the United States, 5000 renal transplant recipients start or restart dialysis because of the unusual propensity of these allografts to develop interstitial fibrosis and tubular atrophy (IF/TA). Although IF/TA often follows one or more identifiable events, our capacity to specifically treat, prevent, or even detect IF/TA at an early stage is poor. These limitations are largely related to our lack of adequate tools to assess graft failure over time. Data accumulated over the past 5 years have demonstrated that tubular epithelial cells may react to certain fibrogenic stimuli to engage in the process of epithelial-to-mesenchymal transition (EMT). In this review, we highlight the current view of EMT with a focus on both its role in the context of renal transplantation and the potential for utilizing markers of EMT to identify patients undergoing early IF/TA.

Contribution of bone microenvironment to leukemogenesis and leukemia progression.

Tumor microenvironment has a major role in cancer progression and resistance to treatment. The bone marrow (BM) is a dynamic network of growth factors, cytokines and stromal cells, providing a permissive environment for leukemogenesis and progression. Both BM stroma and leukemic blasts promote angiogenesis, which is increased in acute lymphoblastic leukemia and acute myeloid leukemia. Growth factors like vascular endothelial growth factor (VEGF), basic fibroblast growth factor and angiopoietins are the main proangiogenic mediators in acute leukemia. Autocrine proleukemic loops have been described for VEGF and angiopoietin in hematopoietic cells. Interactions of stromal cells and extracellular matrix with leukemic blasts can also generate antiapoptotic signals that contribute to neoplastic progression and persistence of treatment-resistant minimal residual disease. High expression of CXC chemokine ligand 4 (CXCR4) by leukemic blasts and activation of the CXCR4-CXCL12 axis is involved in leukemia progression and disruption of normal hematopoiesis. Leukemia-associated bone microenvironment markers could be used as prognostic or predictive indicators of disease progression and/or treatment outcome. Studies related to bone microenvironment would likely provide a better understanding of the treatment resistance associated with leukemia therapy and design of new treatments.

Choosing a mouse model to study the molecular pathobiology of Alport glomerulonephritis.

Alport syndrome, caused by mutations that interfere with the normal assembly of the alpha3alpha4alpha5(IV) collagen network in the glomerular basement membrane (GBM), is the most common inherited glomerular disease leading to renal failure. A detailed knowledge of the underlying pathogenic mechanisms is necessary for developing new, more specific, and effective therapeutic strategies aimed at delaying the onset and slowing disease progression. Studies of several dog and mouse models of Alport syndrome have significantly enhanced our understanding of the disease mechanisms and provided systems for testing potential therapies. In the most widely used Col4a3-/- mouse models of autosomal-recessive Alport syndrome (ARAS), the genetic background strongly affects renal survival. One contributing factor may be the strong ectopic deposition of alpha5alpha6(IV) collagen in the GBM of Col4a3-/- mice on the C57BL/6J background, which is almost undetectable on the 129/Sv background. This isoform 'switch' has not been observed in human ARAS, although it had been reported in the dog model of ARAS. In human patients as well as dog and mouse models of X-linked Alport syndrome, the alpha3-alpha6(IV) collagen chains are absent from the GBM. These biochemical differences among Alport animal models provide an opportunity to determine how the molecular makeup of the GBM affects the glomerular function. At the same time, potentially confounding influences of characteristics unique to a particular strain or model should be carefully considered in the design of studies aiming to define key events underlying the pathobiology of Alport glomerular disease.

Structural basis for the functions of endogenous angiogenesis inhibitors.

Tipping the angiogenic balance between pro- and antiangiogenic stimuli to favor vasculature induction and enhanced angiogenesis is a key event in the growth and progression of tumors. Recently, we demonstrated that the genetic loss of normal physiological levels of individual endogenous inhibitors of angiogenesis leads to a change in the balance between proangiogenic stimulators and their inhibitors, thus favoring enhanced angiogensis and increased tumor growth. Therefore, these endogenous angiogenesis inhibitors provide a physiological threshold against the induction of angiogenesis. The antiangiogenic activities of endostatin, tumstatin, and thrombospondin-1 are evaluated and correlated with their three-dimensional structure and active sites, deriving a structural basis for their activities. Collectively, structural analysis of all three inhibitors demonstrates that the active antiangiogenic sites on these molecules are exposed on the surface and available to bind their putative integrin receptors on proliferating endothelial cells.

Assembly of type IV collagen. Insights from alpha3(IV) collagen-deficient mice.

Type IV collagen includes six genetically distinct polypeptides named alpha1(IV) through alpha6(IV). These isoforms are speculated to organize themselves into unique networks providing mammalian basement membranes specificity and inequality. Recent studies using bovine and human glomerular and testis basement membranes have shown that unique networks of collagen comprising either alpha1 and alpha2 chains or alpha3, alpha4, and alpha5 chains can be identified. These studies have suggested that assembly of alpha5 chain into type IV collagen network is dependent on alpha3 expression where both chains are normally present in the tissue. In the present study, we show that in the lens and inner ear of normal mice, expression of alpha1, alpha2, alpha3, alpha4, and alpha5 chains of type IV collagen can be detected using alpha chain-specific antibodies. In the alpha3(IV) collagen-deficient mice, only the expression of alpha1, alpha2, and alpha5 chains of type IV collagen was detectable. The non-collagenous 1 domain of alpha5 chain was associated with alpha1 in the non-collagenous 1 domain hexamer structure, suggesting that network incorporation of alpha5 is possible in the absence of the alpha3 chain in these tissues. The present study proves that expression of alpha5 is not dependent on the expression of alpha3 chain in these tissues and that alpha5 chain can assemble into basement membranes in the absence of alpha3 chain. These findings support the notion that type IV collagen assembly may be regulated by tissue-specific factors.