PubMed Health⌕ Search

Biomedical subjects

Dominic Cosgrove

Publications and source records attributed to Dominic Cosgrove.

18 recordsLinked to original sources
PubMed2006

Bone-marrow-derived stem cells repair basement membrane collagen defects and reverse genetic kidney disease.

Type IV collagen is a predominant component of basement membranes, and glomeruli of a kidney filter approximately 70-90 liters of plasma every day through a specialized glomerular basement membrane (GBM). In Alport syndrome, a progressive disease primarily affecting kidneys, mutations in GBM-associated type IV collagen genes (COL4A3, COL4A4, or COL4A5) lead to basement membrane structural defects, proteinuria, renal failure, and an absence of all three GBM collagen triple helical chains because of obligatory posttranslational assembly requirements. Here, we demonstrate that transplantation of wild-type bone marrow (BM) into irradiated COL4A3(-/-) mice results in a possible recruitment of BM-derived progenitor cells as epithelial cells (podocytes) and mesangial cells within the damaged glomerulus, leading to a partial restoration of expression of the type IV collagen alpha3 chain with concomitant emergence of alpha4 and alpha5 chain expression, improved glomerular architecture associated with a significant reduction in proteinuria, and improvement in overall kidney histology compared with untreated COL4A3(-/-) mice or irradiated COL4A3(-/-) mice with BM from adult COL4A3(-/-) mice. The alpha3(IV) collagen produced by BM-derived podocytes integrates into the GBM and associates with other alpha-chains to form type IV collagen triple helical networks. This study demonstrates that BM-derived stem cells can offer a viable strategy for repairing basement membrane defects and conferring therapeutic benefit for patients with Alport syndrome.

Animals↗
PubMed2006

Stage-specific action of matrix metalloproteinases influences progressive hereditary kidney disease.

BACKGROUND: Glomerular basement membrane (GBM), a key component of the blood-filtration apparatus in the in the kidney, is formed through assembly of type IV collagen with laminins, nidogen, and sulfated proteoglycans. Mutations or deletions involving alpha3(IV), alpha4(IV), or alpha5(IV) chains of type IV collagen in the GBM have been identified as the cause for Alport syndrome in humans, a progressive hereditary kidney disease associated with deafness. The pathological mechanisms by which such mutations lead to eventual kidney failure are not completely understood. METHODS AND FINDINGS: We showed that increased susceptibility of defective human Alport GBM to proteolytic degradation is mediated by three different matrix metalloproteinases (MMPs)--MMP-2, MMP-3, and MMP-9--which influence the progression of renal dysfunction in alpha3(IV)-/- mice, a model for human Alport syndrome. Genetic ablation of either MMP-2 or MMP-9, or both MMP-2 and MMP-9, led to compensatory up-regulation of other MMPs in the kidney glomerulus. Pharmacological ablation of enzymatic activity associated with multiple GBM-degrading MMPs, before the onset of proteinuria or GBM structural defects in the alpha3(IV)-/- mice, led to significant attenuation in disease progression associated with delayed proteinuria and marked extension in survival. In contrast, inhibition of MMPs after induction of proteinuria led to acceleration of disease associated with extensive interstitial fibrosis and early death of alpha3(IV)-/- mice. CONCLUSIONS: These results suggest that preserving GBM/extracellular matrix integrity before the onset of proteinuria leads to significant disease protection, but if this window of opportunity is lost, MMP-inhibition at the later stages of Alport disease leads to accelerated glomerular and interstitial fibrosis. Our findings identify a crucial dual role for MMPs in the progression of Alport disease in alpha3(IV)-/- mice, with an early pathogenic function and a later protective action. Hence, we propose possible use of MMP-inhibitors as disease-preventive drugs for patients with Alport syndrome with identified genetic defects, before the onset of proteinuria.

Animals↗
PubMed2006

Role for macrophage metalloelastase in glomerular basement membrane damage associated with alport syndrome.

Alport syndrome is a glomerular basement membrane (GBM) disease caused by mutations in type IV collagen genes. A unique irregular thickening and thinning of the GBM characterizes the progressive glomerular pathology. The metabolic imbalances responsible for these GBM irregularities are not known. Here we show that macrophage metalloelastase (MMP-12) expression is >40-fold induced in glomeruli from Alport mice and is markedly induced in glomeruli of both humans and dogs with Alport syndrome. Treatment of Alport mice with MMI270 (CGS27023A), a broad spectrum MMP inhibitor that blocks MMP-12 activity, results in largely restored GBM ultrastructure and function. Treatment with BAY-129566, a broad spectrum MMP inhibitor that does not inhibit MMP-12, had no effect. We show that inhibition of CC chemokine receptor 2 (CCR2) receptor signaling with propagermanium blocks induction of MMP-12 mRNA and prevents GBM damage. CCR2 receptor is expressed in glomerular podocytes of Alport mice, suggesting MCP-1 activation of CCR2 on podocytes may underlie induction of MMP-12. These data indicate that the irregular GBM that characterizes Alport syndrome may be mediated, in part, by focal degradation of the GBM due to MMP dysregulation, in particular, MMP-12. Thus, MMP-12/CCR2 inhibitors may provide a novel and effective therapeutic stra-tegy for Alport glomerular disease.

Animals↗
PubMed2005

Human alpha1 type IV collagen NC1 domain exhibits distinct antiangiogenic activity mediated by alpha1beta1 integrin.

Human noncollagenous domain 1 of the alpha1 chain of type IV collagen [alpha1(IV)NC1], or arresten, is derived from the carboxy terminal of type IV collagen. It was shown to inhibit angiogenesis and tumor growth in vivo; however, the mechanisms involved are not known. In the present study we demonstrate that human alpha1(IV)NC1 binds to alpha1beta1 integrin, competes with type IV collagen binding to alpha1beta1 integrin, and inhibits migration, proliferation, and tube formation by ECs. Also, alpha1(IV)NC1 pretreatment inhibited FAK/c-Raf/MEK/ERK1/2/p38 MAPK activation in ECs but had no effect on the PI3K/Akt pathway. In contrast, alpha1(IV)NC1 did not affect proliferation, migration, or the activation of FAK/c-Raf/MEK1/2/p38/ERK1 MAPK pathway in alpha1 integrin receptor knockout ECs. Consistent with this, alpha1(IV)NC1 elicited significant antiangiogenic effects and tumor growth inhibition in vivo but failed to do the same in alpha1 integrin receptor knockout mice. This suggests a highly specific, alpha1beta1 integrin-dependent antiangiogenic activity of alpha1(IV)NC1. In addition, alpha1(IV)NC1 inhibited hypoxia-induced expression of hypoxia-inducible factor 1alpha and VEGF in ECs cultured on type IV collagen by inhibiting ERK1/2 and p38 activation. This unravels a hitherto unknown function of human alpha1(IV)NC1 and suggests a critical role for integrins in hypoxia and hypoxia-induced angiogenesis. Collectively, the above data indicate that alpha1(IV)NC1 is a potential therapeutic candidate for targeting tumor angiogenesis.

Angiogenesis Inhibitors↗
PubMed2005

Evidence for functional importance of usherin/fibronectin interactions in retinal basement membranes.

Usher syndrome is a genetically heterogeneous disorder characterized by hearing loss with retinitis pigmentosa. Usher syndrome type IIa is the most common of the Usher syndromes, accounting for over half of all cases. The gene encodes a 180 kDa basement membrane glycoprotein called usherin. Here, we demonstrated a specific interaction between usherin and fibronectin in retinal basement membranes. This interaction was confirmed using biochemical, biophysical, and genetic approaches. Surface plasmon resonance assay confirmed that fibronectin binding to usherin is of high affinity and 1:1 stoichiometry. Using a fusion peptide-based co-immunoprecipitation approach, we show that binding to fibronectin occurs at the LE domain of usherin. Recombinant LE domain-specific peptides were engineered that contained single amino acid substitutions corresponding to missense mutations found in humans with Usher syndrome type IIa. Only mutations in loop d of the LE domain abolished the ability of the LE domain to co-immunoprecipitate fibronectin.

Animals↗
PubMed2005

Dysregulation of renal MMP-3 and MMP-7 in canine X-linked Alport syndrome.

Matrix metalloproteinases (MMPs) play an important regulatory role in many biological and pathological processes and their specific role in Alport syndrome (AS) is not yet clearly defined. In this study, the naturally occurring canine X-linked AS was used to demonstrate a potential role for MMP-3 and MMP-7 in Alport renal pathogenesis. Recently, we demonstrated that the expression of MMP-2, MMP-9 and MMP-14 was upregulated in the renal cortex of dogs with a spontaneous form of XLAS. In the present study, we examined necropsy samples of renal cortex from normal and XLAS dogs for MMP-3 and MMP-7 as they have the potential to activate MMP-2 and MMP-9. Immunohistochemical analysis showed strong immunostaining for both MMP-3 and MMP-7 in the interstitial space of XLAS kidneys, while virtually no immunostaining was observed in similar fields from normal dogs. RT-PCR and casein zymography confirmed that both mRNA transcripts and activities of MMP-3 and MMP-7 are elevated in XLAS kidneys. The induction of these MMPs likely contributes to tissue destruction associated with the fibrogenic process, while augmenting the activation of MMP-2 and MMP-9 by MMP-3 and MMP-7 in XLAS. Thus, these data further implicate a role for the MMPs in progressive renal pathogenesis associated with AS.

Animals↗
PubMed2005

Matrix metalloproteinase dysregulation in the stria vascularis of mice with Alport syndrome: implications for capillary basement membrane pathology.

Alport syndrome results from mutations in genes encoding collagen alpha3(IV), alpha4(IV), or alpha5(IV) and is characterized by progressive glomerular disease associated with a high-frequency sensorineural hearing loss. Earlier studies of a gene knockout mouse model for Alport syndrome noted thickening of strial capillary basement membranes in the cochlea, suggesting that the stria vascularis is the primary site of cochlear pathogenesis. Here we combine a novel cochlear microdissection technique with molecular analyses to illustrate significant quantitative alterations in strial expression of mRNAs encoding matrix metalloproteinases-2, -9, -12, and -14. Gelatin zymography of extracts from the stria vascularis confirmed these findings. Treatment of Alport mice with a small molecule inhibitor of these matrix metalloproteinases exacerbated strial capillary basement membrane thickening, demonstrating that alterations in basement membrane metabolism result in matrix accumulation in the strial capillary basement membranes. This is the first demonstration of true quantitative analysis of specific mRNAs for matrix metalloproteinases in a cochlear microcompartment. Further, these data suggest that the altered basement membrane composition in Alport stria influences the expression of genes involved in basement membrane metabolism.

Animals↗
PubMed2004

A domain-specific usherin/collagen IV interaction may be required for stable integration into the basement membrane superstructure.

Usherin is a basement membrane protein encoded by the gene associated with Usher syndrome type IIa, the most common deaf/blind disorder. This report demonstrates a specific interaction between type IV collagen and usherin in the basement membrane, with a 1:1 stoichiometry for binding. Genetic and biochemical approaches were used to explore the role of type IV collagen binding in usherin function. We demonstrate binding occurs between the LE domain of usherin and the 7S domain of type IV collagen. A purified fusion peptide comprising the first four LE modules was shown to compete with full-length recombinant usherin for type IV collagen binding. However, synonymous fusion peptides with single amino acid substitutions resulting from missense mutations that were known to cause Usher syndrome type IIa in humans, failed to compete. Only mutations in loop b of the LE domain abolished binding activity. Co-immunoprecipitation and western blot analysis of testicular basement membranes from the Alport mouse model show a 70% reduction in type IV collagen is associated with a similar reduction in usherin, suggesting the usherin/collagen (IV) interaction stabilizes usherin in the basement membrane. Thus, the domain-specific interaction between usherin and type IV collagen appears essential to usherin stability in vivo, and loss of this interaction may result in Usher pathology in humans.

Animals↗
PubMed2004

Lack of integrin alpha1beta1 leads to severe glomerulosclerosis after glomerular injury.

Severity of fibrosis after injury is determined by the nature of the injury and host genetic susceptibility. Metabolism of collagen, the major component of fibrotic lesions, is, in part, regulated by integrins. Using a model of glomerular injury by adriamycin, which induces reactive oxygen species (ROS) production, we demonstrated that integrin alpha1-null mice develop more severe glomerulosclerosis than wild-type mice. Moreover, primary alpha1-null mesangial cells produce more ROS both at baseline and after adriamycin treatment. Increased ROS synthesis leads to decreased cell proliferation and increased glomerular collagen IV accumulation that is reversed by antioxidants both in vivo and in vitro. Thus, we have identified integrin alpha1beta1 as a modulator of glomerulosclerosis. In addition, we showed a novel pathway where integrin alpha1beta1 modulates ROS production, which in turn controls collagen turnover and ultimately fibrosis. Because integrin alpha1beta1 is expressed in many cell types this may represent a generalized mechanism of controlling matrix accumulation, which has implications for numerous diseases characterized by fibrosis.

Animals↗
PubMed2004

Immunohistochemistry and reverse transcriptase-polymerase chain reaction as methods for diagnostic determination of usher syndrome type IIa.

OBJECTIVES/HYPOTHESIS: Patients having null mutations in the USH2A gene do not produce usherin and therefore are not positive for immunohistochemical staining of the usherin protein. Thus, immunostaining for usherin can serve as a reliable diagnostic tool for Usher syndrome type IIa. STUDY DESIGN: Prospective. METHODS: Immunohistochemical staining for usherin was carried out in basement membrane of minor salivary gland tissue from subjects with confirmed Usher syndrome type IIa and from archival minor salivary gland tissue from patients without Usher syndrome as control samples. Quantitative usherin messenger RNA analysis was performed using minor salivary gland biopsy tissue. RESULTS: Five subjects with Usher syndrome type IIa had no immunostaining in minor salivary gland tissue, whereas control minor salivary gland tissue did stain with usherin antibody. No usherin RNA was detected in biopsy specimens from patients with confirmed Usher syndrome IIa. CONCLUSION: The feasibility was confirmed of diagnosing Usher syndrome type IIa using purified usherin antibody in subjects having two null USH2A mutations.

Basement Membrane↗
PubMed2003

Neutralization of circulating vascular endothelial growth factor (VEGF) by anti-VEGF antibodies and soluble VEGF receptor 1 (sFlt-1) induces proteinuria.

There are about 2.5 million glomeruli in the kidneys each consisting of a barrel of glomerular basement membrane surrounded by glomerular endothelial cells on the inside and glomerular epithelial cells with established foot processes (podocytes) on the outside. Defects in this filtration apparatus lead to glomerular vascular leak or proteinuria. The role of vascular endothelial growth factor (VEGF) in the regulation of glomerular vascular permeability is still unclear. Recent studies indicate that patients receiving anti-VEGF antibody therapy may have an increased incidence of proteinuria. In a different setting, pregnancies complicated by preeclampsia are associated with elevated soluble VEGF receptor 1 protein (sFlt-1), endothelial cell dysfunction and proteinuria. These studies suggest that neutralization of physiologic levels of VEGF, a key endothelial survival factor, may lead to proteinuria. In the present study, we evaluated the potential of anti-VEGF neutralizing antibodies and sFlt-1 in the induction of proteinuria. Our studies demonstrate that anti-VEGF antibodies and sFlt-1 cause rapid glomerular endothelial cell detachment and hypertrophy, in association with down-regulation of nephrin, a key epithelial protein in the glomerular filtration apparatus. These studies suggest that down-regulation or neutralization of circulating VEGF may play an important role in the induction of proteinuria in various kidney diseases, some forms of cancer therapy and also in women with preeclampsia.

Animals↗
PubMed2003

Monocytes may promote myofibroblast accumulation and apoptosis in Alport renal fibrosis.

BACKGROUND: In interstitial fibrosis, monocytes and myofibroblasts have been directly implicated in scarring, apoptosis, and tissue necrosis. While much has been done to explore the role of these cell types individually in fibrosis, the interactive dependency of monocytes and myofibroblasts has been only marginally explored. METHODS: Alport mice were treated or not with a soluble receptor inhibitor for transforming growth factor-beta 1 (TGF-beta 1), which was previously shown to inhibit the accumulation of myofibroblasts, but not monocytes, in the tubulointerstitium. Kidneys were examined for fibrosis using several matrix markers, TGF-beta 1 mRNA expression by in situ hybridization, apoptosis using the terminal deoxynucleotidyl transferase-mediated uridine triphosphate nick end labeling (TUNEL) assay, expression of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPS) by dual immunofluorescence microscopy, MMP activity by gelatin and in situ zymography, MMP mRNA expression by reverse transcription-polymerase chain reaction (RT-PCR), and basement membrane degradation by dual immunofluorescence confocal microscopy and electron microscopy. RESULTS: Treated mice showed a markedly reduced accumulation of matrix proteins. Tissue monocytes express TGF-beta 1 mRNA, and TGF-beta 1 is required for myofibroblast accumulation. The number of apoptotic cells was not influenced by TGF-beta 1 inhibition. Monocytes express MMP-2, MMP-9, TIMP-2, and TIMP-3. MMP activity and mRNA expression is equally up regulated in treated and untreated Alport mice. Tubular basement membranes (TBM) around clusters of monocytes are notably degraded. TGF-beta 1 inhibition does not extend the life of Alport mice. CONCLUSION: These studies demonstrate that monocytes may influence myofibroblast accumulation via TGF-beta1, and that monocytes, and not myofibroblasts, are associated with tubular atrophy in Alport mice. Elevated MMP expression and activity is associated with TBM destruction near monocytes clusters, suggesting an anoikis mechanism may contribute to apoptosis in this model.

Animals↗
PubMed2003

Increased expression of MMP-2, MMP-9 (type IV collagenases/gelatinases), and MT1-MMP in canine X-linked Alport syndrome (XLAS).

BACKGROUND: Alport syndrome is a group of genetic disorders resulting from mutations in either the alpha3(IV), alpha4(IV) or alpha5(IV) collagen chains. The disease is characterized by a progressive glomerulonephritis, usually associated with a high-frequency specific sensorineural hearing loss, dot and fleck retinopathy, and lens abnormalities. Dogs with naturally occurring genetic disorders of basement membrane collagen (type IV) may serve as animal models of Alport syndrome. In this study, a well-characterized naturally occurring canine model was employed to demonstrate a potential role for matrix metalloproteinases (MMPs) in Alport renal disease pathogenesis. METHODS: Adolescent male dogs that developed renal failure were euthanized and necropsied. Clinicopathologic features of the disease were characterized, and kidneys from normal and Alport dogs were analyzed by gelatin zymography, Western blotting, in situ zymography, immunohistology, and by reverse transcription polymerase chain reaction (RT-PCR) for expression of MMP-2, MMP-9, and membrane type 1-MMP (MT1-MMP). RESULTS: Affected dogs developed proteinuria and rapidly progressive juvenile-onset chronic renal failure. The activities of MMP-2 and MMP-9 were significantly induced in Alport kidney. In situ zymography confirmed elevated active metalloproteinases in kidney cryosections of affected dogs. The mRNAs encoding MMP-2, MMP-9 and MT1-MMP were also increased in Alport dogs suggesting that elevated expression of MMPs reflects events in the progression of Alport syndrome in dogs. CONCLUSION: Elevated expression of MMP-2, MMP-9, and MT1-MMP is observed in fibrotic renal cortex from X-linked Alport syndrome dogs. These findings suggest that MMPs may play an important role in matrix accumulation associated with progressive renal scarring in this model.

Animals↗
PubMed2002

Determinants of vascular permeability in the kidney glomerulus.

The human kidneys filter 70 liters of blood plasma every day. The hallmark of almost all kidney diseases, whether acquired or genetic, is the leakage of plasma proteins into the urine because of alterations in the glomerular filtration unit of the kidney. In this regard, the human mutations in nephrin, podocin, alpha-actinin-4, COL4A3, and COL4A5 genes expressed in the glomeruli have been implicated to cause alterations in glomerular filtration apparatus. Nevertheless, the expression of these proteins in relation to each other in mouse models for glomerular vascular leak is unknown. Additionally, within the glomerulus, the central question of whether the primary filtration barrier is the basement membrane or the epithelial slit diaphragm remains ambiguous. Therefore, in this study, we examined the localization and expression of glomerular epithelial slit diaphragm and glomerular basement membrane proteins implicated in glomerular vascular leak using mice deficient in either the alpha3 chain of type IV collagen, the major constituent of glomerular basement membrane, or LMX1B transcription factor, which regulates the expression of key glomerular type IV collagen genes COL4A3 and COL4A4 or nephrin, a glomerular epithelial slit diaphragm-associated protein. This study demonstrates that decreased expression of slit diaphragm protein, nephrin, correlates with a loss of glomerular filter integrity. Additionally, we demonstrate that defects induced by proteins of glomerular basement membrane lead to an insidious plasma protein leak, whereas the defects induced by proteins in the glomerular epithelial slit diaphragms lead to a precipitous plasma protein leak.

Adaptor Proteins, Signal Transducing↗
PubMed2002

Localization and expression of usherin: a novel basement membrane protein defective in people with Usher's syndrome type IIa.

People with Usher's syndrome type IIa have mutations in a novel gene encoding a protein with domains commonly found in many types of extracellular matrix and cell surface receptor proteins. Here we report that this protein, which we refer to as usherin, is a new basement membrane protein. In the mouse, usherin has a broad, but not ubiquitous, tissue distribution. Usherin is found in all of the capillary and structural basement membranes of the human and mouse retina and in the murine inner ear at both post-natal day 0 and in the adult. High levels of usherin are also observed in tissues not affected in Usher's syndrome type IIa, including spleen, testis, oviduct, epididymis, submaxillary gland, and large and small intestines. Many organs are completely devoid of usherin, including the brain, skin, kidney, lung, liver, and skeletal muscle. Expression was observed in the smooth muscle of the small intestine, colon, and oviduct, however, usherin is not present in cardiac smooth muscle. Usherin is critical for normal development and tissue homeostasis in the inner ear and retina, illustrating yet another example of the importance of basement membranes in the development and function of tissues.

Animals↗
PubMed2002

Strial marginal cells play a role in basement membrane homeostasis: in vitro and in vivo evidence.

The interaction of extracellular matrix and receptors plays a role in tissue homeostasis. The thickened strial capillary basement membrane (SCBM) reported in animal models of presbycusis and Alport's syndrome might be secondary to elevated synthesis and/or decreased turnover of specific basement membrane (BM) components. In this study, expression of specific BM proteins, integrin receptors and mediators of matrix turnover in the murine lateral wall were determined using cDNA probes and antibodies. The presence of collagen alpha1 and alpha2(IV) and laminin-8 in the SCBM was verified. The integrin subunits alpha3, alphav and beta1, cell surface receptors for the BM proteins, localized primarily to the SCBM and/or the strial marginal cells as did TIMP-3, a tissue inhibitor of matrix metalloproteinase. The epithelial cell line SV-k1, derived from the lateral wall of the 'immortomouse', showed expression of the same BM proteins as well as demonstrating the presence of markers specific to strial marginal cells, namely Na,K-ATPase alpha1 and beta2 subunits. Thus, the cultured cells are identified as deriving from marginal cells of the stria vascularis. Moreover, these data suggest that a culture system using this marginal cell line will be useful to delineate mechanisms underlying the pathologic accumulation of extracellular matrix in the SCBM.

Animals↗
PubMed2002

Usherin expression is highly conserved in mouse and human tissues.

Usher syndrome is an autosomal recessive disease that results in varying degrees of hearing loss and retinitis pigmentosa. Three types of Usher syndrome (I, II, and III) have been identified clinically with Usher type II being the most common of the three types. Usher type II has been localized to three different chromosomes 1q41, 3p, and 5q, corresponding to Usher type 2A, 2B, and 2C respectively. Usherin is a basement membrane protein encoded by the USH2A gene. Expression of usherin has been localized in the basement membrane of several tissues, however it is not ubiquitous. Immunohistochemistry detected usherin in the following human tissues: retina, cochlea, small and large intestine, pancreas, bladder, prostate, esophagus, trachea, thymus, salivary glands, placenta, ovary, fallopian tube, uterus, and testis. Usherin was absent in many other tissues such as heart, lung, liver, kidney, and brain. This distribution is consistent with the usherin distribution seen in the mouse. Conservation of usherin is also seen at the nucleotide and amino acid level when comparing the mouse and human gene sequences. Evolutionary conservation of usherin expression at the molecular level and in tissues unaffected by Usher 2a supports the important structural and functional role this protein plays in the human. In addition, we believe that these results could lead to a diagnostic procedure for the detection of Usher syndrome and those who carry an USH2A mutation.

Animals↗