Biomedical subjects

Richard J Quigg

Publications and source records attributed to Richard J Quigg.

At least 19 recordsLinked to original sources

PubMed2007

Genetic network and pathway analysis of differentially expressed proteins during critical cellular events in fracture repair.

Bone repair consists of inflammation, intramembranous ossification, chondrogenesis, endochondral ossification, and remodeling. To better understand the translational regulation of these distinct but interrelated cellular events, we used the second generation of BD Clontechtrade mark Antibody Microarray to dissect and functionally characterize proteins differentially expressed between intact and fractured rat femur at each of these cellular events. Genetic network analysis showed that proteins differentially expressed within a given cellular event tend to be physically or functionally correlated. Seventeen such interacting networks were established over five cellular events that were most frequently associated with cell cycle, cell death, cell-to-cell signaling and interaction, and cell growth and proliferation. Eighteen molecular pathways were significantly enriched during the bone repair process, of which ERK/MAPK, NF-kB, PDGF, and T-cell receptor signaling pathways were significant during three or more cellular events. The analyses revealed dynamic temporal expression patterns and cellular-event-specific functions. The inflammation event on Day 1 was characteristic of the cell cycle-related molecular changes. The relative quiet stage of intramembranous ossification on Day 4 and the molecularly most active stage of chondrogenesis on Day 7 were featured by coordinated cell death and cell-proliferation signals. Endochondral ossification on Day 14 experienced a clear transition from the molecular/cellular function to the physiological system development/function. The osteoclast-mediated remodeling on Day 28 was highlighted by the integrin signaling pathway. The distinct changes in protein expression during these cellular events provide a molecular basis for developing cellular event-targeted therapeutic strategy to accelerate bone healing.

Animals↗

PubMed2007

Unrestricted C3 activation occurs in Crry-deficient kidneys and rapidly leads to chronic renal failure.

Deficiency of the C3 convertase regulator Crry is embryonic lethal in mice unless C3 also is absent. For evaluation of the effect of local kidney Crry deficiency in the setting of an intact complement system, Crry(-/-)C3(-/-) mouse kidneys were transplanted into syngeneic C57BL/6 wild-type mice. These Crry-deficient kidneys developed marked inflammatory cell infiltration, tubular damage, and interstitial fibrosis, whereas similar changes were absent in control transplanted kidneys. Strong C3 deposition in the vessels and tubules that correlated significantly with measures of disease supported that complement activation was pathogenic in this model. Microarray studies showed upregulation of a number of chemokine and extracellular matrix genes, which were validated for CCL2 and CXCL10 mRNA and collagen III protein. The functional significance of these pathophysiologic findings was evaluated by removing both native kidneys, so the transplanted kidney alone provided renal function. Within 21 d of transplantation, seven of eight Crry-deficient kidneys in complement-sufficient wild-type hosts failed, compared with two of 13 controls (P = 0.001), with final blood urea nitrogen levels of 133.9 +/- 33.0 and 55.6 +/- 8.3 mg/dl, respectively (P = 0.015). These data show that mouse Crry is a critical complement regulator in the kidney. When absent, unrestricted complement activation occurs and quickly leads to marked inflammation and progressive renal failure, with features relevant to human diseases with underlying defects in complement regulation, such as hemolytic uremic syndrome.

Animals↗

PubMed2006

Systemic lupus erythematosus and the brain: what mice are telling us.

Neuropsychiatric symptoms occur in systemic lupus erythematosus (SLE), a complex, autoimmune disease of unknown origin. Although several pathogenic mechanisms have been suggested to play a significant role in the etiology of the disease, the exact underlying mechanisms still remain elusive. Several inbred strains of mice are used as models to study SLE, which exhibit a diversity of central nervous system (CNS) manifestations similar to that observed in patients. This review will attempt to give a brief overview of the CNS alterations observed in these models, including biochemical, structural and behavioral changes.

Animals↗

PubMed2006

The simple design of complement factor H: Looks can be deceiving.

The complement system is a powerful component of innate immunity which recognizes and facilitates the elimination of pathogens and unwanted host material. Since complement can also lead to host tissue injury and inflammation, strict regulation of its activation is important. One of the key regulators is complement factor H (CFH), a protein with an ever-expanding list of relevant functions. Inherited mutations in CFH can account for membranoproliferative glomerulonephritis (MPGN) type II, atypical hemolytic uremic syndrome, and age-related macular degeneration. The former can be associated with excessive systemic complement activation from dysfunctional CFH, while the latter two are associated with mutations affecting the ability of CFH to bind to anionic surfaces such as on endothelial cells and glomerular and retinal capillary walls. Mice with targeted deletion of CFH can spontaneously develop MPGN and have increased susceptibility to models of GN. In the rodent, CFH on platelets functions as the immune adherence receptor, analogous to CR1 on primate erythrocytes. In mice, platelets lacking CFH are unable to effectively clear immune complexes which results in their accumulation in glomeruli. The same switch also appears to be true in the rodent podocyte where CFH is present in place of CR1 in human podocytes. Thus, CFH has a variety of functions which can affect the diverse roles the complement system plays in health and disease.

Animals↗

PubMed2006

Distinct and separable roles of the complement system in factor H-deficient bone marrow chimeric mice with immune complex disease.

Plasma complement factor H (Cfh) is a potent complement regulator, whereas Cfh on the surface of rodent platelets is responsible for immune complex processing. For dissection between the two, bone marrow chimeras between Cfh-deficient (Cfh(-/-)) and wild-type C57BL/6 mice were created. Platelet Cfh protein was tracked with the Cfh status of the bone marrow donor, indicating that platelet Cfh is of intrinsic origin. In an active model of immune complex disease, Cfh(-/-) mice that were reconstituted with wild-type bone marrow had levels of platelet-associated immune complexes comparable to those of wild-type mice and were protected against the excessive glomerular deposition of immune complexes seen in Cfh(-/-) mice, yet these mice still developed glomerular inflammation. In contrast, wild-type mice with Cfh(-/-) bone marrow had reduced platelet-associated immune complexes and extensive glomerular deposition of complement-activating immune complexes, but they did not develop glomerular pathology. The large quantities of glomerular C3 in wild-type mice with Cfh(-/-) bone marrow were in the form of iC3b and C3dg, whereas active C3b remained in Cfh(-/-) recipients of wild-type bone marrow. These data show that plasma Cfh limits complement activation in the circulation and other accessible sites such as the glomerulus, whereas platelet Cfh is responsible for immune complex processing.

Animals↗

PubMed2005

Complement-dependent apoptosis and inflammatory gene changes in murine lupus cerebritis.

The role of complement activation in the brains of MRL/lpr lupus mice was determined using the potent C3 convertase inhibitor, CR1-related y (Crry), administered both as an overexpressing Crry transgene and as Crry-Ig. Prominent deposition of complement proteins C3 and C9 in brains of MRL/lpr mice was indicative of complement activation and was significantly reduced by Crry. Apoptosis was determined in brain using different independent measures of apoptosis, including TUNEL staining, DNA laddering, and caspase-3 activity, all of which were markedly increased in lupus mice and could be blocked by inhibiting complement with Crry. Complement activation releases inflammatory mediators that can induce apoptosis. The mRNA for potentially proinflammatory proteins such as TNFR1, inducible NO synthase, and ICAM-1 were up-regulated in brains of lupus mice. Crry prevented the increased expression of these inflammatory molecules, indicating that the changes were complement dependent. Furthermore, microarray analysis revealed complement-dependent up-regulation of glutamate receptor (AMPA-GluR) expression in lupus brains, which was also validated for AMPA-GluR1 mRNA and protein. Our results clearly demonstrate that apoptosis is a prominent feature in lupus brains. Complement activation products either directly and/or indirectly through TNFR1, ICAM-1, inducible NO synthase, and AMPA-GluR, all of which were altered in MRL/lpr mouse brains, have the potential to induce such apoptosis. These findings present the exciting possibility that complement inhibition is a therapeutic option for lupus cerebritis.

Animals↗

PubMed2005

Signaling through up-regulated C3a receptor is key to the development of experimental lupus nephritis.

Signaling of the C3a anaphylatoxin through its G protein-coupled receptor, C3aR, is relevant in a variety of inflammatory diseases, but its role in lupus nephritis is undefined. In this study, we show that expression of C3aR was significantly increased in prediseased and diseased kidneys of MRL/lpr lupus mice compared with MRL/+ controls. To investigate the role of C3aR in experimental lupus, a small molecule antagonist of C3aR (C3aRa) was administered continuously to MRL/lpr mice from 13 to 19 wk of age. All 13 C3aRa-treated mice survived during the 6-wk treatment compared with 9 of 14 (64.3%) control animals given vehicle (p = 0.019). Relative to controls, C3aRa-treated animals were protected from renal disease as measured by albuminuria (p = 0.040) and blood urea nitrogen (p = 0.021). In addition, there were fewer neutrophils, monocytes, and apoptotic cells in the kidneys of C3aRa-treated mice. C3aRa treatment also led to reduced renal IL-1beta and RANTES mRNA and phosphorylated phosphatase and tensin homologue deleted on chromosome 10 protein, whereas the mass of phosphorylated protein kinase B/Akt was increased by C3aRa. Thus, C3aR antagonism significantly reduces renal disease in MRL/lpr mice, which further translates into prolonged survival. These data illustrate that C3aR is relevant in experimental lupus nephritis and may be a target for therapeutic intervention in the human disease.

Animals↗

PubMed2005

Early signals for fracture healing.

Fracture healing requires the cooperation of multiple molecular signaling pathways. To better understand this cascade of transcriptional events, we compared the gene expression profiles between intact bone and fractured bone at days 1, 2, and 4 using a rat femur model of bone healing. Cluster analysis identified several groups of genes with dynamic temporal expression patterns and stage-specific functions. The immediate-response genes are highlighted by binding activity, transporter activity, and energy derivation. We consider these activities as critical signals for initiation of fracture healing. The continuously increased genes are characterized by those directly involved in bone repair, thus, representing bone specific forefront workers. The constantly upregulated genes tend to regulate general cell growth and are enriched with genes that are involved in tumorigenesis, suggesting common pathways between two processes. The constantly downregulated genes predominantly involve immune response, the significance of which remains for further investigation. Knowledge acquired through this analysis of transcriptional activities at the early stage of bone healing will contribute to our understanding of fracture repair and bone-related pathological conditions.

Animals↗

PubMed2005

Complement inhibitors targeted to the proximal tubule prevent injury in experimental nephrotic syndrome and demonstrate a key role for C5b-9.

In glomerular diseases of diverse etiologies, dysfunction of the glomerular barrier to protein passage results in proteinuria, and proteinuria is considered an independent risk factor that plays a direct role in inflammation, interstitial fibrosis, and renal failure. The mechanism by which proteinuria leads to nephrotoxic injury is unclear, but a role for complement in mediating interstitial damage appears likely. We describe a strategy for Ag-specific targeting of complement inhibitors using a single chain Ab fragment and show that complement inhibitors targeted to the tubular epithelium protect against tubulointerstitial injury and renal dysfunction in a rat model of puromycin-induced nephrosis. The targeting of systemically administered complement inhibitors markedly enhanced their efficacy and obviated the need to systemically inhibit complement, thus reducing the risk of compromising host defense and immune homeostasis. Targeted inhibition of complement activation by Crry, and of membrane attack complex (MAC) formation by CD59 was equally therapeutic, demonstrating that the MAC plays a key role in proteinuria-induced tubulointerstitial injury. CD59 activity was dependent on its being targeted to the site of complement activation, and this is the first report of specific inhibition of the MAC in vivo after systemic administration of inhibitor. The data establish the MAC is a valid target for pharmaceutical intervention in proteinuric disorders and provide an approach to investigate the role of the MAC in complement-dependent disease under clinically relevant conditions.

Animals↗

PubMed2005

Contrasting roles of complement activation and its regulation in membranous nephropathy.

The complement system is involved in defense against microorganisms, the processing of immune complexes and apoptotic debris, and the development of an appropriate immune response. Along with these physiologic effects, complement activation has the potential to result in tissue pathology. To limit this, various complement regulatory proteins (CRP) are present on host cells, including the glomerular podocyte. Experimental data from the Heymann nephritis (HN) rat model of human membranous nephropathy (MN) have shown that IgG antibodies in subepithelial immune deposits initiate complement activation and C5b-9-mediated damage of the overlying podocyte. Although IgG can activate the classical pathway, there also is evidence that alternative pathway activation occurs in MN, which could occur because of absent, dysfunctional, or inhibited podocyte CRP. Related to this are experimental data in HN showing the presence of antibodies that bind and inhibit podocyte CRP; although such antibodies have not been documented in human MN, a decrease in CR1 quantity on the podocyte has been observed. A s a result of a relative lack of CRP and the exposure of activating complement proteins to tubular cells, alternative complement pathway activation and C5b-9-mediated tubular injury can occur in MN and other proteinuric diseases. Overall, in a disease such as MN, the balance between complement regulation and activation is tipped toward its being activated. Therefore, a number of therapeutic approaches have been developed to counteract this, including recombinant forms of endogenous CRP and complement-inhibitory monoclonal antibodies. There is good reason to be optimistic that approaches to block complement activation will become viable therapy for human MN in the future.

Animals↗

PubMed2005

C5a promotes development of experimental lupus nephritis which can be blocked with a specific receptor antagonist.

The MRL/lpr murine SLE model has widespread complement activation and deposition of complement fragments in affected tissues. The potent anaphylatoxin C5a has the potential to play a key role in the pathogenesis of lupus nephritis. We found that renal expression of C5aR mRNA and protein was significantly increased in MRL/lpr mice compared to control MRL/+ mice. To examine the role of C5a signaling through C5aR, a specific small molecule antagonist (a) of C5aR was administered continuously to MRL/lpr mice from 13 to 19 wks of age. Littermate controls were given vehicle alone. The progressive impairment in renal function exhibited in the control group was prevented by C5aRa treatment. Infiltration of neutrophils and macrophages into kidneys was significantly reduced in animals treated with C5aRa compared to controls. Furthermore, renal expression of IL-1beta and MIP-2 mRNA as well as the extent of apoptosis were significantly decreased with blockade of C5aR, indicating their dependence upon signals delivered through C5aR. Thus, pharmacological blockade of C5aR reduces disease manifestations in experimental lupus nephritis. These data support an important role for the C5a anaphylatoxin in lupus nephritis, and that blockade of C5aR represents a potentially viable treatment for human lupus nephritis.

Age Factors↗

PubMed2005

Treatment of glomerulonephritis: will we ever have options other than steroids and cytotoxics?

Glomerulonephritis refers to a collection of primary renal disorders and those secondary to a systemic disease, all characterized by inflammation within the glomerulus. Given the underlying immunologic nature of these disorders, they are routinely treated with corticosteriods and various cytotoxic agents. Although in many instances such therapies are successful, they are associated with significant morbidity; as such, alternatives are clearly necessary. Our understanding of the pathogenesis of immunologic glomerular diseases has grown remarkably, in large part from the study of rodent disease models. Fundamental to each disorder is the development of an antigen-specific immune response followed by the effector stage of inflammation. To block the immune response, antigen-specific therapy can be used to induce tolerance, such as through the use of double-stranded DNA molecules in lupus nephritis. Since other antigen systems are less well characterized, inducing a more generalized impairment in the immune response by blocking costimulatory molecules CD40-CD154 and CD28-CD80/86 is a growing approach to treat various immunologic disorders and transplantation. To reduce glomerular inflammation, a variety of effector systems have been targeted, including complement, cytokines/chemokines, adhesion molecules, and mediators of cellular proliferation. Of these, antibodies targeting C5 in the complement system, and antibody and receptor antagonists of tumor necrosis factor-alpha (TNF-alpha) have already been used in glomerular disorders with some promise. Less specific blockade of receptor-mediated events stimulated by platelet-derived growth factors and cell cycle proteins may soon be applied to glomerulonephritis. Finally, interruption of fibrosing pathways, which lead to glomerulosclerosis and interstitial fibrosis common to the end-stage of all glomerulonephritis, is the subject of intense effort which may yield effective biologic therapies. In spite of all these advances, we still are dependent on steroids and cytotoxics to treat glomerulonephritis. To get past this, we must devote significant resources to take observations made in basic research laboratories to develop therapeutics and prove their utility in human disease.

Adrenal Cortex Hormones↗

PubMed2005

Experimental membranous nephropathy redux.

Membranous nephropathy (MN) is a common cause of nephrotic syndrome in adults. Active and passive Heymann nephritis (HN) in rats are valuable experimental models because their features so closely resemble human MN. In HN, subepithelial immune deposits form in situ as a result of circulating antibodies. Complement activation leads to assembly of C5b-9 on glomerular epithelial cell (GEC) plasma membranes and is essential for sublethal GEC injury and the onset of proteinuria. This review revisits HN and focuses on areas of substantial progress in recent years. The response of the GEC to sublethal C5b-9 attack is not simply due to disruption of the plasma membrane but is due to the activation of specific signaling pathways. These include activation of protein kinases, phospholipases, cyclooxygenases, transcription factors, growth factors, NADPH oxidase, stress proteins, proteinases, and others. Ultimately, these signals impact on cell metabolic pathways and the structure/function of lipids and key proteins in the cytoskeleton and slit-diaphragm. Some signals affect GEC adversely. Thus C5b-9 induces partial dissolution of the actin cytoskeleton. There is a decline in nephrin expression, reduction in F-actin-bound nephrin, and loss of slit-diaphragm integrity. Other signals, such as endoplasmic reticulum stress, may limit complement-induced injury, or promote recovery. The extent of complement activation and GEC injury is dependent, in part, on complement-regulatory proteins, which act at early or late steps within the complement cascade. Identification of key steps in complement activation, the cellular signaling pathways, and the targets will facilitate therapeutic intervention in reversing GEC injury in human MN.

Animals↗

PubMed2005

An integrated strategy for the optimization of microarray data interpretation.

The completion of a microarray experiment represents just a starting point toward understanding the biology of interest. A follow-up strategy is needed to fully elucidate the functional significance of microarray-derived measurements of differential expression. Given the fact that no single approach can fully unravel the fundamental biology that is typically quite complex, the follow-up strategy must be integrated at multiple levels encompassing bioinformatics, genomics, and proteomics. In this review, we discuss an integrative approach, which can be used to prioritize microarray-derived candidate genes, define their functions, and place them in the context of the biological system being studied.

Animals↗

PubMed2004

Complement factor h limits immune complex deposition and prevents inflammation and scarring in glomeruli of mice with chronic serum sickness.

Factor H is the major complement regulator in plasma. Abnormalities in factor H have been implicated in membranoproliferative glomerulonephritis in both humans and experimental animals. It has been shown that factor H on rodent platelets functions analogously to human erythrocyte complement receptor 1 in its role to traffic immune complexes to the mononuclear phagocyte system. C57BL/6 factor H-deficient mice (Cfh(-/-)) and wild-type (wt) controls were immunized daily for 5 wk with heterologous apoferritin to study the chronic serum sickness GN model. Immunizations were started in 6- to 8-wk-old mice, which was before the development of spontaneous membranoproliferative glomerulonephritis in some Cfh(-/-) animals. Glomerular deposition of IgG immune complexes in glomeruli was qualitatively and quantitatively increased in Cfh(-/-) mice compared with wt mice. Consistent with the increase in glomerular immune complexes and possibly because of alternative pathway complement activation, Cfh(-/-) mice had increased glomerular C3 deposition. Wt mice developed no glomerular pathology. In contrast, Cfh(-/-) mice developed diffuse proliferative GN with focal crescents and glomerulosclerosis. In addition, there was significantly increased expression of collagen IV, fibronectin, and laminin mRNA in Cfh(-/-) glomeruli. These data show a role for platelet-associated factor H to process immune complexes and limit their accumulation in glomeruli. Once deposited in glomeruli, excessive complement activation can lead to glomerular inflammation and the rapid development of a scarring phenotype.

Animals↗

PubMed2004

Role of Toll-like receptor 4 in endotoxin-induced acute renal failure.

Toll-like receptor 4 (TLR4) is present on monocytes and other cell types, and mediates inflammatory events such as the release of TNF after exposure to LPS. C3H/HeJ mice are resistant to LPS-induced mortality, due to a naturally occurring mutation in TLR4. We therefore hypothesized that LPS-induced acute renal failure (ARF) requires systemic TNF release triggered by LPS acting on extrarenal TLR4. We injected C3H/HeJ mice and C3H/HeOuJ controls with 0.25 mg of LPS, and sacrificed them 6 h later for analysis of blood urea nitrogen (BUN) and kidney tissue (n = 8 per group). In contrast to C3H/HeOuJ controls, C3H/HeJ mice were completely resistant to LPS-induced ARF (6-h BUN of 32.3 +/- 1.1 vs 61.7 +/- 5.6 mg/dl). C3H/HeJ mice released no TNF into the circulation at 2 h (0.00 vs 1.24 +/- 0.16 ng/ml), had less renal neutrophil infiltration (6.4 +/- 1.0 vs 11.4 +/- 1.3 neutrophils per high power field), and less renal apoptosis, as assessed by DNA laddering. Transplant studies showed that C3H/HeJ recipients of wild-type kidneys (n = 9) were protected from LPS-induced ARF, while wild-type recipients of C3H/HeJ kidneys (n = 11) developed severe LPS-induced ARF (24-h BUN 44.0 +/- 4.1 vs 112.1 +/- 20.0 mg/dl). These experiments support our hypothesis that LPS acts on extrarenal TLR4, thereby leading to systemic TNF release and subsequent ARF. Renal neutrophil infiltration and renal cell apoptosis are potential mechanisms by which endotoxemia leads to functional ARF.

ADP-Ribosylation Factor 6↗

PubMed2004

CR1/CR2 deficiency alters IgG3 autoantibody production and IgA glomerular deposition in the MRL/lpr model of SLE.

CR1 and CR2 expression is decreased by approximately 50% on B cells of patients with systemic lupus erythematosus (SLE). Expression is also decreased in the MRL/lpr murine model of SLE prior to the development of clinical disease, suggesting that this alteration may play a role in pathogenesis. To determine whether the decrease in receptor levels affects the development of SLE, we analyzed MRL/lpr mice in which CR1/CR2 expression was altered by gene targeting. Mice from each cohort (Cr2+/+, Cr2+/-, and Cr2-/-) were analyzed biweekly for the development of proteinuria and autoantibodies. Kidneys were examined at 12 and 16 weeks for evidence of immune complex deposition and renal disease. Deficiency of CR1/CR2 did not affect survival or development of renal disease as measured by proteinuria. Mice deficient in CR1/CR2 had significantly lower levels of IgG3 rheumatoid factor (RF) and total serum IgG3, suggesting a specific defect in production of IgG3 in response to endogenous autoantigens. Since IgG3 RF has been associated with the development of vasculitis in this model, we examined the mice for alterations in development of this clinical manifestation. Although there was no difference in the development of ear necrosis among the three groups, renal arteritis was not identified in any of the Cr2+/- mice, whereas it was present in 20% of the Cr2+/- and 40% of the Cr2+/+ mice. Finally, significantly higher levels of IgA were seen in the glomeruli of Cr2+/- mice compared to Cr2+/- or Cr2+/+ mice, suggesting that CR1/CR2 are involved in either the regulation of IgA production or the clearance of IgA immune complexes. Together these data support the concept that alterations in CR1/CR2 expression or function affect the regulation of autoantibody production and/or clearance and may have clinical consequences.

Animals↗

PubMed2004

Acute renal failure in endotoxemia is dependent on caspase activation.

In previous work, it was demonstrated that apoptosis occurs in the kidney during LPS-induced acute renal failure (ARF). However, the relative importance of apoptosis in LPS-induced ARF remained unproven. Because the caspase enzyme cascade is responsible for carrying out apoptosis, it was hypothesized that treatment with a caspase inhibitor would protect mice from LPS-induced ARF. C57BL/6 mice received an injection of LPS and were treated with either the broad-spectrum caspase inhibitor z-VAD-fmk or vehicle and compared with unmanipulated mice. LPS induced a significant increase in caspase-3 activity in vehicle-treated mice, which was significantly inhibited by z-VAD. Mice that were treated with z-VAD were protected from ARF and demonstrated significantly less apoptosis as measured by both terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling staining and DNA laddering. Although apoptosis is classically described as a noninflammatory process, z-VAD treatment significantly attenuated multiple markers of inflammation, such as renal neutrophil infiltration and renal expression of the neutrophil chemotactic factor macrophage inflammatory protein-2. Thus, caspase inhibition may protect against LPS-induced ARF not only by preventing apoptotic cell death but also by inhibiting inflammation. These data raise the possibility that apoptotic kidney cells may actually be a source of this local inflammation, contributing to subsequent nonapoptotic renal injury.

Acute Kidney Injury↗