Search for: rheumatoid arthritis methotrexate autoimmune disease biomarker gene expression GWAS HLA genes non-HLA genes
ID | PMID | Title | PublicationDate | abstract |
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11920576 | Regulation of activation-induced receptor activator of NF-kappaB ligand (RANKL) expression | 2002 Apr | Receptor activator of NF-kappaB ligand (RANKL) is a type II membrane protein of the TNF family and plays a critical role in the regulation of osteoclastogenesis. RANKL expressed on osteoblastic stromal cells has been shown to support osteoclast differentiation originated from hematopoietic precursors. Interestingly, RANKL is also expressed on cells of the immune system including T cells and dendritic cells. We have shown that anti-CD3 could induce RANKL expression in T cell hybridoma A1.1 cells and splenic T cells. RANKL expressed on T cells could effectively induce osteoclast formation from the whole population of murine splenocytes. Furthermore, we have found that the induction of RANKL expression is solely dependent on TCR activation-induced Ca2+ mobilization since its expression can be blocked by cyclosporine A and TMB-8, a Ca2+ mobilization inhibitor. Additionally, treatment of A1.1 cells with ionomycin alone also strongly induces RANKL expression, while phorbol myristate acetate by itself does not. Moreover, although inhibition of c-myc has significant effects on anti-CD3-induced Fas ligand (FasL) expression, we have found that the anti-CD3-induced RANKL expression is independent of c-myc. Surprisingly, in contrast to its inhibitory effect on FasL expression, TGF-beta dramatically increased the expression of anti-CD3-induced RANKL expression. In addition to its potential role in immune responses, RANKL expressed on activated T lymphocytes may provide a mechanism for the communication between the immune and skeletal systems during immune responses and disease states such as rheumatoid arthritis. | |
11866530 | Solution structure of a phage-derived peptide antagonist in complex with vascular endothel | 2002 Feb 22 | Vascular endothelial growth factor (VEGF) is a potent endothelial cell-specific mediator of angiogenesis and vasculogenesis. VEGF is involved pathologically in cancer, proliferative retinopathy and rheumatoid arthritis, and as such represents an important therapeutic target. Three classes of disulfide-constrained peptides that antagonize binding of the VEGF dimer to its receptors, KDR and Flt-1, were identified previously using phage display methods. NMR studies of a representative peptide from the most potent class of these peptide antagonists, v107 (GGNECDAIRMWEWECFERL), were undertaken to characterize its interactions with VEGF. v107 has no defined structure free in solution, but binding to VEGF induces folding of the peptide. The solution structure of the VEGF receptor-binding domain-v107 complex was determined using 3940 (1970 per VEGF monomer) internuclear distance and 476 (238 per VEGF monomer) dihedral angle restraints derived from NMR data obtained using samples containing either (13)C/(15)N-labeled protein plus excess unlabeled peptide or (13)C/(15)N-labeled peptide plus excess unlabeled protein. Residual dipolar coupling restraints supplemented the structure determination of the complex and were found to increase significantly both the global precision of VEGF in the complex and the agreement with available crystal structures of VEGF. The calculated ensemble of structures is of high precision and is in excellent agreement with the experimental restraints. v107 has a turn-helix conformation with hydrophobic residues partitioned to one face of the peptide and polar or charged residues at the other face. Contacts between two v107 peptides and the VEGF dimer are mediated by primarily hydrophobic side-chain interactions. The v107-binding site on VEGF overlaps partially with the binding site of KDR and is similar to that for domain 2 of Flt-1. The structure of the VEGF-v107 complex provides new insight into how binding to VEGF can be achieved that may be useful for the design of small molecule antagonists. | |
22649804 | (68)Ga-1,4,7-Triazacyclononane-1,4,7-triacetic acid-c(RGDfK)-human serum albumin-tissue in | 2004 | Extracellular matrix (ECM) adhesion molecules consist of a complex network of fibronectins, collagens, chondroitins, laminins, glycoproteins, heparin sulfate, tenascins, and proteoglycans that surround connective tissue cells, and they are mainly secreted by fibroblasts, chondroblasts, and osteoblasts (1). Cell substrate adhesion molecules are considered essential regulators of cell migration, differentiation, and tissue integrity and remodeling. These molecules play a role in inflammation and atherogenesis, but they also participate in the process of invasion and metastasis of malignant cells in the host tissue (2). Tumor cells adhere to the ECM, which provides a matrix environment for permeation of tumor cells through the basal lamina and underlying interstitial stroma of the connective tissue. Overexpression of matrix metalloproteinases (MMPs) and other proteases by tumor cells allows intravasation of tumor cells into the circulatory system after degrading the basement membrane and ECM (3). Several families of MMPs are involved in atherogenesis, myocardial infarction, angiogenesis, and tumor invasion and metastases (4-7). MMP expression is highly regulated in normal cells, such as trophoblasts, osteoclasts, neutrophils, and macrophages. Elevated levels of MMPs have been found in tumors associated with a poor prognosis for cancer patients (8). There are four members of endogenous tissue inhibitors of metalloproteinases (TIMPs), which regulate the activity of MMPs leading to inhibition of tumor growth and metastasis (9, 10). TIMP-2 (TIMP2) is a bifunctional inhibitor of angiogenesis by inhibition of proteinase activity of MMPs and endothelial cell proliferation via binding to α(3)β(1) (the N-terminal domain) and by MMP-independent anti-angiogenic activity (the C-terminal domain) (11, 12). Kang et al. (13) fused the N-terminal domain of TIMP2 to the C-terminus of human serum albumin (HSA) to form HSA/TIMP2 fusion protein (HSA-TIMP2), which is readily secreted by the transfected yeast Saccharomyces cerevisiae. HSA-TIMP2 retains its anti-angiogenic activity at the C-terminal domain with little MMP inhibitory activity at the N-terminal domain. Lee et al. (14) have evaluated Cy5.5-HSA/TIMP2 (Cy5.5-HSA-TIMP2) for in vivo near-infrared (NIR) fluorescence imaging of rat prostate MLL tumors in nude mice showing maximum tumor accumulation at 2 d after injection. Integrins are a family of heterodimeric glycoproteins on cell surfaces that mediate diverse biological events involving cell–cell and cell–matrix interactions (15). Integrins consist of an α and a β subunit and are important for cell adhesion and signal transduction. The α(v)β(3) integrin is the most prominent receptor affecting tumor growth, tumor invasiveness, metastasis, tumor-induced angiogenesis, inflammation, osteoporosis, and rheumatoid arthritis (16-21). Expression of the α(v)β(3) integrin is strong on tumor cells and activated endothelial cells, whereas expression is weak on resting endothelial cells and most normal tissues. The peptide sequence Arg-Gly-Asp (RGD) has been identified as a recognition motif used by extracellular matrix proteins (vitronectin, fibrinogen, laminin, and collagen) to bind to a variety of integrins, including α(v)β(3). The α(v)β(3) antagonists are being studied as anti-tumor and anti-angiogenic agents (18, 22, 23). Various radiolabeled RGD peptides (antagonists) have been introduced for imaging of tumors and tumor angiogenesis (24). Choi et al. (25) conjugated multiple c(RGDfK) peptides to HSA-TIMP2 to enhance the binding capacity of the protein (RGD-HSA-TIMP2) to tumors and their vasculatures. (68)Ga-NOTA-RGD-HSA-TIMP2 and (68)Ga-NOTA-HSA-TIMP2 have been studied as potential positron emission tomography (PET) probes for imaging α(v)β(3) integrin receptors in nude mice bearing human glioblastoma U87MG tumors. | |
22649803 | (123)I-c(RGDfK)-human serum albumin-tissue inhibitor of matrix metalloproteinase 2 fusion | 2004 | Extracellular matrix (ECM) adhesion molecules consist of a complex network of fibronectins, collagens, chondroitins, laminins, glycoproteins, heparin sulfate, tenascins, and proteoglycans that surround connective tissue cells, and they are mainly secreted by fibroblasts, chondroblasts, and osteoblasts (1). Cell substrate adhesion molecules are considered essential regulators of cell migration, differentiation, and tissue integrity and remodeling. These molecules play a role in inflammation and atherogenesis, but they also participate in the process of invasion and metastasis of malignant cells in the host tissue (2). Tumor cells adhere to the ECM, which provides a matrix environment for permeation of tumor cells through the basal lamina and underlying interstitial stroma of the connective tissue. Overexpression of matrix metalloproteinases (MMPs) and other proteases by tumor cells allows intravasation of tumor cells into the circulatory system after degrading the basement membrane and ECM (3). Several families of MMPs are involved in atherogenesis, myocardial infarction, angiogenesis, and tumor invasion and metastases (4-7). MMP expression is highly regulated in normal cells, such as trophoblasts, osteoclasts, neutrophils, and macrophages. Elevated levels of MMPs have been found in tumors associated with a poor prognosis for cancer patients (8). There are four members of endogenous tissue inhibitors of metalloproteinases (TIMPs), which regulate the activity of MMPs leading to inhibition of tumor growth and metastasis (9, 10). TIMP-2 (TIMP2) is a bifunctional inhibitor of angiogenesis by inhibition of proteinase activity of MMPs and endothelial cell proliferation via binding to α(3)β(1) (the N-terminal domain) and by MMP-independent anti-angiogenic activity (the C-terminal domain) (11, 12). Kang et al. (13) fused the N-terminal domain of TIMP2 to the C-terminus of human serum albumin (HSA) to form HSA/TIMP2 fusion protein (HSA-TIMP2), which is readily secreted by the transfected yeast Saccharomyces cerevisiae. HSA-TIMP2 retains its anti-angiogenic activity at the C-terminal domain with little MMP inhibitory activity at the N-terminal domain. Lee et al. (14) have evaluated Cy5.5-HSA/TIMP2 (Cy5.5-HSA-TIMP2) for in vivo near-infrared (NIR) fluorescence imaging of rat prostate MLL tumors in nude mice showing maximum tumor accumulation at 2 d after injection. Integrins are a family of heterodimeric glycoproteins on cell surfaces that mediate diverse biological events involving cell–cell and cell–matrix interactions (15). Integrins consist of an α and a β subunit and are important for cell adhesion and signal transduction. The α(v)β(3) integrin is the most prominent receptor affecting tumor growth, tumor invasiveness, metastasis, tumor-induced angiogenesis, inflammation, osteoporosis, and rheumatoid arthritis (16-21). Expression of the α(v)β(3) integrin is strong on tumor cells and activated endothelial cells, whereas expression is weak on resting endothelial cells and most normal tissues. The peptide sequence Arg-Gly-Asp (RGD) has been identified as a recognition motif used by extracellular matrix proteins (vitronectin, fibrinogen, laminin, and collagen) to bind to a variety of integrins, including α(v)β(3). The α(v)β(3) antagonists are being studied as anti-tumor and anti-angiogenic agents (18, 22, 23). Various radiolabeled RGD peptides (antagonists) have been introduced for imaging of tumors and tumor angiogenesis (24). Choi et al. (25) conjugated multiple c(RGDfK) peptides to HSA-TIMP2 to enhance the binding capacity of the protein (RGD-HSA-TIMP2) to tumors and their vasculatures. (123)I-RGD-HSA-TIMP2 and (123)I-HSA-TIMP2 have been studied as potential single-photon emission computed tomography (SPECT) probes for imaging α(v)β(3) receptors in nude mice bearing human glioblastoma U87MG tumors. | |
21656984 | Quantum dot800-poly(ethylene glycol)-c(Arg-Gly-Asp-d-Tyr-Lys). | 2004 | Optical fluorescence imaging is increasingly used to monitor biological functions of specific targets in small animals (1-3). However, the intrinsic fluorescence of biomolecules poses a problem when fluorophores that absorb visible light (350–700 nm) are used. Near-infrared (NIR) fluorescence (700–1,000 nm) detection avoids the background fluorescence interference of natural biomolecules, providing a high contrast between target and background tissues. NIR fluorophores have wider dynamic range and minimal background as a result of reduced scattering compared with visible fluorescence detection. They also have high sensitivity, resulting from low infrared background, and high extinction coefficients, which provide high quantum yields. The NIR region is also compatible with solid-state optical components, such as diode lasers and silicon detectors. NIR fluorescence imaging is becoming a noninvasive alternative to radionuclide imaging in small animals (4, 5). Fluorescent semiconductor quantum dots (QDs) are nanocrystals made of CdSe/CdTe-ZnS with radii of 1–10 nm (6-8). They can be tuned to emit signals in a range of wavelengths by changing their sizes and composition, thus providing broad excitation profiles and high absorption coefficients. They have narrow, symmetric emission spectra with long, excited-state lifetimes (20–50 ns) compared with those of fluorescent dyes (1–10 ns). They process good quantum yields of 40%–90% and possess high extinction coefficients. They are more photo-stable than conventional organic dyes. They can be coated and capped with hydrophilic materials for additional conjugation with biomolecules, such as peptides, antibodies, nucleic acids, and small organic compounds, which have been tested in vitro and in vivo (8-12). Although many cells have been labeled with QDs in vitro with little cytotoxicity, there are limited studies of long-term toxicity of QDs in small animals (13-21); little is known about the toxicity and the mechanisms of clearance and metabolism of QDs in humans. The performance of various nanoparticles for biomedical imaging has been reviewed (22, 23). Integrins are a family of heterodimeric, cell-surface glycoproteins that mediate diverse biological events involving cell–cell and cell–matrix interactions (24). Integrins comprise an α and a β subunit, and they are important for cell adhesion and signal transduction. The α(v)β(3) integrin is the most prominent receptor class, affecting tumor growth, tumor invasiveness, metastasis, tumor-induced angiogenesis, inflammation, osteoporosis, and rheumatoid arthritis (25-30). The α(v)β(3) integrin is strongly expressed on tumor cells and activated endothelial cells. In contrast, expression of α(v)β(3) integrin is weak on resting endothelial cells and on most normal tissues. The α(v)β(3) antagonists are being studied as anti-tumor and anti-angiogenic agents, and the agonists are being studied as angiogenic agents for coronary angiogenesis (29, 31, 32). A tripeptide sequence comprising Arg-Gly-Asp (RGD) has been identified as a recognition motif used by extracellular matrix proteins (vitronectin, fibrinogen, laminin, and collagen) to bind to a variety of integrins including α(v)β(3). Various radiolabeled cyclic RGD peptides have been introduced for tumor imaging and tumor angiogenesis (33). QDs that do not contain Cd (non-Cd QDs) have been prepared as an alternative to the more toxic Cd-based QDs. These newly developed non-Cd QDs exhibit improved blood half-life and minimal reticuloendothelial system accumulation with rapid renal clearance (34). These QDs easily extravasate leaky tumor blood vessels because of their enhanced permeability and retention effects. Gao et al. (35) coated non-Cd QD800 (InAs/InP/ZnSe, emission maximum at 800 nm) with di-stearoyl-poly(ethylene glycol)-phosphatidylethanolamine 2000 (DSPE-PEG2000) and c(RGDyK) to form QD800-PEG-RGD for NIR fluorescence imaging of α(v)β(3) integrin expression in the tumor vasculature in mice. QD800-PEG-RGD has been shown to have a high accumulation in tumors with little extravasation and a predominant liver accumulation. | |
20641316 | Pyro-Gly-Pro-Leu-Gly-Leu-Ala-Arg-Lys(BHQ3). | 2004 | Photodynamic therapy (PDT), also known as photochemotherapy, uses light-activated photosensitizers (PS) in the presence of oxygen to kill cells (1). PDT has become a promising modality to treat skin, esophagus, and lung cancers, as well as other diseases such as atherosclerosis, macular degeneration, and rheumatoid arthritis (2). In PDT, light excites the singlet state of the PS, followed by intersystem transition from the singlet state to the triplet state; then, the energy is transferred from the triplet state of the PS to the triplet ground state of oxygen, (3)O(2)(X(3)Σ(g)(-)) ((3)O(2) triplet state quenching) to generate singlet oxygen, (1)O(2)(a(1)Δ(g)) (3). The produced (1)O(2) is a major cytotoxic agent that has a short life time (<200 ns) and an average diffusion range (~20 nm, which is smaller than the diameter of a cell) (2). Such a short diffusion range requires the delivery of target-specific PS agents into subcellular compartments such as cytoskeletal tubulin, lysosomes, mitochondria, plasma membrane, and the nucleus, where they can generate (1)O(2) efficiently (2). A novel type of PS agent, called a photodynamic molecular beacon (PMB) or killer beacon, has been developed to meet this requirement (2, 4). A typical PMB consists of four modular components: a fluorescent PS, a quencher, a linker, and a delivery vehicle. The target-specific linkers keep the fluorescent PS and the quencher within effective distance of the Föster radius (3–6 nm) (2), which allows efficient fluorescence resonance energy transfer between the fluorescent PS and the quencher. As a result, the fluorescent PS is silent until the PMB meets the target, where the enzyme cleaves the linker and activates the fluorescence of the PS (4). Thus, the PS performs two functions by producing (1)O(2) to kill cells and by illuminating detectable fluorescence to image its own therapeutic outcome (5). Matrix metalloproteinases (MMPs) have been pharmaceutical targets for many years because they play important roles in many diseases such as atherosclerosis, lung pulmonary fibrosis, and cancer (4). MMPs in tumors aid the degradation of extracellular matrix, facilitate neoplastic cell motility, and direct cell invasion (4). Also known as matrilysin, MMP subtype-7 (MMP7) is one of only a few MMPs that are actually secreted by tumor cells (6). Pyro-Gly-Pro-Leu-Gly-Leu-Ala-Arg-Lys(BHQ3) (PP(MMP7)B) is a PMB specific for MMP7, and it is detectable with near-infrared (NIR) fluorescence imaging (4). PP(MMP7)B consists of the infrared fluorescence PS pyropheophorbide α (Pyro), a black hole quencher 3 (BHQ3), and a peptide linker (Gly-Pro-Leu-Gly-Leu-Ala-Arg-Lys (GPLGLARK)) (4). The peptide contains the tripeptide motif Pro-Leu-Gly for MMP7 recognition, and the cleavage site is located between the Gly and Leu residues. Pyro acts as the intracellular delivery vehicle and as the PS (absorption, 665 nm; emission, 675 nm and 720 nm) with good (1)O(2) production (50%). Pyro lacks dark toxicity (toxicity in absence of light) because of its low absorption between 450–600 nm. BHQ3 (absorption, 672 nm) can efficiently quench Pyro fluorescence via fluorescence resonance energy transfer (FRET). The cleavage of Pyro-Gly-Asp-Glu-Val-Asp-Gly-Ser-Gly-Lys(BHQ3) (PPB) by MMP7 separates the PS (Pyro) from the quencher (BHQ3) and restores the Pyro fluorescence for detection. | |
20641887 | (64)Cu-Tetraazacyclododecane-N,N’,N’’,N’’’-tetraacetic acid-quantum dot-c(Arg- | 2004 | Optical fluorescence imaging is increasingly used to monitor biological functions of specific targets in small animals (1-3). However, the intrinsic fluorescence of biomolecules poses a problem when fluorophores that absorb visible light (350–700 nm) are used. Near-infrared (NIR) fluorescence (700–1,000 nm) detection avoids the background fluorescence interference of natural biomolecules, providing a high contrast between target and background tissues. NIR fluorophores have a wider dynamic range and minimal background as a result of reduced scattering compared with visible fluorescence detection. They also have high sensitivity, resulting from low fluorescence background, and high extinction coefficients, which provide high quantum yields. The NIR region is also compatible with solid-state optical components, such as diode lasers and silicon detectors. NIR fluorescence (NIRF) imaging is becoming a non-invasive alternative to radionuclide imaging in small animals (4, 5). Fluorescent semiconductor quantum dots (QDs) are nanocrystals made of CdSe/CdTe-ZnS with radii of 1–10 nm (6-8). They can be tuned to emit light in a range of wavelengths by changing their sizes and composition, thus providing broad excitation profiles and high absorption coefficients. They have narrow and symmetric emission spectra with long excited-state lifetimes (20–50 ns) compared with fluorescent dyes (1–10 ns). QDs possess good quantum yields of 40–90% and high extinction coefficients, and they are more photo-stable than conventional organic dyes. They can be coated and capped with hydrophilic materials for additional conjugations with biomolecules, such as peptides, antibodies, nucleic acids, and small organic compounds, which are tested in vitro and in vivo (8-12). Although many cells have been labeled with QDs in vitro with little cytotoxicity, there are only limited studies of long-term toxicity of QDs in small animals (13-21), and little is known about the toxicity and the mechanisms of clearance and metabolism of QDs in humans. Integrins are a family of heterodimeric, cell-surface glycoproteins that mediate diverse biological events involving cell–cell and cell–matrix interactions (22). Integrins comprise an α and a β subunit, and they are important for cell adhesion and signal transduction. The α(v)β(3) integrin is the most prominent receptor class, affecting tumor growth, tumor invasiveness, metastasis, tumor-induced angiogenesis, inflammation, osteoporosis, and rheumatoid arthritis (23-28). The α(v)β(3) integrin is strongly expressed on tumor cells and activated endothelial cells. In contrast, expression of α(v)β(3) integrin is weak on resting endothelial cells and on most normal tissues. The α(v)β(3) antagonists are being studied as anti-tumor and anti-angiogenic agents, and the agonists are being studied as angiogenic agents for coronary angiogenesis (27, 29, 30). A tripeptide sequence comprising Arg-Gly-Asp (RGD) has been identified as a recognition motif used by extracellular matrix proteins (vitronectin, fibrinogen, laminin, and collagen) to bind to a variety of integrins including α(v)β(3). Various radiolabeled cyclic RGD peptides have been introduced for tumor imaging and tumor angiogenesis (31). The multimodality probe (64)Cu-DOTA-QD-c(RGDyK) has been developed for PET and NIRF imaging of tumor vasculature to study in vivo biodistribution of the tracer in tumor-bearing mice (32). (64)Cu-DOTA-QD-c(RGDyK) has been shown to have a high accumulation in the tumor vasculature with little extravasation and a predominant liver and spleen accumulation. | |
14691172 | Chemokines in autoimmune lacrimal gland disease in MRL/MpJ mice. | 2004 Jan | PURPOSE: MRL/MpJ-fas+/fas+ (MRL/+) and MRL/MpJ-fas(lpr)/fas(lpr) (MRL/lpr) mice undergo spontaneous development of inflammation of the lacrimal and salivary glands, similar to that in the human disorder Sjögren's syndrome. Previous work has shown that these lesions appear to be largely T helper (Th)-2-driven, as evidenced by the substantially greater expression of IL-4 than interferon-gamma. The relative contributions of selected chemokines associated with Th1 and Th2 immune responses were assessed. METHODS: Lacrimal glands from MRL/+ and MRL/lpr mice, at ages 1.5 through 9 months were evaluated by immunohistochemistry for the chemokines monocyte chemoattractant protein (MCP)-1 (also known as chemokine ligand [CCL]-2), MCP-5 (CCL12), thymus activation regulated chemokine (TARC; or CCL17), and macrophage-derived chemokine (MDC; or CCL22). Additional lacrimal glands were tested by real-time RT-PCR for chemokines MCP-1 and -5, which are associated with Th2 and Th1 responses, respectively. RESULTS: By immunohistochemistry a significantly greater proportion of mononuclear inflammatory cells in the lacrimal gland lesions stained for MCP-1 (29%-48% depending on age) compared with MCP-5 (1%-3% depending on age) both in MRL/+ (mean difference 34.2%, P < 0.001) and MRL/lpr (mean difference 33.6%, P < 0.001) substrains. Real-time RT-PCR studies showed higher transcript levels of MCP-1 compared with MCP-5, in both MRL/+ (median difference, 37.3; P < 0.0001) and MRL/lpr (median difference, 77.1; P < 0.0001) mice. Relative transcripts of MCP-1 increased with age in both MRL/+ mice (P = 0.02) and MRL/lpr mice (P = 0.03). Staining for TARC was present on lacrimal gland ductular cells but not on the infiltrating lymphocytes, and staining for MDC was present on scattered individual cells throughout the lacrimal gland, but not on infiltrating lymphocytes. CONCLUSIONS: The predominant expression of a Th2-associated chemokine in the lacrimal gland lesions in this murine model of Sjögren's syndrome may contribute to the predominantly Th2-type lymphoid infiltrate in these tissues. | |
12384933 | Involvement of the interferon-gamma-induced T cell-attracting chemokines, interferon-gamma | 2002 Oct | OBJECTIVE: To elucidate the mechanism of the development of T cell infiltrates in the salivary glands of patients with Sjögren's syndrome (SS), we studied T cell-attracting chemokines and their receptors. METHODS: The expression of the T cell-attracting chemokines, interferon-gamma (IFNgamma)-inducible 10-kd protein (IP-10; also called CXCL10), monokine induced by IFNgamma (Mig; also called CXCL9), and stromal cell-derived factor 1 (SDF-1; also called CXCL12), in salivary glands from SS patients was investigated by polymerase chain reaction-enzyme-linked immunosorbent assay (ELISA). Cells that produce chemokines and lymphocytes that express chemokine receptors were identified by immunohistochemistry. The production of IP-10 and Mig proteins by salivary epithelial cells in response to IFNgamma was determined by ELISA. RESULTS: Expression of IP-10 and Mig messenger RNA (mRNA) was significantly up-regulated in SS salivary glands compared with normal salivary glands (both P < 0.01). There was no significant difference in SDF-1 mRNA expression between the SS and normal salivary glands. IP-10 and Mig proteins were predominantly expressed in the ductal epithelium adjacent to lymphoid infiltrates. Most of the CD3+ infiltrating lymphocytes in dense periductal foci expressed CXCR3, the receptor for IP-10 and Mig. IFNgamma induced the production of high levels of IP-10 and Mig proteins from cultured SS salivary epithelial cells. CONCLUSION: These findings suggest that IFNgamma stimulates the production of IP-10 and Mig in the SS ductal epithelium, and that IP-10 and Mig are involved in the accumulation of T cell infiltrates in the SS salivary gland. Chemokines or chemokine receptors could be a rational new therapeutic target in SS. | |
12974767 | Calreticulin binds preferentially with B cell linear epitopes of Ro60 kD autoantigen, enha | 2003 Oct | Calreticulin is a molecular chaperone to newly synthesized polypeptides. Previous studies suggested that calreticulin is probably a protein member of the Ro/La RNP complex. The aims of this study were (a) to investigate whether linear B cell epitopes of the Ro/La RNP complex are bound to calreticulin and (b) if the complex peptide-calreticulin is recognized specifically by anti-Ro autoantibodies. Calreticulin was isolated from either human or pig spleen using a multi-step purification method and found to interact preferentially with biotinylated peptides derived from the sequence of the Ro60 kD 175-184aa(10p) and 216-232aa(17p). The interaction of the peptide-calreticulin complex was favoured by the combination of heat treatment, divalent cations and ATP. La/SSB epitopes did not react with calreticulin. Peptides corresponding to La/SSB epitopes as well as the common epitope of Sm did not interact with calreticulin. Thirty-eight anti-Ro60 KD positive and 23 anti-Ro60 kD negative sera of patients with systemic lupus erythematosus (SLE) and primary Sjögren's syndrome (pSS) were tested. All anti-Ro60 kD positive sera bound the complex calreticulin-17p, while 95% of the same sera had activity against the complex calreticulin - 10p. Tested individually, calreticulin, pep10p and pep17p presented very low reactivity (8%, 11% and 29%, respectively) against anti-Ro60 kD positive sera. Anti-Ro60 KD negative sera did not exhibit significant reactivity either with calreticulin, 10rho and 17rho or with the complexes calreticulin - 10p and calreticulin-17p (<5%). These results suggest that calreticulin can induce conformation-dependent recognition of the Ro60 kD epitopes, leading eventually to their recognition by autoantibodies. This is the first time that such a relationship is shown between a chaperone protein and fragments of an intracellular autoantigen. This work also provides insights into the understanding of mechanisms for autoantibody production. Furthermore, this association can be proved useful for the development of new sensitive assays for autoantibody detection. | |
12358063 | Recombinant OspC from Borrelia burgdorferi sensu stricto, B. afzelii and B. garinii in the | 2002 Sep | Genes for the outer-surface protein C (OspC) from three north European human isolates of Borrelia burgdorferi sensu stricto, B. afzelii and B. garinii were cloned and sequenced. Polyhistidine-tagged recombinant OspC (rOspC) proteins were produced in Escherichia coli and used, after biotinylation, as antigens on streptavidin-coated plates in enzyme-linked immunosorbent assays (ELISA). In IgM ELISA, 30% (5/17) and 35% (6/17) of patients with erythema migrans (EM) in the acute or convalescent phase, respectively, reacted with one to three rOspCs. Of the patients, 53% (8/15) with neuroborreliosis (NB) and 53% (8/15) with Lyme arthritis (LA) had IgM antibodies to OspC. The immunoreactivity was stronger against rOspC from B. afzelii and B. garinii than against rOspC from B. burgdorferi sensu stricto. In early Lyme borreliosis (LB), rOspC and flagella performed equally well in detecting IgM antibodies. Cross-reactive antibodies to rOspC were observed in serum samples from patients with rheumatoid factor positivity and with syphilis or Epstein-Barr virus (EBV) infection. In IgM ELISA, thiocyanate in the serum dilution buffer reduced EBV-associated non-specific positive reactions. Of the patient sera examined in IgG ELISA, 30% (5/17) with EM in the acute phase, 35% (6/17) with EM in the convalescent phase, 33% (5/15) with NB and 60% (9/15) with LA were positive. Because of the heterogeneity of OspC, a polyvalent antigen with several OspC variants from at least B. afzelii and B. garinii is needed to improve the sensitivity of OspC ELISA in the serodiagnosis of LB in Europe. | |
14613276 | Nitrite-mediated protection against hypochlorous acid-induced chondrocyte toxicity: a nove | 2003 Nov | OBJECTIVE: To examine the potential consequences of overproduction of nitric oxide (NO) and nitrite (NO(2) (-)) in the inflamed rheumatoid joint. METHODS: Human articular chondrocytes in culture were exposed to HOCl (hypochlorous acid, a physiologic oxidant formed in increased amounts at sites of chronic inflammation), and assays of cell viability, intracellular ATP and glutathione (GSH), and lactate dehydrogenase (LDH) were performed. HOCl-induced lipid peroxidation and activation of the MAP kinases ERK-1/2, JNK-1/2, and p38 were also measured. The modulatory effects of NO-derived nitrite (NO(2) (-)) and nitrate (NO(3) (-)) on HOCl-mediated chondrocyte toxicity were investigated. RESULTS: Exposure of human articular chondrocytes to HOCl resulted in a concentration- and time-dependent loss of viability, decrease in ATP and GSH levels, LDH leakage, and cell death. HOCl induced significant lipid peroxidation as well as activation of the MAP kinases ERK-1/2 and p38 but not JNK-1/2. However, the presence of NO(2) (-) but not NO(3) (-) substantially decreased HOCl-dependent cellular toxicity even when NO(2) (-) was added at low (microM) concentrations. In sharp contrast, NO(2) (-) (1 mM) did not inhibit superoxide-, hydroxyl radical-, H(2)O(2)-, or peroxynitrite-mediated cytotoxicity. Furthermore, culture media from cells treated with interleukin-1beta (to generate NO and NO(2) (-)) offered significantly more protection against HOCl-mediated cytotoxicity than culture media from untreated cells. CONCLUSION: These data suggest that NO(2) (-) accumulation at chronically inflamed sites where both HOCl and NO are overproduced may be cytoprotective against damage induced by HOCl. Accumulation of NO(2) (-) could represent a novel cytoprotective role of NO in inflamed joints. A mechanism for this is suggested. | |
15134607 | Perioperative complication rate of total ankle replacement is reduced by surgeon experienc | 2004 May | BACKGROUND: Recent studies suggest the perioperative complication rate of total ankle arthroplasty decreases as a surgeon becomes familiar with the procedure. This study tests the hypothesis that the number of perioperative adverse events will decrease as surgeon experience with total ankle replacement increases. METHODS: Ten surgeons completed retrospective chart and radiographic reviews of their first 10 cases as well as 10 subsequent cases of the Scandinavian Total Ankle Replacement (STAR). Not all surgeons completed 10 cases within the allotted time periods, and two patients were excluded for less than 3-month follow-up, resulting in 187 cases for review. The surgeons performed an average of 12.8 (range, 0-61) STARs between these two time periods. Cases were divided into Early Group if they were among the first five STARs a surgeon performed and Late Group if they were after the first five. RESULTS: The average patient age was 60.4 +/- 12.8 years. The etiology of arthrosis included 96 (51%) of 187 posttraumatic, 49 (26%) idiopathic, and 33 (18%) rheumatoid. Patients in Early Group had a 3.1 times greater chance of having a perioperative adverse event (95% CI 1.6-6.1, p <.001), and a 3.2 times greater chance of having a perioperative wound problem (95% CI 1.5-6.8, p =.002) than patients in Late Group. Patients in Early Group took 1 week longer to heal their wounds than patients in Late Group (4.5 vs. 3.5 weeks, p =.046). CONCLUSIONS: This study shows a decrease in the perioperative adverse event rate commensurate with surgeon experience with STAR. In contrast to other reports, this study was unable to show a decrease in the number of perioperative fractures with increasing surgeon experience. This information is important for planning how best to train surgeons new to total ankle replacement and for patient counseling regarding the potential risks of the procedure. | |
12209039 | Inflammatory myositis associated with anti-U1-small nuclear ribonucleoprotein antibodies: | 2002 Sep | OBJECTIVES: Inflammatory myositides are rare chronic disorders which may be either isolated or associated with other conditions such as connective tissue diseases or neoplasia. A large variety of autoantibodies can be detected in patients with myositis, some of which have a diagnostic and/or a prognostic value. Myositis associated with anti-U1-small nuclear ribonucleoprotein antibodies (anti-U1-snRNP Abs) are usually considered as overlapping syndromes, mainly mixed connective tissue diseases (MCTD) in which muscle symptoms occur insidiously during the disease course and are characterized by a favourable outcome. METHODS: The clinical, biological, immunological and pathological findings as well as the outcome of five patients with anti-U1-snRNP-associated myositis were retrospectively analysed. RESULTS: Patients were mainly black females. In all five patients, myositis was the predominant manifestation at presentation. Associated conditions consisted of interstitial lung disease (ILD) (three), arthritis (three) and neurological symptoms (two). No patient presented Raynaud's phenomenon nor met criteria for MCTD. Biological inflammatory features, rheumatoid factor and polyclonal hypergammaglobulinaemia were present in all cases. Besides anti-U1-snRNP Abs, one patient had anti-Ro/SSA and anti-La/SSB Abs at presentation and one additional patient developed anti-double-stranded-DNA and anti-Sm Abs after a follow-up of more than 4 yr. No patient had anti-PM/sclerosis (Scl) nor anti-aminoacyl-tRNA synthetase Abs. All patients dramatically improved with steroids, and reached complete remission (CR) within 3 weeks. Two patients relapsed 18 months after CR. They both reached rapidly second CR using steroids associated or not with oral methotrexate. CONCLUSION: Our data suggest that anti-U1-snRNP Abs may define a subset of myositis characterized by a favourable outcome, though often associated with ILD and/or neurological manifestations. | |
15086406 | Detection of circulating soluble CD28 in patients with systemic lupus erythematosus, prima | 2004 May | The aim of this study was to evaluate the presence and the role of the serum soluble costimulatory molecule CD28 in patients with systemic lupus erythematosus (SLE), primary Sjögren's syndrome (SS), and systemic sclerosis (SSc). Soluble CD28 concentration was determined by ELISA in 45 patients with SLE, 45 patients with primary SS, 30 patients with SSc, and 45 healthy subjects. We also evaluated CD28 mRNA expression by semiquantitative RT-PCR, and the biological activity of recombinant soluble CD28 on T lymphocyte activity. Concentrations of soluble CD28 were significantly higher in patients with SLE, primary SS and SSc than in healthy subjects. Soluble CD28 concentrations were higher in patients with systemic primary SS than in patients with glandular-limited primary SS. PCR analysis suggested that soluble CD28 resulted from the shedding of the membrane form. In vitro assay revealed that soluble CD28 inhibits the anti-CD3 mAb induced T cell proliferation. Soluble CD28, which modulates the proliferation of T lymphocytes, could be associated with disease severity in patients with autoimmune disease, especially primary SS. These results suggest that soluble CD28 could play an important role in the regulation of autoimmune diseases. | |
15353609 | Putative anti-muscarinic antibodies cannot be detected in patients with primary Sjogren's | 2004 Dec | OBJECTIVE: To determine whether autoantibodies directed against muscarinic M3 receptors are present in the serum of patients with primary Sjogren's syndrome (pSS), and if so whether these autoantibodies inhibit secretion by intact salivary acinar cells. METHODS: IgG was purified by affinity chromatography using protein G from sera collected from 15 patients with pSS. Antibody detection was by Western blotting, whole-cell enzyme-linked immunosorbent assay (ELISA) and immunoblotting. The antisecretory activity of the IgG faction was determined using fura-2 microfluorimetry to measure changes in intracellular Ca(2+) activity ([Ca(2+)](i)) in human and mouse salivary gland acinar cells and in Chinese hamster ovary (CHO) cells transfected with human M3 receptors (CHO-M3). RESULTS: We found no specific M3 receptor recognition by the IgG fraction obtained from pSS patients using either Western blotting or ELISA or immunoblot techniques in which epitope conformation were preserved. Chronic exposure to pSS IgG had no effect on agonist-evoked Ca(2+) signals measured in human or mouse submandibular acinar cells or in CHO-M3 cells. However, acute application of IgG from Sjogren's syndrome patients produced a rapidly reversible reduction in the agonist-stimulated elevation in [Ca(2+)](i). CONCLUSION: These data represent the first demonstration of salivary acinar cell inhibition by pSS IgG; however, this inhibition was found to be reversible. Our data also show that pSS IgG binding to M3R cannot be visualized by conventional immunological approaches. | |
15327370 | Complementary peptide epitopes and anti-idiotypic antibodies in autoimmunity. | 2004 Aug | Application of complementary B and T cell epitopes in inducing anti-idiotypic and anti-clonotypic antibodies capable of regulating or suppressing the autoimmune responses in experimental autoimmune myasthenia gravis (EAMG), allergic neuritis (EAN) and allergic encephalomyelitis (EAE) has been the stimulus of many research efforts. Studies on the idiotypic/anti-idiotypic network of anti-La/SSB positive sera from patients with Sjogren's Syndrome (SS) and Systemic Lupus Erythematosus (SLE) and on animals immunized with the complementary epitopes are presented. | |
12969229 | Salivary gland disease in HIV/AIDS and primary Sjögren's syndrome: analysis of collagen I | 2003 Oct | BACKGROUND: Salivary gland disease (SGD) in HIV/AIDS is clinically and histopathologically very similar to Sjögren's Syndrome (SS), although the mechanism of tissue damage is unknown. The aim of this study is to determine the prevalence of SGD in primary SS and in HIV/AIDS in USA and in West African patients, and to seek distinguishing histopathologic features that may help to elucidate underlying mechanisms. METHODS: Histologic sections of minor salivary glands from 164 HIV-positive and -negative patients from Cameroon and the US, and from 17 US patients with primary SS, were evaluated following salivary gland biopsy for inflammatory changes. To confirm the presence of fibrosis, collagen I, which is the most abundant collagen type, was assessed immunohistochemically in H&E-stained sections. RESULTS: Forty-eight per cent of patients with HIV from Cameroon had severe SGD, while it was only in 6% of patients from the US. Patients with HIV in the US had less fibrosis and collagen I deposits than Cameroonians. Seventy-six per cent of US HIV-positive patients had received anti-retroviral therapy, while none of the African patients had. SS and AIDS patients had a tendency for lymphocytes to locate in a perivascular rather than in a periductal distribution. CONCLUSIONS: The prevalence of SGD and the presence of fibrosis and collagen I in Cameroonians with HIV is significantly higher than in HIV-positive American patients, and is similar to US patients with primary SS. The impact of patient selection, anti-retroviral therapy, and pathogenic mechanisms on salivary gland pathology is discussed. | |
12653859 | Levamisole and Chinese medicinal herbs can modulate the serum interleukin-6 level in patie | 2003 Apr | BACKGROUND: Recurrent aphthous ulcerations (RAU) are common oral inflammatory lesions. Interleukin-6 (IL-6) is a pro-inflammatory cytokine that has effects on cellular and humoral immunities. Previous studies have shown that the high serum IL-6 levels in some RAU patients can be reduced by drug treatment. This finding suggests that IL-6 may be a useful marker in evaluating therapeutic effects of RAU. METHODS: In this study, we used a solid phase, two-site sequential chemiluminescent immunometric assay to determine the baseline serum levels of IL-6 in a group of 228 patients with RAU, erythema multiforme (EM), traumatic ulcers (TU), oral submucous fibrosis (OSF), pemphigus vulgaris (PV), or Sjögren's syndrome (SS), and in 77 normal control subjects. Some RAU patients were treated with levamisole plus Chinese medicinal herbs or levamisole only for 0.5-5 months and their serum IL-6 levels were measured after treatment. RESULTS: We found that about 99% of the normal control subjects and the patients with EM, TU, or OSF had a serum IL-6 level within the normal limit of 5.0 pg/ml. However, 24% (48/197) RAU patients, 14% (1/7) EM patients, 43% (3/7) PV patients, and 100% (6/6) SS patients had a serum level of IL-6 greater than 5.0 pg/ml. The mean serum level of IL-6 in patients with RAU (3.6 +/- 3.5 pg/ml, P < 0.001), minor type RAU (2.7 +/- 2.0 pg/ml, P < 0.05), major type RAU (5.2 +/- 4.6 pg/ml, P < 0.001), or herpetiform type RAU (4.1 +/- 3.8 pg/ml, P < 0.01) was higher than that in normal control subjects. The mean serum level of IL-6 in major type (P < 0.001) or in herpetiform type RAU patients (P < 0.05) was higher than that in minor type RAU patients. The mean reduction of serum IL-6 level (10.0 +/- 7.1 pg/ml) in RAU patients after treatment with levamisole plus Chinese medicinal herbs was significantly higher than that (5.1 +/- 3.7 pg/ml) in RAU patients after treatment with levamisole only (P < 0.005), suggesting that the combination therapy is superior to the single therapy of levamisole only. CONCLUSION: We conclude that levamisole and levamisole plus Chinese medicinal herbs can modulate the serum IL-6 level in RAU patients. Although the therapeutic effect of RAU can be assessed by a decrease in the frequency, duration and number of the oral ulcerations, it can also be monitored by a reduction of serum IL-6 level in RAU patients. | |
15027908 | Activation of Epstein-Barr virus by saliva from Sjogren's syndrome patients. | 2004 Feb | The aim of this study was to examine the mechanism of Epstein-Barr virus (EBV) activation by soluble factors from the inflamed salivary glands of patients with Sjogren's syndrome (SS). Saliva from SS patients was used to examine the regulation of EBV activation by an inflammatory salivary microenvironment. Transient transfection of the EBV-negative salivary gland cell line (HSY) with BZLF1, a trans-activating EBV gene promoter-fusion construct (Zp-luc), was used in this study. The results showed that under conditions where the BZLF1 promoter is activated by potent stimuli, SS saliva (from eight of 12 patients) exerts a significant effect on expression of the luciferase gene. A specific inhibitor of protein kinase C did not affect the SS saliva-induced Zp-luc activity, whereas treatment with inhibitors of calmodulin, calcineurin and IP3, dose-dependently decreased this induction. Transforming growth factor beta1 (TGF-beta1), which is known to be expressed in SS salivary glands, dose-dependently induced Zp-luc activity. Hence, these results demonstrate the activation of EBV by SS saliva and suggest that EBV activation at the inflammatory site may occur in the presence of TGF-beta1 via triggering of the mitogen-activated protein kinase (MAPK) kinase signalling pathway. |