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R Bucala

Publications and source records attributed to R Bucala.

At least 55 records · Page 3Linked to original sources
PubMed2000

Involvement of macrophage migration inhibitory factor (MIF) in experimental uric acid nephropathy.

BACKGROUND: Deposition of uric acid in the kidney can lead to progressive tubulointerstitial injury with granuloma formation. We hypothesized that uric acid crystal deposition may induce granuloma formation by stimulating local expression of macrophage migration inhibitory factor (MIF), which is a known mediator of delayed type hypersensitivity (DTH). MATERIALS AND METHODS: A model of acute uric acid nephropathy was induced in rats by the administration of oxonic acid (an inhibitor of uricase), together with uric acid supplements. MIF expression and local cellular response were examined by in situ hybridization and immunohistochemistry. RESULTS: Kidney tissue examined at 35 days posttreatment showed widespread tubulointerstitial damage with intratubular uric acid crystal deposition and granuloma formation. Tubules within the areas of granuloma showed a six-fold increase in MIF mRNA, compared with uninvolved areas by in situ hybridization. Moreover, the areas of increased MIF mRNA expression correlated with sites of dense accumulation of macrophages and T cells, and these cells were activated when assessed by the expression of interleukin-2R (IL-2R) and (MHC) class II. Interestingly, cytoplasmic staining for MIF protein in the uric acid (UA) crystal-associated granulomatous lesions was reduced, indicating a rapid MIF secretion by damaged tubules and macrophages secondary to uric acid crystal stimulation. This was confirmed by the demonstration of a marked increase in urinary MIF protein by Western blot analysis. Control rats fed either a normal diet or only oxonic acid had no discernible evidence of renal disease by routine light microscopy and minimal tubular expression of MIF mRNA and protein. CONCLUSIONS: These data suggest that intrarenal granulomas in urate nephropathy may be the consequence of a crystal induced DTH reaction mediated by MIF.

Animals↗
PubMed1999

Inhibition of macrophage migration inhibitory factor (MIF) tautomerase activity by dopachrome analogs.

A macrophage migration inhibitory factor (MIF) dopachrome tautomerization assay was employed for identification of MIF inhibitors. One group of dopachrome analogs was identified which inhibits MIF tautomerase activity. In particular, the analogs with a leaving group at beta position displayed activity at a concentration of tenfold less than that of the MIF-substrate. These findings could lead to a better understanding of MIF biological activities and the development of agents for the treatment of MIF related diseases.

Indolequinones↗
PubMed1999

A receptor for advanced glycosylation endproducts (AGEs) is colocalized with neurofilament-bound AGEs and SOD1 in motoneurons of ALS: immunohistochemical study.

Neurofilament (NF)-bound AGEs colocalize immunochemically with SOD1 in the motoneurons of patients with ALS. Among three types of AGE receptors reported in the human brain, AGE-R1 (oligosaccharyltransferase family) and AGE-R2 (substrate of protein kinase C) have been found in neurons, while AGE-R3 is restricted to glia. The present study investigates which of these receptors may be responsible for binding AGEs in the NF conglomerates of motoneurons. Immunostaining of paraffin sections from eight ALS patients (five sporadic and three familial) and three control cases was performed with antibodies directed against R1 and R2, in parallel with those against AGEs and SOD1. The sites of AGE-R1 immunoreactivity (IR) in motoneurons were in conformity to those of NF-associated AGE and SOD1 IRs. By contrast, the IR of R2 was negative in NF conglomerates. Negative R2 IR for NF conglomerates was outlined by surrounding coarse R2 immunopositive granules in the perikaryon. No IR for R1 or R2 was found in hyaline or Bunina inclusions. There was no extraneuronal expression of IR for AGE-R1 or AGEs in microglia or astroglia around the NF accumulation. The colocalization of AGE, AGE-R1, and SOD1 at NF conglomerates in motoneurons supports the notion that AGE-mediated oxidative stress and protein aggregation may be implicated in NF conglomeration and ALS pathogenesis.

Amyotrophic Lateral Sclerosis↗
PubMed1999

Cloning and characterization of the murine histone deacetylase (HDAC3).

Histone acetylation modifiers have been described to participate as cofactors in mammalian transcriptional complexes involved in the regulation of cellular proliferation and differentiation. The acetylation of core histone proteins is reversible and regulated by two competing enzymatic activities, histone acetyltransferases (HATs) and histone deacetylases (HDACs). Increasing evidence suggests a connection between histone acetylation and the development of cancer and leukemia. We have recently mapped HDAC3 to mouse chromosome 18B3, a region which is syntenic with human chromosome 5q31, where HDAC3 is imbedded in a group of potential tumor suppressor genes and which has been reported to be the smallest commonly deleted segment in malignant myeloid disease. We report herein the identification and characterization of HDAC3, a yeast RPD3 ortholog in the mouse. Studies on murine HDAC3 may yield important insights on the understanding of myeloproliferative disease in humans.

Amino Acid Sequence↗
PubMed1999

Sustained mitogen-activated protein kinase (MAPK) and cytoplasmic phospholipase A2 activation by macrophage migration inhibitory factor (MIF). Regulatory role in cell proliferation and glucocorticoid action.

Macrophage migration inhibitory factor (MIF) is an important pro-inflammatory mediator with the unique ability to counter-regulate the inhibitory effects of glucocorticoids on immune cell activation. MIF is released from cells in response to glucocorticoids, certain pro-inflammatory stimuli, and mitogens and acts to regulate glucocorticoid action on the ensuing inflammatory response. To gain insight into the molecular mechanism of MIF action, we have examined the role of MIF in the proliferation and intracellular signaling events of the well characterized, NIH/3T3 fibroblast cell line. Both endogenously secreted and exogenously added MIFs stimulate the proliferation of NIH/3T3 cells, and this response is associated with the activation of the p44/p42 extracellular signal-regulated (ERK) mitogen-activated protein kinases (MAP). The MIF-induced activation of these kinases was sustained for a period of at least 24 h and was dependent upon protein kinase A activity. We further show that MIF regulates cytosolic phospholipase A2 activity via a protein kinase A and ERK dependent pathway and that the glucocorticoid suppression of cytokine-induced cytoplasmic phospholipase A2 activity and arachidonic acid release can be reversed by the addition of recombinant MIF. These studies indicate that the sustained activation of p44/p42 MAP kinase and subsequent arachidonate release by cytoplasmic phospholipase A2 are important features of the immunoregulatory and intracellular signaling events initiated by MIF and provide the first insight into the mechanisms that underlie the pro-proliferative and inflammatory properties of this mediator.

3T3 Cells↗
PubMed1999

An inducible gene product for 6-phosphofructo-2-kinase with an AU-rich instability element: role in tumor cell glycolysis and the Warburg effect.

Cancer cells maintain a high glycolytic rate even in the presence of oxygen, a phenomenon first described over 70 years ago and known historically as the Warburg effect. Fructose 2,6-bisphosphate is a powerful allosteric regulator of glycolysis that acts to stimulate the activity of 6-phosphofructo-1-kinase (PFK-1), the most important control point in mammalian glycolysis. The steady state concentration of fructose 2,6-bisphosphate in turn depends on the activity of the enzyme 6-phosphofructo-2-kinase (PFK-2)/fructose-2, 6-bisphosphatase, which is expressed in several tissue-specific isoforms. We report herein the identification of a gene product for this enzyme that is induced by proinflammatory stimuli and which is distinguished by the presence of multiple copies of the AUUUA mRNA instability motif in its 3'-untranslated end. This inducible gene for PFK-2 is expressed constitutively in several human cancer cell lines and was found to be required for tumor cell growth in vitro and in vivo. Inhibition of inducible PFK-2 protein expression decreased the intracellular level of 5-phosphoribosyl-1-pyrophosphate, a product of the pentose phosphate pathway and an important precursor for nucleic acid biosynthesis. These studies identify a regulatory isoenzyme that may be essential for tumor growth and provide an explanation for long-standing observations concerning the apparent coupling of enhanced glycolysis and cell proliferation.

3' Untranslated Regions↗
PubMed1999

Inhibition of advanced glycation endproduct formation by acetaldehyde: role in the cardioprotective effect of ethanol.

Epidemiological studies suggest that there is a beneficial effect of moderate ethanol consumption on the incidence of cardiovascular disease. Ethanol is metabolized to acetaldehyde, a two-carbon carbonyl compound that can react with nucleophiles to form covalent addition products. We have identified a biochemical modification produced by the reaction of acetaldehyde with protein-bound Amadori products. Amadori products typically arise from the nonenzymatic addition of reducing sugars (such as glucose) to protein amino groups and are the precursors to irreversibly bound, crosslinking moieties called advanced glycation endproducts, or AGEs. AGEs accumulate over time on plasma lipoproteins and vascular wall components and play an important role in the development of diabetes- and age-related cardiovascular disease. The attachment of acetaldehyde to a model Amadori product produces a chemically stabilized complex that cannot rearrange and progress to AGE formation. We tested the role of this reaction in preventing AGE formation in vivo by administering ethanol to diabetic rats, which normally exhibit increased AGE formation and high circulating levels of the hemoglobin Amadori product, HbA1c, and the hemoglobin AGE product, Hb-AGE. In this model study, diabetic rats fed an ethanol diet for 4 weeks showed a 52% decrease in Hb-AGE when compared with diabetic controls (P < 0.001). Circulating levels of HbA1c were unaffected by ethanol, pointing to the specificity of the acetaldehyde reaction for the post-Amadori, advanced glycation process. These data suggest a possible mechanism for the so-called "French paradox," (the cardioprotection conferred by moderate ethanol ingestion) and may offer new strategies for inhibiting advanced glycation.

Acetaldehyde↗
PubMed1999

Macrophage migration inhibitory factor in rheumatoid arthritis: evidence of proinflammatory function and regulation by glucocorticoids.

OBJECTIVE: Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine whose involvement in tumor necrosis factor alpha (TNFalpha) synthesis and T cell activation suggests a role in the pathogenesis of rheumatoid arthritis (RA). Antagonism of MIF is associated with marked inhibition of animal models of RA. Uniquely, MIF is inducible by low concentrations of glucocorticoids. We sought to investigate the expression of MIF in RA synovial tissue. METHODS: MIF was demonstrated in human RA synovium by immunohistochemistry, flow cytometry, enzyme-linked immunosorbent assay (ELISA), and reverse transcription-polymerase chain reaction (RT-PCR). Regulation of MIF expression was investigated by treatment of cultured fibroblast-like synoviocytes (FLS) with interleukin-1beta (IL-1beta), TNFalpha, or interferon-gamma (IFNgamma), and dexamethasone (DEX). Mononuclear cell TNFalpha release after exposure to FLS-conditioned medium was measured by ELISA. RESULTS: MIF was present in RA synovial lining CD14+ macrophages and FLS. Constitutive MIF messenger RNA (mRNA) expression was demonstrated by RT-PCR of RNA from unstimulated cultured RA FLS, which also released abundant MIF. Serum, synovial fluid, and FLS intracellular MIF were significantly higher in RA patients than in controls. Synoviocyte MIF was not increased by IL-1beta, TNFalpha, or IFNgamma. In contrast, DEX 10(-7)M significantly reduced synoviocyte MIF, while DEX 10(-10)-10(-12)M induced a significant increase in MIF and MIF mRNA. Peripheral blood mononuclear cell TNFalpha release was induced by culture in RA FLS-conditioned medium, and this induction was significantly abrogated by monoclonal anti-MIF antibody, suggesting that MIF is an upstream regulator of TNFalpha release. CONCLUSION: These data represent the first demonstration of the cytokine MIF in human autoimmune disease and suggest MIF as a potential therapeutic target in RA.

Aged↗
PubMed1999

Detoxification of methylglyoxal by the nucleophilic bidentate, phenylacylthiazolium bromide.

Dicarbonyl-containing compounds such as methylglyoxal (MG) are toxic to cells since they can interact with the nucleophilic centers of macromolecules. MG has been found to accumulate during hyperglycemia, and it has been suggested that this reactive dicarbonyl may contribute to the tissue damage and long-term complications of diabetes. A sensitive bacterial assay for investigating the ability of nucleophilic agents to interact with and detoxify MG has been developed. This assay utilizes the sensitivity of exponential phase cells of an Escherichia coli double mutant lacking the KefB and KefC potassium channels toward MG. The bidentate nucleophile, phenylacylthiazolium bromide (PTB), was found to protect and allow the growth of E. coli cells in the presence of either externally added or endogenously produced MG. In the absence of PTB, growth was completely inhibited and rapid cell death occurred under these conditions. PTB protected E. coli against MG almost as well as aminoguanidine, a compound shown previously to be involved in detoxification. The level of protection by PTB against MG was much greater than for the endogenous nucleophile, glutathione. These data suggested that PTB could interact with and detoxify MG. The mechanism of this interaction was characterized by NMR and mass spectroscopy.

Antiporters↗
PubMed1999

Evidence for a role of macrophage migration inhibitory factor in psoriatic skin disease.

Macrophage migration inhibitory factor (MIF) was originally described as a T-cell-derived lymphokine with the potential to inhibit the random migration of macrophages. However, recent reports have shown a much broader tissue distribution, including the skin. Functionally, MIF appears to act as an antagonist of anti-inflammatory glucocorticoid action. To elucidate the role of MIF in inflammatory skin diseases, we investigated the production and localization of this cytokine in human skin of patients with psoriasis by means of reverse transcription-polymerase chain reaction, immunohistochemistry and Western blot analysis. In normal skin, our results showed a moderate but homogeneous MIF immunoreactivity in all epidermal layers. Endothelial cells and the outer root sheath of hair follicles were also positive for MIF. In lesional psoriatic human skin, a significant increase in MIF immunoreactivity was visible in suprabasal keratinocytes, especially of the spinous layer. In addition, endothelial cells also showed increased immunolabelling for MIF in psoriatic lesions, indicating a cell-specific upregulation of this mediator in untreated psoriasis. Western blot analysis also revealed a clear increase in MIF in homogenates of lesional skin from psoriasis patients. These results suggest a role for MIF in the inflammatory skin disease psoriasis.

Adult↗
PubMed1999

Up-regulation of macrophage migration inhibitory factor in acute renal allograft rejection in the rat.

Recent studies have identified a key role for macrophage migration inhibitory factor (MIF) in a number of immune cell-mediated diseases. The current study investigated the potential role of MIF in acute allograft rejection. Lewis rats underwent bilateral nephrectomy and then received an orthotopic DA renal allograft or an orthotopic Lewis renal isograft. Groups of six animals were killed at day 1 or 5 after transplantation. No immunosuppression was used. Animals receiving a renal allograft exhibited severe rejection on day 5, as shown by high levels of serum creatinine, very low rates of creatinine clearance, and severe tubulitis with a dense macrophage and T cell infiltrate. In contrast, isografts had normal renal function on day 5 with no histological evidence of rejection. Northern blotting showed that renal MIF mRNA expression was unchanged at day 1, but was increased 3.5-fold on day 5. In situ hybridization showed a marked increase in MIF mRNA expression by tubular cells and MIF mRNA expression by many infiltrating mononuclear cells in day 5 allografts. Immunostaining confirmed an increase in tubular MIF protein expression, particularly in areas of severe tubular damage with prominent leucocytic infiltration. Double staining showed that many infiltrating macrophages and T cells expressed the MIF protein in day 5 allografts. There was only a minor increase in MIF expression in day 5 isografts, demonstrating that neither surgical injury nor stress cause significant up-regulation of MIF expression in allograft rejection. In conclusion, this study has demonstrated that local MIF production is specifically increased in acute renal allograft rejection. These results suggest that MIF may play an important role in the cellular immune response mediating acute allograft rejection.

Acute Disease↗
PubMed1999

High resolution physical mapping of human HDAC3, a potential tumor suppressor gene in the 5q31 region.

Histone deacetylases have been described as crucial cofactors of mammalian transcriptional complexes. We have recently identified human histone deacetylase HDAC3 on chromosome 5q31 by fluorescence in situ hybridization (FISH) in a region commonly deleted in malignant myeloid disease. Since HDAC3 carries strong potential to be a tumor suppressor gene, we report herein its exact position between the CD14 and GRIA1 genes within the 5q31.1 subband.

Chromosome Mapping↗
PubMed1999

A switch in the cellular localization of macrophage migration inhibitory factor in the rat testis after ethane dimethane sulfonate treatment.

Macrophage migration inhibitory factor (MIF), one of the first cytokines to be discovered, has recently been localized to the Leydig cells in adult rat testes. In the following study, the response of MIF to Leydig cell ablation by the Leydig cell-specific toxin ethane dimethane sulfonate (EDS) was examined in adult male rats. Testicular MIF mRNA and protein in testicular interstitial fluid measured by ELISA and western blot were only marginally reduced by EDS treatment, in spite of the fact that the Leydig cells were completely destroyed within 7 days. Immunohistochemistry using an affinity-purified anti-mouse MIF antibody localized MIF exclusively to the Leydig cells in control testes. At 7 days post-EDS treatment, there were no MIF immunopositive Leydig cells in the interstitium, although distinct MIF immunostaining was observed in the seminiferous tubules, principally in Sertoli cells and residual cytoplasm, and some spermatogonia. A few peritubular and perivascular cells were also labelled at this time, which possibly represented mesenchymal Leydig cell precursors. At 14 and 21 days, Sertoli cell MIF immunoreactivity was observed in only a few tubule cross-sections, while some peritubular and perivascular mesenchymal cells and the re-populating immature Leydig cells were intensely labeled. At 28 days after EDS-treatment, the MIF immunostaining pattern was identical to that of untreated and control testes. The switch in the compartmentalization of MIF protein at 7 days after EDS-treatment was confirmed by western blot analysis of interstitial tissue and seminiferous tubules separated by mechanical dissection. These data establish that Leydig cell-depleted testes continue to produce MIF, and suggest the existence of a mechanism of compensatory cytokine production involving the Sertoli cells. This represents the first demonstration of a hitherto unsuspected pattern of cellular interaction between the Leydig cells and the seminiferous tubules which is consistent with an essential role for MIF in male testicular function.

Animals↗