Osteopontin is associated with T cells in sarcoid granulomas and has T cell adhesive and cytokine-like properties in vitro

Sarcoidosis is a systemic disease characterized by the accumulation of activated T cells and widespread granuloma formation. In addition, individual genetic predisposition appears to be important in this disease. Osteopontin, a noncollagenous matrix protein produced by macrophages and T lymphocytes, is expressed in the granulomas of tuberculosis, and is associated with genetic susceptibility to intracellular infection. The function of osteopontin in these T cell-mediated responses is unknown. We sought to elucidate the role of osteopontin in granulomatous inflammation by characterizing its expression in different stages of sarcoidosis and its effector function on T cells in vitro. Lymphocyte-associated expression of osteopontin in sarcoidosis was demonstrated by immunohistochemistry, and its expression correlated with granuloma maturity. In addition, osteopontin induced T cell chemotaxis, supported T cell adhesion (an effect enhanced by thrombin cleavage of osteopontin), and costimulated T cell proliferation. These results suggest a novel mechanism by which osteopontin and thrombin modulate T cell recruitment and activation in granulomatous inflammation.     

Adenoviral vector-mediated overexpression of serum amyloid A in apoA-I-deficient mice

Serum amyloid A (SAA) is an acute phase reactant that can become the predominant apolipoprotein of high density lipoprotein (HDL) during severe inflammatory states. However, the function of SAA is unknown. To study the ability of SAA to form HDL in the absence of apolipoprotein A-I, we expressed the mouse SAA pI 6.15 (CE/J) isoform in apolipoprotein A-I knock-out (apoA-I (-/-)) mice using a recombinant adenovirus. As a control, apoA-I (-/-) mice were injected with an adenovirus expressing human apoA-I. High level expression of plasma SAA was obtained in the absence of any endogenous acute phase SAA production. SAA expression increased plasma HDL cholesterol levels about 2-fold, but to a lesser extent than the expression of apoA-I (about 10-fold). The HDL particles isolated by density ultracentrifugation from SAA-expressing mice were heterogeneous in size and composition and rich in free cholesterol as well as apoE and apoA-IV. Of the SAA expressed in the plasma, only a small fraction (4%) was associated with HDL particles in contrast to expressed apoA-I, of which 62% was associated with HDL. We conclude that SAA is unable to substitute for apoA-I in HDL particle formation.     

The primary structure of rabbit serum amyloid A protein isolated from acute phase serum

Serum amyloid A (SAA) protein, a sensitive acute phase protein and the precursor of protein AA in secondary amyloid, was purified from pooled acute phase rabbit serum using two different methods: isolation of protein SAA directly by octyl-Sepharose chromatography of total serum, and dissociation and isolation of apoSAA from acute phase high density lipoprotein (HDL). The protein SAA fraction obtained was further purified using gel filtration and ion exchange chromatography. Rabbit protein SAA has 104 amino acid residues, like human SAA, and has a partially blocked N terminus. The highly conserved region from position 33 to position 63 found in SAA from all species studied was confirmed also in rabbit SAA. No microheterogeneities were observed. The amino acid sequence showed extensive N-terminal homology with the rabbit amyloid A protein, except for the microheterogeneity in position 12 in protein AA. It also showed identical amino acid sequence with that deduced from the rabbit cDNA clone pSAA 55. Complete homologies were found with clone SAA 2, except for positions 22 and 78, clone SA8-1, except for positions 22 and 79 and clone SA7-3, except for position 22. This pSAA 55/SA7-3/SA8-1/SAA2-like protein was the only SAA isotype found both in total serum and in the HDL fraction. Isotypes corresponding to other SAA-like genes could not be found in this pool of acute phase rabbit sera.     

NO forms an adduct with serum albumin that has endothelium-derived relaxing factor-like properties

Recent evidence suggests that sulfhydryl species can react with oxides of nitrogen under physiologic conditions and thereby stabilize endothelium-derived relaxing factor (EDRF) activity, but the presence of a specific in vivo thiol carrier for nitric oxide (NO) remains controversial. The single free sulfhydryl of serum albumin is the most abundant thiol species in plasma (approximately 0.5 mM) and is particularly reactive towards NO. To examine the potential role of serum albumin in endogenous nitric oxide metabolism, we synthesized S-nitroso-BSA (S-NO-BSA), a model S-nitroso-protein, and examined its effects on platelet function and coronary and systemic vascular tone in 16 mongrel dogs. Intravenous bolus S-NO-BSA markedly reduced mean arterial pressure in a dose-dependent manner and proved seven and a half-fold less potent than intravenous nitroglycerin and 10-fold less potent than intravenous S-nitroso-cysteine (half-maximal response of 75 nmol/kg compared to 10 and 7.5 nmol/kg, respectively; P < 0.05); when given by intravenous infusion (half-maximal response = 10 nmol/kg per min), however, S-NO-BSA and nitroglycerin were equipotent. Intravenous bolus S-NO-BSA had a greater duration of action than either nitroglycerin or S-nitroso-cysteine and produced marked prolongation of the template bleeding time associated with dose-dependent inhibition of ex vivo platelet aggregation (half-maximal response approximately 70 nmol/kg). Intracoronary S-NO-BSA increased coronary blood flow (mean +/- SEM) less effectively than nitroprusside, acetylcholine, or S-nitroso-cysteine (165% +/- 24% vs. 315% +/- 82%, 483% +/- 55%, or 475% +/- 66%, respectively; P < 0.05) although with much longer duration of action. On a molar basis, S-nitroso-cysteine proved more effective than S-nitroso-BSA, nitroprusside, or acetylcholine as an epicardial coronary vasodilator. Thus, serum albumin reacts with oxides of nitrogen to form a stable S-nitroso-thiol with properties reminiscent of authentic EDRF supporting the view that protein associated thiol may participate in the action and metabolism of EDRF.     

Evolution of the serum amyloid A (SAA) protein superfamily

The serum amyloid A (SAA) superfamily comprises a number of genes and proteins characterized from a range of mammalian species. The majority of members described to date are dramatically induced during the acute-phase response, suggesting an important short-term beneficial role in the response to tissue injury and inflammation. However, important disease associations have also been proposed for certain SAAs during chronic inflammation. The nomenclature of many of the superfamily members has been the result of comparisons with previously reported sequences implying disease association and/or functional relatedness between such members. The evolutionary relationships of the SAA superfamily members have been investigated by comparisons at both the amino acid and the nucleotide level. The results indicate that all members of the superfamily within a species have been undergoing concerted evolution. This has important implications in ascribing functions and disease associations to individual SAA superfamily members and indicates that designations should not be based on the extent of amino acid identity alone but should be made only following direct experimental observation of the proteins themselves.     

Congo red inhibits proteoglycan and serum amyloid P binding to amyloid beta fibrils

Various data suggest that Alzheimer's disease results from the accumulation of amyloid beta (A beta) peptide fibrils and the consequent formation of senile plaques in the cognitive regions of the brain. One approach to lowering senile plaque burden in Alzheimer's disease brain is to identify compounds that will increase the degradation of existing amyloid fibrils. Previous studies have shown that proteoglycans and serum amyloid P (SAP), molecules that localize to senile plaques, bind to A beta fibrils and protect the amyloid peptide from proteolytic breakdown. Therefore, molecules that prevent the binding of SAP and/or proteoglycans to fibrillar A beta might increase plaque degradation and prove useful in the treatment of Alzheimer's disease. The nature of SAP and proteoglycan binding to A beta is defined further in the present study. SAP binds to both fibrillar and nonfibrillar forms of A beta. However, only the former is rendered resistant to proteolysis after SAP association. It is interesting that both SAP and proteoglycan binding to A beta fibrils can be inhibited by glycosaminoglycans and Congo red. Unexpectedly, Congo red protects fibrillar A beta from breakdown, suggesting that this compound and other structurally related molecules are unlikely to be suitable for use in the treatment of Alzheimer's disease.     

Conformational flexibility of the serum amyloid precursor SAA

The present study on canine left ventricles showed that Emax which we previously proposed as a good index of both ventricular contractility and its pumping capability, decreased from 2.06 to 1.38 kPa/ml (15.5 to 10.4 mmHg/ml) via the sino-aortic baroreceptor reflex. Cerebral ischaemic response increased Emax to 3.83 kPa/ml (28.8 mmHg/ml). Emax decreased to 1.17 kPa/ml (8.8 mmHg/ml) after cardiac denervation.     

Human sexual orientation has a heritable component

We present an overview of behavioral genetics research on homosexual and heterosexual orientation. Family, twin, and adoptee studies indicate that homosexuality and thus heterosexuality run in families. Sibling, twin, and adoptee concordance rates are compatible with the hypothesis that genes account for at least half of the variance in sexual orientation. We note observations of homosexual behavior in animal species, but the analogy to human sexual orientation is unclear. We discuss the reproductive disadvantage of a homosexual orientation and present possible mechanisms that could maintain a balanced polymorphism in human populations.     

Effect of serum amyloid A on selected in vitro functions of isolated human neutrophils

Serum amyloid A (SAA) is an acute phase reactant whose levels in the blood rise as part of the body's response to stress and inflammation. Previous studies have suggested that SAA may carry an anti-inflammatory potential. We evaluated the effects of SAA on human neutrophils activated by N-formyl-methionyl-leucyl-phenylalanine (fMLP) in vitro. At concentrations higher than 10 microg/mL, SAA inhibited neutrophil myeloperoxidase (MPO) release. This effect was located in the N-terminal--that is, amino acid residues 1-14--of the SAA molecule. Directed neutrophil migration was inhibited at the same SAA concentrations. Several amino acid residues (1-14, 15-104, 83-104) contributed to this effect. Neutrophil O2- production was inhibited at low concentrations of SAA (0.1 to 1 microg/ml) and was stimulated at concentrations higher than 50 microg/mL. Neutrophil O2- production induced by phorbol myristate acetate (PMA) and O2- generated by the xanthine-xanthine oxidase reaction were not affected by SAA. These results add to previous data suggesting that SAA, at concentrations recorded in the serum during inflammation, modulates neutrophil function; thus it may play a role in the down-regulation of the inflammatory process.     

Acute inflammation, acute phase serum amyloid A and cholesterol metabolism in the mouse

We describe a patient, RM, who suddenly became amnesic for premorbid autobiographic events in the absence of any known precipitating event. Learning abilities as well as semantic knowledge were normal. Knowledge of famous facts and persons was good, although not perfect. Whether RM suffered from organic or psychogenic isolated retrograde amnesia (IRA) could not be established on the basis of available clinical and neuropsychological elements. Regardless of its aetiology, RM's case respects the boundaries between semantic and episodic memory and so gives further support to the distinction between these two memory systems.     

Catabolism of lipid-free recombinant apolipoprotein serum amyloid A by mouse macrophages in vitro results in removal of the amyloid fibril-forming amino terminus

A series of 101 consecutive patients undergoing pancreatic resection for cancer was retrospectively analyzed to define factors that may affect the immediate postoperative outcome. Overall morbidity and mortality were 28.7% and 10.9%, respectively, although these figures were greatly reduced during the last years; the complication rate dropped from 55.6% (1981-1987) to 20.0% (1993-1995) and the mortality from 16.7% to 6.7%. At univariate statistical analysis the patient characteristics (sex, age, American Society of Anesthesiologists [ASA] class, nutritional status, jaundice), tumor characteristics (site, size, TNM stage, and grading), and type of surgery were found not to affect postoperative morbidity and mortality. In contrast, a significantly lower rate of complications was observed in patients not undergoing gastric resection, in those who received 3 units or less of blood intraoperatively, and in subjects operated more recently (after 1990). At multivariate analysis the period when the operation was performed was the only independent variable that affected the immediate postoperative outcome. Among the examined factors, only the experience acquired over time regarding the intra- and perioperative treatment of these patients seems able to lower the rate of postoperative complications.     

Reduction in amyloid A amyloid formation in apolipoprotein-E-deficient mice

Apolipoproteins have been implicated in the formation of amyloid fibrils. Recent studies have demonstrated that apolipoprotein E (apoE), alone or in combination with apolipoprotein J (apoJ), and other lipoproteins appear to enhance deposition of amyloid fibrils both in systemic and cerebral amyloids, especially Alzheimer's disease (AD). ApoE enhanced the ability of the amyloid beta-protein (1-40) fragment (A beta) to form fibrils in vitro, with apoE4 promoting the greatest fibril formation. ApoE was found associated with both human and mouse amyloid A (AA) deposits. To define the role of apoE in vivo, we utilized mice lacking the apoE gene by gene targeting. We used the AA model in mice to characterize the function of the apoE protein in amyloid fibrillogenesis. ApoE-deficient mice exhibited a decrease in deposition of AA when compared with heterozygous mutant or wild-type animals. In addition, apoE-deficient mice that were injected with an adenovirus that expressed the human apoE3 gene had restored AA deposition and the apoE was associated with the AA fibrils. These results are agreement with the in vitro studies using the beta-peptide and suggest that apoE is not essential for amyloid fibrillogenesis but can promote the development of amyloid deposition.     

The diagnostic capacity of serum amyloid A protein for early recognition of kidney allograft rejection

Chitosan derivatives, sulfated N-acyl-chitosan (S-Cn-chitosan) possessing various lengths of alkyl chain, were prepared, and the properties of their aqueous solutions were examined. The 1H-NMR spectrum of D2O solutions of S-C12-chitosan showed broadening of the proton signals caused by aggregation of the alkyl chain. The solubility of a hydrophobic compounds, azobenzene, was small in the aqueous solutions of S-Cn-chitosan with shorter alkyl chains, but increased with increasing length of the chains above C10, showing that micelles had been formed. The ESR spectrum of a spin probe, TEMPO, in an S-C14-chitosan solution showed the existence of a hydrophobic region in the solution, but this region did not exist in the S-C2-chitosan solution. The rigidity of this region was examined by using a spin probe, 16-doxyl-stearic acid. From these results, it was revealed that S-Cn-chitosan with longer alkyl chains formed a novel type of micelle called a "polymer micelle," which was more stable than the ordinary micelles formed from low-molecular-weight surfactants.     

Enhanced pathologic properties of Dutch-type mutant amyloid beta-protein

Cerebrovascular amyloid beta-protein (Abeta) deposition is a pathological feature of several related disorders including Alzheimer disease and hereditary cerebral hemorrhage with amyloidosis Dutch-type (HCHWA-D). HCHWA-D is caused by a point mutation in the gene that encodes the Abeta precursor and results in a Glu --> Gln substitution at position 22 of Abeta. In comparison to Alzheimer disease, the cerebrovascular Abeta deposition in HCHWA-D is generally more severe, often resulting in intracerebral hemorrhage when patients reach 50 years of age. We recently reported that Abeta(1-42), but not the shorter Abeta(1-40) induces pathologic responses in cultured human leptomeningeal smooth muscle cells including cellular degeneration that is accompanied by a marked increase in the levels of cellular Abeta precursor and soluble Abeta peptide. In the present study, we show that the HCHWA-D mutation converts the normally nonpathologic Abeta(1-40) into a highly pathologic form of the peptide for cultured human leptomeningeal smooth muscle cells. These findings suggest that these altered functional properties of HCHWA-D mutated Abeta may contribute to the early and often severe cerebrovascular pathology that is the hallmark of this disorder.     

7-OH-DPAT has d-amphetamine-like discriminative stimulus properties

A fluorometric procedure has been developed for detection and estimation of laccase activity in fungal broth cultures. Laccase solution was pretreated with catalase for 1 h at 37 degrees C and pH 5. Homovanillic acid was then added and the reaction mixture incubated for a further hour at 37 degrees C. The fluorescence was then developed by addition of 0.1 M glycine buffer at pH 10. Laccase preparations from Pyricularia oryzae, Coriolus hirsutus and Pycnoporus cinnabarinus catalysed formation of a fluorescent product of HVA but the optimum pH values of enzyme activities varied. The culture fluids of several other fungi also catalysed development of fluorescence in solutions containing HVA. p-Hydroxyphenylacetic acid was a poor substrate for all laccases in vivo except that produced by Perennipora tephropora.