Nitric oxide synthase (NOS) has received immense interest as an antimicrobial and antitumoral effector system of mononuclear phagocytes from rodents. Because there is increasing doubt that an analogous system exists in human macrophages, NOS was reexamined in these cells. Under tightly controlled conditions, with murine macrophages as positive controls, human macrophages failed to secrete nitric oxide «0.1 #Lmol/106 cells/24 h), even after activation with endotoxin, intcrferon-v, granulocyte-macrophage colony-stimulating factor, tumor necrosis factor-a, bacteria, or proliferating lymphocytes. The discrepancy between murine and human macrophages depended on neither the anatomic source (blood, peritoneum), the agent used for activation, nor the duration of activation. NOS activity was paralleled by metabolization of L-arginine to L-citrulline. Exogenous tetrahydrobiopterin, an essential cofactor of NOS not synthesized by human macrophages, did not support NOS activity in human macrophages. Also, no NOS activity was found in cellular subfractions of human macrophages. It appears that in humans, the inducible high-output NOS is not conserved as an antimicrobial system of macrophages.Nitric oxide (NO) has recently been brought into focus as an antimicrobial and antitumoral effector system ofmononuclear phagocytes with activity against fungi [1,2], bacteria [3,4], parasites [5][6][7][8], and tumor cells [9][10][11][12][13][14][15]. While there is general agreement that phagocytes from mice and rats synthesize abundant NO from L-arginine [I, 7, 16-19], demonstration of high-output NO synthase (NOS) activity in human mononuclear phagocytes remains controversial. On one hand, it has been proposed that the antimicrobial activity of human blood-derived macrophages, seen after prolonged activation against Mycobacterium aviuni-Mycobacteriurn intracellulare [3] or Trypanosoma cruzii [20], depends on NO. On the other hand, human peritoneal, alveolar, and bloodderived macrophages have not been found to secrete substantial amounts of NO [21][22][23][24], even after treatment with endotoxin (lipopolysaccharide, LPS) and interferon-v (IFN-'Y). Also nitrite, a descendant of NO that is unstable under physiologic conditions, was not detectable in supernatants from human macrophages stimulated with LPS and IFN-')" [23][24][25]. Furthermore, alveolar and peritoneal macrophages have not been found to metabolize appreciable amounts of L-arginine, the substrate of NOS [21].Because of the significance attributed to the effector function of NO produced by mouse and rat macrophages [26][27][28]

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Nitric oxide is a recently discovered biomolecule with a broad range of actions. The present study investigated the regulation of nitric oxide synthase activity by dexamethasone and the cofactor tetrahydrobiopterin in murine macrophages. The influence of the tetrahydrobiopterin biosynthesis inhibitors 2,4-diamino-6-hydroxypyrimidine, an inhibitor of GTP cyclohydrolase I, and phenprocoumon, an inhibitor of sepiapterin reductase, on the synthesis of nitric oxide was investigated. Dexamethasone decreased the nitric oxide production due to direct inhibition of the induction of nitric oxide synthase and of GTP cyclohydrolase. Substitution of tetrahydrobiopterin via sepiapterin could not overcome the dexamethasone-mediated inhibition. 2,4-Diamino-6-hydroxypyrimidine abolished nitric oxide synthesis and synergized with dexamethasone, completely eliminating nitric oxide production. Phenprocoumon inhibited~ production of nitric oxide via interference with later steps of tetrahydrobiopterin biosynthesis. An exogenous supply of tetrahydrobiopterin through sepiapterin led to a further increase of nitric oxide production, even in fully activated macrophages. The 'amount of nitric oxide produced by murine macrophages is therefore limited by the amount of tetrahydrobiopterin present in the cells. Inhibitors of tetrahydrobiopterin biosynthesis could provide a novel approach for therapy of pathological conditions mediated by nitric oxide, such as septic shock.Nitric oxide has recently been brought into focus as an effector molecule involved in non-oxidative killing of bacteria, protozoa, fungi and tumour cells by rodent macrophages [l, 21. Furthermore, NO is suggested to account for the action of endothelium-derived relaxing factor, a product of vascular endothelial cells [3], and as a signaling agent in neurons [4]. NO is synthesized by NO synthase through oxidation of either one of the two equivalent terminal guanidin0 nitrogen atoms from L-arginine, yielding L-citrulline and the active radical. The NO synthases characterized so far exist in two forms, (a) as a constitutive form, which is Ca2+/ calmodulin dependent, expressed in brain [5] and endothelial cells [6]. (b) The NO synthase expressed in activated macrophages [7] is inducible by cytokines and bacterial endotoxin. This form was previously thought to be Ca2+ independent, but was recently shown to contain calmodulin as a tightly . It is endogenously synthesized by the pathway shown in Fig. 1. The complex reaction mechanism of NO synthases, addressing the BH, requirement, has recently been partially elucidated Glucocorticoids have a substantial effect on the natural resistence against a broad range of bacteria and fungi [14]. Recently, it has been reported that dexamethasone reduces the activity of the inducible NO synthase in the murine macrophage-like tumor cell line, J774 [15].The present study investigated the influence of dexamethasone on L-arginine metabolism and biosynthesis of BH, in respect to NO production in murine peritoneal macrophages. The regulatory role o...

BackgroundIncidental hepatocellular carcinoma (iHCC) is a histological finding after liver transplantation (LT) which relevance has been scarcely studied.Aimsto describe the histopathological features of iHCC and to determine its prognostic impact in terms of tumor recurrence and overall survival.MethodsObservational study including 451 consecutive adult LT patients (2000–2013). Patients aged<18, retransplanted or with early postoperative death were excluded. Median follow-up after LT was 58 months. Multiple Cox’s regression was used to assess the prognostic impact of iHCC on tumor recurrence and mortality while controlling for potential confounders.Results141 patients had known HCC before LT (31.3%). Among the remaining 310 patients, the prevalence of iHCC was 8.7% (n = 27). In the explanted liver, 36.2% of patients with known HCC and 25.9% of patients with iHCC trespassed Milan criteria (p = 0.30). Patients with known and iHCC had similar rates of multinodular disease (50.4% vs 55.6%; p = 0.62), macrovascular invasion (6.5% vs 3.7%; p = 0.58), microvascular invasion (12.9% vs 14.8%; p = 0.76) and moderate-poor tumor differentiation (53.9% vs 70.4%; p = 0.09). In the multivariate analysis, iHCC and known HCC had identical recurrence-free survival after controlling for histological features (RR = 1.06, 95%CI 0.36–3.14; p = 0.90). Cumulative 5-year overall survival rates were similar between patients with known and iHCC (65% vs 52.8% respectively; log rank p = 0.44), but significantly inferior as compared with patients without HCC (77.8%) (p = 0.002 and p = 0.007 respectively). Indeed, in the overall cohort, iHCC was an independent predictor of mortality (RR = 3.02; 95%CI 1.62–5.65; p = 0.001).ConclusionThe risk of tumor recurrence after LT is similar in patients with iHCC and known HCC. A close imaging surveillance is strongly recommended for patients awaiting LT in order to detect HCC prior to LT, thus allowing for an adequate selection of candidates, prioritization and indication of bridging therapies.

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Treatment results in our study population were better than those observed in the general population. The long-term response achieved, which was maintained after transplantation, supports the use of IFN for HCV hepatitis in kidney transplant candidates under hemodialysis.

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In the present study we analyzed physiological changes in the relative distribution of FSH isoforms circulating under baseline conditions throughout the ovarian cycle as well as the forms discharged by GnRH stimulation from putative acutely releasable and reserve pituitary pools. Eight normally menstruating women underwent blood sampling on three occasions, once each during the presumptive early or midfollicular phase (FP), late follicular phase to midcycle (preovulatory phase; PO), and mid- to late luteal phase (LP) of the menstrual cycle. Blood samples were withdrawn at 10-min intervals for a total of 10 h before and after the iv administration of 10 and 90 micrograms GnRH. GnRH-stimulated FSH pulses were analyzed for secretory burst mass, secretory burst amplitude, integrated FSH concentrations, and endogenous FSH half-life by deconvolution. Serum FSH isoforms were separated by preparative chromatofocusing in 30 x 1-cm columns and identified by RIA of eluent fractions. The changes observed in serum FSH isoform distribution were then correlated with the corresponding secretory and clearance estimates of the released FSH molecules. In each phase of the menstrual cycle, a significant rise in serum FSH concentrations was observed after administration of the consecutive low and high dose GnRH pulses. The magnitude of the response in terms of secretory burst mass, secretory amplitude, and area of GnRH-induced FSH peaks was significantly higher during the PO. In all cycle phases, but particularly during the FP and PO, administration of the 90-micrograms GnRH dose elicited higher (1.4- to 1.7-fold) FSH secretory responses than the lower dose. Multiple parameter deconvolution of the GnRH-induced FSH pulses revealed that FSH molecules released in response to 10 micrograms GnRH at PO exhibited significantly (P < 0.01) shorter plasma half-lives (108 +/- 11 min) than those released during the follicular and luteal phases of the same menstrual cycles (apparent plasma half-life of FSH released at FP, 222 +/- 37 and 271 +/- 47 min for 10 and 90 micrograms GnRH-induced FSH pulses, respectively; LP, 244 +/- 41 and 198 +/- 40 min; P = NS, FP vs. LP) and in response to the high dose GnRH challenge at PO (276 +/- 40 min). Under all conditions studied, serum FSH charge isoforms were distributed along a pH range of 7.0 to less than 4.0.(ABSTRACT TRUNCATED AT 400 WORDS)