IL-10 inhibits nitric oxide synthesis in murine uterus

OBJECTIVES: Recent reports point to a role for the nitric oxide/nitric oxide synthase (NO/NOS) system in implantation. It has been suggested that inducible NOS expressed at peri-implantation would lead to enhanced NO production, which could promote the attachment of the blastocyst. Short-term administration of NO donors during the pre-implantation period reduced the pregnancy rate in a dose-dependent manner. Thus, it is thought that optimal levels of NO are critical for embryo implantation, so regulation of NOS must be crucial. Taking this into consideration, interleukin-10 (IL-10), synthesized and secreted by the embryo, could be modulating NOS during implantation. In this study we have investigated the in vitro effect of IL-10 on NOS in the uterus. METHODS: To determine the effect of IL-10, slices of uterus from estrogenized mice were pre-incubated for 60 min with different concentrations of IL-10 and NOS activity was measured. RESULTS: IL-10 (50 and 100 ng/ml in vitro) diminished NOS activity. The in vivo administration of lipopolysaccharide (LPS; 8 mg/kg) significantly increased the conversion of arginine into citrulline. This effect was abolished after 60 min of preincubation with IL-10 (100 ng/ml). The stimulatory effect of LPS and estrogen on NOS activity is exerted on the Ca-independent isoform and IL-10 in vitro abolished this increase. We observed that the uterus of pregnant mice on day 5 of gestation synthesized NO. This production was significantly inhibited by preincubation with IL-10 (100 ng/ml). CONCLUSIONS: This report demonstrates that IL-10 is capable of inhibiting NO synthesis in estrogenized, LPS-treated and pregnant rat uterus.     

Fulminant Stenotrophomonas maltophilia soft tissue infection in immunocompromised patients: an outbreak transmitted via tap water

Soft tissue infection caused by Stenotrophomonas maltophilia is uncommon, but nosocomial infections had been reported. We describe herein 2 young female patients, with severe neutropenia, on broad spectrum antimicrobial agents for neutropenic fever, with Hickman-type central venous catheter, who developed mucocutaneous and soft tissue infections with rapidly progressive and devastating course. Cultures from the skin of both patients and from blood of one of them grew S. maltophilia. Both patients died and post mortem examination of the patient with S. maltophilia bacteremia revealed extensive soft tissue necrosis and a vegetation on the mitral valve that grew S. maltophilia. The infection occurred in both patients at the same time and in the same ward. Epidemiological study was done, and surveillance cultures grew the organism from the faucets from the room of 1 patient and also from some of the neighboring rooms in our ward but not from any other ward nor in the water reservoir of the building.     

Significance of skeletal uptake detected on radionuclide renal perfusion studies

Since the introduction of Tc-99m labeled radiopharmaceuticals, flow studies have become a useful and readily performed component of renal nuclear medicine examinations. In addition to detecting aberrations of renal perfusion, a number of extrarenal abnormalities may also be surreptitiously detected. Three cases of hyperperfused skeleton, an extremely unusual finding which complements two previous reports in the literature, have recently been observed. All three of the patients, as well as the two previously described, had underlying neoplastic abnormalities. The appearance of perfused skeleton should therefore be recognized as suggesting the presence of underlying neoplastic disease. Considerable interest as to the etiology and magnitude of the increased skeletal perfusion exists. Measurements of normal marrow and osseous perfusion have been reviewed in order to better appreciate the magnitude of increase required for skeletal visualization on renal flow studies. Some considerations as to the etiology of this increase are offered.     

Mycosis fungoides bullosa

A case of mycosis fungoides bullosa is presented. The results in our study confirmed that the predominant atypical lymphoid cells in the bullae, peripheral blood, and involved lymph nodes expressed the T-cell helper phenotype using immunophenotyping techniques. The literature is reviewed, confirming that our case demonstrated cells of the T-helper phenotype, not only in the skin but also in the blood and lymph node tissue. Bullous lesions in mycosis fungoides are rare.     

Mechanisms of Lysosomal Enzyme Release from Human Leukocytes II. EFFECTS OF cAMP AND cGMP, AUTONOMIC AGONISTS, AND AGENTS WHICH AFFECT MICROTUBULE FUNCTION

Selective release of inflammatory materials from leukocyte lysosomes is reduced by compounds which increase cyclic 3',5'-adenosine monophosphate (cAMP) levels in suspensions of human leukocytes and is augmented by agents which increase cyclic 3',5'-guanosine monophosphate (cGMP) levels in these cell suspensions. Lysosomal enzymes are released in the absence of phagocytosis when cytochalasin B (5 mug/ml) converts polymorphonuclear leukocytes (PMN) to secretory cells: lysosomes merge directly with the plasma membrane upon encounter of PMN with zymosan, and cells selectively extrude substantial proportions of lysosomal, but not cytoplasmic enzymes. beta-Adrenergic stimulation of human leukocytes produced a dose-related reduction in beta-glucuronidase release (blocked by 10(-6) M propranolol) whereas alpha-adrenergic stimulation (phenylephrine plus propranolol) was ineffective. In contrast, the cholinergic agonist carbamylcholine chloride enhanced enzyme secretion, an effect blocked by 10(-6) M atropine. Incubation of cells with exogenous cAMP or with agents that increase endogenous cAMP levels (prostaglandin E1, histamine, isoproterenol, and cholera enterotoxin) reduced extrusion of lysosomal enzymes; in contrast, exogenous cGMP and carbamylcholine chloride (which increases endogenous cGMP levels), increased beta-glucuronidase release. Whereas colchicine (5 x 10(-4) M), a drug which impairs microtubule integrity, reduced selective enzyme release, deuterium oxide, which favors microtubule assembly, enhanced selective release of lyosomal enzymes. The data suggest that granule movement and acid hydrolase release from leukocyte lysosomes requires intact microtubules and may be modulated by adrenergic and cholinergic agents which appear to provoke changes in concentrations of cyclic nucleotides.     

The release of inflammatory mediators from neutrophils

Summary Human polymorphonuclear leukocytes exposed to phagocytic stimuli undergo a series of cellular reactions designed to eliminate foreign invaders such as bacteria, viruses, or fungi. The responses of human granulocytes to the stimuli of phagocytosis are initiated at the cell surface and proceed in the following sequence: a. ligand binding to surface receptors; b. membrane hyperpolarization; c. O ₂ ⁻ generation by a surface oxidase and¹O₂ metabolism; d. generation of thromboxanes and prostaglandins; e. lysosomal enzyme secretion. These events, known as ‘stimulus-secretion coupling’, result in the secretion of toxic substances which, though intended to end up in the phagocytic vacuole, may escape to the extracellular space where they initiate inflammation.     

Amplification of the pulmonary vasodilatory response to inhaled iloprost by subthreshold phosphodiesterase types 3 and 4 inhibition in severe pulmonary hypertension

OBJECTIVE: Aerosolized iloprost causes specific pulmonary vasodilation for about 60 mins in patients with severe primary and secondary pulmonary hypertension. Repeated daily inhalations are currently in use for chronic treatment. The aim of the current study was to evaluate if phosphodiesterase type 3 and 4 inhibition might amplify the prostanoid effect on pulmonary vasodilatation by stabilization of intracellular second messenger cyclic adenosine monophosphate. DESIGN: Uncontrolled clinical trial. SETTING: Medical intensive care unit, Department of Internal Medicine, University Hospital, Giessen, Germany. PATIENTS: A total of 11 patients with precapillary pulmonary hypertension (eight with primary pulmonary hypertension; one with pulmonary hypertension associated with calcinosis, Raynaud phenomenon, esophageal dysfunction, sclerodactyly, and telangiectasia [CREST syndrome]; one with pulmonary hypertension associated with systemic lupus erythematodes, and one with chronic pulmonary embolism) were included. All were classified as New York Heart Association class III or class IV. INTERVENTIONS: During right heart catheterization, a single inhalation with iloprost (1.4 microg per inhalation) was performed, and hemodynamics and gas exchange variables were recorded for the next 2.5 hrs. After the iloprost effects had completely leveled off, the dual-selective phosphodiesterase types 3 and 4 inhibitor tolafentrine was infused in seven patients and aerosolized in five patients at doses that were per se ineffective (80 mg per 2.5 hrs intravenously; approximately 0.8 mg deposited by aerosol), followed by a second iloprost inhalation procedure. MEASUREMENTS: Decrease in pulmonary vascular resistance, duration of drug effect, safety, and tolerability of combined pharmacologic intervention. RESULTS: The decrease in pulmonary vascular resistance to sole iloprost nebulization lasted for approximately 60 mins. This response was enhanced and prolonged to approximately 120 mins in the presence of both infused and aerosolized tolafentrine, without loss of pulmonary selectivity. No adverse events were observed. CONCLUSIONS: These data support the principle that subthreshold selective phosphodiesterase types 3 and 4 inhibition amplifies the lung vasodilatory response to inhaled iloprost, with minute doses being sufficient via the inhalative route.     

Prion strain discrimination in cell culture: the cell panel assay

Prions are thought to consist mainly or entirely of misfolded PrP, a constitutively expressed host protein. Prions associated with the same PrP sequence may occur in the form of different strains; the strain phenotype is believed to be encoded by the conformation of the PrP. Some cell lines can be persistently infected by prions and, interestingly, show preference for certain strains. We report that a cloned murine neuroblastoma cell population, N2a-PK1, is highly heterogeneous in regard to its susceptibility to RML and 22L prions. Remarkably, sibling subclones may show very different relative susceptibilities to the two strains, indicating that the responses can vary independently. We have assembled four cell lines, N2a-PK1, N2a-R33, LD9 and CAD5, which show widely different responses to prion strains RML, 22L, 301C, and Me7, into a panel that allows their discrimination in vitro within 2 weeks, using the standard scrapie cell assay (SSCA).     

Mitochondrial complex II participates in normoxic and hypoxic regulation of α-keto acids in the murine heart

α-Keto acids (α-KAs) are not just metabolic intermediates but are also powerful modulators of different cellular pathways. Here, we tested the hypothesis that α-KA concentrations are regulated by complex II (succinate dehydrogenase=SDH), which represents an intersection between the mitochondrial respiratory chain for which an important function in cardiopulmonary oxygen sensing has been demonstrated, and the Krebs cycle, a central element of α-KA metabolism. SDH subunit D heterozygous (SDHD(+/-)) and wild-type (WT) mice were housed at normoxia or hypoxia (10% O(2)) for 4 days or 3 weeks, and right ventricular pressure, right ventricle/(left ventricle+septum) ratio, cardiomyocyte ultrastructure, pulmonary vascular remodelling, ventricular complex II subunit expression, SDH activity and α-KA concentrations were analysed. In both strains, hypoxia induced increases in right ventricular pressure and enhanced muscularization of distal pulmonary arteries. Right ventricular hypertrophy was less severe in SDHD(+/-) mice although the cardiomyocyte ultrastructure and mitochondrial morphometric parameters were unchanged. Protein amounts of SDHA, SDHB and SDHC, and SDH activity were distinctly reduced in SDHD(+/-) mice. In normoxic SDHD(+/-) mice, α-ketoisocaproate concentration was lowered to 50% as compared to WT animals. Right/left ventricular concentration differences and the hypoxia-induced decline in individual α-KAs were less pronounced in SDHD(+/-) animals indicating that mitochondrial complex II participates in the adjustment of cardiac α-KA concentrations both under normoxic and hypoxic conditions. These characteristics are not related to the hemodynamic consequences of hypoxia-induced pulmonary vascular remodelling, since its extent and right ventricular pressure were not affected in SDHD(+/-) mice albeit right ventricular hypertrophy was attenuated.