Scedosporium apiospermum skeletal infection in an immunocompetent child.

Abstract This is a case of Scedosporium apiospermum skeletal infection in a 10-year-old immunocompetent girl whose chief complaint was left knee swelling and pain. The child had a history of a bicycle accident two months before with a resultant deep penetrating trauma. Systematic administration of broad-spectrum antibiotics for 10 days was used, with no clinical improvement. Magnetic Resonance Imaging and arthrotomy of the affected joint revealed findings suggestive of osteomyelitis. Empirical intravenous antimicrobial therapy was instituted for a total of two months but one month after completion of antibacterial therapy the child returned to the hospital because of persistent knee swelling and pain. Following a new arthrotomy, Scedosporium apiospermum was isolated. The patient was cured with intravenous administration of voriconazole without any side effects and has no evidence of relapse after four years of follow-up.

Multiplex PCR for the discrimination of A. fumigatus, A. flavus, A. niger and A. terreus.

Aspergillus pathogens usually infect immunocompromised patients with lethal outcome. We report a multiplex PCR assay for the discrimination of the most frequent Aspergillus pathogens, A. fumigatus, A. flavus, A. niger and A. terreus, through distinct amplicons of 250 bp, 200 bp, 150 bp and 450 bp respectively, derived from the rDNA gene of A. terreus and the aspergillopepsin genes of the remaining species.

Investigation of serum BDNF levels in drug-naive patients with schizophrenia.

The role of brain-derived neurotrophic factor (BDNF) is to promote and modulate the neuronal responses across neurotransmitter systems in the brain. Therefore, abnormal BDNF signaling may be associated with the pathophysiology of schizophrenia. Decreased BDNF levels in the brain and the serum of patients with psychotic disorders have been reported. In the present study, we assessed serum BDNF levels in a group of 14 drug-naive first-episode patients with schizophrenia (FEP), compared to 15 healthy controls. The serum BDNF levels in the sample of FEP patients was significantly reduced compared to normal controls (23.92+/-5.99 ng/ml vs. 30.0+/-8.43 ng/ml, F=5.01, df=1, p=.034). Negative correlations were shown between serum BDNF levels of the patients and the PANSS Positive and Negative subscale scores. Our findings indicate that BDNF levels at the onset of schizophrenia may reflect associated pathophysiological processes as well as the severity of positive and negative psychotic symptoms.

Oral Candida isolates in patients undergoing radiotherapy for head and neck cancer: prevalence, azole susceptibility profiles and response to antifungal treatment.

Oral pseudomembranous candidiasis and mucositis were assessed in 39 patients receiving a total dose of 39-70 Gy radiotherapy for head and neck cancer. Mucositis was scored using the Radiation Therapy Oncology Group criteria, and oral candidiasis was diagnosed on the basis of clinical evaluation and quantitative laboratory findings. Radiation-induced mucositis was observed in 9/39 patients. Only 3/39 patients discontinued radiotherapy due to acute severe mucosal effects. Candidiasis (colony-forming units 35 to > or = 60/lesion) associated with mucositis was diagnosed in 30/39 patients: the most frequent aetiology of the infection was Candida albicans (n = 23), followed by Candida glabrata (n = 3), Candida krusei (n = 2), Candida tropicalis (n = 1) and Candida kefyr (n = 1). Patients with confirmed oral pseudomembranous candidiasis were treated with either fluconazole 200 mg/day or itraconazole 200 mg/day for 2 weeks. Clinical improvement and concomitant negative Candida cultures (mycologic cure) were the criteria determining a response to antifungal treatment. Etest revealed very low voriconazole MICs (0.004-0.125 microg/ml) for all isolates, and fluconazole resistance for eight C. albicans strains (MIC > 64 microg/ml) and for the C. krusei isolates (MIC > 32 microg/ml). The same strains showed itraconazole susceptibility dose dependence (MIC 0.5 microg/ml). Despite the itraconazole susceptible dose dependent MIC readings, all patients with oral pseudomembranous candidiasis caused by these strains responded to antifungal treatment with 200 mg/day itraconazole. Oral mycologic surveillance of patients undergoing radiotherapy for head and neck malignancies and susceptibility testing of isolates may be indicated in cases with mucositis-associated confirmed oral pseudomembranous candidiasis to ensure prompt administration of targeted antifungal treatment. On the basis of the low MIC values found, clinical evaluation of voriconazole is indicated for management of oral pseudomembranous candidiasis refractory to other azoles.

Activation of the Na+/H+ exchanger by cellular pH and extracellular Na+ in rat adipocytes; inhibition by isoproterenol.

The activity of the Na+/H+ exchanger was studied by measuring the effects of intracellular pH (pHi) and extracellular Na+ [(Na+)o] on pHi recovery and 22Na uptake in rat adipocytes. The resting pHi was acidified from 7.30 +/- 0.02 to 6.99 +/- 0.01 with nigericin in the absence of (Na+)o. pHi recovery induced by 30 mM NaCl was blocked by 100 microM amiloride. The reversibility of the exchanger was studied by Na+ loading, which raised the pHi from 7.30 +/- 0.02 to 7.50 +/- 0.01, and by removing (Na+)o, which decreased pHi to 6.97 +/- 0.01. Both functions of the exchanger, forward and backward, were inhibited by amiloride. The Na+/H+ exchanger was inactive at pHi higher than 7.1 and became increasingly active as pHi decreased to 6.2 (22Na+ uptake, 0.029 +/- 0.003 vs. 0.155 +/- 0.009 nmol/10(5) cells.2.5 min; P < 0.001); this 5-fold stimulation was largely abolished by amiloride (0.025 +/- 0.002; P < 0.001). Na+ influx was also increased as a function of (Na+)o, with an apparent Km of 35 mM. Respective 5- and 44-fold stimulations at 5 mM (0.135 +/- 0.007) and 140 mM (Na+)o (1.228 +/- 0.046 nmol/10(5) cells.2.5 min; P < 0.001) were inhibited by ethylisopropylamiloride. Isoproterenol (Iso; 100 nM) and agents that stimulate cAMP production, such as forskolin (10 microM) and theophyline (1 mM), inhibited the activity of amiloride-sensitive 22Na+ uptake by 85%. Iso inhibited the Na+/H+ exchanger, without affecting the Na+/K(+)-adenosine triphosphatase-dependent and the Na+/K+/Cl- cotransport mechanisms. (Bu)2cAMP (1 mM), a membrane-permeant cAMP analog, mimicked the effects of Iso on the exchanger. The inhibitory effect of Iso was blocked by propranolol, but not by metoprolol, a beta 1-antagonist. In addition, the alpha-adrenergic agonists, phenylephrine (alpha 1) and clonidine (alpha 2), and the alpha-antagonists, prazocin (alpha 1) and yohimbine (alpha 2), did not prevent Iso-induced inhibition of the exchanger. In conclusion, rat adipocytes possess a reversible Na+/H+ exchange mechanism, which is activated by low pHi and normal (Na+)o and is inhibited by Iso via a beta 2-adrenergic receptor stimulation and a cAMP-dependent mechanism.

Regulation of Na+/H+ exchange in rat adipocytes; effects of insulin.

The activity of the Na+/H+ exchanger was examined by acidifying the intracellular pH (pHi) with Na+ propionate (NaP) and monitoring the recovery in the absence of HCO3- in rat adipocytes. Acidification of pHi, monitored with 2',7'-bis-(2-carboxyethyl)-5(6)-carboxyfluorescein, decreased the resting pHi from 7.25 +/- 0.01 to 6.70 +/- 0.01. A spontaneous pHi recovery to 6.90 +/- 0.02 was inhibited by 300 microM amiloride. This effect was Na+ specific, as recovery did not occur in cells exposed to K+ propionate (KP). The addition of NaCl (30 mM) to KP induced pHi alkalinization. Acidification of pHi increased 22Na+ transport from 0.60 +/- 0.12 nmol/10(5) cells.min at resting pHi to 2.893 +/- 0.129 (P < 0.001) and 7.984 +/- 0.312 (P < 0.001) in the first and tenth minutes, respectively. Amiloride inhibited this 5- and 14-fold stimulation by 85% (P < 0.001). Insulin in the presence of 100 microM ouabain stimulated Na+ influx by more than 15% (P < 0.01). Ethylisopropylamiloride (10 microM) inhibited the effect of insulin by 85% (P < 0.001). Intracellular Na+, measured with a Na(+)-specific electrode, increased by 10-fold in acid-loaded cells compared to that in Na(+)-depleted cells (10.750 +/- 0.479 vs. 1.045 +/- 0.100 mM; P < 0.001). Amiloride decreased NaP-stimulated intracellular Na+ by 82% (P < 0.001). To our knowledge, this is the first report showing the presence of an insulin-responsive and amiloride-sensitive Na+/H+ exchanger that regulates pHi by a Na(+)-specific and pHi-dependent mechanism in rat adipocytes.

Insulin-stimulated fluid-phase pinocytosis and internalization of the insulin receptor: differences between the U-937 monocyte and rat adipocyte.

U-937 monocytes, a human cell line, respond acutely to insulin by internalization of the insulin receptor and acceleration of fluid-phase pinocytosis. In the present studies, both processes were shown to require energy and both were dependent on the number of insulin receptors. Monocytes with a reduced number of insulin receptors, ie, down-regulated by a 16-hour insulin treatment, had a markedly reduced response to insulin-stimulation of pinocytosis and a decrease in the amount of insulin receptors internalized. This latter feature resulted, however, from the reduction in the cellular content of insulin receptors. The proportion of receptor internalized during a 30-minute acute treatment with insulin (eg, 59% of the cell surface receptors) was slightly greater than the proportion internalized in control cells. Therefore, down-regulation does not selectively destroy receptors that cycle, leaving only a subpopulation of receptors anchored in the membrane. Apparently, there is only one population of insulin receptors, all of which are equally competent with respect to internalization. Although these results suggest a close relationship between pinocytosis and receptor internalization, it was possible to separate the two systems. The addition of poly-L-lysine produced a marked stimulation of fluid-phase pinocytosis in the absence of any increase in insulin receptor internalization. Thus, movement of the receptor into the internal pool requires more than an increase in the rate of pinocytosis. Rat adipocytes were also studied, and the results differed in several aspects from those of U-937 monocytes.(ABSTRACT TRUNCATED AT 250 WORDS)

Isoproterenol reduces insulin stimulation of hexose uptake by rat adipocytes via a postinsulin binding alteration.

The present studies have investigated the acute changes previously reported in insulin binding and insulin action that occur in fat cells after a 30-min treatment with isoproterenol. We find that the marked reduction in high affinity insulin binding can be explained by a drop in the pH of the incubation medium from 7.4 to 6.9, a change associated with the accelerated production of FFA. The alteration in insulin binding may explain the rightward shift in the insulin dose response for the stimulation of glucose transport. When the incubation medium is modified to prevent the pH change, isoproterenol stimulation fails to reduce insulin binding. In addition, the tyrosine kinase activity of the insulin receptor isolated from isoproterenol-treated cells is not altered, at least as measured by the phosphorylation of tyrosine residues on the artificial substrate, Glu80Tyr20. There are, however, changes produced in the insulin stimulation of 2-deoxy-D-glucose uptake. The response of the cells to a maximum effective concentration of insulin is reduced 35% by a 30-min treatment with 0.1 microM isoproterenol. This change occurs in parallel with a moderate rightward shift in the insulin dose-response curve. Thus, isoproterenol treatment rapidly alters the ability of the adipocyte hexose transport system to respond to insulin, but the responsible alteration(s) is located beyond the insulin-binding site and possibly beyond the insulin receptor itself.

Chemical composition of the extracellular slime glycolipoprotein of Pseudomonas aeruginosa and its relation to gentamicin resistance.

The slime glycolipoproteins (GLPs) extracted from Pseudomonas aeruginosa strain C2 and its laboratory-induced gentamicin-resistant variant were analysed for gross chemical composition. The GLP of the wild-type strain contained significantly greater amounts of neutral sugars, uronic acid and thiobarbituric-reactive material (p less than 0.001) than the GLP of the gentamicin-resistant variant. Also significantly higher (p less than 0.01) was the amino-sugar content of the GLP from the wild-type strain. Paper chromatographic analyses of the hydrolysates of the GLPs revealed that two neutral sugars, rhamnose and mannose, were absent from the GLP of the resistant variant. The GLP of strain C2 contained significantly less protein than the GLP of the gentamicin-resistant variant.

Alterations in the tyrosine kinase activity of the insulin receptor produced by in vitro hyperinsulinemia.

The tyrosine kinase activity of the insulin receptor was examined in fat cells made insulin resistant by hyperinsulinemia. Adipocytes previously treated for 16 h in vitro with 0.1 microM insulin were markedly insensitive to insulin as shown by the requirement of 2.3-fold higher concentration of insulin to stimulate half-maximal activation of glucose metabolism (glucose oxidation and glucose conversion to lipids). A similar rightward shift in the insulin dose-response curve was also evident for an insulin action not dependent on glucose metabolism, i.e. the inhibition of hormone-stimulated lipolysis. The almost 60% loss of insulin sensitivity occurred in conjunction with a 37% loss in insulin-binding activity, an alteration caused by a reduction in the number of insulin receptors. Studies of tyrosine kinase activity demonstrated a preferential alteration in this insulin receptor property, in addition to the receptor loss. The insulin-stimulated Vmax for the phosphorylation of the artificial substrate, Glu80-Tyr20, was significantly reduced by almost 40%, when the activity was expressed per insulin binding and measured in a receptor preparation purified by wheat germ affinity chromatography and immunoprecipitation. An elevation in basal tyrosine kinase activity was also present, which correlated with a lower apparent Km for ATP (0.025 mM) in comparison to the Km of 0.070 mM for the receptors from control cells. These findings indicate the presence of two types of alterations that involve the insulin receptor and hyperinsulinemia: one constituted by a modest reduction in receptor number and a second by modifications in the tyrosine kinase activity of the remaining receptors. Both alterations are required to explain the magnitude of the insulin resistance that develops after 16 h of insulin treatment.

Insulin receptor cycling and insulin action in the rat adipocyte.

The possibility that the insulin receptor of adipocytes undergoes cycling was examined by a method involving pronase digestion at 12 degrees C, followed by insulin binding studies to determine receptor location and quantity. In the absence of insulin treatment, the amount of internal receptors (i.e. protected from pronase) was 10% of total receptor content. Following a 30-min insulin treatment (0.1 microM) at 37 degrees C, the internal receptor content increased 2-fold (206 +/- 12% of control, 100%). This effect was rapid, and maximum internalization was approached by 5 min of insulin treatment. Warming pronase-digested cells to 37 degrees C allowed the internal receptors to move to the cell surface. This movement was rapid also, and expansion of the internal pool by insulin pretreatment provided a 2.4-fold increase in the reinsertion of cell-surface receptors (238 +/- 28% of nontreated cells, 100%). Insulin-pretreated and nontreated cells had approximately 13 and 6%, respectively, of their original cell-surface receptor content, i.e. their content before pronase digestion. These receptors appeared intact after the cycling process, as judged by affinity labeling and sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the receptor and its binding subunit. The ability of the recycled receptor to respond to insulin was examined by studies of glucose incorporation into lipids and the inhibition of isoproterenol-stimulated lipolysis. Cells pretreated with insulin and allowed to recycle (e.g. 13% of normal receptor content) were 2-3-fold more responsive and 7-fold more sensitive to subsequent insulin stimulation than nontreated cells (e.g. 6% of normal receptor content), indicating that the recycled receptors are biologically active and coupled to cellular effector systems.

Hypophyseal metastatic renal cell carcinoma and pituitary adenoma. Case report and review of the literature.

A patient with a remote history of nephrectomy for renal cell carcinoma presented with a visual field defect. At surgery, a metastasis from the hypernephroma and an adenoma were found coexisting in the anterior pituitary gland. Although hypernephromas are known to act as "recipient" tumors in cases in which two primary neoplasms coexist, it is unusual for a renal cell carcinoma to metastasize into another tumor.