Jun Igari

Granulocyte colony-stimulating factor administration modulates the surface expression of effector cell molecules on human monocytes

Granulocyte colony‐stimulating factor (G‐CSF) has been shown to stimulate human neutrophil functions, both in vitro and in vivo. We examined the effects of G‐CSF administration on the surface expression of effector cell molecules on human neutrophils and monocytes. G‐CSF (50 μg/m2/d) was administered subcutaneously to five healthy volunteers once a day for 7 d. Venous blood was obtained immediately before and after the completion of G‐CSF administration and 1 week after the last G‐CSF administration. The surface expression of complement receptors (CR), Fc receptors for IgG (FcR) and cellular adhesion molecules on human neutrophils and monocytes were determined by indirect immunofluorescence using flow cytometry and monoclonal antibodies. The expression of CR1, CR3, FcRI and FcRII on neutrophils increased significantly after G‐CSF administration and then decreased after the last G‐CSF administration. The expression of human leucocyte adhesion molecule‐1 (LAM‐1) on neutrophils reflected the above expression. On the other hand, the administration of G‐CSF increased the expression of CR1, CR3, FcRI and FcRIII on monocytes. The expression of CR1, CR3 and FcRI on monocytes then decreased after the last G‐CSF administration, whereas the expression of FcRIII remained at an increased level. These findings indicate that G‐CSF administration modulates the expression of effector cell molecules on circulating monocytes as well as on neutrophils, resulting in enhanced defence against selected infections or in potentiation of the tumouricidal capacity of phagocytes in cancer patients.

Evaluation of the expression of human CAP18 gene during neutrophil maturation in the bone marrow

To understand the gene expression of CAP18 (18‐kDa cationic antibacterial protein), a member of cathelicidins, we evaluated mRNA and protein expression of CAP18 using human bone marrow cells and mature neutrophils. Northern blot analysis revealed that CAP18 mRNA was expressed more abundantly in bone marrow cells than mature neutrophils, whereas Western blot analysis indicated that CAP18 protein was more abundant in mature neutrophils than bone marrow cells. Consistent with this, in situ hybridization using bone marrow cells demonstrated that the expression of CAP18 mRNA was neutrophil lineage‐specific and was observed primarily in myelocytes (>95%) with limited expression in more immature cells (promyelocytes) and mature cells (metamyelocytes, band cells, and segmented neutrophils). Furthermore, immunohistochemical study indicated that, coincident with the increase of CAP18 mRNA levels, CAP18‐positive cells increased markedly at myelocyte stage, and the increased levels remained almost constant (>95%) in metamyelocytes, band cells, and segmented neutrophils, although the mRNA levels were remarkably reduced in these cells. Together these observations indicate that CAP18 gene transcription likely occurs lineage‐ and stage‐specifically at the myelocyte stage of neutrophil maturation in the bone marrow and results in the synthesis and cytoplasmic accumulation of CAP18, which is present in the subsequent stages of neutrophil maturation. J. Leukoc. Biol. 64:845–852; 1998.

Relative serum amyloid A (SAA) values: the influence of SAA1 genotypes and corticosteroid treatment in Japanese patients with rheumatoid arthritis

OBJECTIVES (1) To determine whether serum concentration of serum amyloid A (SAA) protein is influenced by the SAA1 allele in Japanese patients with rheumatoid arthritis (RA) as previously shown in a healthy control group; and (2) to analyse what factors, based on such an allelic bias, influence the relative SAA values of those patients. METHODS SAA and C reactive protein (CRP) concentrations together with SAA1 genotypes were determined in 316 Japanese patients with RA. The relative SAA values were evaluated as an SAA/CRP ratio. RESULTS Comparison of the three SAA1 homozygote groups showed that the SAA/CRP ratio was highest in the 1.5/1.5 group (mean 9.0, p<0.01v the other two homozygote groups) followed by the 1.3/1.3 group (mean 7.2, NS v the 1.1/1.1 group) and the 1.1/1.1 group (mean 4.0). The SAA/CRP ratio was significantly higher in patients receiving corticosteroids regardless of the presence of allele 1.5. No clear differences in the ratio between patients with or without amyloidosis were found. CONCLUSION The SAA1.5 allele and corticosteroid treatment had a positive influence on SAA concentrations in serum. These findings are important when evaluating SAA concentration in inflammatory diseases and when considering the cause or treatment of amyloidosis.

Altered surface expression of effector cell molecules on neutrophils in myelodysplastic syndromes

The surface expression of effector cell molecules on neutrophils was examined in 18 patients with myelodysplastic syndromes (MDS) and 20 healthy control subjects. The MDS patients were further classified as low clinical risk (L‐MDS, n = 7) and high clinical risk (H‐MDS, n = 11). The expression of Fc receptors for IgG (FcR), complement receptors (CR) and cellular adhesion molecules on neutrophils was determined by flow cytometry and monoclonal antibodies. The effect of granulocyte colony‐stimulating factor (G‐CSF) and tumour necrosis factor‐alpha (TNF) on L‐selectin shedding and CR up‐regulation on neutrophils was also examined. The percentage of FcRI‐positive neutrophils and CD11b/CR3 expression on neutrophils were significantly increased in the H‐MDS patients when compared to the controls. In contrast, the expression of FcRII, FcRIII, L‐selectin, LFA‐1 and CD18 on neutrophils was significantly reduced in the H‐MDS patients compared with the controls. The L‐MDS neutrophils exhibited lower expressions of CR1, L‐selectin, LFA‐1 and CD18 than those of the controls. Neutrophils from some H‐MDS patients showed impaired L‐selectin shedding and CR up‐regulation after stimulation with G‐CSF or TNF, although these were not significantly different when assessed in the whole H‐MDS group. These findings suggest that an altered surface expression of effector cell molecules and an impaired modulation of cellular adhesion molecules on neutrophils may contribute to the increased susceptibility to bacterial infections in MDS patients.

Further characterization of serum amyloid A4 as a minor acute phase reactant and a possible nutritional marker

Abstract A constitutive isotype of human serum amyloid A, serum amyloid A4 (SAA4), is distributed into plasma lipoproteins, primarily in high density lipoproteins. Its physiological function is unknown; its serum concentration has no relationship with those of other major apolipoproteins. In this study, changes in SAA4 concentrations were further characterized. Variations in healthy individuals were negligible. In subjects under-going renal allograft transplantation, SAA4 changed in parallel with acute phase SAA, although its magnitude was not larger than a three-fold increase. This confirmed that SAA4 is a minor acute phase reactant in humans. SAA4 concentrations showed a good agreement with serum pseudocholinesterase activity in healthy subjects and patients with lowered pseudocholinesterase when patients with elevated acute phase SAA were excluded. These results suggest that SAA4 can be an indicator of nutrition or of hepatic protein synthesis in the absence of inflammation.

Antimicrobial Susceptibility of Respiratory Tract Pathogens in Japan During PROTEKT Years 1–5 (1999–2004)

Susceptibility to a range of antimicrobial agents was determined among isolates of Streptococcus pneumoniae, Streptococcus pyogenes, and Haemophilus influenzae collected in 12 centers throughout Japan during years 1-5 (the respiratory seasons of 1999-2004) of the longitudinal Prospective Resistant Organism Tracking and Epidemiology for the Ketolide Telithromycin study. The most frequent source of isolates of S. pneumoniae was from patients with community-acquired pneumonia (CAP) (25.3%). Reduced susceptibility to penicillin or erythromycin resistance was common among S. pneumoniae isolates (30.9-44.5% and 77.2-81.9%, respectively). The macrolide MIC(50) for S. pneumoniae was >or=128 microg/ml (azithromycin and erythromycin) and >or=64 microg/ml (clarithromycin). The erm(B) genotype accounted for the most erythromycin-resistant isolates in each study year. H. influenzae isolates were most commonly derived from patients with CAP (26.2%). The proportion of H. influenzae isolates that were beta-lactamase positive ranged between 4.3% and 9.7%. The prevalence of beta-lactamase-negative ampicillin-resistant isolates increased from 0.4% to 2.6% between years 1 and 4 then to 19.7% in year 5. S. pyogenes isolates were highly susceptible to most antimicrobial agents except macrolides and tetracycline. Telithromycin was highly active against all three pathogens examined throughout the study.

Molecular characterization of epidemic multiresistant Staphylococcus haemolyticus isolates

Fifty-five Staphylococcus haemolyticus specimens isolated from patients and neonatal intensive care unit staff were tested for susceptibility to 12 antimicrobial agents. There were 34 multidrug-resistant isolates which were resistant to oxacillin, ampicillin, cefazolin, cefmetazole, imipenem, and gentamicin. These isolates had a higher frequency of resistance to tobramicin and ofloxacin, and relatively high MICs (2 to 4 micrograms/mL) for vancomycin, although none of the isolates were vancomycin resistant. To investigate hospital-acquired colonization and infection by multiresistant S. haemolyticus, we examined all isolates by pulsed-field gel electrophoresis (PFGE) after SmaI and SstII digestion, and detected an endemic PFGE pattern in multiresistant isolates. The results suggested that local spread of multiresistant S. haemolyticus was hospital acquired, and that the hospital staffs functioned as a reservoir.

Granulocyte colony-stimulating factor administration increases serum concentrations of soluble selectins.

In order to explore the possible role of granulocyte colony‐stimulating factor (G‐CSF) in the inflammatory process, we examined the serum concentrations of soluble selectins (sL‐selectin, sE‐selectin and sP‐selectin) following in vivo administration of G‐CSF to five healthy volunteers and 12 neutropenic patients with haematological malignancies. The serum concentrations of both sL‐selectin and sE‐selectin were slightly but significantly increased after G‐CSF administration in the healthy volunteers. The serum concentrations of all three selectins were significantly increased after G‐CSF administration in the neutropenic patients concomitant with an increase in their neutrophil counts. These findings suggest that G‐CSF may participate in the leucocyte–endothelial cell interactions in vivo.

Complement receptor type 1 (CR1) deficiency on neutrophils in myelodysplastic syndrome

Summary. We present a patient with myelodysplastic syndromes (MDS) whose neutrophils exhibited defective expression of complement receptor type 1 (CR1). A 73‐year‐old man was admitted with an evolution of MDS from RA into RAEBT according to the FAB classification of MDS. The neutrophil alkaline phosphatase (NAP) score was zero. The surface expression of membrance effector melecules on neutrophils was determined by indirect immunofluorescence using flow cytometry and monoclonal antibodies. The expression of CR1 on neutrophils as identified by staining with CD35 was defective in the patient, and the expression of other complement receptors (CR3 and CR4), Fc receptors and adhesion molecules was normal. CR1 deficiency and defective NAP score on neutrophils in the patient might account for impairment of common storage pool, presumably novel intracellular secretory vesicles.

Determination of Sensitivity and Specificity of the Micral-Test II Strip for Detection of Microalbuminuria in Diabetic and Nondiabetic Patients

Accessible online at: http://BioMedNet.com/karger Dear Sir, Microalbuminuria is generally considered an early clinical marker in patients with diabetic nephropathy. The importance of screening for microalbuminuria is now well accepted [1]. Thus, it is necessary to develop sensitive methods for detecting microalbuminuria in patients with diabetic nephropathy. A rapid test is important in all settings where there is no access to a quantitative laboratory test. The Micral-Test II strip represents a modification of the detection principle of the first Micral-Test® strip (Boehringer Mannheim, Germany), a rapid immunological test for the determination of microalbuminuria [2]. The Micral-Test II strip is a gold-labeled, optically readable immunoassay to detect albumin in urine samples [2]. Recently, the results of a multicenter evaluation (eight centers in Europe) of the MicralTest II strip show that this test permits immediate on-site and reliable semiquantitative determination of low albumin concentrations in urine samples [3]. We recently determined the levels of urinary albumin in diabetic and nondiabetic patients using the Micral-Test II strip [2] and an ordinary turbidimetric immunoassay to examine sensitivity and specificity of the test strip. Diagnosis of non-insulin-dependent diabetes mellitus was made according to the criteria of the 75-gram oral glucose tolerance test of the Japanese Diabetic Research Council [4]. Urine samples were obtained from 22 patients with non-insulin-dependent diabetes mellitus and 18 nondiabetic patients in our outpatient clinic. These nondiabetic patients had chronic glomerulonephritis or nephrosclerosis. Patients who shows more than 300 mg of urinary albumin per gram of creatinine were excluded from this study. Sensitivity, specificity, and variability of the Micral-Test II strip were examined [5] and were 88.9, 53.8, and 68.2%, respectively, in patients with diabetic nephropathy. In patients with nondiabetic nephropathy, sensitivity, specificity, and variability were 100, 92.3 and 83.3%, respectively. These levels were significantly higher in patients with nondiabetic than in patients with diabetic nephropathy. The incidence of false-positive reactions in patients with diabetic nephropathy was higher than in patients with nondiabetic nephropathy (46.1 vs. 21.4%). It is postulated that the urine samples of diabetic patients might influence the reaction of the test strip. Further examinations are needed to determine the interference of urine samples in patients with diabetic nephropathy.