Effects of parenteral recombinant human macrophage colony-stimulating factor on monocyte number, phenotype, and antitumor cytotoxicity in nonhuman primates

Recombinant human macrophage colony-stimulating factor (rhM-CSF) was given to cynomolgus monkeys by continuous intravenous infusion or subcutaneous injection, at a dose of 50 to 100 micrograms/kg/d in repetitive 14-day cycles. Starting within 24 to 48 hours of initiation of rhM-CSF, there was a progressive increase in the number of circulating monocytes, from a baseline of 811 +/- 253 cells/microL to a peak of 3,495 +/- 712 cells/microL on day 5 to 7. Many of these cells were large, granular, and extensively vacuolated. The expanded cell population expressed HLA-DR, LFA3, CD11b (904), and CD14 (MY4), and was 77% CD16 (FcRIII) positive by two-color cytofluorometry. In functional assays, fresh monocytes showed little cytotoxicity against cultured human melanoma cells (SKMel-1), with or without prior rhM-CSF treatment. However, after 3 days of in vitro culture in rhM-CSF, monocytes from treated animals mediated efficient antibody-dependent cytotoxicity (ADCC) against SKMel-1 using the murine monoclonal antibody 3F8 (IgG3, anti-ganglioside GD2). Under the same conditions, monocytes from control animals showed little ADCC (17% versus 82%, P less than .05). Antitumor cytotoxicity in the absence of antibody was less efficient and was not significantly different between the two groups. There was a mild decrease in platelet count during rhM-CSF treatment, without clinical symptoms. No abnormalities of serum biochemical parameters were seen. We conclude that parenteral rhM-CSF increases the number of circulating monocytes in nonhuman primates, and that these monocytes mediate increased antitumor ADCC after a brief period of in vitro differentiation. This study has implications for the design of possible future clinical trials combining antitumor monoclonal antibodies and rhM-CSF.     

Therapy of severe aplastic anemia in young adults and children with allogeneic bone marrow transplantation

During an 8-year period, 28 young adults (median age 27 years) and 30 children (median age 10 years) with severe aplastic anemia have received allogeneic bone marrow transplantation (BMT) from major histocompatibility locimatched sibling donors after preparation with cyclophosphamide and total lymphoid irradiation (TLI). All recipients were previously transfused. Comparison of post-bone marrow transplantation events in adults and children reveals equivalent median time to engraftment, median duration of hospitalization, median Karnofsky assessment of activity, and equivalent low rejection rate. Although the incidence of moderate and severe acute graft-v-host disease (GVHD) and of extensive chronic GVHD was greater in adults than in children, the projected survival at 4 years of adults (67%; 95% confidence interval [CI] 49% to 85%) and of children (73%; 95% CI 57% to 89%) was equivalent. All survivors are transfusion-free and have normal peripheral blood counts. One of 28 adults and 2 of 30 children have experienced rejection, and 1 of these patients survives after a second transplant. No malignancies have been identified following transplantation. An unexpectedly high incidence of hypothyroidism has been detected and may be attributable to preparation of recipients with TLI. Therapy of severe aplastic anemia with allogeneic BMT after preparation with cyclophosphamide and TLI offers a high rate of transfusion-free survival and a low rejection rate in previously transfused young adults and children.     

Summer-type relapsing fever (hypersensitivity pneumonitis) secondary to Cladosporium herbarum in the domestic environment

‘Summer-type relapsing fever’ is the most prevalent form of hypersensitivity pneumonitis in Japan. It is usually caused by hypersensitivity to Trichosporon cutaneum – a seasonal mould which thrives in homes with damp, decayed wood, damp mats and bedclothes. The disease has been rarely described outside Japan. We report the first documented case of summer-type hypersensitivity pneumonitis in Europe – in this case caused by hypersensitivity to the mould Cladosporium herbarum.     

Effect of Salt Form on Gelation and Drug Delivery Properties of Diclofenac-Loaded Poloxamer Gels for Delivery to Impaired Skin

Pharmaceutical Research - Treating chronic wounds is a significant clinical challenge, and a topical product would be ideal for pain management. Poloxamer 407, a thermosensitive polymer, would...     

A STUDY OF METHODS USED FOR DISINFECTION OF ANAESTHETIC EQUIPMENT BETWEEN TWO SURGICAL OPERATIONS AT A LARGE TEACHING HOSPITAL

A prospective study on the growth of bacteria on certain commonly used anaesthetic equipment was undertaken in a large teaching hospital with a view to assess the effectiveness of disinfection/sterilization procedures. Samples for microbiological assessment were drawn by the worker using standardised procedures and tested in the laboratory by a microbiologist, blinded to the type of sample. Criteria for growth positivity was taken as > 25 colony forming units. A total of 90 observations were taken. 30 each for ’before use’, ’after use’ and ’after disinfection’. Overall 54.6% of the equipment showed growth “before use” with maximum growth being seen in Suction catheters (66.6%) and Guedal airways (60.0%). On the other hand, the proportion of equipment showing growth “after use” was quite high (84.6%), with suction catheters and endotracheal tubes showing 90.0% growth each. There was significant difference as regards “before” and “after” use growth on Endotracheal tubes, Guedel airways and Face masks (p < 0.05). Analysis of growth “after” disinfection” revealed that the probability of growth remains as high as 70% in suction catheters (95% CI=54% to 86%) and 60% in laryngoscopes (95% CI=43% to 78%). The study revealed gross inadequacies in methods of disinfection being followed at present.KEY WORDS: Anaesthetic equipment, Disinfection     

Detection of Anionic Antimicrobial Peptides in Ovine Bronchoalveolar Lavage Fluid and Respiratory Epithelium

Three small antimicrobial anionic peptides (AP) were originally isolated from an ovine pulmonary surfactant. However, their presence in bronchoalveolar lavage (BAL) fluid and tissues of the respiratory tract is unknown. In this study, we made affinity-purified rabbit polyclonal and mouse monoclonal antibodies to synthetic H-DDDDDDD-OH. Antibody specificity was assessed by a competitive enzyme-linked immunosorbent assay (ELISA), and the exact epitope binding sites were determined with analog peptides synthesized on derivatized cellulose. These antibodies were used to detect AP in BAL fluid by ELISA and in respiratory tissues by Western blot analysis and immunocytochemistry. BAL fluid from 25 sheep contained 0.83 ± 0.33 mM AP (mean ± standard deviation; range, 0.10 to 1.59 mM) and was antimicrobial. The presence of AP in BAL fluid was confirmed by reverse-phase high-pressure liquid chromatography fractionation followed by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry on those fractions which were positive by competitive ELISA and demonstrated antimicrobial activity. In Western blots, polyclonal antibody PAB96-1 and monoclonal antibody 1G9-1C2 (5.0 μg/ml) detected four bands in solubilized turbinate and tracheal epithelial cells (53.7, 31.2, 28.0, and 25.7 kDa) and five bands in lung homogenates (53.5, 37.1, 31.2, 28.0, and 25.7 kDa). Only a single band was seen in solubilized liver and small-intestine homogenates, and no bands were seen in blots containing BAL fluid, albumin, or kidney or spleen homogenates. In pulmonary-tissue sections, both antibodies PAB96-1 and 1G9-1C2 identified accumulated protein in the apical cytoplasm of the bronchial and bronchiolar epithelia, in the cytoplasm of pulmonary endothelial cells, and in an occasional alveolar macrophage. As a first step in identifying a candidate AP precursor gene(s), degenerate oligonucleotides representing all possible coding combinations for H-GADDDDD-OH and H-DDDDDDD-OH were synthesized and used to probe Southern blots of sheep genomic DNA. Following low-stringency washes and a 2-day exposure, strongly hybridizing bands could be identified. One degenerate oligonucleotide, SH87, was used as a hybridization probe to screen a sheep phage genomic library. Two independent phage contained the H-GADDDDD-OH coding sequence as part of a larger predicted protein. AP may originate as part of an intracellular precursor protein, with multistep processing leading to the release of the heptapeptide into mucosal secretions. There it may interact with other innate pulmonary defenses to prevent microbial infection.     

Physical and morphological characteristics of eucaryotic ribosomes and lipopolysaccharide complexes.

Lipopolysaccharides (LPS) from Pasteurella multocida or Brucella abortus were complexed with Aspergillus fumigatus ribosomes by mixing and fixation for 3 days in 3.8% formaldehyde. To investigate the nature of their physical association, ribosomes, LPS, and ribosome-LPS complexes were (i) centrifuged in CsCl gradients to determine buoyant densities, (ii) examined by electron microscopy, and (iii) monitored by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Ribosomes were found to bind to LPS from either P. multocida or B. abortus, producing complexes with densities of 1.45 to 1.50 g/ml. The buoyant density of the fixed ribosomes was 1.54 g/ml, and the buoyant densities of the fixed P. multocida and B. abortus LPS were 1.41 and 1.35 g/ml, respectively. Electron microscopy showed that formaldehyde-fixed ribosomes were attached to the LPS. Complexing of ribosomes to LPS may be of importance as a potentiator or carrier for experimental subunit vaccines.