Maximization of pulmonary hygroscopic aerosol deposition

A newly developed computer model is used to predict the aqueous salt solution concentration, breathing pattern, and inhaled droplet size distribution parameters that will maximize pulmonary deposition of hygroscopic medicinal aerosols. The parameter values providing maximum pulmonary deposition include 1) a NaCl concentration in the aerosolized solution of 0.035 g/ml or higher if the subject can tolerate it, 2) as nearly a monodispersed inhaled aerosol size distribution as possible, 3) an aerosol mass median diameter of 2-3 micron, and 4) slow (7 breaths/min) uninterrupted breathing of 1.5-2 liters of aerosol/breath. With these values, the model predicts that pulmonary deposition can be increased by greater than 100% relative to the deposition achieved in conventional inhalation therapy with isotonic saline-based medications.     

Purification and properties of thyrotrophin releasing hormone (TRH)-containing particles from the rat hypothalamus

The subcellular distribution of the TRH-like immunoreactivity in the rat hypothalamus and brain was studied. In differential centrifugation, the 900 g for 10 min supernatant (S1) of the hypothalamus or brain contained 61--79% of the total TRH. At 11,000 g for 20 min, 51--73% of the TRH in S1 was sedimented. When the hypothalamic S1 was fractioned under non-equilibrium conditions at 25 degrees C, two populations of TRH-containing particles were observed in several types of continuous linear density gradients. Metrizamide and sucrose gradients affected TRH-assay. TRH-particles were very light in Percol-gradients. Isotonic dextran 40,000-sucrose gradients gave the most reproducible results. In these gradients, the large TRH-particles (35%) equilibrated at 1.055--1.060 kg/l and the small ones (23%) at 1.041--1.047 kg/l. Working at 4 degrees C decreased the amount of large TRH-particles. The apparently larger particles contained cytoplasmic and mitochondrial enzymes and were sensitive to hypoosmotic shock like synaptosomes. Electron micrographs confirmed that these particles were synaptosomes. The true nature of the small particles remained unclear but morphologically a part of them were also synaptosomes. Treatment of the animals with reserpine (10 mg/kg i.p., 24 h), with 6-hydroxydopamine (100 microgram/rat i.c.v.) or with 5,7-dihydroxytryptamine (200 microgram/rat i.c.v.) did not affect significantly TRH-recovery or distribution in the hypothalamus.     

Syngeneic anti-idiotypic antisera to murine monoclonal antibodies to monomorphic and polymorphic determinants of HLA class I antigens

BALB/c mice were immunized with syngeneic anti-HLA class I monoclonal antibodies. The latter included the anti-HLA-A2, A28 monoclonal antibody (MoAb) CR11-351, the MoAb Q6/64 to a determinant restricted to HLA-B antigens and the MoAb CR10-215 and CR11-115 to the same (or spatially close) monomorphic determinant. Anti-idiotypic antibodies could be detected in bleedings obtained 3 days after the first booster, increased in titer in bleedings obtained after the second booster, and persisted at high levels in subsequent bleedings. The four anti-HLA class I MoAb did not differ in their ability to elicit syngeneic anti-idiotypic antibodies. Cross-blocking studies with a panel of anti-HLA class I, anti-HLA class II, and anti-human melanoma-associated antigen (MAA) MoAb showed that the anti-MoAb CR10-215 and anti-MoAb CR11-115 antisera contain only antibodies to private idiotopes, whereas the anti-HLA MoAb CR11-351 and anti-MoAb Q6/64 antisera also contain antibodies to public idiotopes. The latter are expressed by the anti-HLA class I MoAb CR11-351, Q1/28, Q6/64, and 6/31, and by the anti-HLA class II MoAb Q5/6, Q5/13, 127, and 441. Public idiotopes were not detected on the nine anti-MAA MoAb tested. Public idiotopes do not interfere with the binding of anti-HLA MoAb with the corresponding antigenic determinants. On the other hand private idiotopes are located within the antigen-combining site, because anti-idiotypic antisera specifically inhibit the binding of the corresponding immunizing anti-HLA class I MoAb to cultured human lymphoid cells in a dose-dependent manner. Analysis by isoelectric focusing of the anti-HLA class I MoAb antisera showed that the spectrotype of the anti-MoAb CR11-351 antiserum comprises four components that focus in the pH 6.9 to 6.2 range, the spectrotype of anti-MoAb Q6/64 antiserum comprises three components that focus in the pH 6.5 to 6.1 range, the spectrotype of the anti-MoAb CR10-215 antiserum comprises three components that focus in the pH 6.4 to 6.1 range, and the spectrotype of the anti-MoAb CR11-115 antiserum comprises three components that focus in the pH 6.6 to 6.4 range.     

Improved assay of coenzyme F420 analogues from methanogenic bacteria

Summary An improved method for separating analogues of coenzyme F₄₂₀ by isocratic reversed-phase high performance liquid chromatography is described. The method offers improved resolution, shorter chromatography runs and requires less complex apparatus.     

Structure of the lysosomal sphingolipid activator protein 1 by homology with influenza virus neuraminidase

The sphingolipid activator protein 1 (SAP-1) increases the rate of hydrolysis of sphingolipids in the lysosome by apparently bringing together the substrate and the corresponding hydrolytic enzyme. This implies specific recognition of both the substrate and enzyme by SAP-1. However, binding domains in SAP-1 and recognition mechanisms involved are unknown. Amino acid sequence comparison of SAP-1 with influenza virus neuraminidase (EC 3.2.1.18, FLU NA) indicates that functional amino acid residues in or near the sialic acid binding site of FLU NA are also found at equivalent positions in the first 48 N-terminal amino acids of SAP-1. This region of homology allows to propose folding of the SAP-1 polypeptide chain by comparison with known crystallographic structure of FLU NA and identify a potential domain for lysosomal enzyme recognition through sialic acid binding. There is also a region of 10 amino acid residues near the C-terminal end of SAP-1 which has a strong propensity to form an alpha-helix with amphiphilic properties of lipid-binding helices. This domain in SAP-1 is probably responsible for the lipid(substrate)-binding function of SAP-1.