Purification and characterization of a human bradykinin binding protein from inflammatory cells.

Bradykinin (BK) is a potent mediator with a broad spectrum of pharmacological and inflammatory actions which are exerted through cell surface receptors. We report here the affinity chromatographic purification of a novel 14 kDa BK binding protein from human blood neutrophils and also peripheral blood mononuclear cells (PBMC), 80% of which are lymphocytes. Radioreceptor crosslinking experiments using bifunctional crosslinkers and radiolabelled BK identified a 14 kDa protein in these cell types both on the cell surface, in glycerol purified plasma membranes and in detergent solubilized cell extracts. Purification by BK affinity chromatography from a variety of BK responsive human cell types i.e. CCD-16Lu lung fibroblasts, HL60 promyelocytes, U937 myelomonocytes and Jurkat T lymphocytes also demonstrated a 14 kDa protein. Purified material obtained from three different BK affinity columns all demonstrated three major proteins at 190, 50 and 14 kDa when eluted with either excess BK or mild acid. Neutrophil fractions from detergent solubilized cell extracts contained an additional 150 kDa protein when eluted with mild acid. Neutrophil and PBMC crude plasma membrane BK affinity column purifications yielded only a single 14 kDa protein. Radioreceptor dot assays of the purified neutrophil eluates containing the 14 kDa protein revealed specific binding to [125I]-BK with a 160 fold excess signal ratio over the original membrane extract. Our data indicates that we have successfully isolated a 14 kDa novel human BK specific binding protein expressed on the surface of inflammatory cells.

[Computerizing the single-breath nitrogen washout test]. / Informatisation du test de rinçage d'azote en respiration unique.

The classical assembly used for the single breath nitrogen washout implies the manipulation of valves by the operator, a perfect coordination of respiratory maneuvers by the subject and a rigid standardization of the interpretation of the tracings by the technicians doing the calculations. We present a computerized analysis of the single breath nitrogen test, based on a microcomputer (Apple II 48K) solving most of the problems above. The opening of the valves is operated by the computer program recorded on disk; the tidal volume and expiratory flow are displayed on the screen on line allowing the subject to comply with the requested limits; the nitrogram is displayed at the end of the test, allowing the technician to explain the errors (if any). The program then goes on computing the slope of phase III, the closing volume, total lung capacity and its subdivisions for each trial, and the average values for two or three trials accepted by the operator. The system worked satisfactorily during a pilot study in healthy adults (variability and reproducibility) and was particularly useful in a field survey of children and adolescents.

[Effects of incomplete inspiration and previous pulmonary volume history on the nitrogen test]. / Effets de l'inspiration incomplète et de l'histoire des volumes pulmonaires antérieurs sur le test d'azote.

The effect of incomplete inspiration and of preceding lung volume history on the different parameters of the expired nitrogen curve after inhaling pure oxygen were studied in 12 healthy non-smokers, aged 21 to 35. The equipment used was attached to a mini-computer enabling the regulation of the measurements and the performance of the tests with immediate calculations on the different indices derived from the curve (VC, RV, TLC, delta N2, CV-closing volume, CV/VC). With each subject three measurements were made: measurements in the normal way (N) after incomplete inspiration (I) and finally after 5 forced inspirations and expirations (previous history - P). The procedure (I) gave values of VC and TLC which were lower than those observed with the other methods and the slope of delta N2 and the CV/VC were higher. After the previous lung volume history (P) VC and TLC were significantly higher, the CV/VC ratio was lower (due to the increase of CV). The measurement technique was not influenced either within sessions or between sessions a week apart.