Bronchoalveolar lavage fluid eosinophils are correlated to natural killer cells in eosinophilic pneumonias.

BACKGROUND: Eosinophilic lung diseases comprise a group of heterogeneous pulmonary disorders linked by increased eosinophils in bronchoalveolar lavage fluid (BALF). There is supporting evidence that natural killer (NK) cells participate in the regulation of eosinophilic inflammation. OBJECTIVE: Our aim was to investigate the relationship between eosinophils and NK cells in BALF in patients with different interstitial lung diseases (ILDs) focusing on eosinophilic pneumonias. METHODS: Of 114 patients who presented with increased BALF eosinophils (>5%), 74 patients were classified into the following groups: 27 had eosinophilic pneumonia (EP), 17 had idiopathic pulmonary fibrosis (IPF), 16 had hypersensitivity pneumonitis (HSP) and 14 had cryptogenic organizing pneumonia (COP/BOOP). Total BALF cells, cell density and cell differential counts were assessed and lymphocyte subsets CD3+, CD4+, CD8+, CD19+, CD3-CD16/56+ (NK) and CD3+CD16/56+ (NKT) were determined by flow cytometry. RESULTS: Significant differences were observed in the percentages of lymphocytes (p < 0.001) and CD3+CD16/56+ cells (p = 0.023) among patient groups. In patients with EP, the percentage of eosinophils correlated positively with the number of CD3-CD16/56+ cells (r = 0.522, p = 0.005), the percentage of CD3-CD16/56+ cells (r = 0.690, p < 0.001), and the absolute count of CD3+CD16/56+ absolute cells (r = 0.609, p = 0.001). However, in patients with IPF, HSP or COP/BOOP, no correlation between the percentage of eosinophils and CD3-CD16/56+ or CD3+CD16/56+ cells was observed. CONCLUSIONS: Eosinophil inflammation seems to develop through a different pathway in EP compared to other ILDs.

Detection of human antibodies to Crimean-Congo haemorrhagic fever virus using expressed viral nucleocapsid protein.

Diagnosis of Crimean-Congo haemorrhagic fever (CCHF) virus infections is hampered by the problems of handling this human pathogen, which requires the highest levels of biological containment. Recombinant antigens were examined for their potential as non-hazardous diagnostic reagents. The nucleocapsid (N) gene of the Greek AP92 isolate of CCHF virus was sequenced from cloned PCR products and the open reading frame was identified by homology to the N protein of a Chinese isolate of CCHF virus. The N protein was expressed to high levels in a baculovirus expression system. Three N protein-derived peptides were expressed in Escherichia coli as fusions with glutathione S-transferase and the antigenicities of these proteins and the baculovirus-expressed protein were tested by ELISA. When tested with laboratory animal sera representing all seven serogroups of nairoviruses, the only reactive sera were those raised to CCHF virus (Greek, Nigerian and Chinese isolates) and, more weakly, Hazara virus. When tested with a panel of known positive and negative human sera, the baculovirus-expressed N protein, and the peptide derived from the central region of the N protein, proved to be the best for identifying CCHF virus-specific IgG.

Expression of the nucleocapsid protein of Dugbe virus and antigenic cross-reactions with other nairoviruses.

The small (S) RNA segment of Dugbe (DUG) virus (Nairovirus, Bunyaviridae) encodes a single protein, the nucleocapsid (N) protein, of M(r) 49.4 kDa. cDNA derived from the complete coding region for the N protein was cloned into Autographa californica nuclear polyhedrosis virus (AcNPV) under control of the polyhedrin promoter and used to infect Spodoptera frugiperda insect cells. Western blotting analysis using monoclonal antibodies demonstrated the production of DUG N protein in the infected cells. Monoclonal and polyclonal antibodies to the N protein of Crimean-Congo haemorrhagic fever (CCHF) virus were found to cross-react weakly with the baculovirus expressed DUG N protein by Western blotting. When used in an enzyme linked immunoassay (ELISA), the DUG N protein reacted with polyclonal mouse immune ascitic fluids raised against either CCHF or Hazara viruses (both members of the CCHF serogroup of nairoviruses). Cross-reactions between DUG virus (Nairobi sheep disease serogroup) and members of other nairovirus serogroups were not detected.

Effect of pyridoxine deficiency on immunological phenomena.

Measurements of serine hydroxymethyltransferase (SHMT) in resting lymphocyte cultures showed that the level of activity of this enzyme was very low. Under the influence of mitogenic stimuli serine hydroxymethyl-transferase activity was induced 5-20-fold. Addition in the cultures of 4-deoxypyridoxine, a potent antagonist of vitamin B6 coenzymes, concurrently with the mitogen, inhibits the induction of serine hydroxymethyltransferase. Thus deoxypyridoxine exerts its effects not only at the level of the enzyme itself by antagonizing the coenzyme but also at the level of its biosynthesis. Synchronous addition of vitamin B6 with deoxypyridoxine effectively reverses the inhibitory effects. The T helper cells of the lymphocyte culture are very sensitive to deoxypyridoxine action in contrast to T suppressor cells. The effect of phytohaemagglutinin or concanavalin A on the production of interleukin-1b, interleukin-2 and interleukin-2 receptors is profoundly affected by deoxypyridoxine. These results give a deeper insight of the mechanisms by which pyridoxine deficiency causes significant reduction of humoral and cellular immune responses to antigenic stimuli. On the basis of the data of this report a detailed illustration of the events that follow the T cell activation is presented. From this investigation the enzyme serine hydroxymethyltransferase emerges as a key element in the processes of immune responses and cell proliferation. Based on this finding we advance the following two propositions for possible future medical application: (i) combination of deoxypyridoxine with immunosuppressive drugs in case of immunosuppressive therapy or organ transplantation. (ii) the enzyme presents an excellent target for chemotherapy and therefore development of special agents directed against its apoprotein may prove to be a very valuable medical tool.