T-cell receptor biases and clonal proliferations in blood and pleural effusions of patients with lung cancer.

We sought evidence that pulmonary carcinomas mediate a cellular immunologic response by analyzing T-cell antigen receptor beta-chain variable gene (TCRBV) repertoires of lymphocytes from peripheral blood (PBL) and malignant pleural effusions (PEL) of five lung cancer patients. Expression levels of 27 TCRBV were quantitated by multiprobe RNase protection assay (RPA), and clonal expansions were identified by sequence enrichment nuclease assay (SENA) and junctional region sequencing. Abnormal TCRBV expansions were identified in all subjects by RPA (mean 6.9 +/- 1.7/patient), and their number closely correlated with elapsed time since initial diagnosis (r = 0.97). SENA, performed in specimens from three patients, confirmed the presence of mono or oligoclonality in 48% of abnormal RPA expansions, and further identified T-cell clones among TCRBV with normal expression levels. The majority of clonal expansions were among PEL, and were nearly equally divided between CD4 and CD8. These data show that T-cell repertoires of lung cancer patients are characterized by marked abnormalities and frequent clonal expansions, most likely representing responses to unique, tumor-specific antigens (TSA). Moreover, this process appears exaggerated among PEL, further suggesting that malignant effusions include local proliferations of tumor reactive T cells. These findings imply the presence of lung cancer TSA capable of eliciting cellular immune responses and raise the possibility that selective immunotherapies can ultimately be developed.

T cell receptor (TCR) BV gene repertoires and clonal expansions of CD4 cells in patients with HIV infections.

Despite extensive investigation, the pathogenesis of T cell depletions that characterize AIDS has not been elucidated. To study this process further, we evaluated T cell antigen receptor beta-chain variable gene (TCRBV) repertoires in peripheral blood lymphocytes (PBL) of 23 HIV-infected patients. Expression levels of 28 TCRBV were determined by multiprobe RNase protection assay after polymerase chain reaction (PCR) amplifications. Abnormal expansions (> 2 s.d. from mean normal values) were frequent in HIV CD4, accounting for 26% of total measured TCRBV in this population. The number and magnitude of abnormalities among individuals were inversely proportional to their CD4 counts (P < 0.012 and P < 0.01, respectively). While abnormalities were not randomly distributed among TCRBV subfamilies, no particular genes were expanded or contracted among all patients. Only 14% of CD8 TCRBV were proportionally expanded (P < 0.01 compared with CD4), and there were limited concordances between paired CD8 and CD4 repertoires among individuals. CDR3 length analyses and TCRBV sequencing showed that most CD4 expansions comprised clonal or oligoclonal populations. Thus, T cell responses in HIV patients are characterized by severe TCRBV biases and clonal expansions among CD4 subsets, and these processes are exaggerated with disease progression. The heterogeneity and oligoclonality of the TCRBV expansions are consistent with responses to HIV-encoded or other conventional antigens rather than superantigenic effects. The presence of CD4 clonal proliferations in these patients may be important in the pathogenesis of HIV, and the absence or reduction of many T cell specificities due to oligoclonal expansions may increase susceptibility to opportunistic infections.

The lesions of cyclosporine-induced autoimmune disease can be equally well elicited by CD4 or CD8 effector T cells.

Lethally irradiated Lewis rats reconstituted with syngeneic bone marrow and given cyclosporine for 4 weeks develop a graft-versus-host-like disease upon withdrawal of CsA. Autoreactive T cells inducing this thymus-dependent autoimmune disease, termed CsA-AI, are demonstrable by adoptive transfer, provided regulatory cells in recipient rats are eliminated. Earlier studies have not unequivocally defined the effector T cells responsible for development of CsA-AI. Some of these studies suggest that both CD4 and CD8 T cells are required, while other studies indicate disease transfer by CD4 or CD8 T cells only. To further clarify this issue, it was necessary to study putative effector T cells in a well-defined setting. Hence, adoptive transfer studies were designed wherein the effect of the T cells of interest could be studied without being influenced by T cells of unwanted origin. Accordingly, recipient rats were thymectomized prior to irradiation, lymph node cells (LNC) from diseased donor rats were depleted of CD4 or CD8 cells before adoptive transfer, and recipients were treated in vivo with CD4- or CD8-depleting mAb. The results showed that CsA-AI developed after adoptive transfer with LNC depleted of either CD4 or CD8 cells. Analysis of PBL and of histologic specimens confirmed the absence of the depleted subset. In both instances, the typical MHC class II expression on keratinocytes and the presence of ED1+ macrophages were identical to the lesions in the primary donors, where both CD4 and CD8 T cells were present. Analysis of the T cell Receptor beta-chain variable region repertoires revealed that their expression patterns in LNC of diseased donors or recipients was comparable to that in normal thymus or LNC--hence, there was no restricted BV repertoire. Taken in toto, our observations indicate that CsA-AI involves both CD4 and CD8 T cells, and that these subsets can generate identical macroscopic and microscopic signs of disease.

T cell receptor biases and clonal proliferations among lung transplant recipients with obliterative bronchiolitis.

Obliterative bronchiolitis (OB) is the most serious late complication of lung transplantation, but the pathogenesis of this disorder has not been elucidated. We sought evidence that OB is mediated by a cellular immunologic response by characterizing T cell antigen receptor beta-chain variable gene (TCRBV) repertoires in lung allograft recipients. Expression levels of 27 TCRBV among recipients were determined by multiprobe RNase protection assay after PCR amplification. In comparison to recipients with no evidence of rejection (n = 9), the PBL TCRBV repertoires of OB subjects (n = 16) exhibited more frequent expansions (16 vs. 9% of all measured TCRBV, P < 0.02), and the magnitudes of these abnormalities were greater (8.2 +/- 0.8 vs. 4.5 +/- 0.3 SD from mean normal values, P < 0.01). TCRBV sequencing showed these expansions were composed of clonal or oligoclonal populations. Thus, T cell responses in the recipients are marked by highly selective clonal expansions, presumably driven by indirect recognition of a limited number of immunodominant alloantigens. These processes are exaggerated among allograft recipients with OB, implying that cognate immune mechanisms are important in the pathogenesis of the disorder. Furthermore, the prominence of finite, distinct TCR phenotypes raise possibilities for development of novel diagnostic modalities and targeted immunotherapies for OB and other manifestations of chronic allograft rejection.

[The value of cytologic and bacteriologic analysis of bronchoscopic specimens with direct microscopy in the diagnosis of pulmonary tuberculosis]. / Vrijednost citoloskih i bakterioloskih analiza bronhoskopskih uzoraka direktnom mikroskopijom u dijagnozi plucne tuberkuloze.

The authors review results for detection of acid-fast bacilli by the method of direct fluorescent microscopy in cytologic specimens obtained by fiberbronchoscopy and in which a granulomatous inflammation was found cytomorphologically. Direct fluorescent microscopy revealed acid-fast bacilli in 30.2% of the specimens. In 10.2% of the patients, acid-fast bacilli detected by fluorescent microscopy were the only positive bacteriological findings which confirmed tuberculosis. The authors suggest this method for all those patients in whose bronchoscopic specimens, suspected cases of tuberculosis infection could be established cytomorphologically.

[Pleural biopsy]. / Biopsija pleure.

From 1975 to December 1990, 391 needle biopsies were performed in 379 patients, aged from 19 to 82 years. The combination of cytological and histological examination of pleural biopsy specimens was done. Histologically useful specimens were obtained in 265 (67.8%) biopsies and cytologically satisfactory in 106 (27.1%). Histology revealed non-specific changes in 136 (51.3%) cases, malignant neoplasm in 61 (23-0%), granulomatous inflammation in 47 (17.8%), chronic inflammation in 20 (7.6%) and rheumatoid pleuritis in 1 (0.3%) case. Cytology showed the presence of malignant neoplastic cells in 60 (56.7%), granulomatous changes in 44 (41.5%) and changes that were cytomorphologically denoted as non-Hodgkin's lymphoma and rheumatoid pleuritis in 2 of the 106 useful biopsy specimens.

[Comparative analysis of bronchoalveolar lavage and transbronchial biopsy in pulmonary sarcoidosis]. / Komparativna analiza bronhoalveolne lavaze i transbronhalne biopsije u plucnoj sarkoidozi.

This study was carried out in 59 patients with histological evidence of pulmonary sarcoidosis. In all patients, bronchoalveolar lavage (BAL) and transbronchial biopsy (TBB) were performed simultaneously. In histologic specimens of TBB, 24/59 (41%) patients had luminal alveolitis, 36/59 (61%) mural alveolitis, 41/59 (69%) granulomas 11/59 (15%) diffuse fibrosis and 24/59 (41%) focal fibrosis. Cytological examinations of BAL revealed 22/59 (37%) patients without alveolitis, 20/59 (34%) with low intensity alveolitis and 17/59 (29%) with high intensity alveolitis. Comparative analysis of these patients showed the association of the alveolitis in BAL and the mural alveolitis or granulomas in TBB (p less than 0.01). The conclusion of this study is that cytological findings in BAL represent interstitial cells very well.

[Intrathoracic extramedullary hematopoiesis]. / Intratorakalna ekstramedularna hematopojeza.

The authors report on 29 cases of intrathoracic localization of extramedullary hematopoiesis. Out of them, 26 cases were confirmed by cytomorphologic examination of the aspirate specimens obtained by transtracheal biopsy of carina while in 3 cases the specimens for cytologic analysis were obtained by transthoracic fine needle aspiration of solitary shadows in the lung parenchyma. The cytomorphologic finding of the hematopoietic cells of the specimens obtained by transtracheal biopsy was unexpected in patients with an active pulmonary tuberculosis, primary bronchial carcinoma, broncho and pleuropneumonia, lung abscess, sarcoidosis, chronic bronchitis, rheumatoid arthritis as well as in patients with hemoptysis.

[Bronchoscopy in the post-therapeutic monitoring of patients with bronchial carcinoma]. / Bronhoskopija u postterapijskom pracenju bolesnika sa karcinomom bronha.

A study of 970 patients treated for bronchogenic carcinoma (operation, cytostatic therapy, irradiation) reveal that only 122 (12.5%) were evaluated by the bronchoscopy. Among 47 (38.5%) out of 122 patients recidivation was confirmed (cytology or histology). The efficacy of radiology and sputum cytology is poor. Only 22/47 (47%) had radiologic evidence of recidivation and 18/47 (38%) had positive sputum cytology. Radiology and sputum cytology reveal recidivation in 27/47 (57%) patients. It means that bronchoscopy is the method of choice in the posttherapeutic monitoring of patients with bronchogenic carcinoma.

Percutaneous transluminal renal angioplasty (PTRA) as a method of therapy for renovascular hypertension in children.

The results of PTRA in treatment of renovascular hypertension in four children aged 5, 7, 13 and 15 are presented. All patients suffered from severe hypertension. Tests showed all of them to have fibromuscular dysplasia stenosis of the main renal artery and significant lateralization of renin levels. One suffered from associated polycystosis of kidneys and in another the renal artery stenosis was bilateral. In all patients successful patency was achieved. In one patient, the arterial pressure after PTRA was normalized, while in the others it was considerably improved. Two patients, tested 8 and 12 months after PTRA, were lost to further follow-up. In one of the remaining two, stenosis and hypertension reappeared 5 years after PTRA. After autotransplantation the patient was normotensive. In the other, also 5 years later, recurrent hypertension appeared related to the associated polycystosis.

[Tuberculosis of the peripheral lymph nodes]. / Tuberkuloza perifernih limfnih cvorova.

An analysis of 2.414 fine needle aspiration specimens, obtained from peripheral lymph nodes, was done in 2.287 patients during the period from 1979 till the end of 1989. This investigation was performed at the Cytologic Laboratory of the Clinic for Lung Diseases "Jordanovac" in Zagreb. Totally, there were 150 (6.2%) aspirates which, cytomorphologically, appeared to have a granulomatous reaction with necrosis. Of 150 aspirates, Mycobacterium tuberculosis was found microscopically and/or by culture in 73 (3.02%) of them. The patients were less than 40 years of age in 63% of the cases. As to the localization, the highest rate of tuberculous lymph nodes (91.8%) was found in the neck and supraclavicular areas.

Qa-1 restricted recognition of foreign antigen by a gamma delta T-cell hybridoma.

Distinct T-lymphocyte subsets recognize antigens in conjunction with different classes of major histocompatibility complex (MHC) glycoproteins using the T-cell receptor (TCR), a disulphide-linked heterodimer associated with the CD3 complex on the cell surface. In general, class I and class II MHC products provide a context for the recognition of foreign antigens by CD8+ and CD4+ T cells, respectively. This recognition seems to be largely dependent on alpha beta TCR heterodimers, whereas the function of the second gamma delta TCR, present on a minor subpopulation of cells, is still unknown. In the mouse, the existence of six cell-surface MHC class I products (K, D, L, Qa-1, Qa-2 and Tla) has been firmly established by serological, biochemical and genetic evidence. So far, only the most polymorphic of them, K, D and L ('classical' class I) have been reported as restriction elements for T-cell recognition of foreign antigens. The function of the relatively invariant Qa and Tla molecules remains unknown. We have made a T-helper cell hybridoma clone (DGT3) that recognizes synthetic copolymer poly(Glu50Tyr50) in the context of Qa-1 cell surface product, and has a CD4-CD8- phenotype. Our studies indicate that DGT3 cells express the gamma delta TCR on the cell surface, implicating its role in Qa-1-restricted antigen recognition. This is the first evidence that T cells can recognize foreign antigen in association with self Qa product, confirming that Qa molecules not only topologically, but also functionally, belong to the MHC.

[Pulmonary alveolar microlithiasis]. / Plucna alveolarna mikrolitijaza.

Essential knowledge of pulmonary alveolar microlithiasis is displayed in connection with the instructive case report that points out how important is to bear in mind also such a rare pulmonary disease in a diagnostic procedure.

[Lymphocytes in sputum]. / Limfociti u sputumu.

The authors analysed 116 hospitalized patients who, in their routine cytologic examination of the sputum, had also a cytomorphologic finding of lymphocytes. The greatest majority of these patients, 63 of them or 54.3% suffered from malignant neoplasm. Out of these 63 patients, 53 of them or 45.7% suffered from primary bronchial carcinoma, whereas 10 patients or 8.6% had non-Hodgkin's lymphoma, metastatic lung cancer of extrathoracic primary localization, Hodgkin's lymphoma, while two patients were supposed to have lung neoplasm. Our study also revealed that 14 patients (out of 116 hospitalized patients) or 12.0% suffered from broncho-pleuropneumonia, 13 or 11.2% from an active pulmonary tuberculosis, 7 or 6.0% from a chronic obstructive bronchitis, 5.1% from sarcoidosis, 3.4% from post tuberculosis pulmonary changes while 2.5% of the patients were found to have a pleural empyema. One case of bronchial asthma, tuberculous pleurisy, bronchiectasis, hamartoma, hemoptysis and a pulmonary infarction were found as well. Due to their own experience the authors conclude that the lymphocytes in the sputum were found to be the most frequent in patients suffering from primary bronchial carcinoma, broncho-pleuropneumonia and pulmonary tuberculosis but that they could also be found in many others pathologic changes of pulmonary parenchyma.

[Hypernephroma cells in pleural effusions]. / Stanice hipernefroma u pleuralnom izljevu.

A rare case of finding of hypernephroma cells in pleural effusion as well as their cytomorphological picture are presented in this paper. Cytomorphologically, the hypernephroma cells in pleural effusion occurred in clusters from three, four to about twenty of cells and rarely appeared singly. The cytoplasm of the cells appeared to be abundant, not sharply bordered and varied in colour from a light-grey to a slight pink-grey while within the cytoplasm, the small pale vacuoles were visible. The nuclei of the cells were regularly round-to-oval shape with a slight anisonucleosis and contained one, rarely two, a good visible blue nucleoli. Of other cells, erythrocytes were also present together with the cells of mesothelial serosa and lymphocytes. Cytomorphologically, the hypernephroma cells in pleural effusion have the same characteristics as have the hypernephroma cells in metastases of lymph nodes in mediastinum or in metastatic nodes of the lung.

[Cytomorphologic changes in bronchial epithelial cells in a patient exposed to an aerosol of the organophosphorus insecticide, Ekalux 25 (quinalphos), 0,0-diethyl-0-quinoxaline-2-phosphorothioate]. / Citomorfoloske promjene na stanicama bronhalnog epitela u bolesnika izlozenog aerosolu organofosfornog insekticida "Ekalux 25" (kvinalfos) 0,0-dietil-0-kvinoksalin-2-il fosforotioat.

In this paper, the authors present cytomorphological changes of the bronchial epithelial cells in the patient who inhaled aerosol organophosphate insecticide "Ekalux 25" 0,0-diethyl-0-quinoxaline-2-il-phosphorothioate, while spraying crops, i.e. Colorado beetle. Our cytomorphologic examination of the sputum (obtained on the 13th and 15th day after the patient's exposure to the insecticide) revealed neutrophil granulocytes, degenerate changed macrophages, degenerate changed bronchial epithelial cells, squamous metaplasia with a poor number of lymphocytes and erythrocytes. Beside this, we also found atypical hypertropic cells of the bronchial epithelium (occurring in single forms or in groups) most of which appeared to be without cilia. Cytoplasm of those cells occurred to be not sharply bordered but poor and had a grey blue colour. Chromatin pattern of the nuclei was more coarse or "wiped", while nucleoli, as rule, appeared to be not discernible. There also occurred some nuclei in mitosis. The number of nuclei in those cells varies from one to several. Sputum specimens obtained later for cytologic analysis as well as bronchoscopic specimens, obtained on the 17th day after the patient's admission to the hospital, did not show any similar cytomorphologic changes. The authors of this paper think that it would be interesting to examine and follow-up the cytomorphologic specimens (bronchial secret, catheter bronchial aspirate smears, brushing and histopathologically) as early as the first days (if it is possible) of a patient's exposure to the organophosphate compounds.