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The effect of metal salts, three nickel and one non-nickel (manganese chloride), was examined on the ability to influence mitogen stimulated normal human blood lymphocytes by means of short term in vitro culture and a tritiated thymidine incorporation test. Purified lymphocytes were incubated for one hour with tissue culture medium containing either one of the nickel salts tested (nickel subsulfide, nickel sulfate or nickel acetate) or manganese chloride. Separate portions of cells were incubated in the metal salt mixtures containing both, nickel and manganese salts. All metal salts were used in predetermined subtoxic concentrations. Two mitogens, phytohemagglutinin (PHA) and concanavalin A (Con A), were used as lymphocyte stimulatory agents. Cells were cultured for 72 hrs. It was found that following incubation with nickel salts, mitogen dependent lymphocyte stimulation was inhibited proportionally to the metal salt concentration. This blocking effect on tritiated thymidine incorporation was stronger for readily soluble nickel salts i.e. sulfate and acetate than for almost insoluble nickel subsulfide with either mitogen used. Manganese chloride used as a single salt resulted in a dose-dependent increase of lymphocyte stimulation as compared to the mitogen stimulated cells without preincubation with either metal (control samples). Cells preincubated with nickel salt-manganese chloride mixtures exhibited an increase of thymidine incorporation but below values for control cells.

Malignant tumors contain numerous macrophages as a major component of the leukocytic infiltrate. Only few studies have evaluated the interaction between products secreted by macrophages and tumor cells. Our objective was to study soluble factors produced by pleural macrophages. We sampled pleural effusions from patients with cancer and used human tumor cell lines as targets. Pleural macrophages were cultured and the supernatants were used as a conditioned medium for cultures of human cell lines A549, HT29, HCT116, SW620, MCF7, MDA-MB231, JURKAT, and HL60. We investigated apoptosis, proliferative activity, and expression of apoptosis regulating proteins Fas, Bcl2, Caspase-3, and survivin of malignant cells cultured in the conditioned medium. Our findings raise the possibility that macrophages from malignant pleural effusions can act as a factor inhibiting apoptosis of malignant cells.

In tumor tissue obtained from a group of 21 patients treated surgically for laryngeal carcinoma, the expression of Fas, Fas-L, CD3, TCR zeta chain and Bcl-2 were estimated. It has been shown that tumor often express: Fas-L, Fas, Bcl-2, the molecules which may influence increased tumor survival. On the other hand lack of MHC class I antigen expression on tumor cells frequently has been observed. As it is known, such tumor cells can not be recognized by cytotoxic T lymphocytes. Frequently observed decreased expression of TCR zeta chain and high level of apoptosis among T cells present in tumor microenvironment seems to depend on direct interactions with molecules expressed by tumor. Understanding of these interactions may be of great importance in evaluation of tumor aggressiveness, patients follow up and the approach to treatment.

The aim of this study was to assess alterations of zeta chain expression and their relation to apoptosis of T lymphocytes. T lymphocytes were obtained from malignant pleural effusions (MPE) of 15 patients. The work focused on TCR-zeta chain expression, apoptosis of T lymphocytes, and on the content of monocyte/macrophages in effusions. Analysis was performed using three color flow cytometry combining CD3, TCR-zeta and TUNEL reaction. The content of tumor and monocyte/macrophage cells has been determined using CD3, CD14, and cytokeratins, as markers of distinct cell subpopulations. Our findings strongly indicate that decreased zeta chain expression in T cells depends on the content of monocyte/macrophage cells, and that the range of the decrease is inversely proportional to the number of monocytes/macrophages in the effusion. Those having low zeta chain expression were the main subpopulation of T cells undergoing apoptosis. These data suggest that decreased zeta chain expression in T cells in MPE may be related to the abundance of monocyte/macrophages in effusions.

Main components of innate and acquired immunity participating in the response to infection were discussed. The role of NF-kappa B transcription factor was stressed as a key regulatory agent governing the immune response to infection. Attention was also paid to the escape mechanisms of microbes to avoid immune reaction of the host.

Cell adhesion molecules (CAM) represent a large group of cell surface protein moieties with distinctive biological functions. In physiological terms they ascertain cell to cell contact such as cell cohesion of epithelia, condition cell migration and transmigration via biological membranes such as blood vessel walls, provide means for homing cells in a new microenvironment etc. These features of CAM are exploited by tumor cells to grow and spread in a tumor bearing host. CD56/N-CAM antigen is 140 kD isoform of neural cell adhesion molecule. N-CAM belongs to the large Ig superfamily of CAMs. CD56 can be traced at various sites, including nervous tissue, neuro-muscular junctions, neuroendocrine and endocrine organs. It is well known as a differentiation antigen of natural killer (NK) cells. Its role and function are far from clear, but its adhesion properties are evident in cell-cell (homophilic) interactions. CD56 has been, however, demonstrated the cells various human malignancies. Tumors of the nervous system such as neuroblastoma, are well known to express this marker. Malignant lymphomas of T-NK cell origin bear CD56, as well as multiple myeloma, melanoma and some cancers of epithelial origin. These data suggest that CD56/N-CAM antigen is, in some unknown manner involved in tumor biology.

Background and Aims: Renal toxicity of first generation protease inhibitors (IP) was not a safety signal in phase III clinical trials, until Mauss et al.’s report in Hepatology. Methods: We retrospectively analyzed 101 HCV patients receiving triple therapy with telaprevir (n = 36) or boceprevir (n = 26) or double therapy (n =39) and a close monitoring of eGFR (MDRD formula) during and after treatment (D0, W4, 8, 12, 16, 24, 36, 48, 60, 72). Changes in eGFR over time were assessed by a linear mixed effects model (LMEM) with search for possible explanatory covariates. Results: Patients treated with telaprevir, presented a significant decrease of eGFR with the same kinetics: initial decrease at W4, nadir at W8 (mean decrease 17.0±18.9ml/min/1.73m2) and in average normalization at W16. The W8eGFR was highly correlated with the D0eGFR (R = 0.49) and was significantly lower in patients with grade II anemia or more (p = 0.031). The LMEM showed that the slope of eGFR versus time was the same for all patients and eGFR nadir could be predicted. In multivariate analysis, eGFR during the first 8 weeks was associated with time, older age, male sex and cryoglobulinemia. Conclusions: The eGFR significantly varied in telaprevir group only. Our model showed that eGFR nadir mainly depended on initial eGFR. As telaprevir was shown to inhibit mostly the human renal drug transporter OCT2 which interacts with creatinin transport, the early decrease of eGFR observed could be a benign phenomenom. However, as true and unpredictable renal toxicity may occur at any time during therapy, we recommend a thorough follow-up of eGFR.

The authors report clinical and neuropathological findings especially immunofluorescent detection of antineuronal antibodies in the case of paraneoplastic syndrome in course of the small-cell lung carcinoma. The clinical symptoms, observed in 48-year-old woman, covered bilateral pyramidal syndrome, cerebellar syndrome, myasthenic syndrome and impairment of the cranial nerves. Neuropathological investigation revealed paraneoplastic encephalopathy in the form of encephalitis. Immunofluorescent analysis showed brightly fluorescent neurons standing out against a dull background.

In search of factors governing the accumulation of tumor infiltrating lymphocytes (TIL), frozen sections from fresh surgical specimens of laryngeal carcinoma (n = 36) were tested by alkaline phosphatase-anti-alkaline phosphatase (APAAP) immunohistochemistry for monomorphic determinants of HLA class I and class II expression on tumor cells and for the distribution of lymphoid cells bearing CD differentiation antigens. Cell subsets were quantitated in two tumor compartments, tumor mass and tumor stroma, by computer-assisted image analysis. In a portion of examined samples lymphoid cell suspension was isolated from cancerous tissues and assessed by flow cytometry. It has been found that T cells, localized mostly in tumor stroma, were predominant cell population in the tumor microenvironment. Their ability to penetrate tumor mass but not tumor stroma, by CD8+ T cells in particular, but also by natural killer (NK) cells, was associated with HLA class I antigen expression on tumor cells. In flow cytometric analysis activated T lymphocytes (CD3+DR+) were abundant in HLA+ tumors as compared to HLA- ones. In 4 year follow up of 20 patients the mortality was higher in HLA- group but the data were not statistically significant. These results show that HLA class I expression on tumor cells favor penetration of cytotoxic lymphoid cells into tumor mass, at least in the laryngeal cancer.

CD56 antigen, an isoform of the neural cell adhesion molecule (NCAM) was previously found by us in human thyroid by APAAP immunohistochemistry in light microscopy on frozen tissue sections. In the current study, it was attempted to trace the antigen in question using another light microscopic immunohistochemical procedure and to validate the results at the ultrastructural level. For light microscopy, cryostat sections of 12 surgical samples of human thyroid were subjected to ABC (preformed avidin-biotin-peroxidase complex) method. For immunoelectron microscopy, immunoperoxidase reaction was carried out on prefixed, small thyroid tissue blocks. Following preliminary inspection of semithin sections, ultrathin sections were examined in the transmission electron microscope. ABC reaction revealed distinct specific CD56 staining of thyrocyte cell membranes. The staining was weak or absent in thyroid papillary carcinoma cells. The results were confirmed in semithin sections by indirect immunoperoxidase. The latter reaction in ultrathin sections at the ultrastructural level has shown that specific reaction product was confined to free and lateral surfaces of thyroid follicular cells. Endothelial cell membranes of thyroid capillary vessels were totally devoid of the reaction product. The reaction was weakly positive in thyroid follicular and papilllary carcinomas but absent from medullary carcinoma.

In a group of 42 patients with surgically treated laryngeal carcinoma, who were divided into those before their 50 and after 60 years of age, main lymphocyte subsets of peripheral blood lymphocytes were assessed by means of flow cytometry. In the group after 60 years of age significantly increased percentage and raised total number of NK (natural killer) cells were found. In the whole group increased total number of all lymphocyte subsets was found. These data indicate that older patients despite an increased number of total to lymphocytes NK cells show higher percentage and total number as compared with normal population at that age.

CD56/N-CAM antigen, 140 kDa isoform of neural cell adhesion molecule (N-CAM) has been previously traced by some of us in follicular epithelium of human thyroid by immunohistochemistry. The reaction product was cell membrane bound, being stronger in hyperactive thyroid as compared to colloid goiter. In the current study, CD56 was searched in other endocrine glands and their tumors including parathyroids, adrenal cortex and parafollicular C cells of the thyroid (TT cell line). The antigen was also examined in the tissue extracts of endocrine and nonendocrine organs by dot blot immunoassay and anti CD56 monoclonal antibody. Besides, some other cell adhesion molecules (CAMs) were looked for in the tissues and cells tested. It has been found that CD56 is expressed in all zones of adrenal cortex, albeit in various intensity. The reaction was cell membrane bound in cortical hyperplasia and adenoma but cytoplasmic in the carcinoma of adrenal cortex. Other endocrine tissues and cells tested were devoid of CD56. Presence of CD56 antigen could be confirmed by dot blot assay with 3M KCl and NP40 extracts of both, thyroid and adrenal glands. Apart from CD56 some other CAMs could be traced in thyroid cell membranes including CD44, VLA-3 integrin and E-cadherin, what was not the case in the adrenal cortex. In parathyroids and parathyroid adenoma, diffuse immunostaining of E-cadherin and irregular, focal expression of CD44 was observed. These results show, apart from CD56, abundance of other CAMs in the thyroid gland and their relative scarcity in other endocrine tissues tested.

In a group of 60 patients with surgically treated laryngeal carcinoma blood lymphocyte subsets were assessed on the day of surgery (day 0) and six weeks thereafter by means of flow cytometry. Blood cells at day 0 were also compared to tumor infiltrating lymphocytes (TIL) isolated from surgical specimens. Significant alterations were found in postoperative period as compared to day 0 manifested by the fall of B cells, increase of activated T lymphocytes and NK cells. There were also marked changes between blood cells at day 0 and TIL, evidence in rise of B cells, of activated T lymphocytes and decline of NK cells within the latter. These data suggest that the assessment of lymphocyte subsets may be of value in immunological monitoring of laryngeal carcinoma patients.

In order to elucidate effects of nickel on human lymphocytes in vitro, peripheral blood mononuclear cells from normal donors were initially tested for viability in the presence of increasing concentrations of two selected nickel salts, sparingly-soluble nickel subsulfide (Ni3S2), and promptly-soluble nickel sulfate (NiSO4). After establishing the toxicity profile, the cells were cultured for 24 h with each compound at three nontoxic concentrations, 0.01 mM, 0.02 mM, and 0.04 mM, to determine its effect on lymphocyte immunophenotype and function. Cells were also cultured in the presence of 0.01-0.04 mM magnesium acetate, Mg(CH3COO)2, while still other cell samples were subjected to a mixture of Mg(CH3COO)2 plus either Ni3S2 or NiSO4 at equimolar concentration. Following the culture, the immunophenotype of the cells was determined by indirect immunofluorescence, using monoclonal antibodies to major differentiation antigens of peripheral blood mononuclear cells, and their natural killer activity toward K562 target cells was measured. Both nickel salts were found to exert distinct effects on lymphocyte phenotype. Exposure of cells to Ni3S2 resulted in the decline of CD4 and natural killer cell populations. NiSO4 diminished the abundance of natural killer cells and, to a limited extent, also of CD4 cells. The nickel salts tested suppressed natural cytotoxicity of peripheral blood mononuclear cells, with Ni3S2 acting more strongly than NiSO4. The addition of Mg(CH3COO)2 to a nickel salt during in vitro culture abolished the above inhibitory effects. Nickel and magnesium salts did not affect CD3, CD8, CD20, and CD11a cell populations. The results indicate that nickel salts have deleterious effects on human peripheral blood mononuclear cells in short-term in vitro culture, but the magnitude of these effects varies, depending on the cell subsets.

In order to analyse migration patterns of donor MHC class II cells out of transplanted kidney and accumulation of host cells within the graft, immunomorphological studies were performed using monoclonal antibodies in rat allogeneic kidney transplantation model. To answer the question of how many donor cells migrate out of the renal cortex MRC 0 x 3 monoclonal antibody (MoAb) against LEW MHC class II antigens was used. In the grafts explanted after 4,24 48 and 73 h, a slow reduction of donor class II cells was observed and some areas in cortex showed only very few, if any, donor cells. At the same time, starting from day 2 after transplantation accumulation of donor cells was found in perivascular spaces. Spleen sections stained at 24, 48, 72 and 96 h after transplantation revealed donor cells present in recipient's spleen. They were detected up to day 3 after surgery. Their numbers, however, decreased after day 2. After 2 and 3 days, accumulations of recipient's cells between tubules were detected. It was found that many cells in infiltrations were stained with anti-T lymphocyte MoAb. Expression of class II antigen on rat kidney cells increases significantly from the day 4 after transplantation.

In order to gain an insight into interactions between human cancer and the surrounding host tissues, surgical samples of lung carcinoma of distinct histological types were examined for the expression of extracellular matrix (ECM) proteins and very late antigen (VLA) integrins, by means of a panel of monoclonal antibodies and immunohistochemistry of frozen tissue sections. It has been found that fibronectin (FN), tenascin (TN) and to a lesser degree collagen IV were abundant in the immediate vicinity of the tumor, but only TN penetrated tumor mass. FN isoforms were scarce or undetectable within the tumor area. The walls of blood vessels in the vicinity of the tumor showed increased an expression of collagen IV and laminin. The latter was occasionally absent within the basal membrane of cancer cells. The expression of EMC proteins was inversely proportional to the intensity of mononuclear tumor infiltrating cells (TIC). VLA integrins were present on both types of the cells: TIC and tumor cells. Percentage of positive TIC varied from 20% to 70%, depending on VLA integrin tested. VLA-3 was demonstrated on most of the cells of squamous carcinoma, but was almost absent on those of anaplastic small cell carcinoma one. In metastatic lymph node, VLA-4 was strongly expressed on tumor cells comparing to lymphoid ones. These data show that VLA integrins and their EMC ligands play apparently an important, but still obscure role in the interactions between lung carcinoma and its host.

Cryosections from surgical tissue blocks of primary laryngeal carcinoma were examined by immunohistochemistry for the distribution of several extracellular matrix (ECM) proteins and of tumor infiltrating cells (TIC). For the assessment of ECM proteins 8 monoclonal antibodies (Moabs) versus fibronectin, its neoplastic isoforms, tenascin, type IV collagen, laminin were used. TIC were evaluated by means of 9 Moabs versus various lymphocyte subsets, NK cells and macrophages. Marked differences were found in overall normal ECM proteins expression between tissues free from neoplastic invasion, those of closest tumor vicinity and of tumor mass. ECM proteins were found to be the most abundant in remote, apparently normal tissue compartments. They were less expressed in direct tumor proximity and almost absent from tumor mass. On the contrary, fibronectin isoforms were absent in tissue areas free from tumor but became demonstrable in tumor mass and its close vicinity. Tumor infiltrating cells were quite abundant in direct tumor vicinity. While comparing an intensity of expression of ECM proteins and accumulation of TIC in laryngeal carcinoma, reverse correlations were noticed. Strong expression of normal ECM proteins was accompanied by relatively scarce lymphocytic infiltrates in areas distant from primary tumor, while in the tissues abundant in TIC accumulation, such as in direct tumor vicinity, only weak ECM protein expression was seen. This was not the case when fibronectin neoplastic isoforms were studied. The latter could be shown in proximity or within tumor mass exclusively.

Among host lymphoid cells engaged in anti-tumor defence, tumor infiltrating cells (TIC) are apparently the most suitable for this purpose, due to feasibility of direct contact with tumor cells. The aim of the present study was to evaluate distribution of TIC which express phenotype of cytotoxic cells, in tissues of laryngeal carcinoma. Cryostat sections of surgical tumor samples were subjected to sensitive APAAP immunohistochemistry, following reaction with one of the panel of monoclonal antibodies (MoAbs) vs. several CD antigens and assessed semiquantitatively. It has been found that the content and distribution of potentially cytotoxic cells are quite heterogenous and vary from case to case in examined cancer. CD8+ cells and those bearing NK cell phenotype were the most frequently encountered, mainly within tumor mass. The cells belonging to NK cell subsets, detected by GL183 and EB6 MoAbs could be demonstrated in tumor proximity. TCR-1+ (tau/delta) T lymphocytes were quite a few in part cases. On the other hand, only a scarce number of cells among TIC expressed interleukin-2 receptor. It is concluded that in the vicinity of laryngeal cancer there are fairly large numbers of potentially cytotoxic cells, but at low or nil state of activation.

Lymphoid cells, isolated from malignant pleural effusions and collected from patients bearing primary lung carcinoma, were examined by means of indirect immunofluorescence and a panel of monoclonal antibodies vs several CD antigens. The percentages of CD4+ T lymphocytes were found to be significantly depressed in malignant effusions as compared to inflammatory ones. In relation to histological type of cancer it was especially evident in squamous cell and anaplastic carcinomas (small and large cell), in comparison to adenocarcinomas. Expression of T cell antigen receptor tau/delta (TCR-1) on T lymphocytes, demonstrated by BB3 MoAb (vs V delta 2), was significantly higher in malignant effusions as compared with non-malignant ones. This was not the case when A13 MoAb (equivalent of TCS 1) was used (vs V delta 1). Percentage values of NK cells, monocyte/granulocyte series activated cells and B lymphocytes did not differ significantly in malignant and non-malignant effusions. It is concluded that these are T lymphocyte subpopulations which are apparently distinct in both effusion groups examined.

SUMMARY Leu-19 (CD56) MoAb is well known to recognize gp220 expressed on natural killer (NK) cells and is widely used as a NK cell marker. The expression of CD56 antigen was tested by means of sensitive alkaline phosphatase-anti-alkaline phosphatase (APAAP) immunohistochemical technique and the above mentioned MoAb as a primary antibody, on frozen sections of various fresh human tissues. Out of 11 organs examined only thyroid gland provided a distinct reaction confined to cell membranes of epithelial follicular cells. The reaction had a diffuse pattern in cases of Graves' disease and colloidal goitre while in Hashimoto's thyroiditis presented as a focal pattern. Other anti-NK cell MoAbs such as VD4 (CD16) and Leu-7 (CD57) reacted only with single cells of thyroid stroma. The results of APAAP staining were confirmed by the cytofluorimetric assessment of isolated thyroid cells. It is speculated that CD56 expression on thyroid cells may have a functional significance, perhaps related to neural–endocrine interactions.

Clinicomorphological analysis has been performed in Schoenlein-Henoch nephropathy. Various clinical symptoms are accompanied by morphological changes of variable type and severity. Electron microscopy is a major tool for evaluating these changes. It relatively frequently modifies the diagnosis made by light microscopy. It mainly concerns class I and VI changes (according to a grading system of the International Study Group of Kidney Diseases in Childhood). It was shown that late prognosis was largely determined by the type and severity of morphological changes. Varying severity of changes in individual glomeruli in the same specimen requires in each case a comparison of results obtained by electron microscopy with those obtained by light microscopy in semithin sections. In three children biopsy was repeated. Progression of morphological changes was found in one child. He developed renal failure. In one child morphological changes on first biopsy did not differ from those on second biopsy. Repeated biopsy was performed due to the presence of hypertension. In one child with persistent proteinuria repeated biopsy showed marked attenuation of morphological changes.

A minor subset of T lymphocytes expresses a CD3-associated TCR composed of gamma and delta chains. The majority of TCR gamma/delta+ cells lack surface CD4 and CD8 antigen and do not react with WT31 mAb. These negative criteria were utilized in early studies to identify TCR gamma/delta+ cells. More recently, mAb to TCR gamma/delta, selected in different laboratories, have permitted the direct identification of TCR gamma/delta+ cells and their subsets. TCR gamma/delta molecules were found to be heterogeneous in size and charge mobility. Two major forms of TCR gamma/delta could be identified that are characterized by the presence or absence of an inter-chain disulphide bond. Biochemical analysis originally suggested that a precise correlation existed between reactivity with BB3 or delta TCS1/A13 mAb and expression of a disulphide (C gamma 1-encoded) or non-disulphide linked (C gamma 2-encoded) form of TCR gamma/delta. However, more recent studies have indicated that these mAb react with the molecular product of V delta 2 or V delta 1, respectively, mAb directed to one or another form of TCR gamma/delta activate the functional program of the cell, leading to intracellular Ca++ mobilization, lymphokine production and triggering of the lytic machinery. Analysis of the target molecules for TCR gamma/delta-mediated recognition revealed that at least some TCR gamma/delta+ cells are capable of specific responses to (allo)antigen and that polymorphic determinants of class I molecules can be recognized (as shown by the specific lysis of P815 cells transfected with HLA-24 allele). Unlike TCR alpha/beta+ cells, TCR gamma/delta+ cells are homogeneously composed of cytolytic precursors, as shown by the analysis of a large panel of clones in both lectin-dependent and redirected killing assays. In spite of their LGL morphology, freshly isolated TCR gamma/delta+ cells do not lyse NK-sensitive targets but do so after exposure to rIL-2. A modest cytolytic activity, however, could be induced also in fresh cells by anti-TCR/CD3 mAb in a redirected killing assay. Analysis of the distribution of the subsets expressing different TCR gamma/delta types showed that BB3+ cells are prevalent in the peripheral blood and virtually absent in the thymus; in contrast, A13+ (delta TCS1+) cells represent the majority of TCR gamma/delta+ thymocytes. Electron microscopic analysis of fresh TCR gamma/delta+ cells showed an extended cytoplasm containing numerous electron-dense granules identifiable as primary lysosomes. Upon stimulation with IL-2, TCR gamma/delta+ cells, similar to other LAK cells, display an increase in their cytoplasmic granules together with a redistribution of cytoskeletal structures.(ABSTRACT TRUNCATED AT 400 WORDS)

The mechanism of pokeweed mitogen (PWM) dependent decreased IgG production by blood lymphocytes from lung cancer patients was studied in comparison to control patients and blood donors. It has been shown that the depletion of monocytes has some influence on IgG synthesis but is not a decisive factor. Also, quantitative alterations in the CD4 and CD8 lymphocyte subsets do not significantly influence the PWM stimulation index for IgG synthesis. The assessment of T lymphocyte suppressor activity in lung cancer patients was performed by means of a co-culture with blood mononuclear cells, while helper activity was evaluated through co-culture with donor B lymphocytes. It has been found that lung cancer patient T lymphocytes have no increased suppressor activity, however, especially in the CD4 subset, display the decrease of helper function for B lymphocytes in PWM-induced IgG synthesis. The weakened helper function of CD4 lymphocytes may explain the suppression of specific antibody synthesis do novo which is evident in patients with lung cancer.

Mononuclear host cells isolated from primary laryngeal carcinoma were assessed by means of indirect immunofluorescence with a panel of monoclonal antibodies against various lymphocyte subsets and macrophages. Tumours of various staging groups were examined in parallel with cells isolated from patient and donor peripheral blood (PBL). It was found that percentage values of cells bearing T3 and T4 phenotype were reduced both in tumour infiltrating cells (TIC) and in PBL population. The fall in T4+ cells in PBL from cancer patients in T3 and T4 staging groups was statistically significant (p less than 0.01) as compared with donor cells. Corresponding values for T8+ cells from TIC were increased in T1 and T2 staging groups of cancer, but showed a gradual fall in advanced stages. The T4+/T8+ cell ratio was decreased in both TIC and PBL cells. The HNK-1+ (NK) cell pattern in TIC was analogous to that for T8+ cells, i.e. the cell percentage decreased with advance in tumour growth. Corresponding values for OKM-1+ were increased in TIC and in patient blood, though in TIC they grew in proportion to tumour growth. Ia+ (HLA-DR+) cells in peripheral blood were significantly increased in patients versus those of donors (p less than 0.01), but only in T3 and T4 staging groups of examined cancer. These results show that subsets of tumour infiltrating cells in laryngeal carcinoma are a complex phenomenon, associated with growth and progression of tumour.

Fluorochrome-labelled Helix pomatia A haemagglutinin (HP) stained T-dependent areas in cryostat tissue sections of human lymph node, tonsil, and spleen and thymocytes within thymic medulla but not within thymic cortex. Neuraminidase treatment of the tissue section was a prerequisite for the positive cell staining. The staining was cell-membrane-associated, and the histological pattern was essentially the same as that observed in parallel sections treated with monoclonal antibodies to T cells (OKT3, Leu-1). In addition, blood vessel walls and connective tissue were also stained by HP conjugates. The latter staining was independent of neuraminidase treatment, resistant to alpha-galactosidase treatment and was not blocked by pretreatment with anti-fibronectin antibody. Body types of staining reaction were specific inasmuch as unlabelled HP and the competitive sugar hapten N-acetyl-D-galactosamine blocked the reaction, Frozen, formalin-fixed tissue sections showed the same stainability and reaction pattern as unfixed sections.

In order to gain some insight into host cell accumulations within primary tumour, frozen sections from surgical specimens of laryngeal carcinoma were subjected to indirect immunofluorescence using a panel of monoclonal antibodies against various human lymphocyte subsets as well as macrophages. In addition, polyclonal antibodies against Ig were used in order to trace B cells. Numerous host cell infiltrates seen at the tumour periphery were composed of T4 (helper) lymphocytes and macrophages. Lymphocytes of OKT8 (suppressor/cytotoxic) and Leu-7 (NK cells) series were intermingled with tumour cells in the case of scanty infiltrates. Infiltrating cells were also linked to the presence of metastases in regional lymph nodes. OKT4-positive abundant infiltrates were usually accompanied by uninvolved nodes, while scanty ones with OKT8 specificity were relatively frequently seen in the patients with evidence of nodal metastases. These differences were not statistically significant, however, B cells as well as plasma cells were infrequently observed and were encountered both in tumour samples with intensive cellular infiltrates as well as in those with scanty ones.

A panel of sera derived from 138 patients with primary bronchogenic carcinoma, non-neoplastic lung conditions and from blood donors was tested for presence of autoantibodies by indirect immunofluorescence on fixed cells of established lung cancer cell line and lung fibroblasts as a substrate. Autoantibodies were detected in 87% and 64% out of patient sera respectively and in 9% of donor sera. Immunofluorescence patterns permitted to distinguish 3 antibody specificities: anti-nucleolar, anti-cytoplasmic and anti-nuclear ones. The major differences were noted in incidence of anti-nucleolar antibodies, which were present in 77% of lung cancer patients and only in 14% of patients with non-neoplastic lung conditions. The autoantibodies in question belonged to IgG and to lesser degree IgA class of immunoglobulin and were not apparently cancer specific because absorption with normal tissue homogenates removed their activity.

Pleural effusions from 15 patients with advanced primary bronchial carcinoma, from 2 patients with metastatic lung cancer and from 6 patients with nonmalignant disease were studied. Immune complexes were found in examined fluids in amounts corresponding to 2.5-210 mg/100 ml of aggregated IgG by means of ELISA solid phase anti C3 and 125ICIq binding radioimmunoassay. Following determination of protein content and salting out by ammonium sulfate of examined fluids, the sediments were subjected to subsequent chromatographic procedure including molecular sieving (Sephadex G-200, Sepharose 4B) and affinity chromatography on Protein A-Sepharose CL-4B. The yield--apparently pure immune complexes--was then split by means of chaotropic agent 2.5 M KSCN. It permitted to obtain 2 fractions: one contained IgG while the other was a non-Ig protein of m. w. = 150 000. The latter isolated from malignant effusions possessed antigenic activity in the leukocyte migration inhibition (LMI) assay. It resulted in inhibition of migration of allogenic peripheral blood leukocytes from lung cancer patients in 87% of cases. It had no activity against leukocytes from nonmalignant disease patients. LMI activity of the final second fraction derived from malignant effusion was significantly different from that of other fractions obtained both from malignant and nonmalignant fluids.

Surface phenotypes of peripheral blood lymphocytes of lung cancer patients and those of two control groups were assessed by means of indirect immunofluorescence with monoclonal antibodies, prior and after 10 day pokeweed mitogen (PWM) in vitro stimulation. There was no significant alteration in pan T cell per cent values prior and after mitogen stimulation in all groups tested. CD4+ cells in lung cancer group were however significantly decreased as compared to blood donor group (37.3% vs 44.9%, p less than 0.05). This decline was even more pronounced in III/IVo stage of tumour progression according to TNM classification (36.8%, p less than 0.05). These changes, however were not cancer specific, because similar decrease of CD4+ cells was seen in a group of patients with nonneoplastic lung conditions (35.7%, p less than 0.01). Following 10 day PWM culture per cent values of CD4+ cells did not change significantly. The assessment of CD8+ lymphocytes has shown marked differences in two subgroups of lung cancer, namely in II (17.4%) and III/IV (26.2%) of tumour progression (p less than 0.05), which returned to normal values following PWM culture. CD4/CD8 ratio was distinctly depressed in cancer patients in relation to donors. The evaluation of surface markers of B lymphocytes activated cells and monocytes did not show significant alterations in all groups examined. Per cent of HNK1+ cells was heightened in cancer group, especially in III/IV stage of tumour progression in relation to donors (21.7% and 22.8% vs 17.3%, p less than 0.05 respectively). PWM stimulation resulted in marked fall of HNK1+ cells to values corresponding to those in donor group. This study indicates some alterations in per cent values of blood lymphocytes subpopulations belonging mainly to T cell lineage in lung cancer patients linked to tumour staging which only partially return to normal following PWM stimulation.

Most mature T lymphocytes express CD3-associated antigen receptor molecules (TCR) formed by alpha and beta chains. Recently, a minor subset has been identified that express a different CD3-associated TCR composed of gamma and delta chains. The cell subset expressing TCR gamma/delta differs from conventional T cells in a number of phenotypic and functional characteristics. The simultaneous lack of both CD4 and CD8 antigens at the cell surface allows one to greatly enrich for TCR gamma/delta + cells (by monoclonal antibodies, mAbs and complement). Cloning of CD4-8- peripheral blood lymphocytes revealed that they are homogeneously composed of cytolytic cells which, in most instances, lyse tumor target cells. TCR gamma/delta + cells proliferated in response to allogeneic cells in mixed lymphocyte culture (MLC) and MLC-derived TCR gamma/delta + cells specifically lysed PHA-induced blast cells bearing the stimulating alloantigens, thus providing the formal proof that they recognize (allo)antigens. The use of different mAbs specific for TCR gamma/delta molecules allowed us to identify two distinct subsets which bound BB3 and delta-TCS-1 mAbs, respectively. The BB3-reactive molecules in peripheral blood-derived TCR gamma/delta + cells were represented by C gamma 1-encoded disulphide-linked heterodimers, whereas delta-TCS-1 reacted with C gamma 2-encoded non disulphide-linked molecules. Both BB3 and delta-TCS-1 mAb induced activation of cloned cells expressing the corresponding antigenic determinants (as assessed by measurements of intracellular Ca++ and/or lymphokine production or cytolytic activity).(ABSTRACT TRUNCATED AT 250 WORDS)

Biliary glycoprotein, I(BGP I) is a carcinoembryonic antigen (CEA) cross-reactive glycoprotein of normal human bile. Its occurrence and localization was studied in normal human gastrointestinal tissues by means of direct immunofluorescence using immunadsorbent purified BGP I antibodies with high selectivity for BGP I, as compared to CEA and 'non-specific cross-reacting antigen' (NCA). As controls fluorescein-labelled CEA and NCA were used. Specific BGP I fluorescence was only found in the biliary tract, i.e. in bile canaliculi, in the lumen of large bile ducts and on the surface of the gall bladder mucosa. No fluorescence was found in the hepatocytes or in the cells lining larger bile ducts or the gall bladder. Fluorescence probably due to cross-reaction with NCA was obtained in the cytoplasm of macrophages in different organs and on the surface of bowel epithelium.