Types Of Immunocompetent Cells Research Articles

Role of NCF2 as a potential prognostic factor and immune infiltration indicator in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the major causes of cancer-related deaths globally. The tumor microenvironment (TME) plays a crucial role in the prognosis and treatment of HCC. Hence, it is important to exploit new biomarkers for survival surveillance and TME estimation of HCC. HCC samples data was collected from The Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) database, and clinical samples were collected from our center. The TME of HCC were explored with ESTIMATE (Estimation of STromal and Immune cells in MAlignant Tumor tissues using Expression data), ssGSEA (single sample Gene Sets Enrichment Analysis) and CIBERSORT algorithm. Differentially expressed genes were analyzed with functional enrichment analysis. Immunohistochemistry was implemented to validate the results. Based on TCGA database, we found that Neutrophil Cytosolic Factor 2 (NCF2) was significantly associated with the prognosis of HCC patients, involved in immune-related biological processes of HCC and closely associated with some types of immunocompetent cells. The survival analysis based on NCF2 expression assessed by immunohistochemistry also confirmed that NCF2-positive group had a shorter relapse free survival (RFS) and overall survival (OS) than NCF2-negative group. Multivariate Cox regression revealed NCF2 expression level and lymphovascular space invasion (LVSI) were independent risk factors for HCC patients. Receiver operating characteristic curves showed that the combination of NCF2 and LVSI had higher predictive efficacy on the 1-year RFS rate and 5-year OS rate than each of them alone. Besides, the expression level of NCF2 was positively associated with M0 and M2 macrophages infiltration. Furthermore, NCF2 expression was positively correlated with CSF1, IL4, IL10, CD206, CD163, CSF1R and TGFβ1. We proposed that higher NCF2 expression predicted an adverse prognosis and more M2 macrophages infiltration in HCC patients.

Hemocytes and fat body cells, the only professional immune cell types in Drosophila, show strikingly different responses to systemic infections.

The fruit fly Drosophila is an excellent model to study the response of different immunocompetent organs during systemic infection. In the present study, we intended to test the hypothesis that the only professional immune organs of the fly, the fat body and hemocytes, show substantial similarities in their responses to systemic infection. However, comprehensive transcriptome analysis of isolated organs revealed highly divergent transcript signatures, with the few commonly regulated genes encoding mainly classical immune effectors from the antimicrobial peptide family. The fat body and the hemocytes each have specific reactions that are not present in the other organ. Fat body-specific responses comprised those enabling an improved peptide synthesis and export. This reaction is accompanied by transcriptomic shifts enabling the use of the energy resources of the fat body more efficiently. Hemocytes, on the other hand, showed enhanced signatures related to phagocytosis. Comparing immune-induced signatures of both cell types with those of whole-body responses showed only a minimal correspondence, mostly restricted again to antimicrobial peptide genes. In summary, the two major immunocompetent cell types of Drosophila show highly specific responses to infection, which are closely linked to the primary function of the respective organ in the landscape of the systemic immune response.

РОЛЬ ЦИТОКИНОВ В РЕМОДЕЛИРОВАНИИ КОСТНОЙ ТКАНИ В НОРМЕ И ПАТОЛОГИИ

Significant successes in the study of physiological and pathophysiological patterns of bone remodeling in recent years have highlighted immune factors important role in bone tissue pathology and significantly revised our ideas about postmenopausal osteoporosis development mechanisms. Advanced osteoimmunology and evidence of immune mechanisms key role in bone remodeling disorders gave us possibility for identification of osteoporosis as chronic immune-mediated disease. Moreover, instead of the term “Osteoporosis”, the term “Immunoporosis” was reasonably used.
 Bone tissue is constantly in state of continuous renewal (remodeling), which is balanced by formation and resorption processes and is achieved through the coordinated functioning of the three main bone cells types. Constant and active interaction between osteocytes, osteoblasts and osteoclasts is ensured by cytokines (RANKL, osteoprotegerin, macrophage colony-stimulating factor, vascular endothelial growth factor, etc) secretion. Moreover, predominantly, bone remodeling regulation is limited by Osteocyte-Osteoblast-Osteoclast system.
 With pathological changes in immune reactivity, which may be caused by deficiency of estrogen, vitamin D, calcium, inflammatory diseases, etc., various types of immunocompetent cells are activated. This is accompanied with increased RANKL production by leukocyte cells, which potentiates processes of maturation, differentiation of osteoclasts, and increase in their activity. In addition to RANKL secretion, activated leukocytes, including T lymphocytes, enhance other osteoclastogenic cytokines production. IL-1, IL-6, IL-17, TNF and TGF-β are main mediators of accelerated bone loss in postmenopausal women.

Dynamics of the cellular composition of lymphoid nodules in the lungs of guinea pigs sensitized with ovalbumin

The article discusses the morphological aspects of the dynamics of the cellular composition of lymphoid nodules in the lungs of guinea pigs as a result of an experimental ovalbumin-induced allergic process. We studied the reactivity of immunocompetent cells of lymphoid formations of the lungs after three times subcutaneous sensitization and subsequent 8-day intranasal aeroallergization with ovalbumin in the early and late stage period of the allergic inflammatory process by microscopic, morphometric and statistical methods. By help of morphometric analysis we demonstrate the general regularity of reactivity of a local specific link of the pulmonary immune system to the action of an allergen, which consists in the elevation of the average amount of immune cells of lymphoid nodules of the lungs, starting from the 30th to the 44th day after the start of the experiment. The maximal coefficient of increase by 5.8 times was observed in counting plasma cells among all types of immunocompetent cells of lymphoid nodules in the lungs during the experiment. It has been statistically proven that the implementation of the ovalbumin-induced allergic inflammatory process in the lungs proceeds according to the humoral type and the duration of its course is not limited by the direct influence of the allergen, it also continues after the end of its action, which is a manifestation of changes in compensatory-adaptive processes in the pulmonary immune system with ovalbumin-induced allergic inflammation.

Dynamics of quantitative changes of diffuse lymphoid tissue cells of bronchi and lungs of guinea pigs sensitized with ovalbumin

Background. There is a progressive increase of respiratory allergic diseases nowadays in the world, made it necessary to study the participation of the components of the immune system in these processes. From the point of view of modern aspects about the organs of the immune system, interesting are the patterns of morphogenesis and function of bronchus associated lymphoid tissue, occupied a special place in the immunological protection of the body due to the large area of contact with various antigens. Morphogenesis and reactive changes in the local immunity in respiratory system in conditions of allergic inflammation remain urgent issue of modern morphology. Objective. To study the changes in diffuse lymphoid tissue of bronchi and lungs of guinea pigs sensitized with ovalbumin. Methods. We have studied the lung of 48 guinea pigs, using histological, immunohistochemical, morphometric, statistical methods, under conditions of experimental ovalbumin-induced allergic inflammation, assessed the average number of lymphocytes, macrophages and plasma cells in the diffuse lymphoid tissue. Results. The average number of lymphocytes in diffuse lymphoid tissue of bronchi and lungs increased from the 23rd day of observation and remained at a high level until the end of the experiment, the maximum was during the early period of the development of allergic inflammation, the increasing coefficient was 4.7. The average number of plasma cells also acquired maximum elevation in the early period of allergic process, the increasing coefficient was 2.0. The most significant average number of macrophages was on the 23rd day of observation with same increasing coefficient. Among all types of immunocompetent cells of diffuse lymphoid tissue in bronchi and lungs, T-lymphocytes prevailed during the experiment elevated almost by 5 times. Conclusions. In the early period of development of experimental ovalbumin-induced allergic inflammation, the specific resistance of the respiratory system manifests itself in the form of activation of local links of cellular and humoral adaptive immunity, as evidenced by the dynamics of changes in the average number of lymphocytes (the maximum increasing coefficient 4.7 in the 1st experimental group), macrophages and plasma cells (maximum increasing coefficient 2.0 in the 1st experimental group) of diffuse lymphoid tissue of bronchi and lungs of guinea pigs.

Positioning of apremilast in treatment of Behçet’s disease

Behçet’s disease is a chronic relapsing inflammatory disorder characterized by oral and genital ulcers, skin lesions, and uveitis. Although the involvement of the gastrointestinal, large vascular, and central nervous systems is less common, it can be fatal at times. To suppress inflammatory exacerbations and recurrences which can cause irreversible organ damage, treatment should be individualized according to the disease phenotype and severity. Apremilast, an oral phosphodiesterase 4 inhibitor, is used to treat psoriasis. The enzyme specific inhibitor increases intracellular cAMP levels, which modulate multiple signaling pathways in both innate and adaptive immunocompetent cells, leading to attenuation of inflammatory responses through the suppression of the functions of various types of immunocompetent cells including Th1 cells, Th17 cells and M1 macrophages. Current evidence shows that apremilast is effective for oral ulcer, the most common symptom of Behçet’s disease. An oral ulcer is considered to represent the basic immunopathology of Behçet’s disease, because the disease susceptibility genes are shared between Behçet’s disease and recurrent aphthous stomatitis. The favorable effects have been documented in other inflammatory diseases with symptoms similar to those of Behçet’s disease. The further studies would shed the light on the potential of apremilast for the treatment of Behçet’s disease.

Features of immune status in patients with various clinical forms of oral lichen planus

Oral mucosal lesions hold one of the lead places in the structure of dental diseases. Oral lichen planus (OLP) considered as a multifactorial disease is of top priority among dermatoses of oral mucosa and red lip border. Diverse putative concepts behind developing lichen planus pathogenesis are discussed including immunoallergic, viral, genetic and membrane-destructive theories. However, an immune theory is thought to play a crucial role in developing lichen planus. Despite documented induction of immune mechanisms, complex interaction between pathological process and normal defense response as well as stirred interest to it, multiple aspects of immunological conflict in lichen planus remain unclear. Few data describing altered immune parameters depending on clinical picture of lichen planus are currently available that suggested to perform the study aimed at examining immune status in patients with various forms of oral lichen planus. There were enrolled 286 oral lichen planus patients (248 females and 38 males), aged 2784 years. Based on clinical picture, all OLP patients were divided into 6 groups: a typical type, exudative-congestive, erosive-ulcerous, hyperkeratotic, atypical, bullous type. An immune status in peripheral blood samples was evaluated by analyzing innate defense mechanisms as well as humoral and cellular immunity. Multiple altered immune parameters characterized by impaired phagocytic capture and metabolic activity, disimmunoglobulinemia, altered ratio of major immunocompetent cell types and subsets were documented during the study. Moreover, OLP patients with typical, hyperkeratotic, exudative hyperemic, atypical and erosive ulcerous forms were found to have increased amount of CD4+ helper T cells associated with a self-sustained immune response due to suppressed elimination of pathogenic agents consequently resulting in developing autoimmune process. While analyzing immune status in OLP patients, it allowed to find a relationship between dysphagocytosis signs, impaired humoral and cellular immunity as well as various clinical forms of the disease. Thus, it suggests imbalanced mechanisms responsible for pathogen elimination might play a role in OLP pathogenesis, including infectious agents being involved in its development.

Peculiarities of cellular composition and microarchitecture of human hemal nodes.

Background. Cellular composition of lymph nodes is well described while in hemal nodes it is still poorly investigated. Objective. To define peculiarities of distribution of major types of immunocompetent cells and stromal elements in human hemal nodes. Methods. Hemal nodes were taken during autopsy from patients who died due to cardiovascular or respiratory pathology. Standard histological and immunohistochemical methods were used to detect different cellular types and stromal elements. Results. Distributions of lymphocytes was investigated with the help of anti-CD20, CD3 and CD8 antibodies. To analyze dendritic cells and macrophages anti S100 and CD68 antibodies were used. Stromal elements were detected with the help of anti SMA and CD34 antibodies. Conclusion. Hemal nodes have the same character of immune cells’ distribution as in lymph nodes, except B-cells, which are equally present in paracortex and medullary cords. Among well-developed intranodal vasculature there were revealed CD34-negatve vessels, which are also known in literature as system of conduits.

Quantitative Trait Locus Analysis Implicates CD4+/CD44high Memory T Cells in the Pathogenesis of Murine Autoimmune Pancreatitis

The mouse strain MRL/MpJ is prone to spontaneously develop autoimmune pancreatitis (AIP). To elucidate the genetic control towards the development of the phenotype and to characterize contributions of immunocompetent cell types, MRL/MpJ mice were interbred with three additional strains (BXD2/TYJ, NZM2410/J, CAST/EIJ) for four generations in an advanced intercross line. Cellular phenotypes were determined by flow cytometric quantification of splenic leukocytes and complemented by the histopathological evaluation of pancreatic lesions. An Illumina SNP array was used for genotyping. QTL analyses were performed with the R implementation of HAPPY. Out of 41 leukocyte subpopulations (B cells, T cells and dendritic cells), only three were significantly associated with AIP: While CD4+/CD44high memory T cells and CD4+/CD69+ T helper (Th) cells correlated positively with the disease, the cytotoxic T cell phenotype CD8+/CD44low showed a negative correlation. A QTL for AIP on chromosome 2 overlapped with QTLs for CD4+/CD44high and CD8+/CD44high memory T cells, FoxP3+/CD4+ and FoxP3+/CD8+ regulatory T cells (Tregs), and CD8+/CD69+ cytotoxic T cells. On chromosome 6, overlapping QTLs for AIP and CD4+/IL17+ Th17 cells and again FoxP3+/CD8+ Tregs were observed. In conclusion, CD4+/CD44high memory T cells are the only leukocyte subtype that could be linked to AIP both by correlation studies and from observed overlapping QTL. The potential role of this cell type in the pathogenesis of AIP warrants further investigations.

The effect of in vivo exposure to zearalenone on cytokine secretion by Th1 and Th2 lymphocytes in porcine Peyer's patches after in vitro stimulation with LPS.

Most research studies investigating the estrogenic effects of zearalenone (ZEN) focus on the mycotoxin's effect on the reproductive system. Since estrogen receptors are present on various types of immunocompetent cells, ZEN can also modify diverse immune functions. This study analyzed immunocompetent cells isolated from Peyer's patches in the ileum of pigs administered ZEN in the estimated daily dose of 8 μg kg(-1) BW (equivalent of 100 μg kg(-1) feed per day(-1)). The objective of the study was to determine whether long-term exposure to low ZEN doses below the NOEL threshold leads to changes in the percentages of lymphocyte subpopulations and cytokine secretion by Th1 (IL-2, IFN-γ) and Th2 (IL-4 and IL-10) lymphocytes in Peyer's patches of the ileum after in vitro stimulation with lipopolysaccharides (LPS). Immunocompetent cells isolated from Payer's patches on experimental days 0, 14, 28 and 42 were cultured in vitro and stimulated with LPS. The presence of IL-2, IFN-γ, IL-4 and IL-10 in culture media was determined by the ELISA method. The results of the study indicate that ZEN inhibits IL-2 and IFN-γ secretion and stimulates IL-4 and IL-10 produc- tion by Th1 and Th2 lymphocytes by shifting the Th1/Th2 balance towards the humoral immune response. The above can promote allergic responses, as demonstrated by the increase in the size of B1 cell populations producing more autoantibodies. ZEN can also lower resistance to viruses and tumors by inhibiting the proliferation of NK cells and IFN-γ secretion.

Liaison between natural killer cells and dendritic cells in human gestation.

A successful pregnancy relies on immunological adaptations that allow the fetus to grow and develop in the uterus, despite being recognized by maternal immune cells. Among several immunocompetent cell types present within the human maternal/fetal interface, DC-SIGN(+) dendritic cells (DCs) and CD56(+) natural killer (NK) cells are of major importance for early pregnancy maintenance, not only generating maternal immunological tolerance but also regulating stromal cell differentiation. Previous reports show the presence of NK-DC cell conjugates in first trimester human decidua, suggesting that these cells may play a role in the modulation of the local immune response within the uterus. While effective immunity is necessary to protect the mother from harmful pathogens, some form of tolerance must be activated to avoid an immune response against fetal antigens. This review article discusses current evidence concerning the functions of DC and NK cells in pregnancy and their liaison in human decidua.

Expression of HLA-G1 at the placental interface of HIV-1 infected pregnant women and vertical transmission of HIV

Objectives The ability of Human Leucocyte Antigen-G (HLA-G) to inhibit the cytolytic effect to immunocompetent cell types, suggests that HLA-G has an immunomodulatory role. In view of this concept the objective of the study was to assess whether the Major Histocompatibility Complex -coded molecule HLA-G mRNA is a risk factor at the placental barrier in HIV-1 positive pregnant women. Design Placental HLA-G1 levels in HIV-1 infected mothers and viral loads in both mothers and their babies were performed on fifty-five participants. Methods Synthesis of complementary deoxyribose nucleic acid (cDNA) was performed using ribose nucleic acid (RNA) extracted from placental tissue samples. Amplification of cDNA using specifically designed primers complementary to the full length HLA-G1 isoform was quantified using real time-polymerase chain reaction (RT-PCR). Viral load assays (Amplicor Version 1.5, Roche Diagnostics) were performed on all plasma samples. Results HLA-G1 primers detected the full length isoform HLA-G1 PCR product at 86.5 °C. Logistic regression calculations indicated that the risk of babies becoming infected increased by 1.3 with every 1 unit increase in HLA-G1 expression. Female babies were 3.7 times more likely to become infected than male. There was a positive correlation between mothers’ log viral load and transmission of infection to the baby ( p = 0.047; 95%CI 1.029–11.499). Conclusion Maternal viral load was a strong predictor of viral transmission. Placental HLA-G1 expression was up-regulated 3.95 times more in placentas of HIV-1 infected mothers with infected babies when compared to uninfected babies.

In vitro studies of interaction of modified silica nanoparticles with different types of immunocompetent cells

Interactions between different types of immune cells and organically-modified silica nanoparticles were studied. The silica particles functionalized with amine groups were prepared by sol-gel technique. Sheep immunoglobulin labeled with fluoresceine isothiocyanate was immobilized by adsorption onto the nanoparticles. The presence of the functional groups was confirmed by infrared absorption measurements. The level of immunocompetent cells interacting with the silica nanoparticles was estimated as the amount of fluorescence-bright cells by flow cytometry method. A low level of interaction of the peripheral blood lymphocytes with the silica nanoparticles was found. On the contrary, the macrophages are actively involved in interaction with the silica nanoparticles. The influence of different size of the silica nanoparticles and incubation time on viability and functional activity of peripheral blood lymphocytes and peritoneal macrophages were investigated.

Inhibition of Transforming Growth Factor-β–Mediated Immunosuppression in Tumor-Draining Lymph Nodes Augments Antitumor Responses by Various Immunologic Cell Types

Tumor-draining lymph nodes (DLN) are the most important priming sites for generation of antitumor immune responses. They are also the location where an immunosuppressive cytokine, transforming growth factor-beta (TGF-beta), plays a critical role in suppressing these antitumor immune responses. We focused on TGF-beta-mediated immunosuppression in DLNs and examined whether local inhibition of TGF-beta augmented antitumor immune responses systemically in tumor-bearing mice models. For inhibition of TGF-beta-mediated immunosuppression in DLNs, C57BL/6 mice subcutaneously bearing E.G7 tumors were administered plasmid DNA encoding the extracellular domain of TGF-beta type II receptor fused to the human IgG heavy chain (TGFR DNA) i.m. near the established tumor. In DLNs, inhibition of TGF-beta suppressed the proliferation of regulatory T cells and increased the number of tumor antigen-specific CD4(+) or CD8(+) cells producing IFN-gamma. Enhancement of antitumor immune responses in DLNs were associated with augmented tumor antigen-specific cytotoxic and natural killer activity in spleen as well as elevated levels of tumor-specific antibody in sera. The growth of the established metastatic as well as primary tumors was effectively suppressed via augmented antitumor immune responses. Inhibition of TGF-beta-mediated immunosuppression in DLNs is significantly associated with augmented antitumor responses by various immunocompetent cell types. This animal model provides a novel rationale for molecular cancer therapeutics targeting TGF-beta.

The Placental Barrier in Allogenic Immune Conflict in Spontaneous Early Abortions: Immunohistochemical and Morphological Study

Morphologic changes in the placental barrier in spontaneous early abortions under the maternal-embryonic immune conflict, and the role of maternal immunoglobulins (Igs) in these changes. We examined chorionic villi and other tissues obtained from 54 aborts between weeks 3.5 and 8 of pregnancy. Material was divided into two groups. Group 1 (control) contained 15 medically recommended and spontaneous early aborts with no signs of inflammations or pathologic immune processes. Group 2 contained 39 spontaneous early aborts with acute chorionic villitis. Immunohistochemical and morphometric methods were used to study the Igs, different types of immunocompetent cells, and apoptosis-related components of the placental barrier. Acute villitis was found to be characterized by the destruction of all components of the chorionic villi, thrombovasculitis with apoptosis of the endothelium of capillaries and erythroblasts, mucous swelling of the basal membrane, and coagulation of the blood proteins. Due to destruction of the capillaries, the number of avasculate villi increased, and the average number of capillaries per villus decreased. The extremely high number of phagolysosomes with IgG and IgA in the villous monocytes in the group 2 indicates an increase in the phagocytic activity of monocytes against maternal Igs and may reflect the presence of mother-embryo immune conflict. Apoptosis of monocytes and a high number of promonocytes were seen accompanied by a high concentration of p53 protein. A large disturbance in the trophoblast occurred with disappearance of bcl-2 and the appearance of Fas ligand. Massive destruction of maternal Igs in embryonic monocytes and acute villitis in the placental barrier are manifested during the mother-embryo immune conflict, and this may be one of the reasons of spontaneous early abortions.

Role of nonsynaptic communication in regulating the immune response

The discovery of nonsynaptic communication in the 1960s and 1970s was an important milestone in investigating the function of the nervous system, and it revolutionized our view about information transmission between neurons. In addition, nonsynaptic communication has a practical importance not only within the nervous system, but in the communication between the peripheral nervous system and other organ systems. Nonsynaptic communication takes place in different immune organs, which are innervated by sympathetic nerve terminals. In addition, the function of microglia, one of the immunocompetent cell types of the brain, can also be affected by neurotransmitters released from axon varicosities. The various functions of immune cells are modulated by released neurotransmitters without any direct synaptic contact between nerve endings and targeted immune cells requiring only functional neurotransmitter receptors on immune cells. Here, we briefly overview the role of the various receptor subtypes mediating nonsynaptic modulation of the function of immunocompetent cells both in the periphery and in the central nervous system.

Interactions between rat colon carcinoma cells and Kupffer cells during the onset of hepatic metastasis.

Liver sinusoids harbor populations of 2 important types of immunocompetent cells, Kupffer cells (KCs) and natural killer (NK) cells, which are thought to play an important role in controlling hepatic metastasis in the first 24 hr upon arrival of the tumor cells in the liver. We studied the early interaction of KCs, NK and CC531s colon carcinoma cells in a syngeneic rat model by confocal laser scanning microscopy. Results showed a minority of KCs (19% periportal and 7% pericentral) involved in the interaction with 94% of tumor cells and effecting the phagocytosis of 92% of them. NK cell depletion decreased the phagocytosis of tumor cells by KCs by 33% over a period of 24 hr, leaving 35% of the cancer cells free, as compared to 6% in NK-positive rats. Surviving cancer cells were primarily located close to the Glisson capsule, suggesting that metastasis would initiate from this region.

Acute laryngotracheitis in the rat induced by Sendai virus: the influx of six different types of immunocompetent cells into the laryngeal mucosa differs strongly between the subglottic and the glottic compartment.

Acute laryngotracheitis is a disease in which mainly the subglottic area is infected, whereas adjacent parts of the larynx, especially the narrow glottic fold, remain unaffected. The reason for the difference between these two directly adjacent regions is unknown. Therefore, in the present study the influx of dendritic cells, neutrophils, T and B lymphocytes, natural killer cells, and macrophages into the mucosa of different laryngeal compartments was investigated after Sendai virus infection in the rat. The aims were to study both the influx of immunocompetent cells and the adhesion of the pathogen and to correlate them to the different reactions of the laryngeal areas during pseudocroup. Acute laryngotracheitis was induced by intranasal application of Sendai virus in brown Norway rats. This virus is exclusively pneumotropic in rodents and belongs to the parainfluenza virus type 1, the main pathogen of acute laryngotracheitis in children. The numbers of dendritic cells, neutrophils, T and B lymphocytes, natural killer cells, and macrophages were determined in the supraglottic, glottic, subglottic, and tracheal mucosa on days 2, 5, 7, and 14 after virus application. Furthermore, the nucleoprotein of the virus and major histocompatibility complex (MHC) Class II expression were detected immunohistologically on the laryngeal epithelium. All cell subsets entered the laryngeal mucosa during inflammation. The highest influx was detected among dendritic cells subglottically. This was accompanied by a strong virus adhesion and MHC Class II expression on the subglottic epithelium. In contrast, only a few immunocompetent cells entered the adjacent glottic mucosa, and on the glottic epithelium staining for virus nucleoprotein and MHC Class II expression was weak. The inflammatory response of the laryngeal mucosa shows great regional differences in this animal model during experimental viral infection. The response was characterized by a strong subglottic and a weak glottic reaction. A possible reason for this difference might be region-specific viral adhesion on the epithelium of the laryngeal areas, as well as differences in MHC Class II expression. Thus, these data agree with the clinical observation during acute laryngotracheitis and may explain why the subglottic part of the larynx is affected preferentially during pseudocroup. The molecular mechanisms mediating the different reactions await clarification.

Identification and Sequencing of Bcl and Apaf1 Genes in the Common Carp, Cyprinus Carpio.

INTRODUCTION. The role of apoptosis in the immune system is being studied in cultured carp, Cyprinus carpio an economically important farmed fish. In intensive culturing systems, stress is suspected of reducing the immune competence of farm animals. Our initial studies investigated the effect of dexamethasone, a corticosteroid analogue, on the different immunocompetent cells in the pronephros and thymus. This is currently being extended to identify and sequence an apoptotic regulator (Bcl-x) and an apoptotic activator gene (APAF1) from the carp cDNA library. METHOD. Cells from the pronephros and thymus were maintained in vitro, in L-15 medium (Sigma) supplemented with 2mM L- glutamine and containing penicillin (100i.u./ml), streptomycin (100µg/ml), and 5% heat- inactivated foetal calf serum, at 20 o C. They were then exposed to different concentrations of dexamethasone (0.1µM to10.0µM 2 ) for up to 48h in time. Control cells were those maintained in the culture medium lacking in dexamethasone. Cell viability and percentage of apoptotic cell death at times 0h, 24h and 48h were determined using nigrosine dye exclusion and acridine orange stain respectively. DNA from approximately 10 6 cells was isolated from each of the above samples and subjected to agarose gel electrophoresis. In the differential experiment, the different types of immunocompetent cells were labelled with monoclonal antibodies WCI04, WCI09 and WCI12 and their responses to 1.0µM dexamethasone over 6 hours was recorded. Degenerate primers for the Bcl-xL equivalent in carp were designed following a homology study on the sequences of Homo sapiens (human) and Xenopus laevis (South African clawed frog). The homology between human APAF1 and that of Danio rerio (Zebrafish) was examined, and primers constructed, for the comparable sequence in the carp. These were used in PCR to obtain sequences from the carp cDNA library. The sequences obtained will now be used as probes to sequence the rest of the gene(s). RESULTS. Statistical analysis reveals that there is a significant difference between the number of control cells undergoing apoptotic cell death and that of cells exposed to the different concentrations of dexamethasone (p<0.001) (Fig.1). This effect is greatest between the 0h and 24h time periods and appears to decrease after this time period, and is similar in the pronephros and the thymus.

The interactive effects of cocaine/gender on immune function in mice. An observation of in vivo acute cocaine exposure

The present experiments used a mouse model including both sexes to study the impact of acute cocaine exposure on the function of four major immunocompetent cell types (T, B cells, NK, and macrophages). Cocaine hydrochloride, 5 mg or 40 mg/kg, was administrated by i.p. injection to C57BL/6 mice. Thymocytes and splenocytes were obtained 24 h after injection. Acute in vivo cocaine exposure inhibited the proliferation of T lymphocytes in response to Con-A in both thymocytes and splenocytes. However, the attenuated IL-2 production was only seen in thymocytes. These effects on T cells were greater in male mice than in female mice. The function of macrophages was also impaired by acute cocaine exposure; however, the impact was greater in female than in male mice. In conclusion, the effects of acute cocaine exposure altered the functions of immunocompetent cells and the effects varied with gender.