Regional Lymph Node Cells Research Articles

Primary gastric small cell carcinoma: report of a case and review of the literature

A 52-year-old man suffering from a pure-type primary gastric small cell carcinoma was treated with surgery and combination chemotherapy. The small cell carcinoma, approximately 6.5 cm in diameter, was situated in the posterior wall of the antrum and there were no distant metastases. Total gastrectomy and regional lymph node dissection was carried out. Histological examination revealed a solid pattern of proliferation of small cells with hyperchromatic, round nuclei and scant cytoplasm. Neoplastic cells infiltrated into the subserosal layer with severe lymphatic and vascular invasion. Regional lymph node cells were mostly replaced by tumor cells that stained positive for Grimelius, neuron-specific enolase (NSE), and synaptophysin. Accumulations of electron-dense core granules in the small neoplastic cells were seen by electron microscopy. Following surgery, the patient was treated with adjuvant chemotherapy consisting of cisplatin and etoposide. The patient is alive and recurrence free 3 years after surgical operation. We review 107 published cases of primary gastric small cell carcinoma, an extremely rare disease first reported in 1976. Small cell carcinoma is an aggressive, malignant tumor. Intensive chemotherapy is essential for patient survival even when curative surgical resection is carried out.

Okkulte lymphatische Tumorzelldisseminierung beim Pankreaskarzinom

The aim of this study was to determine the frequency and effect on prognosis of occult tumor cells in regional lymph nodes judged to be tumor-free in conventional histopathology of pancreatic cancer patients. Among 115 patients who underwent pancreatic resection for pancreatic (n=84) or distal common bile duct malignancy (n=12) or carcinoma of the papilla (n=19), 48 (42%) were staged pN0. Archival paraffin blocks of 271 resected regional lymph nodes of 41 pN0 patients were reevaluated for occult tumor cells using monoclonal antibody Ber-EP4. Cases with or without isolated tumor cells were compared regarding the distribution of various clinicopathological factors. Of 41 pN0 patients, 16 (39%) exhibited single Ber-Ep4 immunoreactive cells or small cell clusters in at least one lymph node. The occurrence of occult tumor cells was not dependent on other clinicopathological factors such as pT stage, grading, or curative resection. However, those cells were encountered more frequently in common bile duct carcinomas (100%) than in pancreatic (36%) or papilla (20%) carcinomas (P=0.009). Occult tumor cells impaired prognosis significantly in uni- and multivariate analyses (estimated 5-year survival 53% for pN0((i-)) vs 10% for pN0((i+)) and 9% for pN1/N2; P=0.0047). Occult tumor cells are frequent in apparently tumor-free lymph nodes of pancreatic cancer patients and often overlooked in conventional histopathology. They are encountered even in limited stages of disease and they impair prognosis, which is comparable to that of patients with true lymphatic metastases. Occult tumor cells in lymph nodes of pancreatic cancer patients could be used to stratify adjuvant therapy.

IL-12 Breaks Dinitrothiocyanobenzene (DNTB)-Mediated Tolerance and Converts the Tolerogen DNTB into an Immunogen

Epicutaneous application of dinitrothiocyanobenzene (DNTB) induces tolerance against its related compound dinitrofluorobenzene (DNFB), because DNTB-pretreated mice cannot be sensitized against the potent hapten DNFB. This tolerance is hapten-specific and transferable. In this study, we demonstrate that IL-12 can break DNTB-mediated tolerance. Furthermore, naive mice treated with IL-12 before DNTB application responded to DNFB challenge with a pronounced ear swelling response without previous sensitization to DNFB, showing that IL-12 can convert the tolerogen DNTB into an immunogen. No differences in numbers or regulatory activity were observed between CD4+CD25+ regulatory T cells isolated from mice treated with DNFB, DNTB, or IL-12 followed by DNTB. However, the number of CD207+ Langerhans cells in regional lymph nodes of DNTB-treated mice was significantly lower than in animals treated with DNFB or IL-12 plus DNTB. Additionally, CD11c+ dendritic cells (DC) isolated from regional lymph nodes of DNTB-treated mice had a significantly lower ability to stimulate T cell proliferation and produced reduced amounts of inflammatory cytokines. Application of both DNFB and DNTB induced apoptotic cell death of DC in the epidermis and the regional lymph nodes. However, the number of apoptotic DC in regional lymph nodes was significantly higher in DNTB-treated animals compared with mice treated with DNFB or IL-12 plus DNTB. Therefore, we conclude that DNTB-mediated tolerance is secondary to inefficient Ag presentation as a result of apoptotic cell death of DC and that IL-12 converts the tolerogen DNTB into an immunogen by preventing DNTB-induced apoptosis of DC.

CCR5-Deficient Mice Develop Experimental Autoimmune Uveoretinitis in the Context of a Deviant Effector Response

Experimental autoimmune uveoretinitis (EAU) is an organ-specific, Th1-cell-mediated disease that targets the neural retina. CCR5 is a chemokine receptor expressed on Th1 cells that promotes their migration. In CCR5-deficient mice, we examined the role of CCR5 in the development of EAU induced by immunization with interphotoreceptor retinoid-binding protein (IRBP) peptide. Wild-type or CCR5-deficient B6 mice were immunized with human IRBP peptide 1-20 (hIRBP-p), and the severity of EAU was assessed clinically and histologically. Splenocytes and cells of regional lymph nodes near the eye were collected and their proliferation and production of IL-6, IL-10, IFN-gamma, and CCL2 (MCP-1) in response to hIRBP-p stimulation were measured. Moreover, the intraocular levels of these cytokines were analyzed. Immunization with hIRBP-p induced EAU in CCR5-deficient mice with a severity comparable to that in wild-type mice. Histologically, T-cell infiltration of the eye was reduced, but granulocyte infiltration was augmented in CCR5-deficient mice. Although splenic T cells from CCR5-deficient mice produced IFN-gamma but not IL-10 on stimulation by hIRBP-p, T cells from the regional lymph nodes failed to produce both cytokines. IL-6 production in the eye and IL-6 and CCL2 production by splenic T cells were predominantly augmented in CCR5-deficient mice. The development of EAU is not prevented in CCR5-deficient mice. Although T-cell infiltration into the eye is apparently reduced in CCR5-deficient mice, the defect is compensated for by granulocyte infiltration, supposedly mediated by augmented intraocular production of IL-6.

Detection of disseminated lung cancer cells in regional lymph nodes by assay of CK19 reverse transcriptase polymerase chain reaction and its clinical significance

To set up a molecular method (i.e. RT-PCR) that can be used to detect disseminated tumor cells (DTCs) in regional lymph nodes (LNs) in patients with lung cancer and to evaluate its clinical significance. Cytokeratin 19 (CK(19)) was used as marker. Serial dilution study for LC-5 cells (a lung squamous cell line) was performed to detect sensitivity of the molecular protocol. Regional LNs (n = 261) and primary lung cancer tissue (n = 40) were obtained from 40 patients with lung cancer who underwent lobectomy or pneumonectomy. They were randomly categorized into two groups: group I (LN-based study, n = 20) and group II (patient-based study, n = 20). Each LN was halved. One half of a LN was subjected to histological examination (HE) and the other half was subjected to RT-PCR amplification of CK(19) mRNA. The effect on survival was analyzed. The cumulative survival was calculated by the Kaplan-Meier method and compared by the log rank test. The Cox model analyzed the prognostic factors. CK(19) mRNA expressed in all tumor tissues as well as LC-5, PAa cells (a lung adenocarcinoma cell line), but not in normal control LNs. Serial dilution study for LC-5 cells demonstrated that CK(19) mRNA was detectable at a concentration as low as 10 LC-5 cells in 1x10(7) LN cells. There was no significant difference between the detecting result of single LN and that of mixed LNs (P > 0.05). In 18 of 40 patients, the metastasis in regional LNs was found by both HE and RT-PCR. Of 22 patients without pathologically involved nodes, six (27%) were found to express CK(19) mRNA in regional LNs. According to the results of regional LNs in 40 patients by molecular assay, the presence of the CK(19) product in LNs was related to tumor size (chi(2) = 5.76, P < 0.025) as well as cell differentiation of the tumor (chi(2) = 7.08, P < 0.01). Following a median observation time of 26 months (range, 4 to 60 months), patients with DTCs in nodes showed significant shorter disease-free survival duration than node-negative patients (log-rank test, P = 0.001). The independence of this prognostic significance was demonstrated by a multivariate analysis (Cox regression model, P = 0.004). The results diagnosed by HE had no significant effect on prognoses (P = 0.455). Comparing with HE, RT-PCR can make more accurate assessment of metastatic status in LNs, which is helpful for screening the patients in whom the early subclinical metastasis exists and disclosing the intrinsic regulation of malignant metastasis. The presence of DTCs in LNs is an independent factor for prognosis. Molecular detection of DTCs in LNs is a supplement for current tumor staging in lung carcinoma.

Detection of disseminated tumor cells in mediastinoscopic lymph node biopsies and lymphadenectomy specimens of patients with NSCLC by quantitative RT-PCR☆

Detection of disseminated tumor cells in mediastinoscopic biopsies could improve staging and might be helpful concerning indications for neoadjuvant therapy regimens. This prospective study was performed to evaluate a simple and observer-independent polymerase chain reaction (PCR)-based method for the detection of disseminated tumor cells in regional lymph nodes. Lymph nodes of 32 consecutive patients without neoadjuvant therapy were removed by systematic lymphadenectomy during resection of primary NSCLC. One hundred of these lymph nodes were cut into two equal halves which were examined using either routine histopathology or quantitative reverse transcriptase PCR (qRT-PCR). qRT-PCR amplification of cytokeratin 19 (CK19) transcripts was applied for the detection of tumor cell-specific RNA. We differentiated between illegitimate marker gene transcription and cancer-specific expression by using a cut-off value that was obtained from the analysis of 18 lymph nodes of patients with benign lung diseases. Subsequent to the evaluation of qRT-PCR, a pilot project with five additional patients was conducted to examine 19 mediastinoscopic biopsies, which were cut into two equal halves and proceeded as described above. Ninety-four (94%) lymph nodes were tumor-free by histopathology. qRT-PCR detected disseminated tumor cells in 26 (28%) of these lymph nodes. All of the remaining six lymph nodes that were judged by the pathologist to contain tumor cells exhibited CK19 transcripts. Twenty-three patients had a pN0 status. qRT-PCR detected disseminated tumor cells in 13 (56%) of these pN0 patients. The mediastinoscopic biopsies showed disseminated tumor cells in four (21%) out of 19 histopathologically tumor-free samples. CK19 qRT-PCR is a sensitive and specific tools for the detection of disseminated tumor cells in regional lymph nodes of patients with operable NSCLC. Further studies are required to asses if this molecular method might improve mediastinoscopic staging.

Morphological Changes in Submandibular Lymph Nodes after Subcutaneous Implantation of Gold Alloys

Functional activity of cells in regional lymph nodes of the major salivary glands was studied in rats after subcutaneous implantation of gold alloys. Alloy 900 modified the function of regional lymph nodes.

The Effects of Rotation Stress on Measures of Immunity. The Role of Opiate Receptors

Experiments were performed on mongrel male rats to study the effects of blockade of delta, mu, and kappa opiate receptors on antibody formation, delayed-type hypersensitivity (DTH), and changes in the numbers of antibody-forming cells (AFC) and nucleated cells in the lymph nodes and spleen in a local form of immune response in a model of rotation stress. These studies showed that rotation stress led to mild suppression of immune inflammation in DTH, significant increases in the numbers of AFC and nucleated cells in regional lymph nodes, but no change in the peripheral blood antibody titer. Blockade of delta, mu, and kappa opiate receptors with naloxone altered these stress-induced effects. It is suggested that abolition of the stimulating effect of rotation stress on the number of AFC and depression of DTH may be associated with blockade of the effects of beta-endorphin and met-enkephalin, which act predominantly via stimulation of delta receptors.

Significance of isolated tumor cells in lymph nodes among gastric cancer patients.

To determine the frequency and prognostic impact of isolated tumor cells (ITC) in regional lymph nodes judged to be tumor free in conventional histopathology among gastric cancer patients. Among 161 patients who underwent gastrectomy and D2-lymphadenectomy, 56 were staged pN0(35%). Archival paraffin blocks of 1148 resected regional lymph nodes of those pN0 patients were reevaluated for ITC using monoclonal antibody Ber-EP4. Patients with and without ITC were compared with regard to the distribution of various clinicopathological factors. Prognostic impact of ITC was tested in uni- and multivariate analysis. Of 56 pN0 patients, 33 (59%) exhibited single Ber-Ep4 immunoreactive cells or small cell clusters in at least one lymph node. The occurrence of ITC was not dependent on other clinicopathological factors. ITC impaired patients' prognoses significantly in uni- as well as multivariate analyses [estimated 5-year survival rate: 82% for pN0((i-))vs 58% for pN0((i+))(p = 0.059) and 15% for pN1/2 (p = 0.0005 and p < 0.0001, respectively)]. ITC are a frequent event in apparently tumor-free lymph nodes of gastric cancer patients and are overlooked by conventional histopathology. They are encountered even in limited stages of disease and impair patients' prognoses. This should be borne in mind when advocating local resection for early gastric cancer.

T-cell receptor Vbeta gene usage by T cells reactive with the tumor-rejection antigen SART-1 in oral squamous cell carcinoma.

We recently described that the SART-1(690-698) peptide could induce HLA-A24-restricted cytotoxic T lymphocytes (CTLs), which recognize the SART-1(259) (+) tumor cells from peripheral blood mononuclear cells (PBMCs) of HLA-A24(+) cancer patients. In our study, in 5 of 14 HLA-A24(+) patients with oral squamous cell carcinomas (SCCs), CTLs could be induced with the SART-1(690-698) peptide from the PBMCs. In 2 of the patients from whom the highest CTL activities were induced, the T-cell receptor (TCR) Vbeta repertoire expressed by the SART-1(690-698)-specific CTLs was found to be restricted and multiple Vbeta families were predominantly expressed in each patient. Although the predominant Vbeta families were different between the 2 patients, Vbeta7 was highly and commonly predominant. The same predominant Vbeta families were also detected in the tumor-infiltrating lymphocytes (TILs) from each patient, and each Vbeta family contained one or more unique T-cell clonotypes. The unique T-cell clonotypes were found to be common between the TILs and SART-1(690-698)-specific CTLs from each patient, and especially 2 T-cell clonotypes with Vbeta7 were identical even in the 2 patients. One of the 2 T-cell clonotypes with Vbeta7 was detected in the TILs from 11 of 14 HLA-A24(+) patients and another was found in those from 8 of HLA-A24(+) patients, while none of 10 HLA-A24(-) patients demonstrated both T-cell clonotypes. These results strongly suggest that the T-cell clonotypes with Vbeta7 are major TCR Vbeta genes expressed by SART-1(690-698)-specific CTLs. Furthermore, autologous tumor cells from one of the HLA-A24(+) patients stimulated the PBMCs and regional lymph node cells (LNCs) to expand the same T-cell clonotypes as those in the SART-1(690-698)-specific CTLs. These results strongly suggest that the SART-1(690-698)-specific CTLs clearly accumulate in vivo, especially in the TILs, as a consequence of in situ antigenic stimulation by autologous tumor cells. The identification of the unique TCR Vbeta genes used by SART-1(259)-specific CTLs should help to improve the diagnosis of the specific immune response in patients with SART-1(259) (+) cancers, especially during anticancer immunotherapy.

Calcium channel blockers suppress the contact hypersensitivity reaction (CHR) by inhibiting antigen transport and presentation by epidermal Langerhans cells in mice.

Since Langerhans cells (LC) are the principal antigen-presenting cells among epidermal cells, treatments suppressing LC function may inhibit CHR. Although calcium channel blockers (CCB) have been shown to suppress the functions of several immunologically active cells, little is known about their effect on LC. In this study we show that pretreatment with topical 1% nifedipine or verapamil HCl significantly suppressed both the sensitization and elicitation phases of a CHR in mice. We then investigated whether CCB affected LC. Flow cytometric analysis of regional lymph node cells obtained 24 h after applying FITC demonstrated that topical CCB treatment significantly reduced the percentage of FITC+ NLDC-145+ cells, suggesting that CCB had suppressed antigen transport by LC. In vitro treatment with nifedipine or verapamil significantly suppressed the antigen-presenting capacity of LC in a dose-dependent manner. In addition, in vitro CCB treatment reduced the percentage of class II MHC antigen-positive epidermal cells and significantly suppressed class II MHC and B7-1 levels in LC, as determined by flow cytometry and reverse transcriptase-polymerase chain reaction, whereas surface expression of B7-2 and mRNA was only weakly reduced. Neither expression of CD45 nor the percentage of CD45+ cells were affected, suggesting that the effects of CCB on LC were not due to cytotoxicity. Our results suggest that CCB inhibit CHR, at least in part, by suppressing the functions of LC.

Altered cutaneous immune parameters in transgenic mice overexpressing viral IL-10 in the epidermis.

IL-10 is a pleiotropic cytokine that inhibits several immune parameters, including Th1 cell-mediated immune responses, antigen presentation, and antigen-specific T cell proliferation. Recent data implicate IL-10 as a mediator of suppression of cell-mediated immunity induced by exposure to UVB radiation (280-320 nm). To investigate the effects of IL-10 on the cutaneous immune system, we engineered transgenic mice that overexpress viral IL-10 (vIL-10) in the epidermis. vIL-10 transgenic mice demonstrated a reduced number of I-A(+) epidermal and dermal cells and fewer I-A(+) hapten-bearing cells in regional lymph nodes after hapten painting of the skin. Reduced CD80 and CD86 expression by I-A(+) epidermal cells was also observed. vIL-10 transgenic mice demonstrated a smaller delayed-type hypersensitivity response to allogeneic cells upon challenge but had normal contact hypersensitivity to an epicutaneously applied hapten. Fresh epidermal cells from vIL-10 transgenic mice showed a decreased ability to stimulate allogeneic T cell proliferation, as did splenocytes. Additionally, chronic exposure of mice to UVB radiation led to the development of fewer skin tumors in vIL-10 mice than in WT controls, and vIL-10 transgenic mice had increased splenic NK cell activity against YAC-1targets. These findings support the concept that IL-10 is an important regulator of cutaneous immune function.

Altered cutaneous immune parameters in transgenic mice overexpressing viral IL-10 in the epidermis

IL-10 is a pleiotropic cytokine that inhibits several immune parameters, including Th1 cell-mediated immune responses, antigen presentation, and antigen-specific T cell proliferation. Recent data implicate IL-10 as a mediator of suppression of cell-mediated immunity induced by exposure to UVB radiation (280-320 nm). To investigate the effects of IL-10 on the cutaneous immune system, we engineered transgenic mice that overexpress viral IL-10 (vIL-10) in the epidermis. vIL-10 transgenic mice demonstrated a reduced number of I-A(+) epidermal and dermal cells and fewer I-A(+) hapten-bearing cells in regional lymph nodes after hapten painting of the skin. Reduced CD80 and CD86 expression by I-A(+) epidermal cells was also observed. vIL-10 transgenic mice demonstrated a smaller delayed-type hypersensitivity response to allogeneic cells upon challenge but had normal contact hypersensitivity to an epicutaneously applied hapten. Fresh epidermal cells from vIL-10 transgenic mice showed a decreased ability to stimulate allogeneic T cell proliferation, as did splenocytes. Additionally, chronic exposure of mice to UVB radiation led to the development of fewer skin tumors in vIL-10 mice than in WT controls, and vIL-10 transgenic mice had increased splenic NK cell activity against YAC-1targets. These findings support the concept that IL-10 is an important regulator of cutaneous immune function.

Phenotypic profile and functional characterization of rat lymph node‐derived γδ T cells: implication in the immune response to cytomegalovirus

Gammadelta T cells are unique, and their localization at sites of infection is considered critical in immune defence. We demonstrate the accumulation of gammadelta T cells in rat regional popliteal lymph nodes (PLNi) starting 2 days after inoculation of cytomegalovirus (CMV) into the footpad. Early-appearance PLNi gammadelta T cells significantly inhibited plaque development and the spread of CMV infection. These gammadelta T cells were negative for CD4 and CD8beta receptors, proliferated in response to interleukin-2 (IL-2) and contained high levels of interferon-gamma (IFN-gamma), the appearance of which correlated with the curing of fibroblasts from virus infection. The addition of anti-IFN-gamma abolished the ability of fibroblast monolayers to be cured from CMV infection. In contrast, this protection was not abolished by the addition of anti-rat IL-2 or anti-rat TNF-alpha, or by the depletion of NKR-P1-bearing cells within gammadelta T cells. In addition, the present study shows that while gammadelta T cells derived from naive and CMV-infected rats are able to kill both YAC-1 targets and CMV-infected syngeneic fibroblasts in vitro, only the latter are able to clear CMV-infected fibroblast monolayers. Finally, our data suggest that the expression of NKR-P1 by gammadelta T cells is critical for cytotoxicity, but its contribution to the curing from CMV infection was limited.

Asymmetry of delayed type hypersensitivity reaction in mice.

The intensity of delayed type hypersensitivity reactions in the left and right paws was studied in mice divided into left- and right-pawed by the motor asymmetry of the brain. The reaction was more pronounced in the left paw in all animals irrespective of motor asymmetry. Motor asymmetry of the brain hemispheres little contributed to the manifestation of differences in the delayed type hypersensitivity reactions in the left and right paws. The authors concluded that asymmetry of cellular immunity is determined by functional asymmetry of cells in regional lymph nodes.

Development of surrogate markers for oral immunisation against Candida albicans

We have developed an oral vaccine using the blastococcoid form of Candida albicans with a murine model of oral candidiasis with infection-resistant (BALB/c) and infection-prone (DBA/2) mice. Previous studies had demonstrated that enhanced clearance was linked to increased production of both Th1 and Th2 cytokines (IFN-γ and IL-4, respectively) in regional lymph nodes, and increased secretion of IFN-γ and NO in saliva. Subsequent study using oral Lactobacillus acidophilus (LAVRI-A1) induced a local cytokine pattern and enhanced clearance of intra-oral C. albicans similar to that noted in animals given killed candida vaccine. Thus, the experiment was repeated with a 2 month gap between completion of a vaccine course and challenge with live C. albicans. Compared to control mice, those given the oral vaccine had a 4 log reduction in colonisation on day 2, associated with twice the levels of secretion of IFN-γ and IL-4 from the regional node cells, five times the level of nitric oxide in saliva, and suppression of NO production with MMLA induced a 2 log increase in oral colonisation. Thus while both IFN-γ and IL-4 appear to contribute to clearance of oral C. albicans, IL-4 appears to act through a paracrine enhancement of NO production. Studies in man are in progress to define similar surrogate markers of immune protection, prior to oral vaccine trial.

Inflammation and cancer, the mastocytoma P815 tumor model revisited: Triggering of macrophage activation in vivo with pro‐tumorigenic consequences

Subcutaneous in vivo injections of cells of the mastocytoma line P815 in syngenic DBA/2 mice induce locally fast growing solid tumors. These have been used extensively as a cancer model to analyze and manipulate the relationship between tumor cells and host's immune defenses. We report that progression of P815 tumors in vivo was accompanied by a burst (Days 5-7) of local inflammatory cells recruitment and angiogenesis observed histologically, corroborated in vivo by MRI with gadolinium, overtranscription of macrophage activation marker genes, secretion of TNF-alpha by regional lymph node cells and concomitant systemic inflammation. No substantial overtranscriptions of either VEGF or IL-10 or TGF-beta genes were observed. Induction of COX-2 gene was a late event. To establish a possible relationship between the tumor-induced local, regional and systemic increase of pro-inflammatory mediators and progression of tumors in vivo, we carried out experiments deliberately modulating the inflammatory status of the recipient animals. Pretreatment of recipient animals by i.p. injection of thioglycolate accelerated P815 tumor growth. At the opposite, treatment of mice with either a COX-1 + COX-2 inhibitor (aspirin, 1 mg/day/mouse) or a specific COX-2 inhibitor (celecoxib, 0.13 mg/day/mouse) for 2 weeks after injection of tumor cells, significantly reduced the size and growth rate of tumors compared to control mice. Experiments carried out in vitro indicated that peritoneal macrophages from untreated animals were strongly activated by live P815 cells and by P815 membrane preparations. The tumor-induced inflammatory reaction could establish a local micro environment favoring tumor progression. The P815 tumor model might be helpful to recognize important factors controlling host/tumor relationship.

Resolution of skeletal muscle inflammation in mdx dystrophic mouse is accompanied by increased immunoglobulin and interferon-gamma production.

Mdx mouse, the animal model of Duchenne muscular dystrophy, develops an X-linked recessive inflammatory myopathy with an apparent sustained capacity for muscle regeneration. We analysed whether changes in the skeletal muscle during myonecrosis and regeneration would correlate with functional alterations in peripheral lymphoid tissues. Here we show that during the height of myonecrosis, mdx mice display marked atrophy of peripheral lymph nodes and extensive muscle inflammation. In contrast, enlargement of draining lymph nodes with accumulation of CD4+ CD44+, CD4+ CD25+, CD8+ CD44+ T lymphocytes and type-2 B cells was consistently observed during amelioration of the muscle lesion. In addition, regeneration of the muscular tissue was accompanied by concomitant increase of immunoglobulin-secreting cells in regional lymph nodes and bone marrow. Double immunolabelling analysis revealed intense B cell proliferation and formation of germinal centre in the follicles of dystrophic regional lymph nodes. Furthermore, lymph node cells produced large amounts of IFN-gamma but not IL-4, IL-6 or IL-10 after in vitro mitogen stimulation with Concanavalin A. As these alterations occurred mainly during the recovery period, we suggested that local activation of the immune system could be an influence which mitigates the myonecrosis of muscular tissue in the mdx dystrophic mouse.

Immunological approach in the evaluation of regional lymph nodes of patients with squamous cell carcinoma of the head and neck.

In cancer, regional lymph node (LN) cells are one of the first components of the immune system to have contact with tumor cells or their products. Therefore, the phenotype and functional properties of hematopoietic cells present within the tumor-draining LN are important to understanding their role in the control of malignant cells. Based on the locoregional metastatic behavior of squamous cell carcinoma of the head and neck (SCCH&N) region, we analyzed tumor-draining lymph nodes from SCCH&N patients to obtain insights into regional tumor immunity. Using a three-color fluorescent labeling technique, surface antigen expression was visualized in mononuclear cells of lymph nodes that were obtained from head and neck cancer patients and compared to mononuclear cells of normal lymph nodes. Cell cycle analyses were performed using propidium iodide. Proliferation after phytohemagglutinin stimulation was measured by a sodium tetrazolium-based assay. LN histology was correlated with flow cytometric findings. Regional lymph nodes of head and neck cancer patients undergo morphologic and functional changes. Flow cytometry revealed a decrease in CD8(+) T cells and in some lymph nodes the presence of second or third populations of larger cells with distinct size and granularity that expressed both T (gammadelta/alphabeta) and different natural killer cell markers. Moreover, cell cycle analyses and proliferation assays showed a diminished response to mitogenic stimuli. These changes were found in both metastatic and hyperplastic lymph nodes from head and neck cancer patients; however, no alterations were found in control lymph nodes or peripheral blood mononuclear cells from noncancer patients. The immune alterations detected in lymphocytes present within the draining lymph nodes of head and neck cancer patients may improve our understanding of how tumor cells escape host immunosurveillance. However, this dysfunction in local draining lymph nodes may not be detected systemically.

Nitric oxide‐enhanced resistance to oral candidiasis

A murine model of oral candidiasis was used to show that nitric oxide (NO) is involved in host resistance to infection with Candida albicans in infection-'resistant' BALB/c and infection-'prone' DBA/2 mice. Following infection, increased NO production was detected in saliva. Postinfection samples of saliva inhibited the growth of yeast in vitro. Treatment with NG-monomethyl-L-arginine (MMLA), an inhibitor of NO synthesis, led to reduced NO production, which correlated with an increase in C. albicans growth. Reduction in NO production following MMLA treatment correlated with an abrogation of interleukin-4 (IL-4), but not interferon-gamma (IFN-gamma), mRNA gene expression in regional lymph node cells. Down-regulation of IL-4 production was accompanied with an increase in IFN-gamma production in infection-'prone' DBA/2 mice. There was a functional relationship between IL-4 and NO production in that mice treated with anti-IL-4 monoclonal antibody showed a marked inhibition of NO production in saliva and in culture of cervical lymph node cells stimulated with C. albicans antigen. The results support previous conclusions that IL-4 is associated with resistance to oral candidiasis and suggest that NO is involved in controlling colonization of the oral mucosal surface with C. albicans.