Ultrasound Biomicroscopy: A Powerful Tool Probing Murine Lymph Node Size in vivo

Invasive cell-counting in lymph node (LN) is the current reference to assess LN changes due to inflammation, immunodeficiency and cancer in murine models. This work evaluates whether ultrasound biomicroscopy (UBM) can measure LN size alterations noninvasively for a large range of sizes (0.1 mm³ to 22 mm³). Correlation was assessed (ρ = 0.91, p < 0.0001) between invasive cell count and LN volume estimated with UBM (24, 2 to 28-week-old, C57BL/6 mice; 13 same-strain, transgenic mice presenting LN hyperplasia). UBM LN modification screening was applied in a skin-graft rejection model and compared with cell-counting (15 mice). UBM LN-size follow-up with fine temporal sampling was demonstrated from 9 d of age (minimum area 0.13 mm²). Reliability (intraclass correlation coefficient [ICC] > 0.84) and variability of UBM evaluations compared favourably with invasive cell count. UBM provides a noninvasive alternative to cell-counting in mice for early detection and longitudinal screening of LN modifications. This can enable significant reduction in the number of mice and exploration of LNs that would be too small to dissect for cell count. (E-mail: Lori.Bridal@upmc.fr or matteo.bosisio@gmail.com)     

Modulation of the HLA class II antigen at a molecular level by maternal serum among cord blood cells and unrelated lymphocytes

Summary A retroplacental serum factor responsible for the downregulation of the MHC class II antigen expression has been described [1]. We found that maternal serum had the same property. The HLA class II modulating activity is however diminished in the presence of maternal serum as compared to retroplacental serum suggesting that the IA like inhibiting factor is released at the fetomaternal interface. After a 3-day incubation period of unrelated lymphocytes and mononuclear cord blood cells in a maternal serum pool, it was shown that the HLADr, the HLADp, and more significantly, the HLADq molecules were modulated. This phenomenon was more pronounced among cord blood cells. When third party lymphocytes and mononuclear cord blood cells were stimulated by Candidine or unrelated mononuclear cells in the presence of retroplacental serum, only the cellular subpopulation belonging to the CD4⁺ subset showed an HLADq downregulation. The molecular constituents of the MHC class II antigen expression characterizing cells belonging to other subsets remained unchanged. When the same stimulation assay was performed in the presence of a control medium (nulliparous serum), we found no changes in the MHC class II molecular constituents. When unrelated mononuclear cells and mononuclear cord blood cells were PHA stimulated in the presence of maternal and nulliparous serum, the HLADq expression of the CD8⁺ subset showed a significant downregulation in the maternal serum mediated stimulation assay as compared to the control stimulation test. The molecular expression of the HLA class II antigen related to the other subpopulation (CD4⁺, CD3⁺) stimulated by a mitogenic lectin remained unchanged. It is suggested that these molecular MHC class II modulations are due to a factor included in the maternal IgG reaction. Retroplacental IgG contains the highest concentration of this factor.     

Apoptotic body-loaded dendritic cells efficiently cross-prime cytotoxic T lymphocytes specific for NA17-A antigen but not for Melan-A/MART-1 antigen

DCs hold promise for cancer immunotherapy due to their functional ambivalence: iDCs internalize antigens, then mDCs trigger naive T-cell activation. However, no consensus has been reached concerning the optimal mode of antigen acquisition for efficient cross-priming of TAA-specific CTLs, and this remains a field of investigation. Here, we used highly purified apobodies derived from an HLA-A*0201-negative melanoma line as a source of tumor antigens for HLA-A*0201 DCs. We compared in vitro mDCs loaded with apobodies to DCs loaded with antigenic peptides, NA17-A(1-9) and Melan-A/MART-1(26-35) A27L analogue, for their capacity to stimulate melanoma antigen-specific T cells from autologous PBLs. Apobody phagocytosis did not induce spontaneous DC maturation, but phagocytic DCs were still responsive to maturation signals, resulting in a functional ability to activate antigen-specific lymphocytes. NA17-A-specific T lymphocytes were activated by both types of stimulation, whereas only peptide-pulsed DCs stimulated the growth of Melan-A/MART-1-specific lymphocytes. We also observed a lack of staining of melanoma-derived apobodies with a Melan-A-specific MAb, suggesting protein alteration during apoptosis induction. After HLA-A*0201/NA17-A multimer sorting, antigen-specific lymphocytes induced by mature DCs loaded with either peptide or apobodies displayed similar functional capacity against peptide-pulsed T2 cells and melanoma cells. Therefore, apobody-loaded DCs can achieve T-cell priming similar to that induced by peptide-pulsed DCs, provided that the apoptotic process allows the preservation of antigen expression.     

Microsatellite alterations in head and neck squamous cell carcinoma and relation to expression of pimonidazole, CA IX and GLUT-1.

BACKGROUND AND PURPOSE: Because the locoregional control for HNSCC is still disappointing, research efforts focus on the exploration of new molecular markers located in both tumour and microenvironment, which could help stratify patients. The aim of the present work was therefore first to assess microsatellite alterations and hypoxia in HNSCC as possible molecular markers. Second, a relation between both was investigated, as hypoxia is known to select for genetic alterations. MATERIAL AND METHODS: Forty-eight patients with advanced HNSCC treated by surgery+/-radiotherapy were included. MSI and LOH were investigated with microsatellite markers using automatic fragment analysis. The presence of hypoxia was assessed by immunohistochemistry for pimonidazole, CA IX and GLUT-1. The mutual relationship between MSI/LOH and hypoxia was evaluated. RESULTS: No MSI was detected in this patient group. LOH occurred mostly on chromosomal arms 3p, 5q, 9p, 17p and 17q. Patients with LOH at D17S799, located in the near environment of p53, showed a higher CA IX expression (p=0.01). CONCLUSIONS: LOH is a possible molecular marker in HNSCC. The positive correlation between LOH at D17S799 and CA IX is in full concordance with previous publications linking hypoxia to selective pressure on the p53 gene.     

Epithelial–myoepithelial carcinoma of the submandibular gland with symptomatic lung metastases treated with chemotherapy

This report concerns a patient with symptomatic lung metastases from an epithelial–myoepithelial carcinoma of the submandibular gland. Although the efficacy of chemotherapy is unknown in this disease, our patient was treated with cisplatin combined with 5-fluorouracil and later with paclitaxel and cyclophosphamide. Chemotherapy allowed disease stabilization and relief of the pulmonary symptoms. This is the first report on the use of chemotherapy in this very rare salivary gland carcinoma.     

Is ventilator-associated pneumonia an independent risk factor for death?

BACKGROUND: Ventilator-associated pneumonia (VAP) has been implicitly accused of increasing mortality. However, it is not certain that pneumonia is responsible for death or whether fatal outcome is caused by other risk factors for death that exist before the onset of pneumonia. The aim of this study was to evaluate the attributable mortality caused by VAP by performing a matched-paired, case-control study between patients who died and patients who were discharged from the intensive care unit after more than 48 h of mechanical ventilation. METHODS: During the study period, 135 consecutive deaths were included in the case group. Case-control matching criteria were as follows: (1) diagnosis on admission that corresponded to 1 of 11 predefined diagnostic groups; (2) age difference within 10 yr; (3) sex; (4) admission within 1 yr; (5) APACHE II score within 7 points; (6) ventilation of control patients for at least as long as the cases. Precise clinical, radiologic, and microbiologic definitions were used to identify VAP. RESULTS: Analysis was performed on 108 pairs that were matched with 91% of success. There were 39 patients (36.1%) who developed VAP in each group. Multivariate analysis showed that renal failure, bone marrow failure, and treatment with corticosteroids but not VAP were independent risk factors for death. There was no difference observed between cases and controls concerning the clinical and microbiologic diagnostic criteria for pneumonia. CONCLUSION: Ventilator-associated pneumonia does not appear to be an independent risk factor for death.     

Cytogenetic and molecular genetic analysis of tumorigenic human bronchial epithelial cells induced by radon alpha particles

To establish a cell culture model for lung carcinogenesis, independent populations of the human papillomavirus 18-immortalized human bronchial epithelial cell line BEP2D were treated with high linear energy transfer radon-simulated alpha-particles, expanded and xenotransplanted into Nu/Nu mice. Six independent cell lines were established from tumors that developed from three separate radiation treatments as follows: treatment (Tx) 1 (30 cGy--two doses), H2BT, Tx 2 (30 cGy-- single dose), R30T1L, R30T2 and R30T3L, Tx 3 (30 cGy--single dose), H1ATN and H1ATBA1. Cytogenetic analysis revealed common changes in all tumor lines: loss of the Y chromosome (ch), one of three copies of ch8, one of three copies of ch14, and one of two copies of ch4p16-pter and ch11p15-pter. Analysis of polymerase chain reaction-amplified short tandem repeats of informative loci confirmed the loss of chY in all lines and loss of heterozygosity (LOH) at eight loci spanning the length of ch8 in all lines from Tx's 1 and 2. Our data support previous studies indicating the presence of tumor suppressor genes on ch8. LOH also was confirmed on ch14 at locus D14S306 in all cell lines from Tx 2 and in one of two lines from Tx 3. This region, 14q12-q13, may contain changes in one of the five known somatostatin receptor genes (SSTR1). No LOH was detected at any of the informative loci tested for on ch4 or ch11.      

Sequential molecular and cellular events during neoplastic progression: a mouse syngeneic ovarian cancer model

Studies performed to identify early events of ovarian cancer and to establish molecular markers to support of early detection and the development of chemopreventive regimens have been hindered by a lack of adequate cell models. Taking advantage of the spontaneous transformation of mouse ovarian surface epithelial (MOSE) cells in culture, we isolated and characterized distinct transitional stages of ovarian cancer as the cells progressed from a premalignant nontumorigenic phenotype to a highly aggressive malignant phenotype. Transitional stages were concurrent with progressive increases in proliferation, anchorage-independent growth capacity, in vivo tumor formation, and aneuploidy. During neoplastic progression, our ovarian cancer model underwent distinct remodeling of the actin cytoskeleton and focal adhesion complexes, concomitant with downregulation and/or aberrant subcellular localization of two tumor-suppressor proteins E-cadherin and connexin-43. In addition, we demonstrate that epigenetic silencing of E-cadherin through promoter methylation is associated with neoplastic progression of our ovarian cancer model. These results establish critical interactions between cellular cytoskeletal remodeling and epigenetic silencing events in the progression of ovarian cancer. Thus, our MOSE model provides an excellent tool to identify both cellular and molecular changes in the early and late stages of ovarian cancer, to evaluate their regulation, and to determine their significance in an immunocompetent in vivo environment.     

Dietetics students perceive themselves as leaders and report they demonstrate leadership in a variety of contexts

A leadership survey was designed and administered to undergraduate dietetics students (n=283) at eight universities to examine leadership actions reported most frequently, the context of these leadership actions, and students' reported perceptions of themselves as leaders. The majority of students perceive themselves as leaders in all context areas. The leadership practice, "Enabling Others to Act," was the most frequently reported. There were no significant differences in leadership behaviors based on college classification status and supervisory experience. Leadership behaviors were more prevalent in students who had previous leadership course-work, were older, or who had previous leadership experience. Dietetics students perceive they demonstrate leadership and do so in a variety of contexts, most frequently in class. Therefore, classroom activities may help strengthen leadership abilities of future dietetics professionals.