Epithelial cell adhesion to a planar substratum: Quantitative studies using a miniaturised parallel-plate shearing apparatus

The nature and quantitative aspects of epithelial cell adhesion are reviewed, and their relevance discussed in relation to the homeostasis of the dento-epithelial junction. The design, construction and testing of a miniaturised parallel-plate shearing apparatus for the measurement of cell adhesion based on the radial flow chamber principle of Fowler & McKay (1980) is described. Cultures of an established epithelial cell line on glass coverslips were exposed to flow conditions for varying times in the shearing chamber at 37°, 8° and 4°C, and subsequently photographed under standardized conditions. The critical shear radius (CSR) was determined by densitometry from a negative film and the minimum distraction force (MDF) at the CSR calculated using the measured flow rate and predetermined viscosity values of the medium. The calculated mean MDF values at 37°C ranged from 1.04 to 1.36 pascals, and was independent of the culture inoculation density (2.5 x 10 5 to 10 6 cells ml-1) and thetime (range: 5 to 20 min) of exposure to shearing conditions. Cell-to-cell adhesion in multilayer cultures was assessed by a cell-separation index (O) which represented the proportion of cells detached in a specific annulus per unit shearing force. The minimal force necessary to separate the upper layer(s) of cells was calculated to be significantly less than cell-to-substratum adhesion (MDF) being in the range of 0.43 to 0.64 Pa. Measurements of cell-to-substratum adhesion at 4°C demonstrated a three to four fold increase of the MDF (6.19 Pa) compared to that at 37°C (1.35 Pa). Part of this adhesion was resistant to mild trypsinisation and was stabilised by low temperature, and by treatment with concanavalin A or colchicine. A classification of cellular adhesion as trypsin-sensitive (TSA) and trypsin-resistant adhesion (TRA) on the basis of the different temperature and protease susceptibility is proposed with the corresponding physiological functions of "frictional" and "tractional" adhesion respectively. The implications of the dual adhesion hypothesis are discussed with respect to the integrity of the dento-gingival junction

Localization of type 1 diabetes susceptibility to the MHC class I genes HLA-B and HLA-A

The major histocompatibility complex (MHC) on chromosome 6 is associated with susceptibility to more common diseases than any other region of the human genome, including almost all disorders classified as autoimmune. In type 1 diabetes the major genetic susceptibility determinants have been mapped to the MHC class II genes HLA-DQB1 and HLA-DRB1 (refs 1-3), but these genes cannot completely explain the association between type 1 diabetes and the MHC region. Owing to the region's extreme gene density, the multiplicity of disease-associated alleles, strong associations between alleles, limited genotyping capability, and inadequate statistical approaches and sample sizes, which, and how many, loci within the MHC determine susceptibility remains unclear. Here, in several large type 1 diabetes data sets, we analyse a combined total of 1,729 polymorphisms, and apply statistical methods - recursive partitioning and regression - to pinpoint disease susceptibility to the MHC class I genes HLA-B and HLA-A (risk ratios >1.5; Pcombined = 2.01 × 10-19 and 2.35 × 10-13, respectively) in addition to the established associations of the MHC class II genes. Other loci with smaller and/or rarer effects might also be involved, but to find these, future searches must take into account both the HLA class II and class I genes and use even larger samples. Taken together with previous studies, we conclude that MHC-class-I-mediated events, principally involving HLA-B*39, contribute to the aetiology of type 1 diabetes. ©2007 Nature Publishing Group

New genetic loci link adipose and insulin biology to body fat distribution.

Body fat distribution is a heritable trait and a well-established predictor of adverse metabolic outcomes, independent of overall adiposity. To increase our understanding of the genetic basis of body fat distribution and its molecular links to cardiometabolic traits, here we conduct genome-wide association meta-analyses of traits related to waist and hip circumferences in up to 224,459 individuals. We identify 49 loci (33 new) associated with waist-to-hip ratio adjusted for body mass index (BMI), and an additional 19 loci newly associated with related waist and hip circumference measures (P < 5 × 10(-8)). In total, 20 of the 49 waist-to-hip ratio adjusted for BMI loci show significant sexual dimorphism, 19 of which display a stronger effect in women. The identified loci were enriched for genes expressed in adipose tissue and for putative regulatory elements in adipocytes. Pathway analyses implicated adipogenesis, angiogenesis, transcriptional regulation and insulin resistance as processes affecting fat distribution, providing insight into potential pathophysiological mechanisms

Critical research gaps and translational priorities for the successful prevention and treatment of breast cancer.

INTRODUCTION: Breast cancer remains a significant scientific, clinical and societal challenge. This gap analysis has reviewed and critically assessed enduring issues and new challenges emerging from recent research, and proposes strategies for translating solutions into practice. METHODS: More than 100 internationally recognised specialist breast cancer scientists, clinicians and healthcare professionals collaborated to address nine thematic areas: genetics, epigenetics and epidemiology; molecular pathology and cell biology; hormonal influences and endocrine therapy; imaging, detection and screening; current/novel therapies and biomarkers; drug resistance; metastasis, angiogenesis, circulating tumour cells, cancer 'stem' cells; risk and prevention; living with and managing breast cancer and its treatment. The groups developed summary papers through an iterative process which, following further appraisal from experts and patients, were melded into this summary account. RESULTS: The 10 major gaps identified were: (1) understanding the functions and contextual interactions of genetic and epigenetic changes in normal breast development and during malignant transformation; (2) how to implement sustainable lifestyle changes (diet, exercise and weight) and chemopreventive strategies; (3) the need for tailored screening approaches including clinically actionable tests; (4) enhancing knowledge of molecular drivers behind breast cancer subtypes, progression and metastasis; (5) understanding the molecular mechanisms of tumour heterogeneity, dormancy, de novo or acquired resistance and how to target key nodes in these dynamic processes; (6) developing validated markers for chemosensitivity and radiosensitivity; (7) understanding the optimal duration, sequencing and rational combinations of treatment for improved personalised therapy; (8) validating multimodality imaging biomarkers for minimally invasive diagnosis and monitoring of responses in primary and metastatic disease; (9) developing interventions and support to improve the survivorship experience; (10) a continuing need for clinical material for translational research derived from normal breast, blood, primary, relapsed, metastatic and drug-resistant cancers with expert bioinformatics support to maximise its utility. The proposed infrastructural enablers include enhanced resources to support clinically relevant in vitro and in vivo tumour models; improved access to appropriate, fully annotated clinical samples; extended biomarker discovery, validation and standardisation; and facilitated cross-discipline working. CONCLUSIONS: With resources to conduct further high-quality targeted research focusing on the gaps identified, increased knowledge translating into improved clinical care should be achievable within five years