'Cytology-on-a-chip' based sensors for monitoring of potentially malignant oral lesions.

UNLABELLED: Despite significant advances in surgical procedures and treatment, long-term prognosis for patients with oral cancer remains poor, with survival rates among the lowest of major cancers. Better methods are desperately needed to identify potential malignancies early when treatments are more effective. OBJECTIVE: To develop robust classification models from cytology-on-a-chip measurements that mirror diagnostic performance of gold standard approach involving tissue biopsy. MATERIALS AND METHODS: Measurements were recorded from 714 prospectively recruited patients with suspicious lesions across 6 diagnostic categories (each confirmed by tissue biopsy -histopathology) using a powerful new 'cytology-on-a-chip' approach capable of executing high content analysis at a single cell level. Over 200 cellular features related to biomarker expression, nuclear parameters and cellular morphology were recorded per cell. By cataloging an average of 2000 cells per patient, these efforts resulted in nearly 13 million indexed objects. RESULTS: Binary "low-risk"/"high-risk" models yielded AUC values of 0.88 and 0.84 for training and validation models, respectively, with an accompanying difference in sensitivity+specificity of 6.2%. In terms of accuracy, this model accurately predicted the correct diagnosis approximately 70% of the time, compared to the 69% initial agreement rate of the pool of expert pathologists. Key parameters identified in these models included cell circularity, Ki67 and EGFR expression, nuclear-cytoplasmic ratio, nuclear area, and cell area. CONCLUSIONS: This chip-based approach yields objective data that can be leveraged for diagnosis and management of patients with PMOL as well as uncovering new molecular-level insights behind cytological differences across the OED spectrum.

A cost-effectiveness analysis of rapid yeast detection kits.

OBJECTIVE: To determine the cost effectiveness of the utilization of over-the-counter yeast infection detection kits in the diagnosis of vaginal candidiasis. METHODS: A cost-benefit analysis based on a group of 70 adult women from a previous prospective study who presented with vaginitis symptoms. By constructing two decision trees, one in which the kits are an option to the women and one in which they are not, we predict the cost for diagnosing vaginal candidiasis in this group of women. RESULTS: For a group of 70 women presenting with vaginitis symptoms, the total cost of diagnosing their infections without the use of kits is predicted to be 7,051.10 dollars. For the same 70 women, the total of cost of diagnosing their infections with the use of kits is predicted to be 5,941.02 dollars. CONCLUSION: We conclude that the use of yeast infection detection kits could reduce the cost of diagnosis by 16%. The introduction of kits could save patients the time, money, and other resources involved in visiting a physician to confirm the diagnosis. Moreover, the sensitivity of yeast kits is superior to the traditional wet mount (77% vs. 52%), so there may be a role for the kits in the physician's office as well.

Calcineurin-dependent lytic granule exocytosis in NK-92 natural killer cells.

Cytotoxic T cells (CTLs) and natural killer cells (NKs) both kill virus-infected cells and tumor cells by releasing the cytoxic contents of their lytic granules. We recently demonstrated a role for calcineurin in lytic granule exocytosis in TALL-104 human leukemic CTLs [M.J. Grybko, J.P. Bartnik, G.A. Wurth, A.T. Pores-Fernando, A. Zweifach, Calcineurin activation is only one calcium-dependent step in cytotoxic T lymphocyte granule exocytosis, J. Biol. Chem. 282 (2007) 18009-18017]. However, whether calcineurin plays a similar role in NK lytic granule release is not known. We tested whether calcineurin is involved in lytic granule exocytosis in human leukemic NK-92 cells using immunosuppressive drugs that block calcineurin function and by overexpressing a constitutively active calcineurin fusion protein. Our results indicate that calcineurin does play a role in lytic granule exocytosis in NK-92 cells, and suggest that, as was the case in TALL-104 cells, there are likely to be multiple calcium-dependent steps.

ERK activation is only one role of PKC in TCR-independent cytotoxic T cell granule exocytosis.

Cytotoxic T cells (CTLs) kill target cells by releasing lytic agents via regulated exocytosis. Three signals are known to be required for exocytosis: an increase in intracellular Ca(2+), activation of protein kinase C (PKC) and activation of extracellular signal regulated signal kinase (ERK). ERK activation required for exocytosis depends on activity of PKC. The simplest possibility is that the sole effect of PKC required for exocytosis is ERK activation. Testing this requires dissociating ERK and PKC activation. We did this using TCR-independent stimulation of TALL-104 human leukemic CTLs. When cells are stimulated with thapsigargin and PMA, agents that increase intracellular Ca(2+) and activate PKC, respectively, PKC-dependent ERK activation is required for lytic granule exocytosis. Expressing a constitutively active mutant MAP kinase kinase activates ERK independent of PKC. However, activating ERK without PKC does not support lytic granule exocytosis, indicating that there are multiple effects of PKC required for granule exocytosis.