Tammy L. Oreskovic

Neurite outgrowth and differentiation of rat cortex progenitor cells are sensitive to lithium chloride at non-cytotoxic exposures.

Neuron-specific in vitro screening strategies have the potential to accelerate the evaluation of chemicals for neurotoxicity. We examined neurite outgrowth as a measure of neuronal response with a commercially available rat cortex progenitor cell model, where cells were exposed to a chemical during a period of cell differentiation. In control cultures, the fraction of beta-III-tubulin positive neurons and their neurite length increased significantly with time, indicating differentiation of the progenitor cells. Expression of glial fibrillary acidic protein, an astrocyte marker, also increased significantly with time. By seeding progenitor cells at varying densities, we demonstrated that neurite length was influenced by cell-cell spacing. After ten days, cultures seeded at densities of 1000 cells/mm(2) or lower had significantly shorter neurites than cultures seeded at densities of 1250 cells/mm(2) or higher. Progenitor cells were exposed to lithium, a neuroactive chemical with diverse modes of action. Cultures exposed to 30 mmol/L or 10 mmol/L lithium chloride (LiCl) had significantly lower metabolic activity than control cultures, as reported by adenosine triphosphate content, and no neurons were observed after ten days of exposure. Cultures exposed to 3 mmol/L, 1 mmol/L, or 0.3 mmol/L LiCl, which encompass lithiums therapeutic range, had metabolic activity similar to control cultures. These cultures exhibited concentration-dependent decreases in neurite outgrowth after ten days of LiCl exposure. Neurite outgrowth results were relatively robust, regardless of the evaluation methodology. This work demonstrates that measurement of neurite outgrowth in differentiating progenitor cell cultures can be a sensitive endpoint for neuronal response under non-cytotoxic exposure conditions.

Three-dimensional hydrogel constructs for exposing cells to nanoparticles

Abstract In evaluating nanoparticle risks to human health, there is often a disconnect between results obtained from in vitro toxicology studies and those from in vivo activity, prompting the need for improved methods to rapidly assess the hazards of engineered nanomaterials. In vitro studies of nanoparticle toxicology often rely on high doses and short exposure periods due to the difficulty of maintaining monolayer cell cultures over extended time periods as well as the difficulty of maintaining nanoparticle dispersions within the culture environment. In this work, tissue-engineered constructs are investigated as a platform for providing doses of nanoparticles over different exposure periods to cells within a three-dimensional environment that can be tuned to mimic in vivo conditions. Uptake of quantum dots (QDs) by model neural cells was first investigated in a high-dose exposure scenario, resulting in a strong concentration-dependent uptake of carboxyl-functionalised QDs. Poly(ethylene glycol) hydrogel scaffolds with varying mesh sizes were then investigated for their ability to support cell survival and proliferation. Cells were co-encapsulated with carboxyl-functionalised poly(ethylene glycol)-coated QDs at a lower dose than is typical for monolayer cultures. Although the QDs leach from the hydrogel within 24 h, they are also incorporated by cells within the scaffold, enabling the use of these constructs in future studies of cell behaviour and function.

Design and Characterization of a BioMEMS Device for In-Vitro Mechanical Stimulation of Single Adherent Cells

This paper presents development of a BioMEMS device to mechanically stimulate single adherent cells by means of electrostatic actuation. The main components of the proposed device include a platform for cell placement and an electrostatic comb drive actuator to provide in-plane motion. A high frequency actuation method was used to enable actuation in aqueous solutions. Displacements greater than 5μm were measured when the device was actuated with a 1 MHz square wave signal with 10V peak amplitude in DI water. Additionally, this device was successfully actuated in ionic solutions up to 50mM NaCl aqueous solution using frequencies greater than 30 MHz. Significant electrolysis and corrosion of the polysilicon and metal layers was observed when the devices were actuated in saline solutions with peak voltages greater than 15V, thus indicating that there is a limit on the maximum actuation voltage that can be used. No noticeable actuation was observed in phosphate buffer solution (PBS) or cell culture medium even when frequencies as high as 50 MHz were used due to ion migration. Theoretical calculations suggest that frequencies of the order of 100-500 MHz will be required for actuation in cell culture media. Currently we are in the process of building an experimental set-up to allow use of such high frequencies. Initial results for cell plating experiments on the cell stretcher platform and other considerations for device implementation are discussed in the end.Copyright

Analysis of ISCD-NIST Survey for Bone Health

In 2007, the National Institute of Standards and Technology and the International Society for Clinical Densitometry designed a survey to prioritize 7 research and standardization action items to improve accuracy and cross-comparability of dual-energy X-ray absorptiometry (DXA) measurements of bone mineral density. In this article, we analyze the 1074 survey responses as one means to define consensus priorities of the community studying bone health and to determine possible correlations between prioritization and demographic data, including geographic location, years of experience practicing DXA, and medical specialty. We find that the distribution of ranks from all respondents is such that we can conclude with statistical confidence that there are perceived distinctions between the relative merits of the 7 action items. Applying a standard vote-counting rule to the data, we determine a complete ranking of the action items. We observe that a consistent ranking of each action item across all demographic subcategories is hard to achieve. When we arrange the 7 action items into 4 groups, however, we can determine a reasonably consistent prioritization. The group containing the development of standard reference databases and phantoms receives the highest priority. In addition, we report consistent themes that emerge from the free-response portion of the survey.