Developing AFM Techniques for Testing PEG Hydrogels

Many instruments are used to find elastic properties of biological samples using methods such as tensile and bending tests, but using the atomic force microscope (AFM) is considered a non-destructive method because it can provide repeated local stiffness information without damaging the sample. It additionally allows the sample to be tested in an aqueous environment, which is optimal for soft materials such as hydrogels. The nanoindentation is performed via cantilever, measuring the deflection of the cantilever during the contact of the sample using a laser. Compared to hard samples, testing soft materials can present more challenges when working with the AFM, creating the need for a refined technique.[1] This study will explore ways to improve the accuracy and feasibility of testing hydrogels, which are significant in biomaterials research as they offer the ability to be altered mechanically and chemically to fit the needs of cells.[2] The technique for testing the hydrogels will be refined through a process moving from dry to wet samples, attempting to repeatedly and successfully obtain topography and elastic properties through high resolution topography scans and force curves. References: Radmacher, M., Tillamnn, R.W., Fritz, M., and Gaub, H.E. (1992), From molecules to cells: imaging soft samples with the atomic force microscope. Science. 257: 1900-1905 Flake, M. M., Nguyen, P. K., Scott, R. A., Vandiver, L. R., Willits, R. K., and Elbert, D. L. Poly(ethylene glycol) microparticles produced by precipitation polymerization in aqueous solution. Biomacromolecules 12 (844-850). 2011

Statistical Issues in Assessing Hospital Performance

From the Preface: The Centers for Medicare and Medicaid Services (CMS), through a subcontract with Yale New Haven Health Services Corporation, Center for Outcomes Research and Evaluation (YNHHSC/CORE), is supporting a committee appointed by the Committee of Presidents of Statistical Societies (COPSS) to address statistical issues identified by the CMS and stakeholders about CMS’s approach to modeling hospital quality based on outcomes. In the spring of 2011, with the direct support of YNHHSC/ CORE, COPSS formed a committee comprised of one member from each of its constituent societies, a chair, and a staff member from the American Statistical Association, and held a preliminary meeting in April. In June, YNHHSC/CORE executed a subcontract with COPSS under its CMS contract to support the development of a White Paper on statistical modeling. Specifically, YNHHSC/CORE contracted with COPSS to “provide guidance on statistical approaches . . .when estimating performance metrics,” and “consider and discuss concerns commonly raised by stakeholders (hospitals, consumer, and insurers) about the use of “hierarchical generalized linear models in profiling hospital quality. The committee convened in June and August of 2011, and exchanged a wide variety of materials. To ensure the committee’s independence, YNHHSC/CORE did not comment on the white paper findings, and CMS pre-cleared COPSS’ publication of an academic manuscript based on the White Paper

Polyethlyene Glycol Microgels to Deliver Bioactive Nerve Growth Factor

Delivery of bioactive molecules is a critical step in fabricating materials for regenerative medicine, yet, this step is particularly challenging in hydrated scaffolds such as hydrogels. Although bulk photocrosslinked poly(ethylene glycol) (PEG) hydrogels have been used for a variety of tissue engineering applications, their capability as drug delivery scaffolds has been limited due to undesirable release profiles and reduction in bioactivity of molecules. To solve these problems, this article presents the fabrication of degradable PEG microgels, which are micron-sized spherical hydrogels, to deliver bioactive nerve growth factor (NGF). NGF release and activity was measured after encapsulation in microgels formed from either 3 kDa or 6 kDa PEG to determine the role of hydrogel mesh size on release. Microgels formed from 6 kDa PEG were statistically larger and had a higher swelling ratio than 3 kDa PEG. The 6 kDa PEG microgels provided a Fickian release with a reduced burst effect and 3 kDa microgels provided anomalous release over ≥20 days. Regardless of molecular weight of PEG, NGF bioactivity was not significantly reduced compared to unprocessed NGF. These results demonstrate that microgels provide easy mechanisms to control the release while retaining the activity of growth factors. As this microgel-based delivery system can be injected at the site of nerve injury to promote nerve repair, the potential to deliver active growth factors in a controlled manner may reduce healing time for neural tissue engineering applications

Dorsal root ganglia neurite outgrowth measured as a function of changes in microelectrode array resistance.

Current research in prosthetic device design aims to mimic natural movements using a feedback system that connects to the patient's own nerves to control the device. The first step in using neurons to control motion is to make and maintain contact between neurons and the feedback sensors. Therefore, the goal of this project was to determine if changes in electrode resistance could be detected when a neuron extended a neurite to contact a sensor. Dorsal root ganglia (DRG) were harvested from chick embryos and cultured on a collagen-coated carbon nanotube microelectrode array for two days. The DRG were seeded along one side of the array so the processes extended across the array, contacting about half of the electrodes. Electrode resistance was measured both prior to culture and after the two day culture period. Phase contrast images of the microelectrode array were taken after two days to visually determine which electrodes were in contact with one or more DRG neurite or tissue. Electrodes in contact with DRG neurites had an average change in resistance of 0.15 MΩ compared with the electrodes without DRG neurites. Using this method, we determined that resistance values can be used as a criterion for identifying electrodes in contact with a DRG neurite. These data are the foundation for future development of an autonomous feedback resistance measurement system to continuously monitor DRG neurite outgrowth at specific spatial locations

High resolution phase contrast photomicrograph of DRG tissue seeded on a microelectrode array after a 2 day culture.

<p>(A). 59 extracellular electrodes were visually classified as having contact with a DRG soma (B) or a DRG neurite (C). For some electrodes it was unclear if there was DRG neurite contact (D) and electrodes devoid of contact with DRG tissue were classified as CLEAN (E). The majority of electrodes classified as CLEAN were located on the opposite side of the array from where the DRG tissue was located (left side of A; scale bar = 200 μm).</p

Electrode resistance values can reflect DRG neurite contact with electrode.

<p>In a single experiment resistance values from electrodes were averaged by classification (DRG neurite, UNCLEAR, or CLEAN). The soma in this experiment was not placed in contact with any electrodes. (A) The change in resistance for electrodes in contact with a DRG neurite (-0.24 ± 0.02 MΩ; n = 16; p < 0.001; F = 69.685) were significantly less than for electrodes classified as UNCLEAR (-0.42 ± 0.02 MΩ; n = 19; p < 0.001) or CLEAN (-0.53 ± 0.01 MΩ; n = 31 electrodes). (B) The distribution of the change in electrode resistances for CLEAN electrodes, UNCLEAR electrodes and electrodes contacted by a DRG neurite were plotted in a frequency curve (0.5 MΩ bins). (C) The average change in resistance of UNCLEAR electrodes was background subtracted across all electrodes in a single experiment. This did not change the relationship between resistance values of electrodes contacted by DRG neurites and CLEAN electrodes. However it allowed for a comparison of resistance values across experiments.</p

The average change in resistance (MΩ) of each classified electrode after background subtraction for all six experiments (SEM in brackets).

<p>The average change in resistance (MΩ) of each classified electrode after background subtraction for all six experiments (SEM in brackets).</p

Thirteen new species of Chilecicada Sanborn, 2014 (Hemiptera: Auchenorrhyncha: Cicadidae: Tibicininae) expand the highly endemic cicada fauna of Chile

The genus Chilecicada Sanborn, 2014 is shown to be a complex of closely related species rather than a monospecific genus. Chilecicada citatatemporaria Sanborn & Cole n. sp., C. culenesensis Sanborn & Cole n. sp., C. curacaviensis Sanborn & Cole n. sp., C. impartemporaria Sanborn & Cole n. sp., C. magna Sanborn & Cole n. sp., C. mapuchensis Sanborn n. sp., C. oraria Sanborn & Cole n. sp., C. parrajaraorum Sanborn n. sp., C. partemporaria Sanborn & Cole n. sp., C. pehuenchesensis Sanborn & Cole n. sp., C. trifascia Sanborn n. sp., C. trifasciunca Sanborn & Cole n. sp., and C. viridicitata Sanborn & Cole n. sp. are described as new. Chilecicada occidentis Walker, 1850 is re-described to facilitate separation of the new species from the only previously known species. Song and cytochrome oxidase I analysis available for most species support the separation of the new taxa from the type species of the genus. Known species distributions and a key to the species of the genus are also provided. The new species increases the known cicada diversity 61.9% to 34 species, 91.2% of which are endemic to Chile