Algorithm Optimization of non-DMSO Cryopreservation Protocols to Improve Mesenchymal Stem Cell Post-Thaw Function

University of Minnesota Ph.D. dissertation. September 2016. Major: Biomedical Engineering. Advisor: Allison Hubel. 1 computer file (PDF); xv, 172 pages.Mesenchymal stem cells (MSCs) are a common transfusion cell therapy that have been used in over 300 clinical trials to treat over 2000 patients with diseases ranging from Crohn’s disease to heart failure. These cells are frequently cryopreserved to better coordinate the timing of cell administration with patient care regimes and to accommodate transport of samples between different sites of collection, processing, and administration. However, cryopreservation with DMSO (the current gold standard) can result in poor cell function post-thaw and adverse reactions upon infusion. We hypothesize that non-DMSO cryopreservative molecules, including sugars, sugar alcohols, amino acids, and other small molecule additives, can be used in combination to protect cell viability and function post-thaw. This research demonstrates that some combinations of non-DMSO cryopreservatives preserve cell functionality better than others, and these effects are dependent not on osmotic or physical changes in solution, but on biological changes that affect the cell during the freezing process. We observe that there is likely a sweet spot concentration combination that produces maximum recovery for each combination of molecules, and demonstrate that an evolutionary algorithm can be used to identify optimized combinations of molecules that yield high cell recovery post-thaw. Additionally, we demonstrate that these novel solutions maintain MSC functionality when evaluated using surface markers, attachment, proliferation, actin alignment, RNA expression, and DNA hydroxymethylation. These advances in cryopreservation can improve cell therapy, and ultimately patient care

Regional Variation in the Density of Essential Genes in Mice

In most species, and particularly in vertebrates, the percentage of genes absolutely required for survival, the essential genes, has not been estimated. To obtain this estimation, we used the mouse as an experimental model to carry out high-efficiency N-ethyl-N-nitrosourea (ENU) mutagenesis screens in two balancer chromosome regions, and compared our results to a third previously published screen. The number of essential genes in each region was predicted based on allele frequencies. We determined that the density of essential genes differs by up to an order of magnitude among genomic regions. This indicates that extrapolating from regional estimates to genome-wide estimates of essential genes has a huge variance. A particularly high density of essential genes on mouse Chromosome 11 coincides with a high degree of regional linkage conservation, providing a possible causal explanation for the density variation. This is the first demonstration of regional variation in essential gene density in the mouse genome

DMSO-free methods of preserving mesenchymal stem cells (MSCs) that retain high levels of post thaw function

A novel, biologically-inspired strategy was developed to improve the preservation of mesenchymal stem cells (MSCs). MSCs are being investigated for the treatment of cardiovascular disorders, diabetes, connective tissue disorders, acute lung injury, amyotrophic lateral sclerosis, kidney diseases and more. To date, over 300 clinical trials involve the use of MSCs, with well over 2000 patients safely treated.Current methods of preserving MSCs are inadequate/ suboptimal. Concerns over poor post thaw function have become so pervasive that it is now common for MSCs to be cultured for 24-72 h prior to administration. These MSCs have a short shelf life (\u3c 24 hours), require special FDA permission, and the process increases cost and reduces access. The research described here utilizes an evolutionary algorithm to identify combinations of naturally occurring osmolytes that yield high cell recovery post thaw and optimize the composition of a DMSO-free, protein-free medium for cryopreservation of the cells. Additionally, we demonstrate that these novel solutions maintain MSC functionality when evaluated using surface markers, attachment, proliferation, actin alignment, RNA expression, and DNA hydroxymethlyation. Please click Additional Files below to see the full abstract

The Ten-Year Road: Joys and Challenges on the Road to Tenure

This paper explores the pre-tenure experiences of five assistant professors employed in the faculty of education of a research-intensive university. Acting as co-researchers, the authors researched their experiences through a critical narrative approach. The analysis, informed by critically-oriented writing that extends Wenger\u27s Communities of Practice, takes as axiomatic the notion that globalized processes of economic restructuring are mediating work in the academy and examines its local manifestations. Discussions explore issues of power, equity, shifting identities, and the need for improved navigational resources. The authors found that the process of critically and collaboratively researching their pre-tenure experiences offered insight into sites of personal and professional agency and also served as the impetus to form the social semiotic spaces that encouraged a sense of community. The Dean, a tenured member, but also a newcomer, serves in the role of critical friend

The Effect of Frequency of Fresh Pasture Allocation on the Feeding Behaviour of High Production Dairy Cows

Publication history: Accepted - 17 January 2022; Published - 20 January 2022.For ruminants, grazing and ruminating activities are essential in nutrient capture and ultimately animal performance however these activities can demand significant time and energy. This study evaluated the effect of three different pasture allocation frequencies (PAF’s; 12, 24 and 36 h) on the feeding behaviour of grazing dairy cows. Eighty-seven spring calving dairy cows were divided into three treatments. Animals were rotationally grazed with fixed paddock sizes of 0.14 ha, 0.28 ha and 0.42 ha paddocks for the 12 h, 24 h and 36 h treatments, respectively. Animals (14 per treatment) were fitted with behaviour halters that monitored feeding activity. Diurnal feeding patterns were evident for all animals irrespective of PAF, concentrating the majority of grazing during daytime (90%) and ruminating activity during night (73%). Treatment significantly affected feeding behavior patterns. Peak grazing activity coincided with fresh pasture allocation in the 12 h and 24 h treatments. In the 36 h treatment, grazing was more evenly distributed over each 24 h period with peak grazing activity witnessed daily between 17:00 and 19:00 regardless of fresh pasture allocation, suggesting lack of anticipation of fresh feed delivery. In the 12 h treatment primiparous animals exhibited greater grazing and ruminating activity relative to multiparous animals in the 12 h treatment highlighting the impact of competition for resources within each feed on lower dominance animals.This research was funded by AgriSearchNI grant number D-88-16. The APC was funded by the Agri-Food and Biosciences Institute

The Effect of Frequency of Fresh Pasture Allocation on Pasture Utilisation and the Performance of High Yielding Dairy Cows

Publication history: Accepted - 19 November 2020; Published - 21 November 2020.Pasture allocation frequency (PAF) can influence pasture availability and grazing behaviour, which subsequently may impact on animal performance. Limited research to-date has investigated grazing management methods to improve the performance of high production dairy cows whilst also achieving high grass utilisation rates. This study evaluated the e ect of three di erent PAF’s (12, 24 and 36 h) on pasture utilisation, the performance of high yielding dairy cows and the interaction with parity. The experiment included two 60-day periods, 90 spring calving dairy cows (27 primiparous animals) in period one and 87 (24 primiparous animals) in period two. The average pre-grazing sward height (11.4 cm) was similar for all treatments in both periods. In period one, pasture utilisation rate was significantly higher (8%) in the 36 h compared to the 12 h treatment. In period two, milk energy output was significantly greater for primiparous animals in the 36 h treatment relative to the other treatments.This research was funded by AgrisearchNI

Creating Livable Neighborhoods in Old Hill and Six Corners, Springfield, Massachusetts

This report documents the work of the Graduate Urban Design Studio in Springfield’s Old Hill and Six Corners neighborhoods with 12,000 inhabitants. Both neighborhoods are challenged by a very low average household income, housing foreclosures, lot vacancies, lack of substantial home ownership, high traffic volumes and poor quality of open space. The primary goal of the project was to develop a vision to renew Old Hill and Six Corners to improve livability for their residents and create new opportunities for housing, recreation, working and commercial activities. Crucial keys to attaining this goal will be creating a stronger sense of place throughout the neighborhood and improved connectivity to the existing public open spaces like the Watershops Pond and the proposed recreational trail “Arc of Recreation”. Four physical design objectives were explored: Creating a vision for livable streets - a more livable streetscape improves walkability, urban ecology, commercial activities and provides an enriching sensual experience. Creating a better physical environment for commercial and business activities – attractive streets create a new identity and support existing businesses. Creating and improving a pedestrian and bicycle oriented circulation and coherent public open space system. Exploring new housing opportunities - new sites for housing integrate the student population in the neighborhoods. We are proud that our proposals were recognized after a tornado hit Springfield in June 2011. REBUILDSPRINGFIELD targeted the tornado-impacted neighborhoods in a collaborative effort with engaged citizens and community groups. Our plans created a valuable planning and design framework to rebuild the City

Troubling meanings of family and competing moral imperatives in the family lives of young people with a parent who is at the end of life

This article draws on a narrative study of young people with a parent who is at the end of life to examine how family lives are troubled by life-limiting parental illness. Young people struggled to reconcile the physical and emotional absence of family members with meanings of ‘family’; the extent to which young people could rely on family to ‘be there’ in these troubling circumstances was of practical, emotional and moral significance. Our discussion is situated in the context of an English end of life care policy predicated on the ideal of a good death as one that takes place at home accompanied by family members. We explore how the shift away from family as a site for nurturing children towards family as a space to care for the dying is experienced by young people, and consider how these competing moral imperatives are negotiated through relational practices of care

Sequencing the genome of the Burmese python (Python molurus bivittatus) as a model for studying extreme adaptations in snakes

The Consortium for Snake Genomics is in the process of sequencing the genome and creating transcriptomic resources for the Burmese python. Here, we describe how this will be done, what analyses this work will include, and provide a timeline