Trehalose Transporter from African Chironomid Larvae Improves Desiccation Tolerance of Chinese Hamster Ovary Cells

Dry preservation has been explored as an energy-efficient alternative to cryopreservation, but the high sensitivity of mammalian cells to desiccation stress has been one of the major hurdles in storing cells in the desiccated state. An important strategy to reduce desiccation sensitivity involves use of the disaccharide trehalose. Trehalose is known to improve desiccation tolerance in mammalian cells when present on both sides of the cell membrane. Because trehalose is membrane impermeant the development of desiccation strategies involving this promising sugar is hindered. We explored the potential of using a high-capacity trehalose transporter (TRET1) from the African chironomid Polypedilum vanderplanki[21] to introduce trehalose into the cytoplasm of mammalian cells and thereby increase desiccation tolerance. When Chinese hamster ovary cells (CHO) were stably transfected with TRET1 (CHO-TRET1 cells) and incubated with 0.4M trehalose for 4h at 37°C, a sevenfold increase in trehalose uptake was observed compared to the wild-type CHO cells. Following trehalose loading, desiccation tolerance was investigated by evaporative drying of cells at 14% relative humidity. After desiccation to 2.60g of water per gram dry weight, a 170% increase in viability and a 400% increase in growth (after 7days) was observed for CHO-TRET1 relative to control CHO cells. Our results demonstrate the beneficial effect of intracellular trehalose for imparting tolerance to partial desiccation

Osmotic Transport across Cell Membranes in Nondilute Solutions: A New Nondilute Solute Transport Equation

The fundamental physical mechanisms of water and solute transport across cell membranes have long been studied in the field of cell membrane biophysics. Cryobiology is a discipline that requires an understanding of osmotic transport across cell membranes under nondilute solution conditions, yet many of the currently-used transport formalisms make limiting dilute solution assumptions. While dilute solution assumptions are often appropriate under physiological conditions, they are rarely appropriate in cryobiology. The first objective of this article is to review commonly-used transport equations, and the explicit and implicit assumptions made when using the two-parameter and the Kedem-Katchalsky formalisms. The second objective of this article is to describe a set of transport equations that do not make the previous dilute solution or near-equilibrium assumptions. Specifically, a new nondilute solute transport equation is presented. Such nondilute equations are applicable to many fields including cryobiology where dilute solution conditions are not often met. An illustrative example is provided. Utilizing suitable transport equations that fit for two permeability coefficients, fits were as good as with the previous three-parameter model (which includes the reflection coefficient, σ). There is less unexpected concentration dependence with the nondilute transport equations, suggesting that some of the unexpected concentration dependence of permeability is due to the use of inappropriate transport equations

Spin-Drying technique for lyopreservation of mammalian cells.

Stabilization of cellular material in the presence of glass-forming sugars at ambient temperatures is a viable approach that has many potential advantages over current cryogenic strategies. Experimental evidence indicates the possibility to preserve biomolecules in glassy matrices of low-molecular mobility using ‘‘glass-forming’’ sugars like trehalose at ambient temperatures. However, when cells are desiccated in trehalose solution using passive drying techniques, a glassy skin is formed at the liquid/vapor interface of the sample. This glassy skin prevents desiccation of the sample beyond a certain level of dryness and induces non-uniformities in the final water content. Cells trapped underneath this glassy skin may degrade due to a relatively high molecular mobility in the sample. This undesirable result underscores the need for development of a uniform, fast drying technique. In the present study, we report a new technique based on the principles of ‘‘spin drying’’ that can effectively address these problems. Forced convective evaporation of water along with the loss of solution due to centrifugal force leads to rapid vitrification of a thin layer of trehalose containing medium that remains on top of cells attached to the spinning glass substrate. The glassy layer produced has a consistent thickness and a small ‘‘surfacearea-to-volume’’ ratio that minimizes any non-homogeneity. Thus, the chance of entrapping cells in a high-mobility environment decreases substantially. We compared numerical predictions to experimental observations of the drying time of 0.2–0.6 M trehalose solutions at a variety of spinning speeds ranging from 1000 to 4000 rpm. The model developed here predicts the formation of sugar films with thicknesses of 200–1000 nm, which was in good agreement with experimental results. Preliminary data suggest that after spin drying cells to about 0.159 ± 0.09 gH2O/gdw (n = 11, ±SE), more than 95% of cells were able to preserve their membrane

Principal hydraulic phenomena in high loaded radial flow hydraulic pumps.

O presente trabalho reúne elementos teóricos e experimentais envolvendo os principais fenômenos hidráulicos causadores de danos em bombas hidráulicas de fluxo radial de grande porte. A preocupação do texto está centrada em proporcionar um aprofundamento técnico acerca destes fenômenos hidráulicos de cavitação, a fim de contribuir na análise e solução de problemas em sistemas de bombeamento. Além disso, este trabalho tem a pretensão de transmitir alguma experiência prática fornecida por um evento em que foi detectada a ocorrência de dois destes fenômenos hidráulicos simultâneos em um sistema de bombeamento e a partir disso, realizou-se a análise completa deste evento, incluindo as propostas de solução aos problemas encontrados, as análises técnica e econômica das alternativas de solução e a análise após a implantação da proposta escolhida. O texto apresenta também uma grande quantidade de gráficos e figuras que buscam facilitar a compreensão das definições e das análises dos fenômenos hidráulicos, além de exemplificar os danos típicos dos fenômenos pela visualização direta destes danos nas bombas.This work presents theoretical and experimental data on the principal hydraulic phenomena responsible for damage in high loaded radial flow hydraulic pumps. The main objective is to provide a technical overview in order to contribute for the analysis and solution on these cavitation pumping systems problems. In addition this work intends to transmit practical experience through a “study of case" where two simultaneous of these hydraulic phenomena were detected. The complete analysis of technical and economical aspects of the alternatives solutions is presented including a great amount of graphs and illustrations that better explain the phenomena of cavitation in hydraulic pumps

Assessing opportunities and challenges for establishing a national program to distribute cord blood for research

Research is needed to enhance cord blood (CB) transplantation outcomes and to develop new clinical applications. Based on quality criteria for transplantation, CB collected by public CB banks (CBBs) is often unsuitable for banking, but may still be valuable for research. Canadian researchers have described a need for a centralized program providing ethically sourced CB for research projects. To meet this need, Canadian Blood Services (CBS), in partnership with The Ottawa Hospital, launched the Cord Blood for Research Program (CBRP) in 2014. The CBRP developed processes for donor research consent and research project approval with oversight from CBS's CBB and appropriate research ethics boards. The CBRP distributes deidentified CB products to research projects across Canada. Since its inception, the CBRP has distributed more than 525 CB units to researchers, supporting 11 research projects. Of the mothers who donate their baby's CB, 77% have chosen to consent to its use for research if it is not bankable. The number of CB units currently available for research via the CBRP exceeds the requests from researchers. The CBRP reliably distributes quality CB products that do not qualify for banking to investigators across Canada in an ethical, legal, and transparent manner. This provides an opportunity for the public to directly support research, helps meet the need expressed by Canada's research community, and maximizes the donor's gift. More research is needed to clarify the factors influencing donor and researcher participation in the CBRP

Obstetrical and neonatal factors associated with optimal public banking of umbilical cord blood in the context of delayed cord clamping

Purpose: To assess the association of specific newborn and maternal factors with indicators of increased blood-forming capacity in umbilical cord blood to inform strategic collection strategies that could augment the quality of units in public cord blood banks. Methods: Data regarding 268 consecutive cord blood units (CBUs) banked by Canadian Blood Services were analyzed. Multivariate analysis was performed to identify factors associated with markers of hematopoietic potency and likelihood of utilization. Results: Delayed clamping of the cord beyond 60 s was associated with reduced volume collected. Any delay in clamping of the cord was associated with reduced total nucleated cell counts. Newborn weight >4,000 g was also associated with greater blood volume in the collection but not with other measures of hematopoietic potency. Cord blood acidosis at birth (p