Cryobiology of Cell and Tissue Cryopreservation: Experimental and Theoretical Analysis

摘要

Preservation of tissue structure, morphology and biomarkers is of utmost importance for pathological examination of biopsy specimens for diagnostic and therapeutic purposes. However current methods employed to evade tissue degradation and preserve biomarkers have several shortcomings that include irreproducibility, morphological artifacts and altered biomarker antigenicity. These artifacts may affect the analysis and subsequent diagnosis of the tissue pathology. This creates need for developing improved preservation methods that reproducibly maintain tissue morphology and biomarker antigenicity and are simple, rapid and inexpensive. Experiments conducted for testing the hypothesis that cryopreservation procedures yield high quality morphology and antigenicity showed that cryopreservation maintains tissue structure, morphology and antigenicity at equivalent or better levels compared to standard freezing techniques. In order to understand the mechanisms of osmotic transport in cellular systems upon exposure to multi-component solutions that are prevalent in virtification protocols, experimental studies were undertaken using microfluidics for single cell manipulation. The experimental data yielded permeability parameters in binary and ternary solutions for MC3T3-E1 murine osteoblasts for the first time. The hydraulic conductivity (L(p)) decreased with increasing concentrations but the solute permeability either increased or decreased with increasing solution concentration. The changes in hydraulic conductivity were consistent with previously published trends and conform to a functional relationship in the form of Arrhenius type relationship between L(p) and solution concentration. Further a theoretical model was developed from principles of linear irreversible thermodynamics to simulate multi--‐‑component mass transport across membrane. The model was successfully validated by comparison with experimental data for murine osteoblasts and showed good agreement between the numerical predictions and experimental observations. The modeling approach can be used to investigate the transport mechanisms, which show that in multicomponent osmotic transport response, the dynamics is dictated by slower moving solute.