Utilization of pyruvate and pyruvate precursors by normal and carcinogen-altered rat tracheal epithelial cells in culture

The metabolism of [14C]pyruvate, [14C]glucose, [14C]glutamine and [14C]alanine was compared between normal rat tracheal epithelial cells and carcinogen-altered cells derived from dimethylbenz(a)anthracene-exposed tracheal implants. Normal primary cultures (NPC) of tracheal cells are distinguished by their need for pyruvate-supplemented medium for growth and survival. The altered cells were selected out by their survival in the unsupplemented medium. Compared to the selected primary cultures (SPC), the NPC showed a three- to four-fold higher incorporation of radioactivity from [2-14C]pyruvate in all the macromolecular fractions, as well as in all the metabolites isolated from the acid soluble fraction and from lactic acid isolated from the medium. [U-14C]glucose was also incorporated at higher levels into lactic acid isolated from the acid soluble fraction and the medium of NPC. These data indicate a higher rate of glycolysis in the normal tracheal cells. This was supported by the findings of a two-fold greater glucose consumption and two-fold higher production of lactic acid isolated from the NPC medium. Lactate dehydrogenase activity was also two-fold higher in NPC. Thus, despite the apparently higher level of pyruvate production in the NPC, exogenous pyruvate is necessary to satisfy the metabolic needs of NPC. The utilization of [U-14C]glutamine or [U-14C]alanine was not markedly different between NPC and SPC. Furthermore, radioactivity from both of the amino acids was recovered in lactic acid in the medium, indicating that both cell types can derive pyruvic acid from either glutamine or alanine. SPC apparently do not use these routes to supply higher levels of pyruvic acid for survival in culture. The oxidation of none of the radioactive metabolites into CO2 was distinctly different between NPC and SPC except for the 1.7-fold higher utilization of [1-14C]glucose along the oxidative arm of the pentose cycle in the normal cells.     

Rheological properties of cross-linked hyaluronan-gelatin hydrogels for tissue engineering

Hydrogels that mimic the natural extracellular matrix (ECM) are used in three-dimensional cell culture, cell therapy, and tissue engineering. A semi-synthetic ECM based on cross-linked hyaluronana offers experimental control of both composition and gel stiffness. The mechanical properties of the ECM in part determine the ultimate cell phenotype. We now describe a rheological study of synthetic ECM hydrogels with storage shear moduli that span three orders of magnitude, from 11 to 3 500 Pa, a range important for engineering of soft tissues. The concentration of the chemically modified HA and the cross-linking density were the main determinants of gel stiffness. Increase in the ratio of thiol-modified gelatin reduced gel stiffness by diluting the effective concentration of the HA component.