Medium perfusion

Butler M, Imamura T, Thomas WG, Thilly M (1983), High yields from microcarrier cultures by medium perfusion, J. Cell. Sci. 61 351-363. 

Figure 1. Time course of medium glucose and lactate concentrations ofLLC-PKI ceiis under conventionai static conditions (A) and with continuous medium renewai (B). (A) LLC-PKi cells were grown to confluence and on 24 well plates, medium was removed from wells every two hours. Two medium replenishment cycles were conducted [r], (B) LLC-PK1 cells were cultured to confluence on micropourus filters and transferred to the EpiFlow perfusion device. Medium perfusion was 2 mi/h. Sampies in the outflowing medium werecoiiectedatreguiarintervais. Giucose and lactate were measured using colorimetric assays. Results represent the mean SEM from 3 experiments. For more information see Felder et. al. 2002 [72].

New cell culture techniques, which may improve the applicability of renal epithelial cultures, are also required. Currenfly there exist two commercially available cell culture perfusion systems, which allow the continuous perfusion of culture media and optimized oxygenation [243]. These systems allow stable longterm culture of quiescent adherent cells [244]. Continuous medium perfusion furthermore may lead to the re-expression of lost functions in continuous cell hues and the maintenance of differentiated properties in primary cells. Recently our laboratory has demonstrated that LLC-PKj cells maintained in a newly developed perfusion system (EpiFlow ) changed from a glycolytic to a more oxidative phenotype [72]. Evidence is also available from experiments in our laboratory that this mode of cultivation helps to prolong the lifetime of primary cultures of proximal tubular cells. Combining perfusion culture with co-culture of a cell type that is an anatomical neighbour in vivo (e.g. epithelial with endothelial, interstitial or immune cells) may improve the state of differentiation of both partner cells and increase the complexity of autocrine and paracrine interaction [73]. 

The Bellco dual overhead drive culture system uses a similar stirring system to the Techne and additionally permits continuous medium perfusion... 

Growth substrate characteristics influence epithelial cell orientation and phenotypic expression due to the interaction of receptor sites on the cell surface with specific sites in the substrate/matrix. For example, growth of the Xenopus laevis kidney cell hne A6 on microporous substrates could induce the expression of vasopressin receptors absent in solid support grown monolayers [157]. Similar observations have been reported when rabbit primary cultures of renal cortical collecting ducts were maintained on microporous supports under constant medium perfusion [158]. 

Grayson, W.L., Marolt, D., Bhumiratana, S., Frohhch, M., Guo, X.E., Vunjak-Novakovic, G., 2011. Optimizing the medium perfusion rale in bone tissue engineering bioreactors. Bio-technol. Bioeng. 108,1159-1170. 

Gingival atrophy, where the root is exposed leading to tooth sensitivities and caries, is a prevalent disorder that requires better graft options [109]. The Santerre group recently published that a D-PHl PU is compatible with HGFs cultured in a medium-perfused bioreactor [110], The coculture of HUVECs with HGFs was subsequently tested to see if HUVECs could vascularize the PU scaffolds. HUVEC clusters could be seen at the end of the 28-day test and the cocultures had modulated cytokine activities depending on the ratio of HUVEC HGF [105]. 

Mizuno, S., Allemann, R, and Glowacki, J. Effects of medium perfusion on matrix production by bovine chondrocytes in three-dimensional collagen sponges. /. Biomed. Mater. Res., 56,368,2001. 

Cartmell, S.H. et al. Effects of medium perfusion rate on cell seeded three-dimensional bone... 

Acidification of the medium perfusing the PG in vitro has little or no effect on the activities recorded from the CSN or the GPB. Recordings from cultured cat PG neurons show that local acidification, achieved by pressure pulse ejection of acidified (pH 6.5) medium from a micropipette, has no effect on the discharge of PG neurons. However, in most neurons (81%), aeidification blocked the action potentials evoked... 

Hosseinkhani, H., Inatsugu, Y, Hiraoka, Y, Inoue, S., Shimokawa, H., and Tabata, Y. 2005. Impregnation of plasmid Dna into three-dimensional scaffolds and medium perfusion enhance in vitro Dna expression of mesenchymal stem cells. Tissue Engineering, 11,1459-1475. 

Medium perfusion was employed to produce stress stimulation at the interior of the construct. In cultivating chondrocyte-seeded PGA scaffold, the perfusion system increased the amount of GAGs by 180% compared to the static conditions with a more organized and homogenous matrix (Pazzano et al. 2000). 

Radisic, M., L. Yang, J. Boublik, R. J. Cohen, R. Langer, L. E. Freed, and G. Vunjak-Novakovic. 2004. Medium perfusion enables engineering of compact and contractile cardiac tissue. Am J Physiol Heart Circ Physiol 286 H507-H516. 

Culture and differentiation of mESCs in a perfused 3D fibrous matrix has also been reported (Li et al., 2003). In this study, perfusion led to a higher growth rate and final cell density in relation to static conditions. A polyethylene terepthalate (PET) matrix was applied for construction of the scaffold, which provided a larger surface area for adhesion, growth, and reduced contact inhibition. A bioprocess for efficient ESC-derived cardiomyocyte production was also developed (Bauwens et al., 2005). This system was capable of monitoring and control oxygen tension and pH in 500-mL vessels with continuous medium perfusion. Oxygen tension was shown to he a culture parameter that can be manipulated to improve cardiomyocyte yield. 

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