Mechanical Dissociation of Swine Liver to Produce Organoid Units for Tissue Engineering and In Vitro Disease Modeling
ABSTRACT The complex intricate architecture of the liver is crucial to hepatic function. Standard protocols used for enzymatic digestion to isolate hepatocytes destroy tissue structure and result in significant loss of synthetic, metabolic, and detoxification processes. We describe a process using mechanical dissociation to generate hepatic organoids with preserved intrinsic tissue architecture from swine liver. Oxygen-supplemented perfusion culture better preserved organoid viability, morphology, serum protein synthesis, and urea production, compared with standard and oxygen-supplemented static culture. Hepatic organoids offer an alternative source for hepatic assist devices, engineered liver, disease modeling, and xenobiotic testing.
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ABSTRACT: Many oxygen mass-transfer modeling studies have been performed for various bioartificial liver (BAL) encapsulation types; yet, to our knowledge, there is no experimental study that directly and noninvasively measures viability and metabolism as a function of time and oxygen concentration. We report the effect of oxygen concentration on viability and metabolism in a fluidized-bed NMR-compatible BAL using in vivo (31)P and (13)C NMR spectroscopy, respectively, by monitoring nucleotide triphosphate (NTP) and (13)C-labeled nutrient metabolites, respectively. Fluidized-bed bioreactors eliminate the potential channeling that occurs with packed-bed bioreactors and serve as an ideal experimental model for homogeneous oxygen distribution. Hepatocytes were electrostatically encapsulated in alginate (avg. diameter, 500 μm; 3.5×10(7) cells/mL) and perfused at 3 mL/min in a 9-cm (inner diameter) cylindrical glass NMR tube. Four oxygen treatments were tested and validated by an in-line oxygen electrode: (1) 95:5 oxygen:carbon dioxide (carbogen), (2) 75:20:5 nitrogen:oxygen:carbon dioxide, (3) 60:35:5 nitrogen:oxygen:carbon dioxide, and (4) 45:50:5 nitrogen:oxygen:carbon dioxide. With 20% oxygen, β-NTP steadily decreased until it was no longer detected at 11 h. The 35%, 50%, and 95% oxygen treatments resulted in steady β-NTP levels throughout the 28-h experimental period. For the 50% and 95% oxygen treatment, a (13)C NMR time course (∼5 h) revealed 2-(13)C-glycine and 2-(13)C-glucose to be incorporated into [2-(13)C-glycyl]glutathione (GSH) and 2-(13)C-lactate, respectively, with 95% having a lower rate of lactate formation. (31)P and (13)C NMR spectroscopy is a noninvasive method for determining viability and metabolic rates. Modifying tissue-engineered devices to be NMR compatible is a relatively easy and inexpensive process depending on the bioreactor shape.Tissue Engineering Part C Methods 07/2012; DOI:10.1089/ten.TEC.2011.0629 · 4.64 Impact Factor
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ABSTRACT: Cardiovascular disease is the leading cause of morbidity and mortality in the Western World. The inability of fully differentiated, load-bearing cardiovascular tissues to in vivo regenerate and the limitations of the current treatment therapies greatly motivate the efforts of cardiovascular tissue engineering to become an effective clinical strategy for injured heart and vessels. For the effective production of organized and functional cardiovascular engineered constructs in vitro, a suitable dynamic environment is essential, and can be achieved and maintained within bioreactors. Bioreactors are technological devices that, while monitoring and controlling the culture environment and stimulating the construct, attempt to mimic the physiological milieu. In this study, a review of the current state of the art of bioreactor solutions for cardiovascular tissue engineering is presented, with emphasis on bioreactors and biophysical stimuli adopted for investigating the mechanisms influencing cardiovascular tissue development, and for eventually generating suitable cardiovascular tissue replacements.09/2013; 4(3):329-70. DOI:10.1260/2040-2222.214.171.1249
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ABSTRACT: Nano-and microscale technologies have made a marked impact on the development of drug delivery systems. The loading efficiency and particle size of nano/micro particles can be better controlled with these new technologies than conventional methods. Moreover, drug delivery systems are moving from simple particles to smart particles and devices with programmable functions. These technologies are also contributing to in vitro and in vivo drug testing, which are important to evaluate drug delivery systems. For in vitro tests, lab-on-a-chip models are potentially useful as alternatives to animal models. For in vivo test, nano/micro-biosensors are developed for testing chemicals and biologics with high sensitivity and selectivity. Here, we review the recent development of nanoscale and microscale technologies in drug delivery including drug delivery systems, in vitro and in vivo tests.Journal of Mechanics in Medicine and Biology 04/2011; 21(11):337-367. DOI:10.1142/S021951941100406X · 0.80 Impact Factor