Immobilisation of Lactate Oxidase and Deoxyribonuclease I for use within a Bio-Artificial Liver Assist Device for the Treatment of Acute Liver Failure

Constraints of cell supply indicate that proliferating cell lines are likely to be essential components of Bio-Artificial Liver support devices (BAL) for the treatment of acute liver failure. The Liver Group BAL employs clones of cells derived from the HepG2 cell line, which in common with many tumour derived cells, are predominantly dependent on anaerobic glycolysis for energy supply, leading to production of lactate within the bioreactor. The BAL system requires prolonged culture of alginate encapsulated HepG2 cells, and lactate accumulation presents a potential hazard in this system: at ~15 mM, accumulated lactate becomes toxic to the cells in the bioreactor, and also compromises alginate bead integrity by chelating the calcium ions necessary for alginate polymerisation. Furthermore, the tumour lineage of the cells could prove a potential threat to patient safety should any HepG2 DNA enter the patient’s system. It was hypothesised that inclusion of immobilised Lactate oxidase (LOx) to catalyse degradation of lactate into pyruvate could offset these limitations whilst simultaneously providing a potential energy source utilisable by HepG2 cells. In a similar fashion, immobilised Deoxyribonuclease I (DNase I) could be utilised to remove non-patient DNA during the treatment phase of the BAL system. Here it is demonstrated that functionalised glass beads are a feasible method of immobilising LOx and DNase I. Enzymatic activity was retained even after prolonged incubation at 37°C in the presence of human plasma, offering a means of reducing lactate levels during HepG2 culture, and potentially removing circulating DNA below practically detectable levels, thus facilitating cellular performance and BAL efficiency as a safe and effective potential therapy for acute liver failure

The Role of the BMP Signaling Antagonist Noggin in the Development of Prostate Cancer Osteolytic Bone Metastasis

Members of the BMP and Wnt protein families play a relevant role in physiologic and pathologic bone turnover. Extracellular antagonists are crucial for the modulation of their activity. Lack of expression of the BMP antagonist noggin by osteoinductive, carcinoma-derived cell lines is a determinant of the osteoblast response induced by their bone metastases. In contrast, osteolytic, carcinoma-derived cell lines express noggin constitutively. We hypothesized that cancer cell-derived noggin may contribute to the pathogenesis of osteolytic bone metastasis of solid cancers by repressing bone formation. Intra-osseous xenografts of PC-3 prostate cancer cells induced osteolytic lesions characterized not only by enhanced osteoclast-mediated bone resorption, but also by decreased osteoblast-mediated bone formation. Therefore, in this model, uncoupling of the bone remodeling process contributes to osteolysis. Bone formation was preserved in the osteolytic lesions induced by noggin-silenced PC-3 cells, suggesting that cancer cell-derived noggin interferes with physiologic bone coupling. Furthermore, intra-osseous tumor growth of noggin-silenced PC-3 cells was limited, most probably as a result of the persisting osteoblast activity. This investigation provides new evidence for a model of osteolytic bone metastasis where constitutive secretion of noggin by cancer cells mediates inhibition of bone formation, thereby preventing repair of osteolytic lesions generated by an excess of osteoclast-mediated bone resorption. Therefore, noggin suppression may be a novel strategy for the treatment of osteolytic bone metastases