Search

Background aims This article describes the development of a small-scale model for Ficoll-based cell separation as part of process development of an advanced therapy medicinal product and its qualification. Because of the complexity of biological products, their manufacturing process as well as characterization and control needs to be accurately understood. Likewise, scale-down models serve as an indispensable tool for process development, characterization, optimization and validation. This scale-down model represents a cell processor device widely used in advance therapies. This approach is inteded to optimise resources and to focus its use on process characterisation studies under the paradigm of the Quality by design. A scale-down model should reflect the large manufacturing scale. Consequently, this simplified system should offer a high degree of control over the process parameters to depict a robust model, even considering the process limitations. For this reason, a model should be developed and qualified for the intended purpose. Methods Process operating parameters were studied, and their resulting performance at full scale was used as a baseline to guide scale-down model development. Once the model was established, comparability runs were performed by establishing standard operating conditions with bone marrow samples. These analyses showed consistency between the bench and the large scale. Additionally, statistical analyses were employed to demonstrate equivalence. Results The process performance indicators and assessed quality attributes were equivalent and fell into the acceptance ranges defined for the large-scale process. Conclusions This scale-down model is suitable for use in process characterization studies.

The beneficial effects of atorvastatin are based on both cholesterol-dependent and independent mechanisms. The latter probably include the ability of the estatin to enhance the expression of endothelial nitric oxide synthase (eNOS) and to cause a vasodilatation. In turn, the antioxidant and anti-inflammatory actions of melatonin are related to its vascular protection. In the present study, we investigated the efficacy of the combination of melatonin plus atorvastatin against endothelial cell damage induced by inflammation and oxidative stress injury. Human umbilical vein endothelial cells (HUVEC) were cultured with bacterial lipopolysaccharide (LPS) in the presence or absence of melatonin and/or atorvastatin. LPS inhibited eNOS mRNA and protein expression, which was reversed by atorvastatin and, to a lesser extent, by melatonin. Together, melatonin + atorvastatin induced higher eNOS protein expression than either compound alone. Melatonin, but not atorvastatin, reduced free radical generation, lipid peroxidation, and interleukin-6 levels induced by LPS. In the presence of atorvastatin, the effects of melatonin were maintained or even improved. These data suggest that melatonin improves the beneficial effects of atorvastatin and reduces its side effects in endothelial cells during inflammation and under conditions of oxidative stress.

The beneficial effects of atorvastatin are based on both cholesterol-dependent and independent mechanisms. The latter probably include the ability of the estatin to enhance the expression of endothelial nitric oxide synthase (eNOS) and to cause a vasodilatation. In turn, the antioxidant and anti-inflammatory actions of melatonin are related to its vascular protection. In the present study, we investigated the efficacy of the combination of melatonin plus atorvastatin against endothelial cell damage induced by inflammation and oxidative stress injury. Human umbilical vein endothelial cells (HUVEC) were cultured with bacterial lipopolysaccharide (LPS) in the presence or absence of melatonin and/or atorvastatin. LPS inhibited eNOS mRNA and protein expression, which was reversed by atorvastatin and, to a lesser extent, by melatonin. Together, melatonin + atorvastatin induced higher eNOS protein expression than either compound alone. Melatonin, but not atorvastatin, reduced free radical generation, lipid peroxidation, and interleukin-6 levels induced by LPS. In the presence of atorvastatin, the effects of melatonin were maintained or even improved. These data suggest that melatonin improves the beneficial effects of atorvastatin and reduces its side effects in endothelial cells during inflammation and under conditions of oxidative stress.