A resazurin-based,nondestructive assay for monitoring cell proliferation during a scaffold-based 3D culture process

Development of viable cell estimation method without sacrificing proliferation and functions of cells cultured on regenerative biomaterials is essential for regenerative engineering.Cytotoxicity and depletion of resazurin are critical but often overlooked limitations that hindered applications of resazurin in viable cell estimation.The present work found that cytotoxicity and depletion of resazurin depended on cell concentration,resazurin concentration and resazurin incubation time.A simple strategy which only allowed cells to incubate with resazurin during each measurement was developed to eliminate negative effects of resazurin.This strategy was verified by monitoring proliferation of MC3T3-E1 preosteoblasts on poly(D,L-lactic acid)scaffold during a continuous 3D culture process for up to 21 days,comparing the accuracy with MTT assay which is a destructive assay with high sensitivity and accuracy and commonly used in regenerative engineering and comparing viability,proliferation and differentiation functions of MC3T3-E1,which were treated with/without this strategy for nondestructive evaluation.This method showed comparable linearity of standard curve and characteristics of growth curve to MTT assay.No major negative effects of this method on MC3T3-E1 viability and functions were found.Our work highlighted the importance of the concentration and incubation time of resazurin in designing application-specific nondestructive viability assay and would be helpful in improving the implanted medical devices as well as in regenerative engineering.

Comparative study on hemodynamic environments around patient-specific carotid atherosclerotic plaques with different symmetrical features

Atherosclerotic plaque near the carotid sinus is a key risk factor for ischemic stroke.Although disturbed hemodynamic environments around atherosclerotic plaque had been investigated in many studies,the effect of their symmetrical features,especially longitudinal asymmetrical characteristics,had not been comparatively studied.In this study,three-dimensional carotid bifurcation models were established based on CT images of three patients with 50%stenosis and with different symmetrical features of carotid atherosclerotic plaques including concentric plaque,eccentric plaque,or eccentric longitudinal asymmetrical plaque.A healthy subject was chosen as the control.Wall shear stress(WSS)and oscillating shear index in the regions of upstream,downstream,downstream shoulders and regions around the point of maximum stenosis were analyzed.The results indicated that the maximum WSS around the eccentric longitudinal asymmetrical plaque were 8.6 times larger than that of the eccentric plaque and 1.9 times larger than that of the concentric plaque.And the distributions of WSS and OSI around the eccentric longitudinal asymmetric plaque were more disordered than those of the others.Our results highlighted the risk of the eccentric longitudinal asymmetrical plaque on plaque development and rupture and might provide a new index for evaluating the development of atherosclerosis.