Significance: Cardiac conduction disorders are potentially fatal illnesses caused by abnormalities in the heart’s electrical conduction system. Current treatments for these disorders, such as pacemakers, are effective but are bulky, rigid, and invasive. Here we develop a method to optically modulate cardiac beating frequency in primary cultured cardiomyocytes and adult rat hearts ex vivo, to a specified target frequency. Specifically, we use a low-irradiance moving laser stimulus and a biocompatible polymer–silicon nanowire composite material to achieve this modulation. This work has implications for future bioelectric studies of the cardiac conduction system as well as therapeutics for cardiac conduction disorders in the clinic.

In a previous study we had shown that membrane cholesterol removal induced unregulated lysosomal exocytosis events leading to the depletion of lysosomes located at cell periphery. However, the mechanism by which cholesterol triggered these exocytic events had not been uncovered. In this study we investigated the importance of cholesterol in controlling mechanical properties of cells and its connection with lysosomal exocytosis. Tether extraction with optical tweezers and defocusing microscopy were used to assess cell dynamics in mouse fibroblasts. These assays showed that bending modulus and surface tension increased when cholesterol was extracted from fibroblasts plasma membrane upon incubation with MβCD, and that the membrane-cytoskeleton relaxation time increased at the beginning of MβCD treatment and decreased at the end. We also showed for the first time that the amplitude of membrane-cytoskeleton fluctuation decreased during cholesterol sequestration, showing that these cells become stiffer. These changes in membrane dynamics involved not only rearrangement of the actin cytoskeleton, but also de novo actin polymerization and stress fiber formation through Rho activation. We found that these mechanical changes observed after cholesterol sequestration were involved in triggering lysosomal exocytosis. Exocytosis occurred even in the absence of the lysosomal calcium sensor synaptotagmin VII, and was associated with actin polymerization induced by MβCD. Notably, exocytosis triggered by cholesterol removal led to the secretion of a unique population of lysosomes, different from the pool mobilized by actin depolymerizing drugs such as Latrunculin-A. These data support the existence of at least two different pools of lysosomes with different exocytosis dynamics, one of which is directly mobilized for plasma membrane fusion after cholesterol removal.

Cholesterol regulates numerous cellular processes. Depleting its synthesis in skeletal myofibers induces vacuolization and contraction impairment. However, little is known about how cholesterol reduction affects cardiomyocyte behavior. Here, we deplete cholesterol by incubating neonatal cardiomyocytes with methyl-beta-cyclodextrin. Traction force microscopy shows that lowering cholesterol increases the rate of cell contraction and generates defects in cell relaxation. Cholesterol depletion also increases membrane tension, Ca²⁺ spikes frequency and intracellular Ca²⁺ concentration. These changes can be correlated with modifications in caveolin-3 and L-Type Ca²⁺ channel distributions across the sarcolemma. Channel regulation is also compromised since cAMP-dependent PKA activity is enhanced, increasing the probability of L-Type Ca²⁺ channel opening events. Immunofluorescence reveals that cholesterol depletion abrogates sarcomeric organization, changing spacing and alignment of α-actinin bands due to increase in proteolytic activity of calpain. We propose a mechanism in which cholesterol depletion triggers a signaling cascade, culminating with contraction impairment and myofibril disruption in cardiomyocytes.

Introduction: Metastasis is the main cause of cancer-associated death in colorectal cancer (CRC). The presence of circulating tumor cells (CTC) in the blood is associated with an increased risk of recurrence and poor prognosis. The clinical significance of CTCs as a novel biomarker has been extensively studied in the last decade. It has been shown that CTC detection applies to early cancer detection. The presence of CTCs is associated with metastatic spread and poor survival and is also useful as a marker for therapy response. Areas covered: We summarize the role of CTC in CRC, their clinical significance, current methods for CTC detection and challenges as well as future perspectives of CTC research. Expert commentary: The clinical significance of CTC in CRC patients is well established. Although insightful, the available marker-based approaches hampered our understanding of the CTCs and their biology, as such approaches undermined the heterogeneity of these cell populations. New technologies expand the marker-based detection to multi biomarker-based approaches together with recent technological advances in microfluidics for single cell enrichment and analysis.