Therapeutic Applications of Curcumin and Derivatives in Colorectal Cancer.

Curcumin (CUR), a natural phenolic compound, has been increasingly investigated in several malignancies due to its safe profile and ability to affect a wide range of oncogenic targets. With the ability to affect metastasis, apoptosis, and angiogenesis in colorectal cancer (CRC) and its tolerability at high doses, CUR is an attractive target for study. However, poor bioavailability and unfavorable pharmacokinetics and pharmacodynamics have hampered CUR's efficacy in clinical trials. Development of its derivatives and alternative delivery methods have shown the potential to overcome its inherent bioavailability issues. Recent analyses of various derivatives and nanoparticle encapsulation of CUR have demonstrated increased effectiveness in CRC studies. A major advantage of CUR has been its synergistic effects when used in combination with various chemotherapeutic agents. CUR offers a unique treatment option in terms of patient safety and its ability to be used in combination with current treatments for CRC. Further development of its derivatives and alternative delivery options offer potential new avenues of treatment that could outperform previous efforts to establish CUR as a CRC therapy.

TRPM7 is critical for short-term synaptic depression by regulating synaptic vesicle endocytosis.

Transient receptor potential melastatin 7 (TRPM7) contributes to a variety of physiological and pathological processes in many tissues and cells. With a widespread distribution in the nervous system, TRPM7 is involved in animal behaviors and neuronal death induced by ischemia. However, the physiological role of TRPM7 in central nervous system (CNS) neuron remains unclear. Here, we identify endocytic defects in neuroendocrine cells and neurons from TRPM7 knockout (KO) mice, indicating a role of TRPM7 in synaptic vesicle endocytosis. Our experiments further pinpoint the importance of TRPM7 as an ion channel in synaptic vesicle endocytosis. Ca2+ imaging detects a defect in presynaptic Ca2+ dynamics in TRPM7 KO neuron, suggesting an importance of Ca2+ influx via TRPM7 in synaptic vesicle endocytosis. Moreover, the short-term depression is enhanced in both excitatory and inhibitory synaptic transmissions from TRPM7 KO mice. Taken together, our data suggests that Ca2+ influx via TRPM7 may be critical for short-term plasticity of synaptic strength by regulating synaptic vesicle endocytosis in neurons.

Phosphatidylserine is critical for vesicle fission during clathrin-mediated endocytosis.

Phosphatidylserine (PS), a negatively charged phospholipid present predominantly at the inner leaflet of the plasma membrane, has been widely implicated in many cellular processes including membrane trafficking. Along this line, PS has been demonstrated to be important for endocytosis, however, the involved mechanisms remain uncertain. By monitoring clathrin-mediated endocytosis (CME) of single vesicles in mouse chromaffin cells using cell-attached capacitance measurements that offer millisecond time resolution, we demonstrate in the present study that the fission-pore duration is reduced by PS addition, indicating a stimulatory role of PS in regulating the dynamics of vesicle fission during CME. Furthermore, our results show that the PS-mediated effect on the fission-pore duration is Ca2+ -dependent and abolished in the absence of synaptotagmin 1 (Syt1), implying that Syt1 is necessary for the stimulatory role of PS in vesicle fission during CME. Consistently, a Syt1 mutant with a defective PS-Syt1 interaction increases the fission-pore duration. Taken together, our study suggests that PS-Syt1 interaction may be critical in regulating fission dynamics during CME.

The juxtamembrane region of synaptotagmin 1 interacts with dynamin 1 and regulates vesicle fission during compensatory endocytosis in endocrine cells.

Synaptotagmin 1 (Syt1) is a synaptic vesicle protein that is important for the kinetics of both exocytosis and endocytosis, and is thus a candidate molecule to link these two processes. Although the tandem Ca(2+)-binding C2 domains of Syt1 have important roles in exocytosis and endocytosis, the function of the conserved juxtamembrane (jxm) linker region has yet to be determined. We now demonstrate that the jxm region of Syt1 interacts directly with the pleckstrin homology (PH) domain of the endocytic protein dynamin 1. By using cell-attached capacitance recordings with millisecond time resolution to monitor clathrin-mediated endocytosis of single vesicles in neuroendocrine chromaffin cells, we find that loss of this interaction prolongs the lifetime of the fission pore leading to defects in the dynamics of vesicle fission. These results indicate a previously undescribed interaction between two major regulatory proteins in the secretory vesicle cycle and that this interaction regulates endocytosis.

Methods for cell-attached capacitance measurements in mouse adrenal chromaffin cell.

Neuronal transmission is an integral part of cellular communication within the brain. Depolarization of the presynaptic membrane leads to vesicle fusion known as exocytosis that mediates synaptic transmission. Subsequent retrieval of synaptic vesicles is necessary to generate new neurotransmitter-filled vesicles in a process identified as endocytosis. During exocytosis, fusing vesicle membranes will result in an increase in surface area and subsequent endocytosis results in a decrease in the surface area. Here, our lab demonstrates a basic introduction to cell-attached capacitance recordings of single endocytic events in the mouse adrenal chromaffin cell. This type of electrical recording is useful for high-resolution recordings of exocytosis and endocytosis at the single vesicle level. While this technique can detect both vesicle exocytosis and endocytosis, the focus of our lab is vesicle endocytosis. Moreover, this technique allows us to analyze the kinetics of single endocytic events. Here the methods for mouse adrenal gland tissue dissection, chromaffin cell culture, basic cell-attached techniques, and subsequent examples of individual traces measuring singular endocytic event are described.

Actin polymerization does not provide direct mechanical forces for vesicle fission during clathrin-mediated endocytosis.

Actin polymerization is important for vesicle fission during clathrin-mediated endocytosis (CME), and it has been proposed that actin polymerization may promote vesicle fission during CME by providing direct mechanical forces. However, there is no direct evidence in support of this hypothesis. In the present study, the role of actin polymerization in vesicle fission was tested by analyzing the kinetics of the endocytic tubular membrane neck (the fission-pore) with cell-attached capacitance measurements to detect CME of single vesicles in a millisecond time resolution in mouse chromaffin cells. Inhibition in dynamin GTPase activity increased the fission-pore conductance (Gp), supporting the mechanical role of dynamin GTPase in vesicle fission. However, disruptions in actin polymerization did not alter the fission-pore conductance Gp, thus arguing against the force-generating role of actin polymerization in vesicle fission during CME. Similar to disruptions of actin polymerization, cholesterol depletion results in an increase in the fission-pore duration, indicating a role for cholesterol-dependent membrane reorganization in vesicle fission. Further experiments suggested that actin polymerization and cholesterol might function in vesicle fission during CME in the same pathway. Our results thus support a model in which actin polymerization promotes vesicle fission during CME by inducing cholesterol-dependent membrane reorganization.

Synaptotagmin 1 is necessary for the Ca2+ dependence of clathrin-mediated endocytosis.

The role of Ca²âº in synaptic vesicle endocytosis remains uncertain due to the diversity in various preparations where several forms of endocytosis may contribute variably in different conditions. Although recent studies have demonstrated that Ca²âº is important for clathrin-mediated endocytosis (CME), the mechanistic role of Ca²âº in CME remains to be elucidated. By monitoring CME of single vesicles in mouse chromaffin cells with cell-attached capacitance measurements that offer millisecond time resolution, we demonstrate that the dynamics of vesicle fission during CME is Ca²âº dependent but becomes Ca²âº independent in synaptotagmin 1 (Syt1) knock-out cells. Our results thus suggest that Syt1 is necessary for the Ca²âº dependence of CME.