Gelsolin regulates receptor-mediated and fluid-phase endocytosis in platelets.

BACKGROUND AND OBJECTIVE: Endocytosis is the process by which platelets incorporate extracellular molecules into their secretory granules. Endocytosis is mediated by the actin cytoskeleton in nucleated cells, however, the endocytic mechanisms in platelets are undefined. To better understand platelet endocytosis, we studied gelsolin (Gsn), an actin-severing protein that promotes actin assembly. METHODS: Mouse platelets from gelsolin-null (Gsn-/-) and wild-type (WT) controls were used. The uptake of fluorescent cargo molecules was compared as a measure of their endocytic efficiency. Receptor-mediated endocytosis was measured by the uptake of fibrinogen and transferrin; fluid-phase endocytosis was monitored by the uptake of fluorescent dextrans. RESULTS: ADP-stimulated WT platelets readily internalized both receptor-mediated and fluid-phase cargo. In contrast, Gsn-/- platelets showed a severe defect in the endocytosis of both types of cargo. The treatment of WT platelets with the actin-disrupting drugs cytochalasin D and jasplankinolide also reduced endocytosis. Notably, the individual and combined effects of Gsn deletion and drug treatment were similar for both receptor-mediated and fluid-phase endocytosis, indicating that Gsn mediates endocytosis via its action on the actin cytoskeleton. CONCLUSION: Our study demonstrates that Gsn plays a key role in the uptake of bioactive mediators by platelets.

CRISPR-Cas9 Gene Editing Protects from the A53T-SNCA Overexpression-Induced Pathology of Parkinson's Disease In Vivo.

Mutations in specific genes, including synuclein alpha (SNCA) that encodes the α-synuclein protein, are known to be risk factors for sporadic Parkinson's disease (PD), as well as critical factors for familial PD. In particular, A53T-mutated SNCA (A53T-SNCA) is a well-studied familial pathologic mutation in PD. However, techniques for deletion of the mutated SNCA gene in vivo have not been developed. Here, we used the CRISPR-Cas9 system to delete A53T-SNCA in vitro as well as in vivo. Adeno-associated virus carrying SaCas9-KKH with a single-guide RNA targeting A53T-SNCA significantly reduced A53T-SNCA expression levels in vitro. Furthermore, we tested its therapeutic potential in vivo in a viral A53T-SNCA-overexpressing rat model of PD. Gene deletion of A53T-SNCA significantly rescued the overexpression of α-synuclein, reactive microgliosis, dopaminergic neurodegeneration, and parkinsonian motor symptoms. Our findings propose CRISPR-Cas9 system as a potential prevention strategy for A53T-SNCA-specific PD.

Filamin A in platelets: Bridging the (signaling) gap between the plasma membrane and the actin cytoskeleton.

Platelets are anucleate cells that are essential for hemostasis and wound healing. Upon activation of the cell surface receptors by their corresponding extracellular ligands, platelets undergo rapid shape change driven by the actin cytoskeleton; this shape change reaction is modulated by a diverse array of actin-binding proteins. One actin-binding protein, filamin A (FLNA), cross-links and stabilizes subcortical actin filaments thus providing stability to the cell membrane. In addition, FLNA binds the intracellular portion of multiple cell surface receptors and acts as a critical intracellular signaling scaffold that integrates signals between the platelet's plasma membrane and the actin cytoskeleton. This mini-review summarizes how FLNA transduces critical cell signals to the platelet cytoskeleton.

The Effects of Educational Programs on Knowledge, International Normalized Ratio, Warfarin Adherence, and Warfarin-Related Complications in Patients Receiving Warfarin Therapy: An Integrative Review.

BACKGROUND: Education is considered a crucial element in anticoagulation management for patients with atrial fibrillation, valvular disease, ischemic heart disease, and venous thromboembolism. However, the effects of education on the patients prescribed warfarin are seldom investigated. OBJECTIVES: This integrative review was conducted to explore the effects of educational programs on patients prescribed warfarin for the aforementioned cardiovascular diseases and to identify the components of effective programs. METHODS: A systematic search of clinical trials was performed in 8 databases from inception to August 2020. Two reviewers performed the eligibility assessment, methodological evaluation, and data extraction. A total of 9 studies were included and analyzed via narrative synthesis. RESULTS: Nine studies involving a combined total of 1335 patients were included in the review. The findings suggest that educational programs have potential benefits related to international normalized ratio control and warfarin knowledge. However, their effects on major bleeding and thromboembolic events are unremarkable. Stronger evidence is recommended to confirm these findings, and the limited evidence examining the effects of education on warfarin adherence, minor bleeding, abnormal international normalized ratio, readmission rate, and warfarin-related mortality requires further exploration. Verbal education supported by written materials was the main educational delivery mode. A lecture length of approximately 45 minutes was likely appropriate. Notably, the integration of educational strategies, application of follow-up interventions and monitoring measures, adoption of psychological theories in program development, and inclusion of nurses or pharmacists in program conduction seemed to be effective program components. CONCLUSIONS: The effects of educational programs on patients prescribed warfarin mainly for atrial fibrillation, valvular disease, ischemic heart disease, and venous thromboembolism remain inconclusive. Further research using randomized controlled trials is warranted.

Near-infrared phototherapy for patient-derived orthotopic xenograft model of hepatocellular carcinoma in combination with indocyanine green.

BACKGROUND: Hepatocellular carcinoma notably takes up and retains indocyanine green (ICG). Here, we investigated whether patient-derived orthotopic xenograft of hepatocellular carcinoma could accumulate ICG and show full remission via phototherapy. METHODS: NIR light and ICG were tested for cytotoxicity in cancerous cell lines (Huh-7, Hep3B). Patient-derived orthotopic xenograft (PDoX) mice were subjected to phototherapy comprising of daily NIR exposure (0.5-1.75 W/cm2) and intravenous injection of ICG (5-20 mg/kg2). Moreover, NIR laser was flashed on individual mouse until hepatocellular carcinoma completely loss the fluorescence, as determined by NIR camera. RESULTS: Cytotoxicity increased in response to the input energy, but insufficient energy (< 150 joule/cm2) was irresponsive at all irradiances. NIR irradiance in the range of 0.5-1.75 W/cm2 took 5-7 days to elicit complete remission from PDoX mice in combination with 20 mg/kg ICG. In contrast, phototherapy could completely ablate hepatocellular carcinoma at 5-15 mg/kg ICG. CONCLUSIONS: ICG could potentiate the tumoricidal ability of NIR light in a dose-dependent manner, and vice versa. Regardless of ICG dosage, however, phototherapy treated group showed a relatively high survival rate compared to the non-treated group. Notably, real-time phototherapy could halve the effective ICG dosage for full remission of deep-seated tumor.

Free will: A case study in reconciling phenomenological philosophy with reductionist sciences.

Phenomenology aspires to philosophical analysis of humans' subjective experience while it strives to avoid pitfalls of subjectivity. The first step towards naturalizing phenomenology - making phenomenology scientific - is to reconcile phenomenology with modern physics, on the one hand, and with modern cellular and molecular neuroscience, on the other hand. In this paper, free will is chosen for a case study to demonstrate the feasibility. Special attention is paid to maintain analysis with mathematical precision, if possible, and to evade the inherent deceptive power of natural language. Laplace's determinism is re-evaluated along with the concept of microscopic reversibility. A simple and transparent version of proof demonstrates that microscopic reversibility is irreconcilably incompatible with macroscopic irreversibility, contrary to Boltzmann's claim. But the verdict also exalts Boltzmann's statistical mechanics to the new height of a genuine paradigm shift, thus cutting the umbilical cord linking it to Newtonian mechanics. Laplace's absolute determinism must then be replaced with a weaker form of causality called quasi-determinism. Biological indeterminism is also affirmed with numerous lines of evidence. The strongest evidence is furnished by ion channel fluctuations, which obey an indeterministic stochastic phenomenological law. Furthermore, quantum indeterminacy is shown to be relevant in biology, contrary to the opinion of Erwin Schrödinger. In reconciling phenomenology of free will with modern sciences, three issues - alternativism, intelligibility and origination - of free will must be accounted for. Alternativism and intelligibility can readily be accounted for by quasi-determinism. In order to account for origination of free will, the concept of downward causation must be invoked. However, unlike what is commonly believed, there is no evidence that downward causation can influence, shield off, or overpower low-level physical forces already known to physicists. Quasi-determinism offers an escape route: The possibility that downward causation arising from hierarchical organization of biological structures can modify dispersions of physical laws remains open. Empirical evidence in support of downward causation is scanty but nevertheless exists. Still, origination of free will must be considered an unsolved problem at present. It is demonstrated that objectivity does not guarantee scientific rigor in the study of complex phenomena, such as human creativity. In its replacement, universality and overall consistency between a theory and empirical evidence must be maintained. Visual thinking is proposed as a reasoning tool to ensure universality and overall consistency through inference to the best explanation.

The role of pattern recognition in creative problem solving: a case study in search of new mathematics for biology.

Rosen classified sciences into two categories: formalizable and unformalizable. Whereas formalizable sciences expressed in terms of mathematical theories were highly valued by Rutherford, Hutchins pointed out that unformalizable parts of soft sciences are of genuine interest and importance. Attempts to build mathematical theories for biology in the past century was met with modest and sporadic successes, and only in simple systems. In this article, a qualitative model of humans' high creativity is presented as a starting point to consider whether the gap between soft and hard sciences is bridgeable. Simonton's chance-configuration theory, which mimics the process of evolution, was modified and improved. By treating problem solving as a process of pattern recognition, the known dichotomy of visual thinking vs. verbal thinking can be recast in terms of analog pattern recognition (non-algorithmic process) and digital pattern recognition (algorithmic process), respectively. Additional concepts commonly encountered in computer science, operations research and artificial intelligence were also invoked: heuristic searching, parallel and sequential processing. The refurbished chance-configuration model is now capable of explaining several long-standing puzzles in human cognition: a) why novel discoveries often came without prior warning, b) why some creators had no ideas about the source of inspiration even after the fact, c) why some creators were consistently luckier than others, and, last but not least, d) why it was so difficult to explain what intuition, inspiration, insight, hunch, serendipity, etc. are all about. The predictive power of the present model was tested by means of resolving Zeno's paradox of Achilles and the Tortoise after one deliberately invoked visual thinking. Additional evidence of its predictive power must await future large-scale field studies. The analysis was further generalized to constructions of scientific theories in general. This approach is in line with Campbell's evolutionary epistemology. Instead of treating science as immutable Natural Laws, which already existed and which were just waiting to be discovered, scientific theories are regarded as humans' mental constructs, which must be invented to reconcile with observed natural phenomena. In this way, the pursuit of science is shifted from diligent and systematic (or random) searching for existing Natural Laws to firing up humans' imagination to comprehend Nature's behavioral pattern. The insights gained in understanding human creativity indicated that new mathematics that is capable of handling effectively parallel processing and human subjectivity is sorely needed. The past classification of formalizability vs. non-formalizability was made in reference to contemporary mathematics. Rosen's conclusion did not preclude future inventions of new biology-friendly mathematics.

Towards physical dynamic tolerance: an approach to resolve the conflict between free will and physical determinism.

This paper attempts to resolve the conflict between free will and determinism. The problem is approached by demonstrating that: (a) some well-established experimental observations indicate that irreversibility persists at the molecular level, (b) microscopic reversibility is not fully compatible with macroscopic irreversibility, (c) an overall consistency can be maintained if microscopic reversibility is regarded only as an excellent approximation, whereas microscopic irreversibility together with chaos can account for macroscopic reversibility, and (d) endogenous noise serves a vital function of nerve excitation. Thus, the mean of position and momentum specified by a non-deterministic law of motion gives the law its superficially deterministic behavior and predictability, whereas its dispersion grants dynamic tolerance and irreversibility. Therefore, causality is preserved while a limited degree of freedom allows for the exercise of free will. However, it is argued that free will can never be proven or disproven by a conventional behavioral experiment.