Microfluidic characterization of single-cell biophysical properties and the applications in cancer diagnosis.

Single-cell biophysical properties play a crucial role in regulating cellular physiological states and functions, demonstrating significant potential in the fields of life sciences and clinical diagnostics. Therefore, over the last few decades, researchers have developed various detection tools to explore the relationship between the biophysical changes of biological cells and human diseases. With the rapid advancement of modern microfabrication technology, microfluidic devices have quickly emerged as a promising platform for single-cell analysis offering advantages including high-throughput, exceptional precision, and ease of manipulation. Consequently, this paper provides an overview of the recent advances in microfluidic analysis and detection systems for single-cell biophysical properties and their applications in the field of cancer. The working principles and latest research progress of single-cell biophysical property detection are first analyzed, highlighting the significance of electrical and mechanical properties. The development of data acquisition and processing methods for real-time, high-throughput, and practical applications are then discussed. Furthermore, the differences in biophysical properties between tumor and normal cells are outlined, illustrating the potential for utilizing single-cell biophysical properties for tumor cell identification, classification, and drug response assessment. Lastly, we summarize the limitations of existing microfluidic analysis and detection systems in single-cell biophysical properties, while also pointing out the prospects and future directions of their applications in cancer diagnosis and treatment.

A microfluidic generator of dynamic shear stress and biochemical signals based on autonomously oscillatory flow.

Biological cells in vivo typically reside in a dynamic flowing microenvironment with extensive biomechanical and biochemical cues varying in time and space. These dynamic biomechanical and biochemical signals together act to regulate cellular behaviors and functions. Microfluidic technology is an important experimental platform for mimicking extracellular flowing microenvironment in vitro. However, most existing microfluidic chips for generating dynamic shear stress and biochemical signals require expensive, large peripheral pumps and external control systems, unsuitable for being placed inside cell incubators to conduct cell biology experiments. This study has developed a microfluidic generator of dynamic shear stress and biochemical signals based on autonomously oscillatory flow. Further, based on the lumped-parameter and distributed-parameter models of multiscale fluid dynamics, the oscillatory flow field and the concentration field of biochemical factors has been simulated at the cell culture region within the designed microfluidic chip. Using the constructed experimental system, the feasibility of the designed microfluidic chip has been validated by simulating biochemical factors with red dye. The simulation results demonstrate that dynamic shear stress and biochemical signals with adjustable period and amplitude can be generated at the cell culture chamber within the microfluidic chip. The amplitudes of dynamic shear stress and biochemical signals is proportional to the pressure difference and inversely proportional to the flow resistance, while their periods are correlated positively with the flow capacity and the flow resistance. The experimental results reveal the feasibility of the designed microfluidic chip. Conclusively, the proposed microfluidic generator based on autonomously oscillatory flow can generate dynamic shear stress and biochemical signals without peripheral pumps and external control systems. In addition to reducing the experimental cost, due to the tiny volume, it is beneficial to be integrated into cell incubators for cell biology experiments. Thus, the proposed microfluidic chip provides a novel experimental platform for cell biology investigations.

A microfluidic system for precisely reproducing physiological blood pressure and wall shear stress to endothelial cells.

To reproduce hemodynamic stress microenvironments of endothelial cells in vitro is of vital significance, by which one could exploit the quantitative impact of hemodynamic stresses on endothelial function and seek innovative approaches to prevent circulatory system diseases. Although microfluidic technology has been regarded as an effective method to create physiological microenvironments, a microfluidic system to precisely reproduce physiological arterial hemodynamic stress microenvironments has not been reported yet. In this paper, a novel microfluidic chip consisting of a cell culture chamber with on-chip afterload components designed by the principle of input impedance to mimic the global hemodynamic behaviors is proposed. An external feedback control system is developed to accurately generate the input pressure waveform. A lumped parameter hemodynamic model (LPHM) is built to represent the input impedance to mimic the on-chip global hemodynamic behaviors. Sensitivity analysis of the model parameters is also elaborated. The performance of reproducing physiological blood pressure and wall shear stress is validated by both numerical characterization and flow experiment. Investigation of intracellular calcium ion dynamics in human umbilical vein endothelial cells is finally conducted to demonstrate the biological applicability of the proposed microfluidic system.

Multifunctional exosome-mimetics for targeted anti-glioblastoma therapy by manipulating protein corona.

Targeted drug delivery to the glioblastoma (GBM) overcoming blood-brain barrier (BBB) has been challenging. Exosomes are promising vehicles for brain tumor drug delivery, but the production and purification hinder its application for nanomedicine. Besides, the formation of protein corona (PC) may affect the behaviour of nanocarriers. Here, multifunctional exosomes-mimetics (EM) are developed and decorated with angiopep-2 (Ang) for enhancing GBM drug delivery by manipulating PC. Docetaxel (DTX)-loaded EM with Ang modification (DTX@Ang-EM) show less absorption of serum proteins and phagocytosis by macrophages. Ang-EM show enhanced BBB penetration ability and targeting ability to the GBM. Ang-EM-mediated delivery increase the concentration of DTX in the tumor area. The multifunctional DTX@Ang-EM exhibits significant inhibition effects on orthotopic GBM growth with reduced side effects of the chemotherapeutic. Findings from this study indicate that the developed DTX@Ang-EM provide a new strategy for targeted brain drug delivery and GBM therapy.

Artificial exosomes for translational nanomedicine.

Exosomes are lipid bilayer membrane vesicles and are emerging as competent nanocarriers for drug delivery. The clinical translation of exosomes faces many challenges such as massive production, standard isolation, drug loading, stability and quality control. In recent years, artificial exosomes are emerging based on nanobiotechnology to overcome the limitations of natural exosomes. Major types of artificial exosomes include 'nanovesicles (NVs)', 'exosome-mimetic (EM)' and 'hybrid exosomes (HEs)', which are obtained by top-down, bottom-up and biohybrid strategies, respectively. Artificial exosomes are powerful alternatives to natural exosomes for drug delivery. Here, we outline recent advances in artificial exosomes through nanobiotechnology and discuss their strengths, limitations and future perspectives. The development of artificial exosomes holds great values for translational nanomedicine.

Precise generation of dynamic biochemical signals by controlling the programmable pump in a Y-shaped microfluidic chip with a "christmas tree" inlet.

The generation of dynamic biochemical signals in a microfluidic control system is of importance for the study of the interaction between biological cells and their niches. However, most of microfluidic control systems are not able to provide dynamic biochemical signals with high precision and stability due to inherent mechanical vibrations caused by the actuators of the programmable pumps. In this paper, we propose a novel microfluidic feedback control system integrating an external feedback control system with a Y-shaped microfluidic chip with a "Christmas tree" inlet. The Proportional Integral Derivative (PID) controller is implemented to reduce the influence of vibrations. In order to regulate the control parameters efficiently, a mathematical model is built to describe the actuator of the programmable pump, in which a fractional-order model is utilized. Both simulation and experimental studies are carried out, confirming that the microfluidic feedback control system can precisely and stably generate desired dynamic biochemical signals.

Separation of micro and sub-micro diamagnetic particles in dual ferrofluid streams based on negative magnetophoresis.

In the present study, we numerically demonstrate an approach for separation of micro and sub-micro diamagnetic particles in dual ferrofluid streams based on negative magnetophoresis. The dual streams are constructed by an intermediate sheath flow, after which the negative magnetophoretic force induced by an array of permanent magnets dominates the separation of diamagnetic particles. A simple and efficient numerical model is developed to calculate the motions of particles under the action of magnetic field and flow field. Effects of the average flow velocity, the ratio of sheath fluid flow to sample fluid flow, the number of the magnet pair as well as the position of magnet pair are investigated. The optimal parametric condition for complete separation is obtained through the parametric analysis, and the separation principle is further elucidated by the force analysis. The separation of smaller micro and sub-micro diamagnetic particles is finally demonstrated. This study provides an insight into the negative magnetophoretic phenomenon and guides the fabrication of feasible, low-cost diagnostic devices for sub-micro particle separation.

Antineuroinflammatory therapy: potential treatment for autism spectrum disorder by inhibiting glial activation and restoring synaptic function.

Autism spectrum disorder (ASD) is a neurodevelopmental disorder that is characterized by deficits in social interactions and perseverative and stereotypical behavior. Growing evidence points toward a critical role for synaptic dysfunction in the onset of ASD, and synaptic function is influenced by glial cells. Considering the evidence that neuroinflammation in ASD is mediated by glial cells, one hypothesis is that reactive glial cells, under inflammatory conditions, contribute to the loss of synaptic functions and trigger ASD. Ongoing pharmacological treatments for ASD, including oxytocin, vitamin D, sulforaphane, and resveratrol, are promising and are shown to lead to improvements in behavioral performance in ASD. More importantly, their pharmacological mechanisms are closely related to anti-inflammation and synaptic protection. We focus this review on the hypothesis that synaptic dysfunction caused by reactive glial cells would lead to ASD, and discuss the potentials of antineuroinflammatory therapy for ASD.

Transport of dynamic biochemical signals in a microfluidic single cell trapping channel with varying cross-sections.

Dynamic biochemical signal control in vitro is important in the study of cellular responses to dynamic biochemical stimuli in microenvironment in vivo. To this end, we designed a microfluidic single cell trapping channel with varying cross-sections. In this work, we analyzed the transport of dynamic biochemical signals in steady and non-reversing pulsatile flows in such a microchannel. By numerically solving the 2D time-dependent Taylor-Aris dispersion equation, we studied the transport mechanism of different signals with varying parameters. The amplitude spectrum in steady flow shows that the trapping microchannel acts as a low-pass filter due to the longitudinal dispersion. The input signal can be modulated nonlinearly by the pulsatile flow. In addition, the nonlinear modulation effects are affected by the pulsatile flow frequency, the pulsatile flow amplitude and the average flow rate. When the flow frequency is much smaller or larger than that of the biochemical signal, the signal can be transmitted more efficiently. Besides, smaller pulsatile flow amplitude and larger average flow rate can decrease the nonlinear modulation and promote the signal transmission. These results demonstrate that in order to accurately load a desired dynamic biochemical signal to the trapped cell to probe the cellular dynamic response to the dynamic biochemical stimulus, the transport mechanism of the signals in the microchannel should be carefully considered.

Association between parental body mass index and autism spectrum disorder: a systematic review and meta-analysis.

Studies have examined the association between parental body mass index (BMI) and autism spectrum disorder (ASD) in offspring, with inconsistent results, especially regarding maternal obesity, overweight and underweight. Cochrane Library, EMBASE, PubMed and PsycINFO databases were searched up to March 2018 for relevant observational studies with no language restriction. Our literature search identified 13 eligible studies for meta-analysis (involving 943,293 children and 30,337 cases). For maternal BMI (13 studies), both maternal obesity [OR 1.41 (95% CI 1.19-1.67)] and maternal overweight [OR 1.16 (95% CI 1.05-1.27)] were significantly associated with ASD, while maternal underweight was not associated with ASD [OR 1.08 (95% CI 0.98-1.20)]. For paternal BMI (three studies), no association was found (paternal obesity: OR 1.28, 95% CI 0.94-1.74; overweight: OR 1.07, 95% CI 0.99-1.15; underweight: OR 1.12, 95% CI 0.87-1.44). Pooled estimates were robust in sensitivity analysis and subgroup analyses. Publication bias may exist for studies assessing maternal BMI and ASD risk, but the filled estimates were not altered. Relative to normal weight, maternal obesity and overweight were significantly associated with increased ASD risk, while maternal underweight was not associated with ASD. Although no association between paternal BMI and ASD was found, current evidence is limited (three studies). Future studies are warranted to address more confounding factors and to identify potential mediators of the association, but pre-pregnancy weight control is suggested.

Supplement intervention associated with nutritional deficiencies in autism spectrum disorders: a systematic review.

PURPOSE: Nutritional supplements have been used for correction of deficiencies that may occur in patient with autism spectrum disorder (ASD) and to improve core symptoms. We aim to provide current best evidence about supplements for nutritional deficiencies and core symptoms in children with ASD and to evaluate the effectiveness and safety. METHODS: A systematic literature search of scientific databases was performed to retrieve relevant randomized controlled trials. Risk of bias was assessed for each study. RESULTS: 18 randomized controlled trials of five supplements were included. B6/Mg was not helpful for improving ASD symptoms (seven RCTs). Two RCTs of methyl B12 reported some improvement in ASD severity but the effects on the correction of deficiencies were inconclusive. Two RCTs of vitamin D3 both reported increased levels of mean 25(OH)D in serum but inconsistent results in behavioral outcomes. Omega-3 fatty acid supplementation did not affect ASD behaviors but may correct deficiencies (six RCTs). One RCT of folinic acid reported positive results in improving ASD symptoms measured by various behavioral scales. CONCLUSIONS: Current evidence for the use of supplements for correcting nutritional deficiencies in children with ASD and to improve the symptoms is little. More studies are needed.

A microemulsion of puerarin-phospholipid complex for improving bioavailability: preparation, in vitro and in vivo evaluations.

Puerarin is a phytochemical with various pharmacological effects, but poor water solubility and low oral bioavailability limited usage of puerarin. The purpose of this study was to develop a new microemulsion (ME) based on phospholipid complex technique to improve the oral bioavailability of puerarin. Puerarin phospholipid complex (PPC) was prepared by a solvent evaporation method and was characterized by X-ray diffraction and infrared spectroscopy. Pseudo-ternary phase diagrams were constructed to investigate the effects of different oil on the emulsifying performance of the blank ME. Intestinal mucosal injury test was conducted to evaluate safety of PPC-ME, and no sign of damage on duodenum, jejunum and ileum of rats was observed using hematoxylin-eosin staining. In pharmacokinetic study of PPC-ME, a significantly greater Cmax (1.33 µg/mL) was observed when compared to puerarin (Cmax 0.55 µg/mL) or PPC (Cmax 0.70 µg/mL); the relative oral bioavailability of PPC-ME was 3.16-fold higher than puerarin. In conclusion, the ME combined with the phospholipid complex technique was a promising strategy to enhance the oral bioavailability of puerarin.

Hemodynamics of ventricular-arterial coupling under enhanced external counterpulsation: An optimized dual-source lumped parameter model.

BACKGROUND AND OBJECTIVE: Enhanced external counterpulsation (EECP) is a mechanically assisted circulation technique widely used in the rehabilitation and management of ischemic cardiovascular diseases. It contributes to cardiovascular functions by regulating the afterload of ventricle to improve hemodynamic effects, including increased diastolic blood pressure at aortic root, increased cardiac output and enhanced blood perfusion to multiple organs including coronary circulation. However, the effects of EECP on the coupling of the ventricle and the arterial system, termed ventricular-arterial coupling (VAC), remain elusive. We aimed to investigate the acute effect of EECP on the dynamic interaction between the left ventricle and its afterload of the arterial system from the perspective of ventricular output work. METHODS: A neural network assisted optimization algorithm was proposed to identify the ordinary differential equation (ODE) relation between aortic root blood pressure and flow rate. Based on the optimized order of ODE, a lumped parameter model (LPM) under EECP was developed taking into consideration of the simultaneous action of cardiac and EECP pressure sources. The ventricular output work, in terms of aortic pressure and flow rate cooperated with the LPM, was used to characterize the VAC of ventricle and its afterload. The VAC subjected to the principle of minimal ventricular output work was validated by solving the Euler-Poisson equation of cost function, ultimately determining the waveforms of aortic pressure and flow rate. RESULTS: A third-order ODE can precisely describe the hemodynamic relationship between aortic pressure and flow rate. An optimized dual-source LPM with three energy-storage elements has been constructed, showing the potential in probing VAC under EECP. The LPM simulation results demonstrated that the VAC in terms of aortic pressure and flow rate yielded to the minimal ventricular output work under different EECP pressures. CONCLUSIONS: The ventricular-arterial coupling under EECP is subjected to the minimal ventricular output work, which can serve as a criterion for determining aortic pressure and flow rate. This study provides insight for the understanding of VAC and has the potential in characterizing the performance of the ventricular and arterial system under EECP.

Biomimetic nanodecoys deliver cholesterol-modified heteroduplex oligonucleotide to target dopaminergic neurons for the treatment of Parkinson's disease.

Parkinson's disease (PD) is the second most common neurodegenerative disorder characterized by the accumulation of α-synuclein (α-syn) aggregates called Lewy bodies leading to the gradual loss of dopaminergic (DA) neurons in the substantia nigra. Although α-syn expression can be attenuated by antisense oligonucleotides (ASOs) and heteroduplex oligonucleotide (HDO) by intracerebroventricular (ICV) injection, the challenge to peripheral targeted delivery of oligonucleotide safely and effectively into DA neurons remains unresolved. Here, we designed a new DNA/DNA double-stranded (complementary DNA, coDNA) molecule with cholesterol conjugation (Chol-HDO (coDNA)) based on an α-syn-ASO sequence and evaluated its silence efficiency. Further, Chol-HDO@LMNPs, Chol-HDO-loaded, cerebrovascular endothelial cell membrane with DSPE-PEG2000-levodopa modification (L-DOPA-CECm)-coated nanoparticles (NPs), were developed for the targeted treatment of PD by tail intravenous injection. CECm facilitated the blood-brain barrier (BBB) penetration of NPs, together with cholesterol escaped from reticuloendothelial system uptake, as well as L-DOPA was decarboxylated into dopamine which promoted the NPs toward the PD site for DA neuron regeneration. The behavioral tests demonstrated that the nanodecoys improved the efficacy of HDO on PD mice. These findings provide insights into the development of biomimetic nanodecoys loading HDO for precise therapy of PD. STATEMENT OF SIGNIFICANCE: The accumulation of α-synuclein (α-syn) aggregates is a hallmark of PD. Our previous study designed a specific antisense oligonucleotide (ASO) targeting human SNCA, but the traumatic intracerebroventricular (ICV) is not conducive to clinical application. Here, we further optimize the ASO by creating a DNA/DNA double-stranded molecule with cholesterol-conjugated, named Chol-HDO (coDNA), and develop a DA-targeted biomimetic nanodecoy Chol-HDO@LMNPs by engineering cerebrovascular endothelial cells membranes (CECm) with DSPE-PEG2000 and L-DOPA. The in vivo results demonstrated that tail vein injection of Chol-HDO@LMNPs could target DA neurons in the brain and ameliorate motor deficits in a PD mouse model. This investigation provides a promising peripheral delivery platform of L-DOPA-CECm nanodecoy loaded with a new Chol-HDO (coDNA) targeting DA neurons in PD therapy.

Transport of dynamic biochemical signals in steady flow in a shallow Y-shaped microfluidic channel: effect of transverse diffusion and longitudinal dispersion.

Dynamic biochemical signal control is important in in vitro cell studies. This work analyzes the transportation of dynamic biochemical signals in steady and mixing flow in a shallow, Y-shaped microfluidic channel. The characteristics of transportation of different signals are investigated, and the combined effect of transverse diffusion and longitudinal dispersion is studied. A method is presented to control the widths of two steady flows in the mixing channel from two inlets. The transfer function and the cutoff frequency of the mixing channel as a transmission system are presented by analytically solving the governing equations for the time-dependent Taylor-Aris dispersion and molecular diffusion. The amplitude and phase spectra show that the mixing Y-shaped microfluidic channel acts as a low-pass filter due to the longitudinal dispersion. With transverse molecular diffusion, the magnitudes of the output dynamic signal are reduced compared to those without transverse molecular diffusion. The inverse problem of signal transportation for signal control is also solved and analyzed.

A mathematical model for intracellular NO and ROS dynamics in vascular endothelial cells activated by exercise-induced wall shear stress.

Vascular endothelial cells (ECs) residing in the innermost layer of blood vessels are exposed to dynamic wall shear stress (WSS) induced by blood flow. The intracellular nitric oxide (NO) and reactive oxygen species (ROS) in ECs modulated by the dynamic WSS play important roles in endothelial functions. Mathematical modeling is a popular methodology for biophysical studies. It can not only explain existing cell experiments, but also reveal the underlying mechanism. However, the previous mathematical models of NO dynamics in ECs are limited to the static WSS induced by constant flow, while arterial blood flow is a periodic pulsatile flow with varying amplitude and frequency at different exercise intensities. In this study, a mathematical model of intracellular NO and ROS dynamics activated by dynamic WSS based on the in vitro cell experiments is developed. With the hypothesis of the viscoelastic body, the Kelvin model is adopted to simulate the mechanosensors on EC. Thus, the NO dynamics activated by dynamic shear stresses induced by constant flow, pulsatile flow, and oscillatory flow are analyzed and compared. Moreover, the roles of ROS have been considered for the first time in the modeling of NO dynamics in ECs based on the analysis of cell experiments. The predictions of the proposed model coincide fairly well with the experimental data when ECs are subjected to exercise-induced WSS. The mechanism is elucidated that WSS induced by moderate-intensity exercise is most favorable to NO production in ECs. This study can provide valuable insights for further study of NO and ROS dynamics in ECs and help develop appropriate exercise regimens for improving endothelial functions.

Fabricating a multi-component microfluidic system for exercise-induced endothelial cell mechanobiology guided by hemodynamic similarity.

Generating precise in vivo arterial endothelial hemodynamic microenvironments using microfluidics is essential for exploring endothelial mechanobiology. However, a hemodynamic principle guiding the fabrication of microfluidic systems is still lacking. We propose a hemodynamic similarity principle for quickly obtaining the input impedance of the microfluidic system in vitro derived from that of the arterial system in vivo to precisely generate the desired endothelial hemodynamic microenvironments. First, based on the equivalent of blood pressure (BP) and wall shear stress (WSS) waveforms, we establish a hemodynamic similarity principle to efficiently map the input impedance in vivo to that in vitro, after which the multi-component microfluidic system is designed and fabricated using a lumped parameter hemodynamic model. Second, numerical simulation and experimental studies are carried out to validate the performance of the designed microfluidic system. Finally, the intracellular Ca2+ responses after exposure to different intensities of exercise-induced BP and WSS waveforms are measured to improve the reliability of EC mechanobiological studies using the designed microfluidic system. Overall, the proposed hemodynamic similarity principle can guide the fabrication of a multi-component microfluidic system for endothelial cell mechanobiology.

Recent progress of macrophage vesicle-based drug delivery systems.

Nanoparticle drug delivery systems (NDDSs) are promising platforms for efficient delivery of drugs. In the past decades, many nanomedicines have received clinical approval and completed translation. With the rapid advance of nanobiotechnology, natural vectors are emerging as novel strategies to carry and delivery nanoparticles and drugs for biomedical applications. Among diverse types of cells, macrophage is of great interest for their essential roles in inflammatory and immune responses. Macrophage-derived vesicles (MVs), including exosomes, microvesicles, and those from reconstructed membranes, may inherit the chemotactic migration ability and high biocompatibility. The unique properties of MVs make them competing candidates as novel drug delivery systems for precision nanomedicine. In this review, the advantages and disadvantages of existing NDDSs and MV-based drug delivery systems (MVDDSs) were compared. Then, we summarized the potential applications of MVDDSs and discuss future perspectives. The development of MVDDS may provide avenues for the treatment of diseases involving an inflammatory process.

Nonviral delivery systems for antisense oligonucleotide therapeutics.

Antisense oligonucleotides (ASOs) are an important tool for the treatment of many genetic disorders. However, similar to other gene drugs, vectors are often required to protect them from degradation and clearance, and to accomplish their transport in vivo. Compared with viral vectors, artificial nonviral nanoparticles have a variety of design, synthesis, and formulation possibilities that can be selected to accomplish protection and delivery for specific applications, and they have served critical therapeutic purposes in animal model research and clinical applications, allowing safe and efficient gene delivery processes into the target cells. We believe that as new ASO drugs develop, the exploration for corresponding nonviral vectors is inevitable. Intensive development of nonviral vectors with improved delivery strategies based on specific targets can continue to expand the value of ASO therapeutic approaches. Here, we provide an overview of current nonviral delivery strategies, including ASOs modifications, action mechanisms, and multi-carrier methods, which aim to address the irreplaceable role of nonviral vectors in the progressive development of ASOs delivery.