Polydopamine-Based Interfacial Engineering of Extracellular Matrix Hydrogels for the Construction and Long-Term Maintenance of Living Three-Dimensional Tissues.

Diverse biological processes in the body rely on the ability of cells to exert contractile forces on their extracellular matrix (ECM). In three-dimensional (3D) cell culture, however, this intrinsic cellular property can cause unregulated contraction of ECM hydrogel scaffolds, leading to a loss of surface anchorage and the resultant structural failure of in vitro tissue constructs. Despite advances in the 3D culture technology, this issue remains a significant challenge in the development and long-term maintenance of physiological 3D in vitro models. Here, we present a simple yet highly effective and accessible solution to this problem. We leveraged a single-step surface functionalization technique based on polydopamine to drastically increase the strength of adhesion between hydrogel scaffolds and cell culture substrates. Our method is compatible with different types of ECM and polymeric surfaces and also permits prolonged shelf storage of functionalized culture substrates. The proof-of-principle of this technique was demonstrated by the stable long-term (1 month) 3D culture of human lung fibroblasts. Furthermore, we showed the robustness and advanced application of the method by constructing a dynamic cell stretching system and performing over 100 000 cycles of mechanical loading on 3D multicellular constructs for visualization and quantitative analysis of stretch-induced tissue alignment. Finally, we demonstrated the potential of our technique for the development of microphysiological in vitro models by establishing microfluidic 3D co-culture of vascular endothelial cells and fibroblasts to engineer self-assembled, perfusable 3D microvascular beds.

Microphysiological Engineering of Self-Assembled and Perfusable Microvascular Beds for the Production of Vascularized Three-Dimensional Human Microtissues.

The vasculature is an essential component of the circulatory system that plays a vital role in the development, homeostasis, and disease of various organs in the human body. The ability to emulate the architecture and transport function of blood vessels in the integrated context of their associated organs represents an important requirement for studying a wide range of physiological processes. Traditional in vitro models of the vasculature, however, largely fail to offer such capabilities. Here we combine microfluidic three-dimensional (3D) cell culture with the principle of vasculogenic self-assembly to engineer perfusable 3D microvascular beds in vitro. Our system is created in a micropatterned hydrogel construct housed in an elastomeric microdevice that enables coculture of primary human vascular endothelial cells and fibroblasts to achieve de novo formation, anastomosis, and controlled perfusion of 3D vascular networks. An open-top chamber design adopted in this hybrid platform also makes it possible to integrate the microengineered 3D vasculature with other cell types to recapitulate organ-specific cellular heterogeneity and structural organization of vascularized human tissues. Using these capabilities, we developed stem cell-derived microphysiological models of vascularized human adipose tissue and the blood-retinal barrier. Our approach was also leveraged to construct a 3D organotypic model of vascularized human lung adenocarcinoma as a high-content drug screening platform to simulate intravascular delivery, tumor-killing effects, and vascular toxicity of a clinical chemotherapeutic agent. Furthermore, we demonstrated the potential of our platform for applications in nanomedicine by creating microengineered models of vascular inflammation to evaluate a nanoengineered drug delivery system based on active targeting liposomal nanocarriers. These results represent a significant improvement in our ability to model the complexity of native human tissues and may provide a basis for developing predictive preclinical models for biopharmaceutical applications.

Determination of Optimal Heart Rate Variability Features Based on SVM-Recursive Feature Elimination for Cumulative Stress Monitoring Using ECG Sensor.

Routine stress monitoring in daily life can predict potentially serious health impacts. Effective stress monitoring in medical and healthcare fields is dependent upon accurate determination of stress-related features. In this study, we determined the optimal stress-related features for effective monitoring of cumulative stress. We first investigated the effects of short- and long-term stress on various heart rate variability (HRV) features using a rodent model. Subsequently, we determined an optimal HRV feature set using support vector machine-recursive feature elimination (SVM-RFE). Experimental results indicate that the HRV time domain features generally decrease under long-term stress, and the HRV frequency domain features have substantially significant differences under short-term stress. Further, an SVM classifier with a radial basis function kernel proved most accurate (93.11%) when using an optimal HRV feature set comprising the mean of R-R intervals (mRR), the standard deviation of R-R intervals (SDRR), and the coefficient of variance of R-R intervals (CVRR) as time domain features, and the normalized low frequency (nLF) and the normalized high frequency (nHF) as frequency domain features. Our findings indicate that the optimal HRV features identified in this study can effectively and efficiently detect stress. This knowledge facilitates development of in-facility and mobile healthcare system designs to support stress monitoring in daily life.

Localization of ultrasound waveform for low intensity ultrasound-induced neuromodulation in a mouse model.

A collimator design was investigated to localize ultrasound stimulation using a flat ultrasound transducer for ultrasound-induced neuromodulation in a mouse model. In brain stimulation, the specific location of stimulation must be specified, as the region responsible for motor or sensory function is clustered in a narrow brain area. To localize ultrasound stimulation, three types of collimator design were simulated to determine the optimal collimator design. The performance of the simulated optimal collimator was compared to that of an unmounted collimator in a transducer in both in vivo and in vitro experiments. Throughout the experiments, the localized ultrasound waveform was shaped using the optimized collimator, which elicited neural spike activity in the targeted motor cortex. The optimized collimator shows potential for controlling a localized ultrasound waveform for ultrasound-induced neuromodulation in a small animal model.

Differential heart rate variability and physiological responses associated with accumulated short- and long-term stress in rodents.

In this study, we tested the hypothesis that chronic stress has cumulative effects over time on heart rate variability (HRV) and physiological responses in a rodent model of chronic mild stress. Rats were exposed to either short-term (2weeks) or long-term (4weeks) stress, followed by a 1-week recovery period. Controls were normally housed rats that did not undergo the stress procedure. For electrocardiogram recordings, transmitters were implanted in all rats 10days before the onset of the experiment to allow recovery from surgery. To investigate physiological responses, body weight, temperature, sucrose preference, and serum corticosterone levels were measured weekly. Rats exposed to both short- and long-term stress showed significant reductions in body weight, which did not normalize after the recovery period. A significant difference was observed between short- and long-term stress groups in serum corticosterone levels, with long-term stress significantly increasing serum corticosterone levels, which remained elevated after the recovery period (P<0.05). HRV analysis indicated that long-term stress significantly decreased time-domain indexes, whereas significantly increased frequency-domain indexes were observed in the low-frequency range (0.1-1Hz). These results may represent dysfunction in parasympathetic/vagal modulation with hyperactivation of the sympathetic nervous system after long-term exposures to stress. In addition, prolonged Q-to-T wave (QT) intervals were observed in rats exposed to long-term stress, which did not return to baseline levels after the recovery period. These findings are consistent with the view that chronic stress is associated with cardiac autonomic disorders and emphasize the importance of monitoring stress in our daily life since the effects of even mild stress may be cumulative.

Ankle-Angle Estimation from Blind Source Separated Afferent Activity in the Sciatic Nerve for Closed-Loop Functional Neuromuscular Stimulation System.

Cuff electrode recording has been proposed as a solution to obtain robust feedback signals for closed-loop controlled functional neuromuscular stimulation (FNS) systems. However, single-channel cuff electrode recording requires several electrodes to obtain the feedback signal related to each muscle. In this study, we propose an ankle-angle estimation method in which recording is conducted from the proximal nerve trunk with a multichannel cuff electrode to minimize cuff electrode usage. In experiments, muscle afferent signals were recorded from a rabbit's proximal sciatic nerve trunk using a multichannel cuff electrode, and blind source separation and ankle-angle estimation were performed using fast independent component analysis (PP/FastICA) combined with dynamically driven recurrent neural network (DDRNN). The experimental results indicate that the proposed method has high ankle-angle estimation accuracy for both situations when the ankle motion is generated by position servo system or neuromuscular stimulation. Furthermore, the results confirm that the proposed method is applicable to closed-loop FNS systems to control limb motion.

An implantable wireless optogenetic stimulation system for peripheral nerve control.

An implantable wireless optogenetic stimulation system with an LED-based optical stimulation cuff electrode was developed for peripheral nerve control. The proposed system consisted of a battery-powered optical cuff electrode, optical stimulation controller, and wireless communication system. The optical cuff electrode had a polydimethylsiloxane (PDMS) structure was designed to illuminate the entire sciatic nerve. The wireless communication system was designed to comply with medical implant communication service (MICS) regulations. To evaluate the proposed system, optogenetic stimulation was performed in optogenetic transgenic mice (Thy1::ChR2). The optical cuff electrode was implanted on the sciatic nerve, and movement was elicited during optical stimulation. The experimental results show that ankle movement can be generated wirelessly using optical stimulation pulse parameters.

Characteristics of the neuronal firing patterns in the subthalamic nucleus with graded dopaminergic cell loss in the nigrostriatal pathway.

The aim of this study was to evaluate the neuronal firing changes in the subthalamic nucleus (STN) in a graded mouse model of Parkinson's disease. Unilateral graded dopaminergic cell loss in the substantia nigra pars compacta was achieved by injecting different concentrations of 6-hydroxydopamine (6-OHDA) in the right medial forebrain bundle. Electrophysiological analysis of neuronal firing patterns in the STN revealed an increased firing rate, burst index, and interspike interval coefficient of variation in groups treated with higher 6-OHDA concentrations. The results of this study suggest the detailed pathophysiological characteristics of Parkinson's disease in a mouse model.