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Microfluidics for Drug Development: From Synthesis to Evaluation

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... However, the application of these devices was limited by their complexity, high production cost, toxic byproducts, difficulty in mixing multiple components, and restricted gas permeability. Later on, soft lithography for chip production emerged, and microfluidic technology entered a period of rapid development [25,26]. Soft lithography enables the fabrication of various polymer-based microfluidic devices. ...
... Drug compounds are synthesized either to obtain a medicinal and biologically significant product or to modify an existing compound to make it more clinically valuable [26,[29][30][31]. Drug discovery involves designing the molecular structure of the drug, lead compound synthesis, and lead optimization for pharmacological or clinical application. ...
... Traditionally, chemical synthesis is uncontrollable, inefficient, error prone, and requires large amounts of time and reagents. The advent of microfluidic technology offers benefits, such as less reagent use, faster reaction speeds, lower embodied expenditures of energy, smaller equipment, greater selectivity of reaction concerning products, and more secure reactions [26]. This section describes microreactors for drug synthesis, such as the microchannels reactor and the droplet microreactor. ...
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While there are many clinical drugs for prophylaxis and treatment, the search for those with low or no risk of side effects for the control of infectious and non-infectious diseases is a dilemma that cannot be solved by today’s traditional drug development strategies. The need for new drug development strategies is becoming increasingly important, and the development of new drugs from traditional medicines is the most promising strategy. Many valuable clinical drugs have been developed based on traditional medicine, including drugs with single active ingredients similar to modern drugs and those developed from improved formulations of traditional drugs. However, the problems of traditional isolation and purification and drug screening methods should be addressed for successful drug development from traditional medicine. Advances in microfluidics have not only contributed significantly to classical drug development but have also solved many of the thorny problems of new strategies for developing new drugs from traditional drugs. In this review, we provide an overview of advanced microfluidics and its applications in drug development (drug compound synthesis, drug screening, drug delivery, and drug carrier fabrication) with a focus on its applications in conventional medicine, including the separation and purification of target components in complex samples and screening of active ingredients of conventional drugs. We hope that our review gives better insight into the potential of traditional medicine and the critical role of microfluidics in the drug development process. In addition, the emergence of new ideas and applications will bring about further advances in the field of drug development.
... For example, Huang et al. 159 constructed an arrayed geometrically enhanced mixing chip with individual function zones to quickly and accurately evaluate the potency of five drugs. In addition, with the rapid development of digital manufacturing techniques, 3DP-assisted microfluidic chip devices have proved advantageous in improving the efficiency, speed, and control of pharmaceutical analysis 160 . ...
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Pharmaceutical analysis is a discipline based on chemical, physical, biological, and information technologies. At present, biotechnological analysis is a short branch in pharmaceutical analysis; however, bioanalysis is the basis and an important part of medicine. Biotechnological approaches can provide information on biological activity and even clinical efficacy and safety, which are important characteristics of drug quality. Because of their advantages in reflecting the overall biological effects or functions of drugs and providing visual and intuitive results, some biotechnological analysis methods have been gradually applied to pharmaceutical analysis from raw material to manufacturing and final product analysis, including DNA super-barcoding, DNA-based rapid detection, multiplex ligation-dependent probe amplification, hyperspectral imaging combined with artificial intelligence, 3D biologically printed organoids, omics-based artificial intelligence, microfluidic chips, organ-on-a-chip, signal transduction pathway-related reporter gene assays, and the zebrafish thrombosis model. The applications of these emerging biotechniques in pharmaceutical analysis have been discussed in this review.
... After more than 30 years of development [1], microfluidic technology has made great progress in the fields of scientific research and industry. As a useful tool for researchers wishing to handle fluids of small volume, microfluidics has proven itself in various applications such as drug development/screening [2,3], tissue engineering [4], point-of-care testing [5], and chemical [6] and material synthesis [7,8]. In addition to academic research, microfluidic technologies have shown great potential in diverse industries and found use in various commercial devices such as inkjet printheads and digital polymerase chain reaction (dPCR) instruments [9]. ...
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This review mainly studies the development status, limitations, and future directions of modular microfluidic systems. Microfluidic technology is an important tool platform for scientific research and plays an important role in various fields. With the continuous development of microfluidic applications, conventional monolithic microfluidic chips show more and more limitations. A modular microfluidic system is a system composed of interconnected, independent modular microfluidic chips, which are easy to use, highly customizable, and on-site deployable. In this paper, the current forms of modular microfluidic systems are classified and studied. The popular fabrication techniques for modular blocks, the major application scenarios of modular microfluidics, and the limitations of modular techniques are also discussed. Lastly, this review provides prospects for the future direction of modular microfluidic technologies.
... Nevertheless, traditional digital microfluidics platforms can only perform multiplex or parallel biochemical reactions due to the use of a small number of droplets (from several up to tens) [27]. In addition, traditional digital microfluidics platforms have not been leveraged for 3D cell culture with continuous perfusion [28]. Therefore, the development of an advanced digital organ-on-a-chip platform with good parallelism, simple preparation and operation would be a significant step in the screening of anticancer drugs and toxicity testing, enabling a more efficient drug screening pipeline for liver cancer. ...
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Organ-on-a-chip systems have been increasingly recognized as attractive platforms to assess toxicity and to develop new therapeutic agents. However, current organ-on-a-chip platforms are limited by a “single pot” design, which inevitably requires holistic analysis and limits parallel processing. Here, we developed a digital organ-on-a-chip by combining a microwell array with cellular microspheres, which significantly increased the parallelism over traditional organ-on-a-chip for drug development. Up to 127 uniform liver cancer microspheres in this digital organ-on-a-chip format served as individual analytical units, allowing for analysis with high consistency and quick response. Our platform displayed evident anti-cancer efficacy at a concentration of 10 μM for sorafenib, and had greater alignment than the “single pot” organ-on-a-chip with a previous in vivo study. In addition, this digital organ-on-a-chip demonstrated the treatment efficacy of natural killer cell-derived extracellular vesicles for liver cancer at 50 μg/mL. The successful development of this digital organ-on-a-chip platform provides high-parallelism and a low-variability analytical tool for toxicity assessment and the exploration of new anticancer modalities, thereby accelerating the joint endeavor to combat cancer. Graphic abstract
... They deliver drugs to the target sites, ensure maximum efficacy of drugs and increase the duration of drug release. Drug evaluation mainly focuses on the toxicity, safety, pharmacokinetics and physicochemical properties of newly developed compounds [3]. Drug toxicity affects the safety of drugs in vivo, and toxicity screening is a key step in drug evaluation. ...
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Microfluidic technology has been highly useful in nanovolume sample preparation, separation, synthesis, purification, detection and assay, which are advantageous in drug development. This review highlights the recent developments and trends in microfluidic applications in two areas of drug development. First, we focus on how microfluidics has been developed as a facile tool for the fabrication of drug carriers including microparticles and nanoparticles. Second, we discuss how microfluidic chips could be used as an independent platform or integrated with other technologies in drug toxicity screening. Challenges and future perspectives of microfluidic applications in drug development have also been provided considering the present technological limitations.
... Microfluidics is a technique for accurately processing and manipulating small amounts of fluid in a microscale device with channels ranging from tens to hundreds of micrometers [1]. Some of the main advantages of microfluidics are the low consumption of reagents, high repeatability, fast reaction rates, and accurate control of physical/chemical properties [1][2][3][4][5][6]. One application of microfluidics is the capture and release of biomolecules and drug delivery on the micrometer scale [7]. ...
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Stimuli-responsive hydrogels have a wide range of potential applications in microfluidics, which has drawn great attention. Double cross-linked hydrogels are very well suited for this application as they offer both stability and the required responsive behavior. Here, we report the integration of poly(N-isopropylacrylamide) (PNiPAAm) hydrogel with a permanent cross-linker (N,N′-methylenebisacrylamide, BIS) and a redox responsive reversible cross-linker (N,N′-bis(acryloyl)cystamine, BAC) into a microfluidic device through photopolymerization. Cleavage and re-formation of disulfide bonds introduced by BAC changed the cross-linking densities of the hydrogel dots, making them swell or shrink. Rheological measurements allowed for selecting hydrogels that withstand long-term shear forces present in microfluidic devices under continuous flow. Once implemented, the thiol-disulfide exchange allowed the hydrogel dots to successfully capture and release the protein bovine serum albumin (BSA). BSA was labeled with rhodamine B and functionalized with 2-(2-pyridyldithio)-ethylamine (PDA) to introduce disulfide bonds. The reversible capture and release of the protein reached an efficiency of 83.6% in release rate and could be repeated over 3 cycles within the microfluidic device. These results demonstrate that our redox-responsive hydrogel dots enable the dynamic capture and release of various different functionalized (macro)molecules (e.g., proteins and drugs) and have a great potential to be integrated into a lab-on-a-chip device for detection and/or delivery.
... It can deliver an efficient concentration of the drugs to the tumor site with a lower level of injection, and less side effects would be achieved. Accordingly, new technology such as microfluidics organ/tumoron-a chip and microfluidic cell arrays can help to assess the performance of this candidate carrier on the cells or colon tumor prior to the in-vivo test or clinical trials [61][62][63]. ...
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Bone is an active organ that continuously undergoes an orchestrated process of remodeling throughout life. Bone tissue is uniquely capable of adapting to loading, hormonal, and other changes happening in the body, as well as repairing bone that becomes damaged to maintain tissue integrity. On the other hand, diseases such as osteoporosis and metastatic cancers disrupt normal bone homeostasis leading to compromised function. Historically, the ability to investigate processes related to either physiologic or diseased bone tissue is limited by traditional models that fail to emulate the complexity of native bone. Organ‐on‐a‐chip models are based on technological advances in tissue engineering and microfluidics, enabling the reproduction of key features specific to tissue microenvironments within a microfabricated device. Compared to conventional in vitro and in vivo bone models, microfluidic models, and especially organ‐on‐a‐chip platforms, provide more biomimetic tissue culture conditions, with increased predictive power for clinical assays. In this review, microfluidic and organ‐on‐a‐chip technologies designed for understanding the biology of bone as well as bone‐related diseases and treatments are reported. Finally, the authors discuss the limitations of the current models and point toward future directions for microfluidics and organ‐on‐a‐chip technologies in bone research. Compared to conventional in vitro and in vivo bone models, microfluidic models and especially organs‐on‐a‐chip platforms, provide more biomimetic tissue culture conditions, with increased predictive power for clinical assays. In this progress report, microfluidic and organ‐on‐a‐chip technologies designed for understanding the biology of bone as well as bone‐related diseases and treatments are covered.
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Significance Isolation of sufficient numbers of circulating tumor cells (CTCs) in cancer patients could provide an alternative to invasive tumor biopsies, providing multianalyte cell-based biomarkers that are not available from current plasma circulating tumor DNA sequencing. Given the average prevalence at one CTC per billion blood cells, very large blood volumes must be screened to provide enough CTCs for reliable clinical applications. By creating an ultrahigh-throughput magnetic sorter, we demonstrate the efficient removal of leukocytes from near whole blood volume equivalents. Combined with leukapheresis to initially concentrate blood mononuclear cells, this LP CTC-iChip platform will enable noninvasive sampling of cancer cells in sufficient numbers for clinical applications, ranging from real-time pharmacokinetic monitoring of drug response to tissue-of-origin determination in early-stage cancer screening.
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Cardiovascular disease is a chronic disease that leads to impaired cardiac function and requires long-term management to control its progression. Despite the importance of hydrogels for therapeutic applications, a contradiction between the size of a hydrogel and the amount of loaded drug has been encountered when using conventional fabrication methods. In this study, biocompatible reservoir microcapsules (diameter ~100 µm) with a large liquid core and polymeric shell were fabricated via a one-step phase separation of poly(ethylene glycol) diacrylate (PEGDA) and dextran within pre-gel droplets through microfluidics. By controlling the process of phase separation, high drug-loading efficiency (~80%) for long-term release (30 days) of vascular endothelial growth factor (VEGF) and platelet-derived growth factor (PDGF) was achieved. Drug molecules were dispersed within the liquid core at a concentration above saturation solubility for sustained delivery via regulation of the shells. Effective therapeutic enhancement of human umbilical vein endothelial cell (HUVEC) and umbilical artery smooth muscle cell (SMC) proliferation and tube formation in vitro promoted rapid cell proliferation and increased the number of migrated cells by ~1.7 times. Moreover, in vivo blood vessel regeneration for cardiovascular control induced by sustained dual-drug (VEGF and PDGF) delivery to the rat heart was achieved, showing the effectiveness of long-term protein delivery in improving cardiac function and significantly reducing ventricular wall thickness and fibrosis of the infarct region. The ratio of heart tissue scarring was reduced to 11.2% after microcapsule treatment compared with 21.4% after saline treatment in the rat model. By using these reservoir microcapsules, similar sustained delivery of proteins, mRNAs and biologic drugs could be developed for the treatment of a range of long-term chronic diseases and regenerative medicine.
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Techniques that enable the spatial arrangement of living cells into defined patterns are broadly applicable to tissue engineering, drug screening, and cell–cell investigations. Achieving large‐scale patterning with single‐cell resolution while minimizing cell stress/damage is, however, technically challenging using existing methods. Here, a facile and highly scalable technique for the rational design of reconfigurable arrays of cells is reported. Specifically, microdroplets of cell suspensions are assembled using stretchable surface‐chemical patterns which, following incubation, yield ordered arrays of cells. The microdroplets are generated using a microfluidic‐based aerosol spray nozzle that enables control of the volume/size of the droplets delivered to the surface. Assembly of the cell‐loaded microdroplets is achieved via mechanically induced coalescence using substrates with engineered surface‐wettability patterns based on extracellular matrices. Robust cell proliferation inside the patterned areas is demonstrated using standard culture techniques. By combining the scalability of aerosol‐based delivery and microdroplet surface assembly with user‐defined chemical patterns of controlled functionality, the technique reported here provides an innovative methodology for the scalable generation of large‐area cell arrays with flexible geometries and tunable resolution.
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Drug penetration into solid tumours remains a major challenge in the effective treatment of cancer. Microbubble (MB) mediated sonoporation offers a potential solution to this by enhancing the uptake of drugs into cells. Additionally, in using an ultrasound (US) trigger, drug delivery can be localised to the tumour, thus reducing the off-site toxicity associated with systemic delivery. The majority of in vitro studies involving the observation of MB-enhanced drug efficacy have been conducted on 2D monolayer cell cultures, which are known to be poor models for in vivo tumours. 3D spheroid cultures allow for the production of multicellular cultures complete with extracellular matrix (ECM) components. These cultures effectively recreate many of the physiological features of the tumour microenvironment and have been shown to be far superior to previous 2D monolayer models. However, spheroids are typically handled in well-plates in which the fluid environment is static, limiting the physiological relevance of the model. The combination of 3D cultures and microfluidics would allow for the production of a dynamic system in which spheroids are subjected to in vivo like fluid flow and shear stressesThis study presents a microfluidic device containing an array of spheroid traps, into which multiple pre-grown colorectal cancer (CRC) spheroids were loaded. Reservoirs interfaced with the chip use hydrostatic pressure to passively drive flow through the system and subject spheroids to capillary like flow velocities. The use of reservoirs also enabled multiple chips to be run in parallel, allowing for the screening of multiple therapeutic treatments (n = 690 total spheroids analysed). This microfluidic platform was used to investigate MB enhanced drug delivery and showed that co-delivery of 3 μM doxorubicin (DOX) + MB + US reduced spheroid viability to 48 ± 2%, compared to 75 ± 5% observed with 3 μM DOX alone. Delivery of drug loaded MBs (DLMBs), in which DOX-loaded liposomes (DOX-LS) were conjugated to MBs, reduced spheroid viability to 62 ± 3%, a decrease compared to the 75 ± 3% viability observed with DOX-LS in the absence of MBs + US.
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Inertial microfluidics has been broadly investigated, resulting in the development of various applications, mainly for particle or cell separation. Lateral migrations of these particles within a microchannel strictly depend on the channel design and its cross-section. Nonetheless, the fabrication of these microchannels is a continuous challenging issue for the microfluidic community, where the most studied channel cross-sections are limited to only rectangular and more recently trapezoidal microchannels. As a result, a huge amount of potential remains intact for other geometries with cross-sections difficult to fabricate with standard microfabrication techniques. In this study, by leveraging on benefits of additive manufacturing, we have proposed a new method for the fabrication of inertial microfluidic devices. In our proposed workflow, parts are first printed via a high-resolution DLP/SLA 3D printer and then bonded to a transparent PMMA sheet using a double-coated pressure-sensitive adhesive tape. Using this method, we have fabricated and tested a plethora of existing inertial microfluidic devices, whether in a single or multiplexed manner, such as straight, spiral, serpentine, curvilinear, and contraction-expansion arrays. Our characterizations using both particles and cells revealed that the produced chips could withstand a pressure up to 150 psi with minimum interference of the tape to the total functionality of the device and viability of cells. As a showcase of the versatility of our method, we have proposed a new spiral microchannel with right-angled triangular cross-section which is technically impossible to fabricate using the standard lithography. We are of the opinion that the method proposed in this study will open the door for more complex geometries with the bespoke passive internal flow. Furthermore, the proposed fabrication workflow can be adopted at the production level, enabling large-scale manufacturing of inertial microfluidic devices.
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Mining the antibody repertoire of plasma cells and plasmablasts could enable the discovery of useful antibodies for therapeutic or research purposes1. We present a method for high-throughput, single-cell screening of IgG-secreting primary cells to characterize antibody binding to soluble and membrane-bound antigens. CelliGO is a droplet microfluidics system that combines high-throughput screening for IgG activity, using fluorescence-based in-droplet single-cell bioassays2, with sequencing of paired antibody V genes, using in-droplet single-cell barcoded reverse transcription. We analyzed IgG repertoire diversity, clonal expansion and somatic hypermutation in cells from mice immunized with a vaccine target, a multifunctional enzyme or a membrane-bound cancer target. Immunization with these antigens yielded 100–1,000 IgG sequences per mouse. We generated 77 recombinant antibodies from the identified sequences and found that 93% recognized the soluble antigen and 14% the membrane antigen. The platform also allowed recovery of ~450–900 IgG sequences from ~2,200 IgG-secreting activated human memory B cells, activated ex vivo, demonstrating its versatility. Millions of primary IgG-secreting cells from mouse and human are characterized for activity and antibody sequence at the single-cell level.
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A microfluidic protein aggregation device (microPAD) that allows the user to perform a series of protein incubations with various concentrations of two reagents is demonstrated. The microfluidic device consists of 64 incubation chambers to perform individual incubations of the protein at 64 specific conditions. Parallel processes of metering reagents, stepwise concentration gradient generation, and mixing are achieved simultaneously by pneumatic valves. Fibrillation of bovine insulin was selected to test the device. The effect of insulin and sodium chloride (NaCl) concentration on the formation of fibrillar structures was studied by observing the growth rate of partially folded protein, using the fluorescent marker Thioflavin-T. Moreover, dual gradients of different NaCl and hydrochloric acid (HCl) concentrations were formed, to investigate their interactive roles in the formation of insulin fibrils and spherulites. The chip-system provides a bird’s eye view on protein aggregation, including an overview of the factors that affect the process and their interactions. This microfluidic platform is potentially useful for rapid analysis of the fibrillation of proteins associated with many misfolding-based diseases, such as quantitative and qualitative studies on amyloid growth.
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Microfluidics offers numerous advantages for the synthesis of short-lived radiolabeled imaging tracers: performing ¹⁸F-radiosyntheses in microliter-scale droplets has exhibited high efficiency, speed, and molar activity as well as low reagent consumption. However, most reports have been at the preclinical scale. In this study we integrate a [¹⁸F]fluoride concentrator and a microdroplet synthesizer to explore the possibility of synthesizing patient doses and multi-patient batches of clinically-acceptable tracers. In the integrated system, [¹⁸F]fluoride (up to 41 GBq [1.1 Ci]) in [¹⁸O]H2O (1 mL) was first concentrated ∼80-fold and then efficiently transferred to the 8 μL reaction chip as a series of small (∼0.5 μL) droplets. Each droplet rapidly dried at the reaction site of the pre-heated chip, resulting in localized accumulation of large amounts of radioactivity in the form of dried [¹⁸F]TBAF complex. The PET tracer [¹⁸F]fallypride was synthesized from this concentrated activity in an overall synthesis time of ∼50 min (including radioisotope concentration and transfer, droplet radiosynthesis, purification, and formulation), in amounts up to 7.2 GBq [0.19 Ci], sufficient for multiple clinical PET scans. The resulting batches of [¹⁸F]fallypride passed all QC tests needed to ensure safety for clinical injection. This integrated technology enabled for the first time the impact of a wide range of activity levels on droplet radiosynthesis to be studied. Furthermore, this substantial increase in scale expands the applications of droplet radiosynthesis to the production of clinically-relevant amounts of radiopharmaceuticals, and potentially even centralized production of clinical tracers in radiopharmacies. The overall system could be applied to fundamental studies of droplet-based radiochemical reactions, or to the production of radiopharmaceuticals labeled with a variety of isotopes used for imaging and/or targeted radiotherapeutics.
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Current drug development techniques are expensive and inefficient, partially due to the use of preclinical models that do not accurately recapitulate in vivo drug efficacy and cytotoxicity. To address this challenge, we report on an integrated, in vitro multi-organoid system that enables parallel assessment of drug efficiency and toxicity on multiple 3D tissue organoids. Built in a low-cost, adhesive film-based microfluidic device, these miniaturized structures require less than 200 µL fluid volume and are amenable to both matrix-based 3D cell culture and spheroid aggregate integration, each supported with an in situ photocrosslinkable hyaluronic acid hydrogel. Here, we demonstrate this technology first with a three-organoid device consisting of liver, cardiac, and lung constructs. We show that these multiple tissue types can be kept in common circulation with high viability for 21 days and validate the platform by investigating liver metabolism of the prodrug capecitabine into 5-fluorouracil (5-FU) and observing downstream toxicity in lung and cardiac organoids. Then we expand the integrated system to accommodate six humanized constructs, including liver, cardiac, lung, endothelium, brain, and testes organoids. Following a 14-day incubation in common media, we demonstrate multi-tissue interactions by metabolizing the alkylating prodrug ifosfamide in the liver organoid to produce chloroacetaldehyde and induce downstream neurotoxicity. Our results establish an expandable, multi-organoid body-on-a-chip system that can be fabricated easily and used for the accurate characterization of drug interactions in vitro. Statement of Significance The use of 3-dimensional (3D) in vitro models in drug development has advanced over the past decade. However, with several exceptions, the majority of research studies using 3D in vitro models, such as organoids, employ single tissue types, in isolated environments with no “communication” between different tissues. This is a significant limiting factor because in the human body there is significant signaling between different cells, tissues, and organs. Here we employ a low-cost, adhesive film-based microfluidic device approach, paired with a versatile extracellular matrix-derived hyaluronic acid hydrogel to support integrated systems of 3 and 6 3D organoid and cell constructs. Moreover, we demonstrate an integrated response to drugs, in which downstream toxicity is dependent on the presence of liver organoids.
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Controlling the mobility of liquids along surfaces is widely exploited in various technologies to achieve self‐lubrication, phase‐change heat transfer, and microfluidics. Despite commendable progress in directional liquid transport on peristome‐mimetic surfaces, liquid merely spreads directionally with a wetted trail remaining. It is a challenge to achieve directional contracting of spreading liquid at the rear side and ultimately unidirectional motion in bulk from one site to another. Here it is shown that liquids resting on the peristome‐mimetic surfaces can crawl directionally and rapidly in an inchworms‐like way under the action of sudden spontaneous bubbles levitation. Vacuuming or chemical reaction induces sudden nucleation, growth, coalescence (Ostwald ripening process), and rupture of bubbles in the asymmetric microcavities of the peristome‐mimetic surface with directional overpressure beneath the liquid, resulting in the guided contracting and spreading of the liquid. Bubbles regulate this new mode of liquid directional motion. The strategy offers opportunities for liquids directional motion for various applications, such as in microfluidic devices, oil–water separation, and water collection systems. A general strategy is presented for inchworm‐like unidirectional motion of liquids on peristome‐mimetic surfaces. In situ X‐ray imaging clearly visualizes the liquids moving mechanism. Vacuuming or chemical reaction induces the sudden nucleation, growth, coalescence, and rupture of bubbles in the asymmetric microcavities. This new mode of liquid directional motion offers opportunities for biofluidic, microfluidic devices, and water collection systems.
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Tumor progression, including metastasis, is significantly influenced by factors in the tumor microenvironment (TME) such as mechanical force, shear stress, chemotaxis, and hypoxia. At present, most cancer studies investigate tumor metastasis by conventional cell culture methods and animal models, which are limited in data interpretation. Although patient tissue analysis, such as human patient-derived xenografts (PDX), can provide important clinical relevant information, they may not be feasible for functional studies as they are costly and time-consuming. Thus, in vitro three-dimensional (3D) models are rapidly being developed that mimic TME and allow functional investigations of metastatic mechanisms and drug responses. One of those new 3D models is tumor-on-a-chip technology that provides a powerful in vitro platform for cancer research, with the ability to mimic the complex physiological architecture and precise spatiotemporal control. Tumor-on-a-chip technology can provide integrated features including 3D scaffolding, multicellular culture, and a vasculature system to simulate dynamic flow in vivo. Here, we review a select set of recent achievements in tumor-on-a-chip approaches and present potential directions for tumor-on-a-chip systems in the future for areas including mechanical and chemical mimetic systems. We also discuss challenges and perspectives in both biological factors and engineering methods for tumor-on-a-chip progress. These approaches will allow in the future for the tumor-on-a-chip systems to test therapeutic approaches for individuals through using their cancerous cells gathered through approaches like biopsies, which then will contribute toward personalized medicine treatments for improving their outcomes.
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Drug development suffers from a lack of predictive and human-relevant in vitro models. Organ-on-chip (OOC) technology provides advanced culture capabilities to generate physiologically appropriate, human-based tissue in vitro, therefore providing a route to a predictive in vitro model. However, OOC technologies are often created at the expense of throughput, industry-standard form factors, and compatibility with state-of-the-art data collection tools. Here we present an OOC platform with advanced culture capabilities supporting a variety of human tissue models including liver, vascular, gastrointestinal, and kidney. The platform has 96 devices per industry standard plate and compatibility with contemporary high-throughput data collection tools. Specifically, we demonstrate programmable flow control over two physiologically relevant flow regimes: perfusion flow that enhances hepatic tissue function and high-shear stress flow that aligns endothelial monolayers. In addition, we integrate electrical sensors, demonstrating quantification of barrier function of primary gut colon tissue in real-time. We utilize optical access to the tissues to directly quantify renal active transport and oxygen consumption via integrated oxygen sensors. Finally, we leverage the compatibility and throughput of the platform to screen all 96 devices using high content screening (HCS) and evaluate gene expression using RNA sequencing (RNA-seq). By combining these capabilities in one platform, physiologically-relevant tissues can be generated and measured, accelerating optimization of an in vitro model, and ultimately increasing predictive accuracy of in vitro drug screening.
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Micro total analytical systems (μTAS) are attractive to multiple fields that include chemistry, medicine and engineering due to their portability, low power usage, potential for automation, and low sample and reagent consumption, which in turn results in low waste generation. The development of fully-functional μTAS is an iterative process, based on the design, fabrication and testing of multiple prototype microdevices. Typically, microfabrication protocols require a week or more of highly-skilled personnel time in high-maintenance cleanroom facilities, which makes this iterative process cost-prohibitive in many locations worldwide. Rapid-prototyping tools, in conjunction with the use of polydimethylsiloxane (PDMS), enable rapid development of microfluidic structures at lower costs, circumventing these issues in conventional microfabrication techniques. Multiple rapid-prototyping methods to fabricate PDMS-based microfluidic devices have been demonstrated in literature since the advent of soft-lithography in 1998; each method has its unique advantages and drawbacks. Here, we present a tutorial discussing current rapid-prototyping techniques to fabricate PDMS-based microdevices, including soft-lithography, print-and-peel and scaffolding techniques, among other methods, specifically comparing resolution of the features, fabrication processes and associated costs for each technique. We also present thoughts and insights towards each step of the iterative microfabrication process, from design to testing, to improve the development of fully-functional PDMS-based microfluidic devices at faster rates and lower costs.
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Conventional 2D or even 3D in vitro culture models for human reproductive organs cannot properly recapitulate the bidirectional endocrine crosstalk between the uterine endometrium and the ovary. This crosstalk is essential for maintaining the various physiological features and functions of each tissue. Moreover, most in vitro models for the female reproductive tract also fail to mimic its multicellular structure. We therefore developed a novel 'dual reproductive organ on a chip' that reflects the bidirectional endocrine cross-talk and the complex multicellular structures by integrating various cellular components of both the human uterine endometrium and the ovary with several biodegradable natural polymers. Indeed, the bidirectional endocrine crosstalk between these two tissues is achieved through media sharing between channels, and it can markedly improve the viability of loaded cells within each chamber of the chip platform. In addition, we also identified a reliable reproductive toxicity marker, SERPINB2, which is significantly increased in response to various toxic exposures in both endometrial and ovarian follicular cells. Based on these findings, we next established a SERPINB2 luciferase reporter system that was specifically designed for detecting and quantifying the toxicity of certain substances. By introducing this SERPINB2 luciferase reporter system into the loaded cells within the chip platform, we ultimately developed an effective 'dual reproductive organ-on-chip' that was successfully used to predict the reproductive toxicity of various hazardous materials.
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Retinal cells within neurovascular units generate the blood-retinal barrier (BRB) to regulate the local retinal microenvironment and to limit access to inflammatory cells. Breakdown of the endothelial junctional complexes in the BRB negatively affects neuronal signaling and ultimately causes vision loss. As new therapeutics are being developed either to prevent barrier disruption or to restore barrier function, access to physiologically relevant human in vitro tissue models that recapitulate important features of barrier biology is essential for disease modeling, target validation, and toxicity assessment. Here, a tunable organ-on-a-chip model of the retinal microvasculature using human retinal microvascular endothelial cells with integrated flow is described. Automated imaging and image analysis methods are employed for facile screening of leakage mediators and cytokine inhibitors on barrier properties. The developed retinal microvasculature-on-a-chip will enable improved understanding of BRB biology and provide an additional tool for drug discovery.
Article
The silk fibroin (SF) prepared by Bombyx mori silkworms is one of the mainly abundant natural fiber and can be obtained simply and economically. SF as bio-material has superior bio-compatibility and bio-degradability. The current review provides an inclusive outline of up to date and novel developments on SF as bio-material based applications in tissue engineering and various drug delivery. SF as bio-materials was comprehensively reviewed, demonstrating the characteristics and applications of SF bio-materials in tissue engineering and drug delivery systems. Convenient regeneration, superb bio-compatibility, significant mechanical properties and versatile bio-degradability of SF has been investigated for the preparation of a range of articles such as films, spongy matrices, hydrogels, etc., and has been examined for use in a choice of tissue engineering utilization. Also, SF nanoparticles have been effectively designed and are competent to manage the release rate of biomolecules in a continuous approach with high stability. Therefore, the present review comprehensively covered the advancement made on SF based drug delivery, in vitro engineering and rejuvenation determines possibilities for additional progress in these areas.
Article
Traditional drug screening models are often unable to faithfully recapitulate human physiology in health and disease, motivating the development of microfluidic organs-on-a-chip (OOC) platforms that can mimic many aspects of human physiology and in the process alleviate many of the discrepancies between preclinical studies and clinical trials outcomes. Linsitinib, a novel anti-cancer drug, showed promising results in pre-clinical models of Ewing Sarcoma (ES), where it suppressed tumor growth. However, a Phase II clinical trial in several European centers with patients showed relapsed and/or refractory ES. We report an integrated, open setting, imaging and sampling accessible, polysulfone-based platform, featuring minimal hydrophobic compound binding. Two bioengineered human tissues - bone ES tumor and heart muscle - were cultured either in isolation or in the integrated platform and subjected to a clinically used linsitinib dosage. The measured anti-tumor efficacy and cardiotoxicity were compared with the results observed in the clinical trial. Only the engineered tumor tissues, and not monolayers, recapitulated the bone microenvironment pathways targeted by linsitinib, and the clinically-relevant differences in drug responses between non-metastatic and metastatic ES tumors. The responses of non-metastatic ES tumor tissues and heart muscle to linsitinib were much closer to those observed in the clinical trial for tissues cultured in an integrated setting than for tissues cultured in isolation. Drug treatment of isolated tissues resulted in significant decreases in tumor viability and cardiac function. Meanwhile, drug treatment in an integrated setting showed poor tumor response and less cardiotoxicity, which matched the results of the clinical trial. Overall, the integration of engineered human tumor and cardiac tissues in the integrated platform improved the predictive accuracy for both the direct and off-target effects of linsitinib. The proposed approach could be readily extended to other drugs and tissue systems.
Article
Pickering emulsions are surfactant-free emulsions stabilized by solid particles. Their unique structure endows them with good stability, excellent biocompatibility, and environmental friendliness. Pickering emulsions have displayed great potential in oral drug delivery. Several-fold increases in the oral bioavailability or bioaccessibility of poorly soluble drugs, such as curcumin, silybin, puerarin, and rutin, were achieved by using Pickering emulsions, whereas controlled release was found for indomethacin and caffeine. The shell of the interfacial particle stabilizers provides enhanced gastrointestinal stability to the cargos in the oil core. Here, we also discuss general considerations concerning particle stabilizers and design strategies to control lipid digestion.
Article
Organs-on-chips (OoCs), also known as microphysiological systems or 'tissue chips' (the terms are synonymous), have attracted substantial interest in recent years owing to their potential to be informative at multiple stages of the drug discovery and development process. These innovative devices could provide insights into normal human organ function and disease pathophysiology, as well as more accurately predict the safety and efficacy of investigational drugs in humans. Therefore, they are likely to become useful additions to traditional preclinical cell culture methods and in vivo animal studies in the near term, and in some cases replacements for them in the longer term. In the past decade, the OoC field has seen dramatic advances in the sophistication of biology and engineering, in the demonstration of physiological relevance and in the range of applications. These advances have also revealed new challenges and opportunities, and expertise from multiple biomedical and engineering fields will be needed to fully realize the promise of OoCs for fundamental and translational applications. This Review provides a snapshot of this fast-evolving technology, discusses current applications and caveats for their implementation, and offers suggestions for directions in the next decade.
Article
In the past five years, droplet microfluidic techniques have unlocked new opportunities for the high-throughput genome-wide analysis of single cells, transforming our understanding of cellular diversity and function. However, the field lacks an accessible method to screen and sort droplets based on cellular phenotype upstream of genetic analysis, particularly for large and complex cells. To meet this need, we developed Dropception, a robust, easy-to-use workflow for precise single-cell encapsulation into picoliter-scale double emulsion droplets compatible with high-throughput screening via fluorescence-activated cell sorting (FACS). We demonstrate the capabilities of this method by encapsulating five standardized mammalian cell lines of varying size and morphology as well as a heterogeneous cell mixture of a whole dissociated flatworm (5 - 25 μm in diameter) within highly monodisperse double emulsions (35 μm in diameter). We optimize for preferential encapsulation of single cells with extremely low multiple-cell loading events (<2% of cell-containing droplets), thereby allowing direct linkage of cellular phenotype to genotype. Across all cell lines, cell loading efficiency approaches the theoretical limit with no observable bias by cell size. FACS measurements reveal the ability to discriminate empty droplets from those containing cells with good agreement to single-cell occupancies quantified via microscopy, establishing robust droplet screening at single-cell resolution. High-throughput FACS screening of cellular picoreactors has the potential to shift the landscape of single-cell droplet microfluidics by expanding the repertoire of current nucleic acid droplet assays to include functional phenotyping.
Article
Designing strategies to utilize the synergistic effect of probiotics and prebiotics is a promising way in treating metabolic-related diseases. Here, inspired by the mutually promotable but mutually incompatible characteristics of Yin and Yang, dual-core microcapsules that encapsulate Lactobacillus and Bacillus subtilis into separate compartments were presented through electrostatically-driven microfluidics. The microcapsules showed acid resistance and preserved probiotics activity. They also fostered the proliferation of probiotics while creating an anaerobic environment and promoted lactic acid fermentation without affecting each other. It has been demonstrated that the microcapsules could reduce inflammation, improve fat metabolism and restore intestinal barrier function, thus contributing to the treatment of metabolic syndrome in vivo. These features make the dual-core microcapsules an ideal candidate for treating metabolic syndrome and related diseases.
Article
Epilepsies are a group of neurological disorders characterised by recurrent epileptic seizures. Seizures, defined as abnormal transient discharges of neuronal activity, can affect the entire brain circuitry or remain more focal in the specific brain regions and neuronal networks. Human pluripotent stem cell (hPSC)-derived neurons are a promising option for modelling epilepsies, but as such, they do not model groups of connected neuronal networks or focal seizures. Our solution is a Modular Platform for Epilepsy Modelling In Vitro (MEMO), a lab-on-chip device, in which three hPSC-derived networks are separated by a novel microfluidic cell culture device that allows controlled network-to-network axonal connections through microtunnels. In this study, we show that the neuronal networks formed a functional circuitry that was successfully cultured in MEMO for up to 98 days. The spontaneous neuronal network activities were monitored with an integrated custom-made microelectrode array (MEA). The networks developed spontaneous burst activity that was synchronous both within and between the axonally connected networks, i.e. mimicking both local and circuitry functionality of the brain. A convulsant, kainic acid, increased bursts only in the specifically treated networks. The activity reduction by an anticonvulsant, phenytoin, was also localised to treated networks. Therefore, modelling focal seizures in human neuronal networks is now possible with the developed chip. Link to the article: https://authors.elsevier.com/a/1bfTK3PVtpkTKl
Article
Droplet microfluidics is a powerful platform for high-throughput single-molecule protein analysis. However, the issues of coalescence and crosstalk of droplets compromise the accuracy of detection and hinder its wide application. To address these limitations, a novel colloidosome-based method was presented by combining a Pickering emulsion with droplet microfluidics for single-molecule protein analysis. Utilizing the self-assembly of easily synthesized colloidal surfactant F-SiO2 NPs at the water/oil interface, the colloidosomes are rigidly stabilized and can effectively avoid the leakage of fluorescent molecules. The crosstalk-free colloidosomes enable high-throughput single-molecule protein analysis, including heterogenous dynamic studies and digital detection. As a robust and accurate method, colloidosome-based microfluidics is promising as a powerful tool for a wide variety of applications, such as directed enzyme evolution, digital enzyme-linked immunosorbent assay (ELISA), and screening of antibiotics.
Article
Cell signaling greatly affected by complicated and temporally dynamic extracellular microenvironments controls most of the physiological functions in vivo. To reconstruct or simulate such microenvironments in vitro represents a fundamental approach for revealing the underlying mechanisms of those sophisticated processes. Recent advances in microfluidics have added a new dimension to cell signaling analysis, for examples, concentration gradient generators (amplitude aspect) or hydrodynamic gating strategy (frequency aspect), but it is still challengeable to capture single-cell dynamic signaling in response to mimicked extracellular microenvironment with varied stimuli waveforms of different amplitude and frequency in a high-throughput manner. In this article, we proposed a novel microfluidic strategy coupling multi-channel synchronous hydrodynamic gating with microfluidic concentration gradient generators (μMHG-CGG) to probe dynamic signaling of single cells with high throughput. The μMHG-CGG allows rapid delivery of dynamic chemical signals in both high frequency (as high as 670 mHz) and multiple amplitude domains at the same time, and simultaneously high-throughput probing cell dynamics at single-cell resolution in real-time. By applying the proposed system, the mechanisms for encoding/decoding systems (termed “frequency coding” or “amplitude coding”) via GPCRs mediated signaling pathways responding to histamine (HA) and adenosine triphosphate (ATP) in single HeLa cells were investigated. The optimal drug concentrations of single-cells response to HA and ATP individually or in combination were also successfully discussed, allowing us to obtain both single-cell heterogeneity and statistics from the cell population.
Article
The role of skin in the human body is indispensable, serving as a barrier, moderating homeostatic balance, and representing a pronounced endpoint for cosmetics and pharmaceuticals. Despite the extensive achievements of in vitro skin models, they do not recapitulate the complexity of human skin; thus, there remains a dependence on animal models during preclinical drug trials, resulting in expensive drug development with high failure rates. By imparting a fine control over the microenvironment and inducing relevant mechanical cues, skin‐on‐a‐chip (SoC) models have circumvented the limitations of conventional cell studies. Enhanced barrier properties, vascularization, and improved phenotypic differentiation have been achieved by SoC models; however, the successful inclusion of appendages such as hair follicles and sweat glands and pigmentation relevance have yet to be realized. The present Review collates the progress of SoC platforms with a focus on their fabrication and the incorporation of mechanical cues, sensors, and blood vessels.
Article
Drug delivery systems are developed to maximize drug efficacy and minimize side effects. As drug delivery technologies improve, the drug becomes safer and more comfortable for patients to use. During the last seven decades, extraordinary progress has been made in drug delivery technologies, such as systems for long-term delivery for months and years, localized delivery, and targeted delivery. The advances, however, will face a next phase considering the future technologies we need to overcome many physicochemical barriers for new formulation development and biological unknowns for treating various diseases. For immediate and long-term progress into the future, the drug delivery field should use time and resources for more translatable research ideas. The drug delivery discipline has to continue working on basic, applied, translational, and clinical research in a concerted manner to produce drug delivery systems that work for patients. It is a time to focus our attention on things that matter. It is also a time to develop realistic research goals and outcomes, diversify drug delivery technologies, and take the collective responsibility for our actions.
Article
Immobilizing liposomes in the hydrogel matrix network has become an effective strategy to protect liposomes against rapid clearance in the body, thus facilitating their localized drug delivery in a controlled and sustained manner for a long period. However, the low efficiency of drug delivery and short joint retention time seriously weaken its therapeutic efficacy. Herein, an efficient delivery platform was designed to anchor liposomes by integrating them with photo-crosslinkable GelMA matrix, rapidly forming monodisperse [email protected] hybrid microgels under ultraviolet light using a one-step innovational microfluidics technology. The liposomes were firmly anchored within microgels by the physical network hindrance and non-covalent interaction. Due to the double impediment from the lipid membrane and the hydrogel matrix network, kartogenin (KGN) encapsulated in [email protected] microgel presented notable extended release kinetics. Compared with KGN-loaded liposomes ([email protected]), KGN-loaded [email protected] microgels ([email protected]@KGN) could extend KGN release for over three weeks and remarkably promote chondrocyte differentiation of bone marrow mesenchymal stem cells(BMSCs)in vitro. Furthermore, the in vivo study demonstrated that [email protected]@KGN, with enhanced joint residence over five weeks, could effectively reduce osteophyte burden and prevent articular cartilage degeneration as well as subchondral bone changes when intraarticularly injected in a surgically induced rat osteoarthritis model. Collectively, [email protected] microgel, as an innovative extended delivery platform, holds tremendous potential for osteoarthritis treatment.
Article
The technology of organ-on-a-chip tries to mimic the complexity of native tissues in vitro. Important progress has been made recently in using this technology to study the gut with and without microbiota. These in vitro models can serve as an alternative to animal models for studying physiology, pathology, and pharmacology. While these models have greater physiological relevance compared to two-dimensional (2D) cell systems in vitro, endocrine and immunological functions in gut-on-a-chip models are still poorly represented. Furthermore, the construction of complex models, in which different cell types and structures interact, remains a challenge. Generally, gut-on-chip models have the potential to advance our understanding of the basic interactions found within the gut and lay the foundation for future applications in understanding pathophysiology, developing drugs, and personalizing medical treatments.
Article
Tumor has always been a major threat to human health due to its high morbidity and mortality. Among many cancer treatments, drug delivery microcarriers as a newly emerging method has attracted much attention. Here, we present a novel kind of multifunctional drug-delivery microcarriers formed from silk fibroin inverse opal scaffold and temperature-responsive poly (N-isopropylacrylamide) (PNIPAM) hydrogel for controllable and sustained drug release. Because of their uniform porous microstructure and interconnected nanopores, the PNIPAM hydrogel loaded with therapeutic drug can fill into the inverse opal scaffolds. It was demonstrated that these microcarriers had the ability to release drugs controllably and sustainably with temperature changes, which could not only reduce the drug waste but improve the effect of oncotherapy as well. In addition, due to the excellent biocompatibility of the silk fibroin, the inverse opal scaffolds could also support the adhesion and growth of normal cells, which was contributed to the tissue regeneration in the lesions. These features indicate that the designed drug-delivery microcarriers are ideal for oncotherapy and tissue engineering.
Article
The vascular network is a central component of the organ‐on‐a‐chip system to build a 3D physiological microenvironment with controlled physical and biochemical variables. Inspired by ubiquitous biological systems such as leaf venation and circulatory systems, a fabrication strategy is devised to develop a biomimetic vascular system integrated with freely designed chambers, which function as niches for chamber‐specific vascularized organs. As a proof of concept, a human‐on‐leaf‐chip system with biomimetic multiscale vasculature systems connecting the self‐assembled 3D vasculatures in chambers is fabricated, mimicking the in vivo complex architectures of the human cardiovascular system connecting vascularized organs. Besides, two types of vascularized organs are built independently within the two halves of the system to verify its feasibility for conducting comparative experiments for organ‐specific metastasis studies in a single chip. Successful culturing of human hepatoma G2 cells (HepG2s) and mesenchymal stem cells (MSCs) with human umbilical vein endothelial cells (HUVECs) shows good vasculature formation, and organ‐specific metastasis is simulated through perfusion of pancreatic cancer cells and shows distinct cancer encapsulation by MSCs, which is absent in HepG2s. Given good culture efficacy, study design flexibility, and ease of modification, these results show that the bioinspired human‐on‐leaf‐chip possesses great potential in comparative and metastasis studies while retaining organ‐to‐organ crosstalk.
Article
Significance Heavier objects usually sink in a less dense fluid. Water-walking anthropods and biomimetic water-walking robots harness surface tension in order to overcome this tendency, floating on top of liquids. By hanging a coacervate-encased droplet of a denser aqueous dextran solution from the surface of an aqueous solution of poly(ethylene glycol) (PEG), we harness the binding of dense droplets to interfaces by surface tension to build two-dimensional ensembles of structurally complex droplets. Applications ranging from reaction vessels with selective transport to motors and robotics are enabled by our findings.
Article
The nano-sized drug is an alternative strategy for enhancing the dissolution rate of pharmaceutical ingredients with poor solubility. Microfluidic technology has been widely applied for the preparation of nanoparticles because of its controllability. Combining the above two concerns, this study proposed an anti-solvent precipitation method to obtain itraconazole (ITZ) nanoparticles using a continuous flow droplet-based microreactor. In addition to the experimental study of the generation of different shapes of droplets, the influencing factors of nanoparticle preparation such as the flow rate ratio of the dispersed phase to the continuous phase, the residence time, the drug initial concentration and the stabilizer were further analyzed. The advantages of the droplets system were fully demonstrated in comparison with the conventional laminar flow system in the form of the T microfluidic device. The results showed that smaller nanoparticles of itraconazole with narrower size distribution were prepared in droplets and addressed that the particle agglomeration and growth would be restrained in droplets under the increased residence time, higher initial concentration or amphiphilic stabilizers.
Article
This study was to co-encapsulate a chemokine (stromal cell-derived factor-1, SDF-1) and a chondroinductive molecule (kartogenin, KGN) within microspheres via microfluidics, and to incorporate them into a hyaluronic acid (HA) injectable scaffold for articular cartilage defect repair. HA injectable scaffold, as a cartilage-friendly microenvironment, was prepared by crosslinking HA with 1,4-butanediol diglycidyl ether. A microfluidic device was set up to prepare monodisperse PLGA microspheres (49 μm) to load SDF-1 and KGN. An in vivo model of full-thickness articular cartilage defects in rabbits was applied to evaluate the reparative capacity of the current package. The SDF-1 and KGN were co-encapsulated simultaneously within the core and shell area of the microsphere with high loading efficiency and sustained release profiles of more than 2 months. The release profiles of them were highly matched and well fitted to a first-order mathematical model. These microspheres when incorporated into HA injectable scaffold were demonstrated to heal the full-thickness articular cartilage defects in rabbits. The regenerated tissue had the typical cartilage histological characters and integrated well with the surrounding tissue at 12w. This developed cell-free system could serve as an efficient therapy for articular cartilage defects treatment, serving as a supplementary way to cell based therapies.
Article
Significance Droplet manipulation has received great attention and research for its potential applications in various fields. Wettability surfaces, especially tunable wettability surfaces, are efficient way to manipulate the pinning and sliding of droplets. However, these surfaces only demonstrate uniform wettability control behavior and thus were difficult to accurately manipulate droplets to a certain location or multiple locations. In this paper, inspired by the microstructure of stomata distributed on plants, we present programmable wettability arrays for droplets manipulation by using microfluidic emulsification templates. Based on the intelligent composite system, controllable droplet sliding on programmable wettability pathways and effective droplet transfer for printing with mask integration have been demonstrated.
Article
The traditional regulatory drug approval paradigm comprising discrete phases of clinical testing that culminate in a large randomized superiority trial has historically been predominant in oncology. However, this approach has evolved in the current era of drug development, with multiple other development pathways now being utilized. Indeed, treatment approaches designed on the basis of an improved understanding of cancer biology have led to unprecedented responses in early phase trials, sometimes resulting in drug approvals in the absence of large-scale trials. At the same time, improved molecular diagnostic technologies have led to the identification of ever-smaller patient subgroups for molecularly targeted therapy. Moreover, new FDA regulatory paradigms have enabled the rapid review and accelerated approval of certain drugs in the absence of survival data. Regulatory approvals based on large-cohort trials with surrogate or intermediate clinical end points or on non-inferiority trials, as well as new tumour-agnostic indications, also set important precedents in the field. In this Viewpoint, we asked two leading oncologists involved in clinical drug development, an expert in regulatory science and prescription drug policy and a prominent patient advocate, to provide their opinions on the implications of these changes in regulatory practices for patient care.