Human iPSC and CRISPR targeted gene knock-in strategy for studying the somatic TIE2L914F mutation in endothelial cells.

Induced pluripotent stem cell (iPSC) derived endothelial cells (iECs) have emerged as a promising tool for studying vascular biology and providing a platform for modelling various vascular diseases, including those with genetic origins. Currently, primary ECs are the main source for disease modelling in this field. However, they are difficult to edit and have a limited lifespan. To study the effects of targeted mutations on an endogenous level, we generated and characterized an iPSC derived model for venous malformations (VMs). CRISPR-Cas9 technology was used to generate a novel human iPSC line with an amino acid substitution L914F in the TIE2 receptor, known to cause VMs. This enabled us to study the differential effects of VM causative mutations in iECs in multiple in vitro models and assess their ability to form vessels in vivo. The analysis of TIE2 expression levels in TIE2L914F iECs showed a significantly lower expression of TIE2 on mRNA and protein level, which has not been observed before due to a lack of models with endogenous edited TIE2L914F and sparse patient data. Interestingly, the TIE2 pathway was still significantly upregulated and TIE2 showed high levels of phosphorylation. TIE2L914F iECs exhibited dysregulated angiogenesis markers and upregulated migration capability, while proliferation was not affected. Under shear stress TIE2L914F iECs showed reduced alignment in the flow direction and a larger cell area than TIE2WT iECs. In summary, we developed a novel TIE2L914F iPSC-derived iEC model and characterized it in multiple in vitro models. The model can be used in future work for drug screening for novel treatments for VMs.

Highly scalable and standardized organ-on-chip platform with TEER for biological barrier modeling.

The development of new therapies is hampered by the lack of predictive, and patient-relevant in vitro models. Organ-on-chip (OOC) technologies can potentially recreate physiological features and hold great promise for tissue and disease modeling. However, the non-standardized design of these chips and perfusion control systems has been a barrier to quantitative high-throughput screening (HTS). Here we present a scalable OOC microfluidic platform for applied kinetic in vitro assays (AKITA) that is applicable for high, medium, and low throughput. Its standard 96-well plate and 384-well plate layouts ensure compatibility with existing laboratory workflows and high-throughput data collection and analysis tools. The AKITA plate is optimized for the modeling of vascularized biological barriers, primarily the blood-brain barrier, skin, and lung, with precise flow control on a custom rocker. The integration of trans-epithelial electrical resistance (TEER) sensors allows rapid and repeated monitoring of barrier integrity over long time periods. Together with automated liquid handling and compound permeability testing analyses, we demonstrate the flexibility of the AKITA platform for establishing human-relevant models for preclinical drug and precision medicine's efficacy, toxicity, and permeability under near-physiological conditions.

A Study on the Simulation and Experiment of Evaporative Condensers in an R744 Air Conditioning System.

The heat transfer characteristics of evaporative condensers in an R744 air conditioning system were evaluated using the numerical and the experimental methods. Two configurations of condensers were studied: Case 1 with five layers of tubes and Case 2 with eight layers of tubes. In order to evaluate the heat transfer characteristics, the temperature field, the phase change, the pressure distribution, and thermodynamic parameters were considered. For Case 2, it indicated the capability of R744 condensation from the superheated status to the liquified status by analyzing the outlet temperature of the condenser changed from 28.7 °C to 30.3 °C with a change in condensation pressure from 72.6 bar to 68.5 bar. In this study, R744 mass flow rate increases from 14.34 kg/h to 46.08 kg/h, and the pressure drop also increases from 0.23 bar to 0.47 bar for the simulation and 0.4 bar to 0.5 bar for the experiment, respectively. The results indicate that the five-layer configuration causes a higher pressure drop and lower COP than those obtained from the eight-layer one (splitting into two sets for smaller pressure drop). Furthermore, the evaporative condensers using mini tubes that are flooded in the cooling water tank are suitable for the subcritical R744 air conditioning system. In addition, the results obtained from the experimental data are in good agreement with those obtained from the numerical results, with a deviation of less than 5%.

Low-Cost, Accessible Fabrication Methods for Microfluidics Research in Low-Resource Settings.

Microfluidics are expected to revolutionize the healthcare industry especially in developing countries since it would bring portable, easy-to-use, self-contained diagnostic devices to places with limited access to healthcare. To date, however, microfluidics has not yet been able to live up to these expectations. One non-negligible factor can be attributed to inaccessible prototyping methods for researchers in low-resource settings who are unable to afford expensive equipment and/or obtain critical reagents and, therefore, unable to engage and contribute to microfluidics research. In this paper, we present methods to create microfluidic devices that reduce initial costs from hundreds of thousands of dollars to about $6000 by using readily accessible consumables and inexpensive equipment. By including the scientific community most embedded and aware of the requirements of healthcare in developing countries, microfluidics will be able to increase its reach in the research community and be better informed to provide relevant solutions to global healthcare challenges.