Recapitulation of complex transport and action of drugs at tumor microenvironment using tumor-microenvironment-on-chip

Targeted delivery aims to selectively distribute drugs to targeted tumor tissue but not to healthy tissue. This can address many of clinical challenges by maximizing the efficacy but minimizing the toxicity of anti-cancer drugs. However, complex tumor microenvironment poses various barriers hindering the transport of drugs and drug delivery systems. New tumor models that allow for the systematic study of these complex environments are highly desired to provide reliable test beds to develop drug delivery systems for targeted delivery. Recently, research efforts have yielded new in vitro tumor models, the so called tumor-microenvironment-on-chip, that recapitulate certain characteristics of the tumor microenvironment. These new models show benefits over other conventional tumor models, and have the potential to accelerate drug discovery and enable precision medicines. However, further research is warranted to overcome their limitations and to properly interpret the data obtained from these models. In this article, key features of the in vivo tumor microenvironment that are relevant to drug transport processes for targeted delivery was discussed, and the current status and challenges for developing in vitro transport model systems was reviewed

Reliable detection of subchromosomal deletions and duplications using cell-based noninvasive prenatal testing

Objective To gather additional data on the ability to detect subchromosomal abnormalities of various sizes in single fetal cells isolated from maternal blood, using low-coverage shotgun next-generation sequencing for cell-based noninvasive prenatal testing (NIPT). Method Fetal trophoblasts were recovered from approximately 30 mL of maternal blood using maternal white blood cell depletion, density-based cell separation, immunofluorescence staining, and high-resolution scanning. These trophoblastic cells were picked as single cells and underwent whole genome amplification for subsequent genome-wide copy number analysis and genotyping to confirm the fetal origin of the cells. Results Applying our fetal cell isolation method to a series of 125 maternal blood samples, we detected on average 4.17 putative fetal cells/sample. The series included 15 cases with clinically diagnosed fetal aneuploidies and five cases with subchromosomal abnormalities. This method was capable of detecting findings that were 1 to 2 Mb in size, and all were concordant with the microarray or karyotype data obtained on a fetal sample. A minority of fetal cells showed evidence of genome degradation likely related to apoptosis. Conclusion We demonstrate that this cell-based NIPT method has the capacity to reliably diagnose fetal chromosomal abnormalities down to 1 to 2 Mb in size

Validation Studies for Single Circulating Trophoblast Genetic Testing as a Form of Noninvasive Prenatal Diagnosis

It has long been appreciated that genetic analysis of fetal or trophoblast cells in maternal blood could revolutionize prenatal diagnosis. We implemented a protocol for single circulating trophoblast (SCT) testing using positive selection by magnetic-activated cell sorting and single-cell low-coverage whole-genome sequencing to detect fetal aneuploidies and copy-number variants (CNVs) at ∼1 Mb resolution. In 95 validation cases, we identified on average 0.20 putative trophoblasts/mL, of which 55% were of high quality and scorable for both aneuploidy and CNVs. We emphasize the importance of analyzing individual cells because some cells are apoptotic, in S-phase, or otherwise of poor quality. When two or more high-quality trophoblast cells were available for singleton pregnancies, there was complete concordance between all trophoblasts unless there was evidence of confined placental mosaicism. SCT results were highly concordant with available clinical data from chorionic villus sampling (CVS) or amniocentesis procedures. Although determining the exact sensitivity and specificity will require more data, this study further supports the potential for SCT testing to become a diagnostic prenatal test

AN ENHANCER-TRAP AND SPLICE-TRAP SCREEN FOR REGULATORS OF ENDOCRINE FUNCTIONS IN THE CENTRAL NERVOUS SYSTEM AND THE EPITRACHEAL GLAND OF DROSOPHILA MELANOGASTER

Peptide hormones play important roles in a wide variety of biological processes, such as learning and memory, body weight homeostasis, circadian rhythms, and addiction. In Drosophila, over 20 peptide hormones are produced in the central nervous system (CNS) by distinct groups of specialized neurons, peptidergic neurons. Another important endocrine function is performed by the Inka cell, responsible for the production and secretion of the ecdysis triggering hormone (ETH), which is critical for the ecdysis process of the animal. Although many genes encoding peptide hormones have been identified, our knowledge about factors that regulate the production and secretion of peptide hormones is still very limited. In order to identify more endocrine regulators, we performed an enhancer trap and splice trap screen for genes that are expressed in peptidergic neurons in the CNS and the epitracheal gland, where the ETH-expressing Inka cell is located. From a screen of 545 splice trap lines and 287 enhancer trap lines, we obtained 28 insertions in 25 genes that are expressed in peptidergic neurons of the CNS, and 14 insertions in 14 genes that are expressed in the epitracheal gland. For lines that are expressed in the CNS, we further mapped the expression patterns by immunostaining with antisera to several neuropeptides, including Bursicon, Drosophila insulin-like peptide 2, crustacean cardioactive peptide, and leucokinin, each of which is expressed in distinct groups of neurons. Among the 28 lines identified, the BG00836 insertion in the gene alan shepard (shep) was extremely interesting to us because of its restricted reporter gene expression in peptidergic neurons. shepBG00836 mutant adults showed defects in wing expansion and decreased resistance to starvation, and these phenotypes were reverted by precise excision of the P element. However, it was later shown that these may be neomorphic phenotypes brought by the combination of the shep mutation and unknown factors in the genetic background. Follow-up experiments on shep showed that it is involved in the control of neurite outgrowth in the CNS. Therefore, shep may function as a general developmental regulator for peptidergic neurons. Lines with reporter gene expression in the epitracheal glands may reveal genes that participate in the regulation of ETH expression or secretion. We examined the functions of the trapped loci by knocking down gene expression specifically in the Inka cells with transgenic RNA interference (RNAi). Expression of UAS-taiman (tai)RNAi in the Inka cells led to ecdysis defects and reduced ETH expression. TAI is a known ecdysone receptor coactivator, and expression levels of TAI were correlated with the circulating steroid hormone titer. Previous research results in our lab and the Cherbas lab suggested that the basic-leucine zipper transcription factor Cryptocephal (CRC) and the EcR/USP heterodimeric ecdysone receptor together form a complex on the ETH promoter and activate the expression of ETH. Based on these findings, we propose a model in which a complex of TAI, CRC, and EcR/USP binds to the ETH promoter and activates ETH transcription

Induction of Apoptosis in HepaRG Cell Line by Aloe-Emodin through Generation of Reactive Oxygen Species and the Mitochondrial Pathway

Background/Aims: Aloe-emodin (1,8-dihydroxy-3-hydroxymethyl-anthraquinone), an anthraquinone active compounds, is isolated from some traditional medicinal plants such as Rheum palmatum L. and Cassia occidentalis, which induce hepatotoxicity in rats. The aim of this study was to determine potential cytotoxic effects of aloe-emodin on HepaRG cells and to define the underlying mechanism. Methods: MTT was used to evaluate cell viability. Apoptotic cell death was analyzed via Annexin V-FITC/PI double staining. Intracellular reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) were determined by flow cytometry, while the expression of apoptosis-related proteins was determined by Western blot analysis. Results: Treatment with aloe-emodin significantly reduced cell viability and induced apoptosis in HepaRG cells in a dose- and time-dependent manner. It provoked ROS generation and depolarization of MMP in HepaRG cells when compared with controls. Aloe-emodin dose-dependently increased release of mitochondrial cytochrome c, and levels of Fas, p53, p21, Bax/Bcl-2 ratio, as well as activation of caspase-3, caspase-8, caspase-9, and subsequent cleavage of poly(ADP-ribose)polymerase (PARP). It also induced S-phase cell cycle arrest by increasing the expression of p21 and cyclin E proteins while significantly decreasing the expression of cyclin A and CDK2. Conclusion: These results suggest that aloe-emodin inhibits cell proliferation and induces apoptosis in HepaRG cells, most probably through a mechanism involving both Fas death pathway and the mitochondrial pathway by generation of ROS. These findings underscore the need for risk assessment of human exposure to aloe-emodin

Comparison of LASSO and random forest models for predicting the risk of premature coronary artery disease

Abstract Purpose With the change of lifestyle, the occurrence of coronary artery disease presents a younger trend, increasing the medical and economic burden on the family and society. To reduce the burden caused by this disease, this study applied LASSO Logistic Regression and Random Forest to establish a risk prediction model for premature coronary artery disease(PCAD) separately and compared the predictive performance of the two models. Methods The data are obtained from 1004 patients with coronary artery disease admitted to a third-class hospital in Liaoning Province from September 2019 to December 2021. The data from 797 patients were ultimately evaluated. The dataset of 797 patients was randomly divided into the training set (569 persons) and the validation set (228 persons) scale by 7:3. The risk prediction model was established and compared by LASSO Logistic and Random Forest. Result The two models in this study showed that hyperuricemia, chronic renal disease, carotid artery atherosclerosis were important predictors of premature coronary artery disease. A result of the AUC between the two models showed statistical difference (Z = 3.47, P < 0.05). Conclusions Random Forest has better prediction performance for PCAD and is suitable for clinical practice. It can provide an objective reference for the early screening and diagnosis of premature coronary artery disease, guide clinical decision-making and promote disease prevention

A Novel Gel-Forming Solution Based on PEG-DSPE/Solutol HS 15 Mixed Micelles and Gellan Gum for Ophthalmic Delivery of Curcumin

Curcumin (Cur) is a naturally hydrophobic polyphenol with potential pharmacological properties. However, the poor aqueous solubility and low bioavailability of curcumin limits its ocular administration. Thus, the aim of this study was to prepare a mixed micelle in situ gelling system of curcumin (Cur-MM-ISG) for ophthalmic drug delivery. The curcumin mixed micelles (Cur-MMs) were prepared via the solvent evaporation method, after which they were incorporated into gellan gum gels. Characterization tests showed that Cur-MMs were small in size and spherical in shape, with a low critical micelle concentration. Compared with free curcumin, Cur-MMs improved the solubility and stability of curcumin significantly. The ex vivo penetration study revealed that Cur-MMs could penetrate the rabbit cornea more efficiently than the free curcumin. After dispersing the micelles in the gellan gum solution at a ratio of 1:1 (v/v), a transparent Cur-MM-ISG with the characteristics of a pseudoplastic fluid was formed. No obvious irritations were observed in the rabbit eyes after ocular instillation of Cur-MM-ISG. Moreover, Cur-MM-ISG showed a longer retention time on the corneal surface when compared to Cur-MMs using the fluorescein sodium labeling method. These findings indicate that biocompatible Cur-MM-ISG has great potential in ophthalmic drug therapy

Tradeoff of CO2 and CH4 emissions from global peatlands under water-table drawdown

International audienceThe climate impact of water-table drawdown in peatlands is unclear as carbon dioxide emissions increase and methane emissions decrease due to drying. This study shows decreasing water-table depth results in net greenhouse gas emissions from global peatlands, despite reducing methane emissions.Water-table drawdown across peatlands increases carbon dioxide (CO2) and reduces methane (CH4) emissions. The net climatic effect remains unclear. Based on global observations from 130 sites, we found a positive (warming) net climate effect of water-table drawdown. Using a machine-learning-based upscaling approach, we predict that peatland water-table drawdown driven by climate drying and human activities will increase CO2 emissions by 1.13 (95% interval: 0.88-1.50) Gt yr(-1) and reduce CH4 by 0.26 (0.14-0.52) GtCO(2)-eq yr(-1), resulting in a net increase of greenhouse gas of 0.86 (0.36-1.36) GtCO(2)-eq yr(-1) by the end of the twenty-first century under the RCP8.5 climate scenario. This drops to 0.73 (0.2-1.2) GtCO(2)-eq yr(-1) under RCP2.6. Our results point to an urgent need to preserve pristine and rehabilitate drained peatlands to decelerate the positive feedback among water-table drawdown, increased greenhouse gas emissions and climate warming