Uniform sized cancer spheroids production using hydrogel-based droplet microfluidics: a review.

Three-dimensional (3D) cell culture models have been extensively utilized in various mechanistic studies as well as for drug development studies as superior in vitro platforms than conventional two-dimensional (2D) cell culture models. This is especially the case in cancer biology, where 3D cancer models, such as spheroids or organoids, have been utilized extensively to understand the mechanisms of cancer development. Recently, many sophisticated 3D models such as organ-on-a-chip models are emerging as advanced in vitro models that can more accurately mimic the in vivo tissue functions. Despite such advancements, spheroids are still considered as a powerful 3D cancer model due to the relatively simple structure and compatibility with existing laboratory instruments, and also can provide orders of magnitude higher throughput than complex in vitro models, an extremely important aspects for drug development. However, creating well-defined spheroids remain challenging, both in terms of throughputs in generation as well as reproducibility in size and shape that can make it challenging for drug testing applications. In the past decades, droplet microfluidics utilizing hydrogels have been highlighted due to their potentials. Importantly, core-shell structured gel droplets can avoid spheroid-to-spheroid adhesion that can cause large variations in assays while also enabling long-term cultivation of spheroids with higher uniformity by protecting the core organoid area from external environment while the outer porous gel layer still allows nutrient exchange. Hence, core-shell gel droplet-based spheroid formation can improve the predictivity and reproducibility of drug screening assays. This review paper will focus on droplet microfluidics-based technologies for cancer spheroid production using various gel materials and structures. In addition, we will discuss emerging technologies that have the potential to advance the production of spheroids, prospects of such technologies, and remaining challenges.

Endocrine-disrupting compounds and their impact on human placental function: evidence from placenta organ-on-chip studies.

The effects of endocrine-disrupting compounds (EDCs) on the placenta, a critical gestational organ for xenobiotic protection, are well reported; however, models to determine the role of EDCs in placental disruption are limited. An advanced 2nd-trimester human placenta organ-on-chip model (2TPLA-OOC) was developed and validated, with six representative cells of the maternal and the fetal interface interconnected with microchannels. Various EDCs (150 ng mL-1 each of bisphenol A, bisphenol S, and polybrominated diphenyl ethers-47 and -99) were gradually propagated across the chip for 72 hours, and their various effects were determined. Cigarette smoke extract (CSE), an environmental risk factor, was used as a positive control. EDCs produced overall oxidative stress in the placental/decidual cells, induced cell-specific endocrine effects, caused limited (<10%) apoptosis/necrosis in trophoblasts and mesenchymal cells, induced localized inflammation but an overall anti-inflammatory shift, did not change immune cell migration from stroma to decidua, and did not affect placental nutrient transport. Overall, (1) the humanized 2TPLA-OOC recreated the placental organ and generated data distinct from the trophoblast and other cells studied in isolation, and (2) at doses associated with adverse pregnancies, EDCs produced limited and localized insults, and the whole organ compensated for the exposure.

Development of oxidative stress-associated disease models using feto-maternal interface organ-on-a-chip.

Oxidative stress (OS) and inflammation arising from cellular derangements at the fetal membrane-decidual interface (feto-maternal interface [FMi]) is a major antecedent to preterm birth (PTB). However, it is impractical to study OS-associated FMi disease state during human pregnancy, and thus it is difficult to develop strategies to reduce the incidences of PTB. A microfluidic organ-on-chip model (FMi-OOC) that mimics the in vivo structure and functions of FMi in vitro was developed to address this challenge. The FMi-OOC contained fetal (amnion epithelial, mesenchymal, and chorion) and maternal (decidua) cells cultured in four compartments interconnected by arrays of microchannels to allow independent but interconnected co-cultivation. Using this model, we tested the effects of OS and inflammation on both fetal (fetal → maternal) and maternal (maternal → fetal) sides of the FMi and determined their differential impact on PTB-associated pathways. OS was induced using cigarette smoke extract (CSE) exposure. The impacts of OS were assessed by measuring cell viability, disruption of immune homeostasis, epithelial-to-mesenchymal transition (EMT), development of senescence, and inflammation. CSE propagated (LC/MS-MS analysis for nicotine) over a 72-hour period from the maternal to fetal side, or vice versa. However, they caused two distinct pathological effects on the maternal and fetal cells. Specifically, fetal OS induced cellular pathologies and inflammation, whereas maternal OS caused immune intolerance. The pronounced impact produced by the fetus supports the hypothesis that fetal inflammatory response is a mechanistic trigger for parturition. The FMi disease-associated changes identified in the FMi-OOC suggest the unique capability of this in vitro model in testing in utero conditions.

A Microphysiological Device to Model the Choriodecidual Interface Immune Status during Pregnancy.

During human pregnancy the chorion (fetal) lines decidua (maternal) creating the feto-maternal interface. Despite their proximity, resident decidual immune cells remain quiescent during gestation and do not invade the chorion. Infection and infiltration of activated immune cells toward the chorion are often associated with preterm birth. However, the mechanisms that maintain choriodecidual immune homeostasis or compromise immune barrier functions remain unclear. To understand these processes, a two-chamber microphysiological system (MPS) was created to model the human choriodecidual immune interface under normal and infectious conditions in vitro. This MPS has outer (fetal chorion trophoblast cells) and inner chambers (maternal decidual + CD45+ cells [70:30 ratio]) connected by microchannels. Decidual cells were treated with LPS to mimic maternal infection, followed by immunostaining for HLA-DR and HLA-G, immune panel screening by imaging cytometry by time of flight, and immune regulatory factors IL-8 and IL-10, soluble HLA-G, and progesterone (ELISA). LPS induced a proinflammatory phenotype in the decidua characterized by a decrease in HLA-DR and an increase in IL-8 compared with controls. LPS treatment increased the influx of immune cells into the chorion, indicative of chorionitis. Cytometry by time of flight characterized immune cells in both chambers as active NK cells and neutrophils, with a decrease in the abundance of nonproinflammatory cytokine-producing NK cells and T cells. Conversely, chorion cells increased progesterone and soluble HLA-G production while maintaining HLA-G expression. These results highlight the utility of MPS to model choriodecidual immune cell infiltration and determine the complex maternal-fetal crosstalk to regulate immune balance during infection.

Identification of Microorganisms that Bind Specifically to Target Materials of Interest Using a Magnetophoretic Microfluidic Platform.

Discovery of microorganisms and their relevant surface peptides that specifically bind to target materials of interest can be achieved through iterative biopanning-based screening of cellular libraries having high diversity. Recently, microfluidics-based biopanning methods have been developed and exploited to overcome the limitations of conventional methods where controlling the shear stress applied to remove cells that do not bind or only weakly bind to target surfaces is difficult and the overall experimental procedure is labor-intensive. Despite the advantages of such microfluidic methods and successful demonstration of their utility, these methods still require several rounds of iterative biopanning. In this work, a magnetophoretic microfluidic biopanning platform was developed to isolate microorganisms that bind to target materials of interest, which is gold in this case. To achieve this, gold-coated magnetic nanobeads, which only attached to microorganisms that exhibit high affinity to gold, were used. The platform was first utilized to screen a bacterial peptide display library, where only the cells with surface peptides that specifically bind to gold could be isolated by the high-gradient magnetic field generated within the microchannel, resulting in enrichment and isolation of many isolates with high affinity and high specificity toward gold even after only a single round of separation. The amino acid profile of the resulting isolates was analyzed to provide a better understanding of the distinctive attributes of peptides that contribute to their specific material-binding capabilities. Next, the microfluidic system was utilized to screen soil microbes, a rich source of extremely diverse microorganisms, successfully isolating many naturally occurring microorganisms that show strong and specific binding to gold. The results show that the developed microfluidic platform is a powerful screening tool for identifying microorganisms that specifically bind to a target material surface of interest, which can greatly accelerate the development of new peptide-driven biological materials and hybrid organic-inorganic materials.

Testing of drugs using human feto-maternal interface organ-on-chips provide insights into pharmacokinetics and efficacy.

Objectives: To improve preclinical drug testing during pregnancy, we developed multiple microfluidic organ-on-chip (OOC) devices that represent the structure, functions, and responses of the two feto-maternal interfaces (FMis) in humans (fetal membrane [FMi-OOC] and placenta [PLA-OOC]). This study utilized feto-maternal interface OOCs to test the kinetics and efficacy of drugs during pregnancy. Study design: The FMi-OOC contained amnion epithelial, mesenchymal, chorion trophoblast, and decidual cells. The PLA-OOC contained cytotrophoblasts (BeWo), syncytiotrophoblasts (BeWo + forskolin), and human umbilical vein endothelial cell lines. Therapeutic concentrations of either pravastatin or rosuvastatin (200 ng mL-1), a model drug for these experiments, were applied to either decidua (in FMi-OOC) and syncytiotrophoblasts (in PLA-OOC) chambers under normal and oxidative stress conditions (induced by cigarette smoke extract [CSE 1 : 25]) to evaluate maternal drug exposure during normal pregnancy or oxidative stress (OS) associated pathologies, respectively. We determined statin pharmacokinetics and metabolism (LC-MS/MS), drug-induced cytotoxicity (LDH assay), and efficacy to reduce OS-induced inflammation (multiplex cytokine assay). Results: Both OOCs mimicked two distinct human feto-maternal interfaces. The drugs tested permeated the maternal-fetal cell layers of the FMi-OOC and PLA-OOC within 4 hours and generated cell and time-specific statin metabolites from various cell types without causing any cytotoxicity. OS-induced pro-inflammatory cytokines were effectively reduced by statins by increasing anti-inflammatory cytokine response across the FMi-OOC and PLA-OOC. Conclusion: Two distinct feto-maternal interface OOCs were developed, tested, and validated for their utility to conduct preclinical trials during pregnancy. We demonstrated that the placenta and fetal membranes-decidual interface both are able to transport and metabolize drugs and that the safety and efficacy of a drug can be determined using the anatomical structures recreated on OOCs.

Stress signaler p38 mitogen-activated kinase activation: a cause for concern?

Oxidative stress (OS) induced activation of p38 mitogen-activated kinase (MAPK) and cell fate from p38 signaling was tested using the human fetal membrane's amnion epithelial cells (AEC). We created p38 KO AEC using the CRISPR/Cas9 approach and tested cell fate in response to OS on an AEC-free fetal membrane extracellular matrix (ECM). Screening using image CyTOF indicated OS causing epithelial-mesenchymal transition (EMT). Further testing revealed p38 deficiency prevented AEC senescence, EMT, cell migration, and inflammation. To functionally validate in vitro findings, fetal membrane-specific conditional KO (cKO) mice were developed by injecting Cre-recombinase encoded exosomes intra-amniotically into p38αloxP/loxP mice. Amnion membranes from p38 cKO mice had reduced senescence, EMT, and increased anti-inflammatory IL-10 compared with WT animals. Our study suggested that overwhelming activation of p38 in response to OS inducing risk exposures can have an adverse impact on cells, cause cell invasion, inflammation, and ECM degradation detrimental to tissue homeostasis.

Chronic health effects associated with electronic cigarette use: A systematic review.

Introduction: Over the last decade, e-cigarette use has been on the rise but with growing health concerns. The objective of this systematic review was to update findings for chronic health outcomes associated with e-cigarette use from the 2018 National Academies of Sciences, Engineering, and Medicine (NASEM) report. Methods: Three bibliographic databases were searched to identify studies comparing the chronic health effects of e-cigarette users (ECU) to non-smokers (NS), smokers, and/or dual users indexed between 31 August 2017 and 29 January 2021. Two independent reviewers screened abstracts and full texts. Data were extracted by one reviewer and verified by a second one. Outcomes were synthesized in a narrative manner using counts and based on statistical significance and direction of the association stratified by study design and exposure type. Risk of bias and certainty of evidence was assessed. The protocol was prospectively registered on Open Science Framework https://osf.io/u9btp. Results: A total of 180 articles were eligible. This review focused on 93 studies for the 11 most frequently reported outcomes and from which 59 reported on daily e-cigarette use. The certainty of evidence for all outcomes was very low because of study design (84% cross-sectional) and exposure type (27% reported on exclusive ECU, i.e., never smoked traditional cigarettes). Overall, the summary of results for nearly all outcomes, including inflammation, immune response, periodontal and peri-implant clinical parameters, lung function, respiratory symptoms, and cardiovascular disease, suggested either non-significant or mixed results when daily ECU was compared to NS. This was also observed when comparing exclusive ECU to NS. The only notable exception was related to oral health where most (11/14) studies reported significantly higher inflammation among daily ECU vs. NS. Compared to the smokers, the exclusive-ECUs had no statistically significant differences in inflammation orperiodontal clinical parameters but had mixed findings for peri-implant clinical parameters. Conclusions: This review provides an update to the 2018 NASEM report on chronic health effects of e-cigarette use. While the number of studies has grown, the certainty of evidence remains very low largely because of cross-sectional designs and lack of reporting on exclusive e-cigarette exposure. There remains a need for higher quality intervention and prospective studies to assess causality, with a focus on exclusive e-cigarette use.

ZrO2/ZnO/TiO2 Nanocomposite Coatings on Stainless Steel for Improved Corrosion Resistance, Biocompatibility, and Antimicrobial Activity.

The ultrathin nanocomposite coatings made of zirconium oxide (ZrO2), zinc oxide (ZnO), and titanium oxide (TiO2) on stainless steel (SS) were prepared by the radio frequency sputtering method, and the effects of the nanocomposite coating on corrosion protection and antibacterial activities of nanocomposite coated SS were investigated. Scanning electron microscopy was conducted to observe surface morphology of nanocomposite coatings with distinct distribution of grains with the formation on SS substrate. From the electrochemical impedance spectroscopy results, ZrO2/ZnO/TiO2 nanocomposite coating showed excellent corrosion protection performance at 37 °C during immersion in simulated body fluid and saliva solution for 12 and 4 weeks, respectively. The impedance of ZrO2/ZnO/TiO2 (40/10/50) nanocomposite coated SS exhibited values about 5 orders of magnitude higher than that of uncoated SS with polarization at the low-frequency region. Cell viability of ZrO2/ZnO/TiO2 nanocomposite coated SS was examined under mouse fibroblasts culture (L929), and it was observed that the nanocomposite coating improves proliferation through effective cellular attachment compared to uncoated SS. From the antimicrobial activity results, ZrO2/ZnO/TiO2 nanocomposite-coated SS showed killing efficiency of 81.2% and 72.4% against Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus, respectively.

Molecular mechanisms of environmental toxin cadmium at the feto-maternal interface investigated using an organ-on-chip (FMi-OOC) model.

Human labor is associated with feto-maternal-derived signals that coordinate to initiate delivery. Exposure to environmental chemicals can prematurely trigger labor-initiating signals at the feto-maternal interface (FMi: decidua, amniochorion), leading to spontaneous preterm birth (PTB). Testing the association between environmental chemical exposure and PTB is difficult due to many limitations in vivo or in vitro. Physiological organ-on-chips (OOCs) are potential alternatives for studying mechanisms leading to PTB. The presented study tested the effect of maternal exposure to cadmium (Cd), an environmental toxin, using the FMi-OOC that incorporates maternal decidua cells and three different fetal cells (chorion, amnion mesenchymal, and amnion epithelial cells). Cd transport through the FMi and its impact on cell cycle, cell death, and inflammation were analyzed. Cd treatment resulted in significant cell death and a pro-inflammatory environment in the maternal decidua, but had minimal effect on the fetal chorion cells, and no effect in the fetal amnion cells compared to controls. The maternal response, but lack of fetal response, indicates that Cd-mediated adverse effects originate from maternal pathophysiology rather than fetal-derived triggers of preterm labor. This study demonstrates that the FMi-OOC can indeed predict the response of FMi upon exposure to chemicals, opening the possibility for using OOC models for environmental toxin screens.

Generation and characterization of human Fetal membrane and Decidual cell lines for reproductive biology experiments†.

Human fetal membrane and maternal decidua parietalis form one of the major feto-maternal interfaces during pregnancy. Studies on this feto-maternal interface is limited as several investigators have limited access to the placenta, and experience difficulties to isolate and maintain primary cells. Many cell lines that are currently available do not have the characteristics or properties of their primary cells of origin. Therefore, we created, characterized the immortalized cells from primary isolates from fetal membrane-derived amnion epithelial cells, amnion and chorion mesenchymal cells, chorion trophoblast cells and maternal decidua parietalis cells. Primary cells were isolated from a healthy full-term, not in labor placenta. Primary cells were immortalized using either a HPV16E6E7 retroviral or a SV40T lentiviral system. The immortalized cells were characterized for the morphology, cell type-specific markers, and cell signalling pathway activation. Genomic stability of these cells was tested using RNA seq, karyotyping, and short tandem repeats DNA analysis. Immortalized cells show their characteristic morphology, and express respective epithelial, mesenchymal and decidual markers similar to that of primary cells. Gene expression of immortalized and primary cells were highly correlated (R = 0.798 to R = 0.974). Short tandem repeats DNA analysis showed in the late passage number (>P30) of cell lines matched 84-100% to the early passage number (

A scalable system for generation of mesenchymal stem cells derived from induced pluripotent cells employing bioreactors and degradable microcarriers.

Human mesenchymal stem cells (hMSCs) are effective in treating disorders resulting from an inflammatory or heightened immune response. The hMSCs derived from induced pluripotent stem cells (ihMSCs) share the characteristics of tissue derived hMSCs but lack challenges associated with limited tissue sources and donor variation. To meet the expected future demand for ihMSCs, there is a need to develop scalable methods for their production at clinical yields while retaining immunomodulatory efficacy. Herein, we describe a platform for the scalable expansion and rapid harvest of ihMSCs with robust immunomodulatory activity using degradable gelatin methacryloyl (GelMA) microcarriers. GelMA microcarriers were rapidly and reproducibly fabricated using a custom microfluidic step emulsification device at relatively low cost. Using vertical wheel bioreactors, 8.8 to 16.3-fold expansion of ihMSCs was achieved over 8 days. Complete recovery by 5-minute digestion of the microcarriers with standard cell dissociation reagents resulted in >95% viability. The ihMSCs matched or exceeded immunomodulatory potential in vitro when compared with ihMSCs expanded on monolayers. This is the first description of a robust, scalable, and cost-effective method for generation of immunomodulatory ihMSCs, representing a significant contribution to their translational potential.

Fabrication methods for a gel-based separation-free device for whole blood glucose detection.

In this paper, we describe two fabrication methods (well array-based and biopsy punching-based) of gel disks to construct a gel-based point-of-care (POC) diagnosis device for direct colorimetric measurement of human whole blood glucose without any extra blood separation step. The gel disks are made of Polyethylene glycol (PEG) diacrylate (PEG-DA) containing immobilized glucose colorimetric assay reagents. The performances of three types of PEG-DA gel (molecular weight: 575, 3,400, and 10,000) based sensors as well as the two fabrication methods were investigated.•The fabricated devices enabled colorimetric whole blood glucose sensing assay without the need for blood cell separation•The biopsy punching-based gel disk fabrication method provided less variation on the fabricated gel disks.

Organ-on-chip of the cervical epithelial layer: A platform to study normal and pathological cellular remodeling of the cervix.

Damage to the cervical epithelial layer due to infection and inflammation is associated with preterm birth. However, the individual and/or collective roles of cervical epithelial layers in maintaining cervical integrity remain unclear during infection/inflammation. To determine the intercellular interactions, we developed an organ-on-chip of the cervical epithelial layer (CE-OOC) composed of two co-culture chambers connected by microchannels, recapitulating the ectocervical and endocervical epithelial layers. Further, we tested the interactions between cells from each distinct region and their contributions in maintaining cervical integrity in response to LPS and TNFα stimulations. The co-culture of ectocervical and endocervical cells facilitated cellular migration of both epithelial cells inside the microchannels. Compared to untreated controls, both LPS and TNFα increased apoptosis, necrosis, and senescence as well as increased pro-inflammatory cytokine productions by cervical epithelial cells. In summary, the CE-OOC established an in vitro model that can recapitulate the ectocervical and the endocervical epithelial regions of the cervix. The established CE-OOC may become a powerful tool in obstetrics and gynecology research such as in studying cervical remodeling during pregnancy and parturition and the dynamics of cervical epithelial cells in benign and malignant pathology in the cervix.

Organic Anion Transporting Polypeptide 2B1 in Human Fetal Membranes: A Novel Gatekeeper for Drug Transport During Pregnancy?

Current intervention strategies have not been successful in reducing the risks of adverse pregnancy complications nor maternal and fetal morbidities associated with pregnancy complications. Improving pregnancy and neonatal outcomes requires a better understanding of drug transport mechanisms at the feto-maternal interfaces, specifically the placenta and fetal membrane (FM). The role of several solute carrier uptake transporter proteins (TPs), such as the organic anion transporting polypeptide 2B1 (OATP2B1) in transporting drug across the placenta, is well-established. However, the mechanistic role of FMs in this drug transport has not yet been elucidated. We hypothesize that human FMs express OATP2B1 and functions as an alternate gatekeeper for drug transport at the feto-maternal interface. We determined the expression of OATP2B1 in term, not-in-labor, FM tissues and human FM cells [amnion epithelial cell (AEC), chorion trophoblast cell (CTC), and mesenchymal cells] using western blot analyses and their localization using immunohistochemistry. Changes in OATP2B1 expression was determined for up to 48 h after stimulation with cigarette smoke extract (CSE), an inducer of oxidative stress. The functional role of OATP2B1 was determined by flow cytometry using a zombie violet dye substrate assay. After OATP2B1 gene silencing, its functional relevance in drug transport through the feto-maternal interface was tested using a recently developed feto-maternal interface organ-on-a-chip (OOC) system that contained both FM and maternal decidual cells. Propagation of a drug (Rosuvastatin, that can be transported by OATP2B1) within the feto-maternal interface OOC system was determined by mass spectrometry. FMs express OATP2B1 in the CTC and AEC layers. In FM explants, OATP2B1 expression was not impacted by oxidative stress. Uptake of the zombie violet dye within AECs and CTCs showed OATP2B1 is functionally active. Silencing OATP2B1 in CTCs reduced Rosuvastatin propagation from the decidua to the fetal AEC layer within the feto-maternal interface-OOC model. Our data suggest that TPs in FMs may function as a drug transport system at the feto-maternal interface, a function that was previously thought to be performed exclusively by the placenta. This new knowledge will help improve drug delivery testing during pregnancy and contribute to designing drug delivery strategies to treat adverse pregnancy outcomes.

Calreticulin is a Critical Cell Survival Factor in Malignant Neoplasms.

Calreticulin (CRT) is a high-capacity Ca2+ protein whose expression is up-regulated during cellular transformation and is associated with disease progression in multiple types of malignancies. At the same time, CRT has been characterized as an important stress-response protein capable of inducing immunogenic cell death (ICD) when translocated to the cell surface. It remains unclear why CRT expression is preserved by malignant cells during the course of transformation despite its immunogenic properties. In this study, we identify a novel, critical function of CRT as a cell survival factor in multiple types of human solid-tissue malignancies. CRT knockdown activates p53, which mediates cell-death response independent of executioner caspase activity and accompanied full-length poly ADP ribose polymerase (PARP) cleavage. Mechanistically, we show that down-regulation of CRT results in mitochondrial Ca2+ overload and induction of mitochondria permeability transition pore (mPTP)-dependent cell death, which can be significantly rescued by the mPTP inhibitor, Cyclosporin A (CsA). The clinical importance of CRT expression was revealed in the analysis of the large cohort of cancer patients (N = 2,058) to demonstrate that high levels of CRT inversely correlates with patient survival. Our study identifies intracellular CRT as an important therapeutic target for tumors whose survival relies on its expression.

Amnion membrane organ-on-chip: an innovative approach to study cellular interactions.

The amnion membrane that lines the human intrauterine cavity is composed of amnion epithelial cells (AECs) connected to an extracellular matrix containing amnion mesenchymal cells (AMCs) through a basement membrane. Cellular interactions and transitions are mechanisms that facilitate membrane remodeling to maintain its integrity. Dysregulation of cellular remodeling, primarily mediated by oxidative stress (OS), is often associated with preterm birth. However, the mechanisms that maintain membrane homeostasis remain unclear. To understand these mechanisms, we developed an amnion membrane organ-on-chip (AM-OOC) and tested the interactive and transition properties of primary human AECs and AMCs under normal and OS conditions. AM-OOC contained 2 chambers connected by type IV collagen-coated microchannels, allowing independent culture conditions that permitted cellular migration and interactions. Cells grown either independently or coculture were exposed to OS inducing cigarette smoke extract, antioxidant N-acetyl-l-cysteine (NAC), or both. When grown independently, AECs transitioned to AMCs and migrated, whereas AMCs migrated without transition. OS caused AECs' transition but prevented migration, whereas AMCs' migration was unhindered. Coculture of cells facilitated transition, migration, and eventual integration in the contiguous population. OS cotreatment in both chambers facilitated AECs' transition, prevented migration, and increased inflammation, a process that was prevented by NAC. AM-OOC recapitulated cellular mechanisms observed in utero and enabled experimental manipulation of cells to determine their roles during pregnancy and parturition.-Richardson, L., Jeong, S., Kim, S., Han, A., Menon, R. Amnion membrane organ-on-chip: an innovative approach to study cellular interactions.

A continuous-flow acoustofluidic cytometer for single-cell mechanotyping.

The biophysical properties of cells such as their compressibility have been found to be closely related to disease progression such as cancer development and metastasis. As cancer cells are heterogeneous, rapid and high-throughput evaluation of cell biophysical properties at single-cell resolution is needed to assess their potential as biomarkers for cancer staging and prognosis. Acoustofluidics has shown promise as a contactless method for accurately measuring cell biophysical properties; however, previously reported methods had relatively low throughput due to their requirement of no-flow conditions. This work presents a high-throughput continuous flow-based acoustofluidic cell mechanotyping method at single-cell resolution that retains the advantage of simplicity and low-cost.

Antibody Therapy Targeting CD47 and CD271 Effectively Suppresses Melanoma Metastasis in Patient-Derived Xenografts.

The high rate of metastasis and recurrence among melanoma patients indicates the existence of cells within melanoma that have the ability to both initiate metastatic programs and bypass immune recognition. Here, we identify CD47 as a regulator of melanoma tumor metastasis and immune evasion. Protein and gene expression analysis of clinical melanoma samples reveals that CD47, an anti-phagocytic signal, correlates with melanoma metastasis. Antibody-mediated blockade of CD47 coupled with targeting of CD271(+) melanoma cells strongly inhibits tumor metastasis in patient-derived xenografts. This therapeutic effect is mediated by drastic changes in the tumor and metastatic site immune microenvironments, both of whichwhich exhibit greatly increased density of differentiated macrophages and significantly fewer inflammatory monocytes, pro-metastatic macrophages (CCR2(+)/VEGFR1(+)), and neutrophils, all of which are associated with disease progression. Thus, antibody therapy that activates the innate immune response in combination with selective targeting of CD271(+) melanoma cells represents a powerful therapeutic approach against metastatic melanoma.

A droplet microfluidics platform for rapid microalgal growth and oil production analysis.

Microalgae have emerged as a promising source for producing future renewable biofuels. Developing better microalgal strains with faster growth and higher oil production rates is one of the major routes towards economically viable microalgal biofuel production. In this work, we present a droplet microfluidics-based microalgae analysis platform capable of measuring growth and oil content of various microalgal strains with single-cell resolution in a high-throughput manner. The platform allows for encapsulating a single microalgal cell into a water-in-oil emulsion droplet and tracking the growth and division of the encapsulated cell over time, followed by on-chip oil quantification. The key feature of the developed platform is its capability to fluorescently stain microalgae within microdroplets for oil content quantification. The performance of the developed platform was characterized using the unicellular microalga Chlamydomonas reinhardtii and the colonial microalga Botryococcus braunii. The application of the platform in quantifying growth and oil accumulation was successfully confirmed using C. reinhardtii under different culture conditions, namely nitrogen-replete and nitrogen-limited conditions. These results demonstrate the capability of this platform as a rapid screening tool that can be applied to a wide range of microalgal strains for analyzing growth and oil accumulation characteristics relevant to biofuel strain selection and development. Biotechnol. Bioeng. 2016;113: 1691-1701. © 2016 Wiley Periodicals, Inc.