A tissue-engineered model of the blood-tumor barrier during metastatic breast cancer.

Metastatic brain cancer has poor prognosis due to challenges in both detection and treatment. One contributor to poor prognosis is the blood-brain barrier (BBB), which severely limits the transport of therapeutic agents to intracranial tumors. During the development of brain metastases from primary breast cancer, the BBB is modified and is termed the 'blood-tumor barrier' (BTB). A better understanding of the differences between the BBB and BTB across cancer types and stages may assist in identifying new therapeutic targets. Here, we utilize a tissue-engineered microvessel model with induced pluripotent stem cell (iPSC)-derived brain microvascular endothelial-like cells (iBMECs) and surrounded by human breast metastatic cancer spheroids with brain tropism. We directly compare BBB and BTB in vitro microvessels to unravel both physical and chemical interactions occurring during perivascular cancer growth. We determine the dynamics of vascular co-option by cancer cells, modes of vascular degeneration, and quantify the endothelial barrier to antibody transport. Additionally, using bulk RNA sequencing, ELISA of microvessel perfusates, and related functional assays, we probe early brain endothelial changes in the presence of cancer cells. We find that immune cell adhesion and endothelial turnover are elevated within the metastatic BTB, and that macrophages exert a unique influence on BTB identity. Our model provides a novel three-dimensional system to study mechanisms of cancer-vascular-immune interactions and drug delivery occurring within the BTB.

Discovery of 4'-trifluoromethylchalcones as novel, potent and selective hCYP1B1 inhibitors without concomitant AhR activation.

Human cytochrome P450 1B1 (hCYP1B1), an extrahepatic cytochrome P450 enzyme over-expressed in various tumors, has been validated as a promising target for preventing and treating cancers. Herein, two series of chalcone derivatives were synthesized to discover potent hCYP1B1 inhibitors without AhR agonist effect. Structure-activity relationship (SAR) studies demonstrated that 4'-trifluoromethyl on the B-ring strongly enhanced the anti-hCYP1B1 effects, identifying A9 as a promising lead compound. Further SAR analysis on A9 derivatives (modified A-ring of 4'-trifluoromethylchalcone) showed that introducing 2-methoxyl improved the anti-hCYP1B1 effect and selectivity, while introducing a methoxyl at the C-4 site was beneficial for avoiding AhR activation. Ultimately, five 4'-trifluoromethyl chalcones were identified as potent hCYP1B1 inhibitors (IC50 < 10 nM), while B18 exhibits the most potent anti-hCYP1B1 effect (IC50 = 3.6 nM), suitable metabolic stability and good cell-permeability. B18 also acted as an AhR antagonist and could down-regulate hCYP1B1 in living systems. Mechanistic studies showed that B18 potently inhibited hCYP1B1 in a competitive inhibition manner (Ki = 3.92 nM), while docking simulations revealed that B18 could tightly bind to the catalytic cavity of hCYP1B1 mainly via hydrophobic and hydrogen-bonding interactions. Furthermore, B18 could potently inhibit hCYP1B1 in living cells and showed remarkable anti-migration ability on MFC-7 cells. Taken together, this study deciphered the SARs of chalcones as hCYP1B1 inhibitors and provided several potent hCYP1B1 inhibitors as promising candidates for the development of more efficacious anti-migration agents.

Effect of Cooking Method and Doneness Degree on Volatile Compounds and Taste Substance of Pingliang Red Beef.

This study used gas chromatography-ion mobility spectrometry (GC-IMS) and high-performance liquid chromatography (HPLC) methods to examine the impact of cooking methods and doneness on volatile aroma compounds and non-volatile substances (fatty acids, nucleotides, and amino acids) in Pingliang red beef. The flavor substances' topographic fingerprints were established, and 45 compounds were traced to 71 distinct signal peaks. Pingliang red beef's fruity flavor was enhanced thanks to the increased concentration of hexanal, styrene, and 2-butanone that resulted from instant boiling. The levels of 3-methylbutanal, which contributes to the characteristic caramel-chocolate-cheese aroma, peaked at 90 min of boiling and 40 min of roasting. The FFA content was reduced by 28.34% and 27.42%, respectively, after the beef was roasted for 40 min and instantly boiled for 10 s (p > 0.05). The most distinctive feature after 30 min of boiling was the umami, as the highest levels of glutamate (Glu) (p < 0.05) and the highest equivalent umami concentration (EUC) values were obtained through this cooking method. Additionally, adenosine-5'-monophosphate (AMP) and inosine-5'-monophosphate (IMP) decreased with increasing doneness compared to higher doneness, indicating that lower doneness was favorable in enhancing the umami of the beef. In summary, different cooking methods and doneness levels can affect the flavor and taste of Pingliang red beef, but it is not suitable for high-doneness cooking.

Engineering the human blood-brain barrier at the capillary scale using a double-templating technique.

In vitro blood-brain barrier (BBB) models have played an important role in studying processes such as immune cell trafficking and drug delivery, as well as contributing to the understanding of mechanisms of disease progression. Many biological and pathological processes in the cerebrovasculature occur in capillaries and hence the lack of robust hierarchical models at the capillary scale is a major roadblock in BBB research. Here we report on a double-templating technique for engineering hierarchical BBB models with physiological barrier function at the capillary scale. We first demonstrate the formation of hierarchical vascular networks using human umbilical vein endothelial cells. We then characterize barrier function in a BBB model using brain microvascular endothelial-like cells (iBMECs) differentiated from induced pluripotent stem cells (iPSCs). Finally, we characterize immune cell adhesion and transmigration in response to perfusion with the inflammatory cytokine tumor necrosis factor-alpha, and show that we can recapitulate capillary-scale effects, such as leukocyte plugging, observed in mouse models. Our double-templated hierarchical model enables the study of a wide range of biological and pathological processes related to the human BBB.

Optical Cellular Micromotion: A New Paradigm to Measure Tumor Cells Invasion within Gels Mimicking the 3D Tumor Environments.

Measuring tumor cell invasiveness through 3D tissues, particularly at the single-cell level, can provide important mechanistic understanding and assist in identifying therapeutic targets of tumor invasion. However, current experimental approaches, including standard in vitro invasion assays, have limited physiological relevance and offer insufficient insight into the vast heterogeneity in tumor cell migration through tissues. To address these issues, here the concept of optical cellular micromotion is reported on, where digital holographic microscopy is used to map the optical nano- to submicrometer thickness fluctuations within single-cells. These fluctuations are driven by the dynamic movement of subcellular structures including the cytoskeleton and inherently associated with the biological processes involved in cell invasion within tissues. It is experimentally demonstrated that the optical cellular micromotion correlates with tumor cells motility and invasiveness both at the population and single-cell levels. In addition, the optical cellular micromotion significantly reduced upon treatment with migrastatic drugs that inhibit tumor cell invasion. These results demonstrate that micromotion measurements can rapidly and non-invasively determine the invasive behavior of single tumor cells within tissues, yielding a new and powerful tool to assess the efficacy of approaches targeting tumor cell invasiveness.

Self-Organization Formation of Multicellular Spheroids Mediated by Mechanically Tunable Hydrogel Platform: Toward Revealing the Synergy of Chemo- and Noninvasive Photothermal Therapy against Colon Microtumor.

Three-dimensional (3D) tumor cell culture offers a more tissue-recapitulating model in cancer treatment evaluation. However, conventional models based on cell-substrate adhesion deprivation are still of insufficient real tumor mimic. In this work, a novel method is proposed for inducing multicellular spheroids (MCSs) formation based on hydrogel with tunable microenvironmental properties. Colon tumor cells DLD1 cultured on hydrogel substrate with proper physical stimulation form MCSs via self-organization. Chemotherapy based on clinical drug and far-infrared photothermal therapy is evaluated with DLD1 MCSs obtained by this method. The synergism of chemotherapy and noninvasive photothermal therapy based on graphene device is further verified in MCSs model and it is believed this method holds potential in in vitro anti-tumor strategies evaluation for precision medicine.

[Time-series characteristics of drought and flood in spring soybean growing season and its effect on soybean yield in Heilongjiang Province, China]. / é»‘é¾™æ±Ÿçœå¤§è±†ç”Ÿé•¿å­£æ—±æ¶æ—¶åºç‰¹å¾åŠå ¶å¯¹äº§é‡çš„å½±å“.

Under the background of climate change, the spatial-temporal distribution of precipita-tion in Heilongjiang Province is uneven, and drought and flood frequently change, which is not conducive to the safety of soybean production for the province. To clarify the influence mechanism of drought and flood in the growing season on soybean yield in Heilongjiang Province, we analyzed the time-series characteristics of drought and flood in soybean growing season and its effect on soybean yield in different growth stages, based on data of daily precipitation from 60 meteorological stations during 1961 to 2018 and soybean yield in the same period, with the standardized precipitation index (SPI) as the drought and flood evaluation index. The results showed that, from 1961 to 2018, the influence range of drought in soybean growing season in Heilongjiang Province showed a weak decreasing trend, while that of flood showed a weak increasing trend. In the same period, the intensity of both drought and flood showed a weak increasing trend, with slightly stronger role of flood intensity. The probability of the co-occurrence of drought and flood accounted for 60.3%. The soybean growing season in Heilongjiang Province may become wetter. From 2012 to 2018, the influence range and occurrence intensity of flood were significantly higher than that of drought, six years of the whole or regional flood occurred, in which five years were moderate degrees. The effects of drought and flood on soybean yield differed across regions in soybean growing season. The effect of flood on soybean yield was significantly stronger than that of drought in the Northwest, North and East, and were similar in the Midland, while in the Southwest, South and Southeast, the effect of drought was much greater than that of flood. The fluctuation of soybean yield was closely related to drought and flood during bloom-seed-filling period. Among them, in the Northwest, Southwest, Midland, South and Southeast of Heilongjiang, soybean yield would reach a high level when there was a little bit more precipitation, but the moderate and above-moderate levels of flood would cause the reduction. In the North, the fluctuation of soybean yield was mainly affected by flood, while in the East, the effects of drought and flood on soybean yield were similar.

Effects of iron-catalyzed and metmyoglobin oxidizing systems on biochemical properties of yak muscle myofibrillar protein.

The objective of this study was to compare the effects of two oxidation systems on the biochemical properties of yak myofibrillar protein (MP). Oxidation was induced by incubating MP with either an iron-catalyzed oxidizing system (IOS) or a metmyoglobin-oxidizing system (MOS). The following indicators of protein oxidation and protein degradation were analyzed. The carbonyl, disulfide bonds, dityrosine, and ß-sheet content increased markedly with oxidant concentration in both systems(P < .05), whereas the total sulfhydryl, surface hydrophobicity and α-helix content decreased significantly(P < .05). Furthermore, the MOS carbonyl formation rate was significantly faster than the IOS rate, and the MOS significantly affected the formation of disulfide bonds and inhibited the exposure of hydrophobic amino acids. Both oxidative systems promoted cross-linking of myosin heavy chains (MHCs) and action, but the degree of cross-linking in IOS was greater than that in MOS. MOS also promoted cross-linking of myosin light chains (MLCs). IOS and MOS produced mainly 20-25-kDa and 20-17-kDa MLC degradation products, respectively. In conclusion, oxidation caused cross-linking in MHCs or MLCs through disulfide bonds, but the extent of such cross-linking was oxidant dose-dependent and specific to each oxidizing system.

A systematic investigation of the effect of the fluid shear stress on Caco-2 cells towards the optimization of epithelial organ-on-chip models.

Epithelial cells experience constant mechanical forces, including fluid shear stress (FSS) on their apical surface. These forces alter both structure and function. While precise recapitulation of the complex mechanobiology of organs remains challenging, better understanding of the effect of mechanical stimuli is necessary towards the development of biorelevant in vitro models. This is especially relevant to organs-on-chip models which allow for fine control of the culture environment. In this study, the effects of the FSS on Caco-2 cell monolayers were systematically determined using a microfluidic device based on Hele-Shaw geometry. This approach allowed for a physiologically relevant range of FSS (from ∼0 to 0.03 dyn/cm2) to be applied to the cells within a single device. Exposure to microfluidic FSS induced significant phenotypical and functional changes in Caco-2 cell monolayers as compared to cells grown in static conditions. The application of FSS significantly altered the production of mucus, expression of tight junctions, vacuolization, organization of cytoskeleton, formation of microvilli, mitochondrial activity and expression of cytochrome P450. In the context of the intestinal epithelium, this detailed understanding of the effects of the FSS will enable the realization of in vitro organs-on-chip models with well-defined and tailored characteristics to a specific purpose, including for drug and nanoparticle absorption studies. The Hele-Shaw approach used in this study could be readily applied to other cell types and adapted for a wide range of physiologically relevant FSS.

Thermo-Sensitive PLGA-PEG-PLGA Tri-Block Copolymer Hydrogel as Three-Dimensional Cell Culture Matrix for Ovarian Cancer Cells.

Thermo-sensitive hydrogels which could encapsulate cells and provide a three dimensional (3D) microenvironment have great potential in building new cell culture models in vitro. In this study, a thermal responsive hydrogel based on PLGA-PEG-PLGA tri-block copolymers was developed as matrix for 3D ovarian cancer cell culturing. The gelation of PLGA-PEG-PLGA tri-block copolymer was concentration-dependent. SEM images showed the pores were suitable for the formation of 3D cell structures. Cell morphological results showed that large aggregates of ovarian cancer cells (HO8910) were formed after cultured for 10 days. Therefore, hydrogel based on PLGA-PEG-PLGA tri-block copolymers hold potential as in vitro cell culture matrix for ovarian cancer cells.

High Quality Multicellular Tumor Spheroid Induction Platform Based on Anisotropic Magnetic Hydrogel.

In recent years, multicellular spheroid (MCS) culture has been extensively studied both in fundamental research and application fields since it inherits much more characteristics from in vivo solid tumor than conventional two-dimensional (2D) cell culture. However, anticell adhesive MCS culture systems such as hanging drop allow certain cell lines only to form loose, irregular aggregates rather than MCS with physiological barriers and pathophysiological gradients, which failed to mimic in vivo solid tumor in these aspects. To address this issue, we improved our previously established anisotropic magnetic hydrogel platform, enabling it to generate multicellular spheroids with higher efficiency. The qualities of multicellular tumor spheroids (MCTSs) obtained on our platform and from classic 3D culture systems were compared in terms of morphology, biological molecule expression profiles, and drug resistance. In this novel platform, mature MCTSs with necrotic cores could be observed in 1 week. And results of molecular biological assays with real time-PCR and western-blot confirmed that MCTSs obtained from our platform performed higher cell pluripotency than those obtained from the hanging drop system. Moreover, a lower cell apoptosis ratio and better viability of cancer cells were observed on our platform both under culturing and drug treatment. In conclusion, higher quality of MCTSs obtained from this anisotropic magnetic hydrogel than classic hanging drop system validate its potential to be an in vitro platform of inducing tumor MCTS formation and drug efficacy evaluation.

Sliced Magnetic Polyacrylamide Hydrogel with Cell-Adhesive Microarray Interface: A Novel Multicellular Spheroid Culturing Platform.

Cell-adhesive properties are of great significance to materials serving as extracellular matrix mimics. Appropriate cell-adhesive property of material interface can balance the cell-matrix interaction and cell-cell interaction and can promote cells to form 3D structures. Herein, a novel magnetic polyacrylamide (PAM) hydrogel fabricated via combining magnetostatic field induced magnetic nanoparticles assembly and hydrogel gelation was applied as a multicellular spheroids culturing platform. When cultured on the cell-adhesive microarray interface of sliced magnetic hydrogel, normal and tumor cells from different cell lines could rapidly form multicellular spheroids spontaneously. Furthermore, cells which could only form loose cell aggregates in a classic 3D cell culture model (such as hanging drop system) were able to be promoted to form multicellular spheroids on this platform. In the light of its simplicity in fabricating as well as its effectiveness in promoting formation of multicellular spheroids which was considered as a prevailing tool in the study of the microenvironmental regulation of tumor cell physiology and therapeutic problems, this composite material holds promise in anticancer drugs or hyperthermia therapy evaluation in vitro in the future.

Fabrication of hydrogel with cell adhesive micropatterns for mimicking the oriented tumor-associated extracellular matrix.

For mimicking the fibrous extracellular matrix (ECM), a facile method for patterning anticell adhesive substrate was novelly applied on agarose hydrogel. Without using masks or templates for etching, we applied the magnetic field-induced colloidal assembly of magnetic nanoparticles on the flat agarose hydrogel to form cell-adhesive micropatterns. Meanwhile, tuning the hydrogel substrate's modulus to fit real tissue was experimentally demonstrated. Magnetic nanobeads were also assembled on this hydrogel surface and formed more complete and regular patterns. The patterned hydrogel substrate could actually influence behaviors of different cancer cells, including adhesion, growth, and migration.