Single-cell landscape of the ecosystem in early-relapse hepatocellular carcinoma

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Effects of detergents on catalytic activity of human endometase/matrilysin 2, a putative cancer biomarker

Matrix metalloproteinases (MMPs) are a family of hydrolytic enzymes that play significant roles in development, morphogenesis, inflammation, and cancer invasion. Endometase (matrilysin 2 or MMP-26) is a putative early biomarker for human carcinomas. The effects of the ionic and nonionic detergents on catalytic activity of endometase were investigated. The hydrolytic activity of endometase was detergent concentration dependent, exhibiting a bell-shaped curve with its maximum activity near the critical micelle concentration (CMC) of nonionic detergents tested. The effect of Brij-35 on human gelatinase B (MMP-9), matrilysin (MMP-7), and membrane-type 1 MMP (MT1-MMP) was further explored. Their maximum catalysis was observed near the CMC of Brij-35 (∼ 90 μM). Their IC₅₀ values were above the CMC. The inhibition mechanism of MMP-7, MMP-9, and MT1-MMP by Brij-35 was a mixed type as determined by Dixon’s plot; however, the inhibition mechanism of endometase was noncompetitive with a K_i value of 240 μM. The catalytic activities of MMPs are influenced by detergents. Monomer of detergents may activate and stabilize MMPs to enhance catalysis, but micelle of detergents may sequester enzyme and block the substrate binding site to impede catalysis. Under physiological conditions, a lipid or membrane microenvironment may regulate enzymatic activity.     

Glioma-Derived miRNA-Containing Extracellular Vesicles Induce Angiogenesis by Reprogramming Brain Endothelial Cells

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Three-dimensional perfused cell culture

Compelling evidence suggests the limitation and shortcomings of the current and well established cell culture method using multi-well plates, flasks and Petri dishes. These are particularly important when cell functions are sensitive to the local microenvironment, cell–cell and cell–extracellular matrix interactions. There is a clear need for advanced cell culture systems which mimic in vivo and more physiological conditions. This review summarises and analyses recent progress in three dimensional (3D) cell culture with perfusion as the next generation cell culture tools, while excluding engineered tissue culture where three dimensional scaffold has to be used for structural support and perfusion for overcoming mass transfer control. Apart from research activities in academic community, product development in industry is also included in this review.     

Fluid shear stress regulates HepG2 cell migration though time-dependent integrin signaling cascade

Hepatocellular carcinoma (HCC) is a subtype of malignant liver cancer with poor prognosis and limited treatment options. It is noteworthy that mechanical forces in tumor microenvironment play a pivotal role in mediating the behaviors and functions of tumor cells. As an instrumental type of mechanical forces in vivo, fluid shear stress (FSS) has been reported having potent physiologic and pathologic effects on cancer progression. However, the time-dependent mechanochemical transduction in HCC induced by FSS remains unclear. In this study, hepatocellular carcinoma HepG2 cells were exposed to 1.4 dyn/cm² FSS for transient duration (15s and 30s), short duration (5 min, 15 min and 30 min) and long duration (1h, 2h and 4h), respectively. The expression and translocation of Integrins induced FAK-Rho GTPases signaling events were examined. Our results showed that FSS endowed HepG2 cells with higher migration ability via reorganizing cellular F-actin and disrupting intercellular tight junctions. We further demonstrated that FSS regulated the expression and translocation of Integrins and their downstream signaling cascade in time-dependent patterns. The FSS downregulated focal adhesion components (Paxillin, Vinculin and Talin) while upregulated the expression of Rho GTPases (Cdc42, Rac1 and RhoA) in long durations. These results indicated that FSS enhanced tumor cell migration through Integrins-FAK-Rho GTPases signaling pathway in time-dependent manners. Our in vitro findings shed new light on the role of FSS acting in physiologic and pathological processes during tumor progression, which has emerged as a promising clinical strategy for liver carcinoma.     

Investigating the effect of cell substrate on cancer cell stiffness by optical tweezers

The mechanical properties of cells are influenced by their microenvironment. Here we report cell stiffness alteration by changing the cell substrate stiffness for isolated cells and cells in contact with other cells. Polydimethylsiloxane (PDMS) is used to prepare soft substrates with three different stiffness values (173, 88 and 17 kPa respectively). Breast cancer cells lines, namely HBL-100, MCF-7 and MDA-MB-231 with different level of aggressiveness are cultured on these substrates and their local elasticity is investigated by vertical indentation of the cell membrane. Our preliminary results show an unforeseen behavior of the MDA-MB-231 cells. When cultured on glass substrate as isolated cells, they are less stiff than the other two types of cells, in agreement with the general statement that more aggressive and metastatic cells are softer. However, when connected to other cells the stiffness of MDA-MB-231 cells becomes similar to the other two cell lines. Moreover, the stiffness of MDA-MB-231 cells cultured on soft PDMS substrates is significantly higher than the stiffness of the other cell types, demonstrating thus the strong influence of the environmental conditions on the mechanical properties of the cells.     

Culture of Amelanchier x grandiflora in a programmable micropropagation apparatus

A micropropagation system was developed to test concepts for automation and microenvironment alteration. Amelanchier x grandiflora Rehd. Princess Diana (serviceberry) shoot cultures were grown in seven-liter polycarbonate containers. Through computer control, the apparatus intermittently applied culture medium to the plant material according to a selected schedule. Shoot growth in the programmable system was compared with four micropropagation treatments: gelled and liquid medium in baby food jars and gelled and non-cycling liquid medium in a seven-liter vessel. Plants cultured in continuous contact with liquid medium became increasingly vitrified. Significantly greater shoot number, shoot length, shoot weight, and culture weight occurred in the programmable system than in jars with gelled medium. The combination of liquid medium, 7-liter vessel, and intermittent contact with medium caused a significantly higher proliferation rate than any combination of jar/vessel and gelled/liquid medium tested.     

PYK2 is overexpressed in chronic lymphocytic leukaemia: A potential new therapeutic target

Chronic Lymphocytic Leukaemia (CLL) is the most common adult B-cell leukaemia and despite improvement in patients' outcome, following the use of targeted therapies, it remains incurable. CLL supportive microenvironment plays a key role in both CLL progression and drug resistance through signals that can be sensed by the main components of the focal adhesion complex, such as FAK and PYK2 kinases. Dysregulations of both kinases have been observed in several metastatic cancers, but their role in haematological malignancies is still poorly defined. We characterized FAK and PYK2 expression and observed that PYK2 expression is higher in leukaemic B cells and its overexpression significantly correlates with their malignant transformation. When targeting both FAK and PYK2 with the specific inhibitor defactinib, we observed a dose–response effect on CLL cells viability and survival. In vivo treatment of a CLL mouse model showed a decrease of the leukaemic clone in all the lymphoid organs along with a significant reduction of macrophages and of the spleen weight and size. Our results first define a possible prognostic value for PYK2 in CLL, and show that both FAK and PYK2 might become putative targets for both CLL and its microenvironment (e.g. macrophages), thus paving the way to an innovative therapeutic strategy.     

Pan-Cancer analyses of Necroptosis-Related genes as a potential target to predict immunotherapeutic outcome

Necroptosis is a unique programmed death mechanism of necrotic cells. However, its role and specific mechanism in cancer remain unclear, and a systematic pan-cancer analysis of necroptosis is yet to be conducted. Thus, we performed a specific pan-cancer analysis using The Cancer Genome Atlas and Genotype-Tissue Expression databases to analyse necroptosis expression in terms of cancer prognosis, DNA methylation status, tumour mutative burden, microsatellite instability, immune cell infiltration in different types of cancer and molecular mechanisms. For the first time, we explored the correlation between necroptosis and immunotherapy prognosis. Thus, our study provides a relatively comprehensive understanding of the carcinogenicity of necroptosis in different types of cancer. It is suggested that necroptosis can be used to evaluate the sensitivity of different patients to immunotherapy and may become a potential target for tumour immunotherapy.     

Tissue microenvironment and the genetic control of hair pigment patterns in mice

This study was conducted to assess microenvironmental variability within integumental tissue of genetically identical mice with respect to a specific cellular response: cyclic synthesis of yellow and black pigment by hair bulb melanocytes. Crosses were performed within and between inbred strains of mice that were isogenic with the exception of a single gene substitution at the agouti locus. Agouti locus genes included the A^vy, A^w, A, a^td, a^t, a^x, a^m, and a alleles. The pigment patterns of dorsal, flank, and ventral hairs of the first and third hair generations and of hairs growing in special integumentary areas such as the pinna, tail, and hind foot were studied. It was found that the amount of yellow pigment synthesized by hair bulb melanocytes within genetically identical mice is both agedependent and conditioned by the integumentary environment. Furthermore, the special integumentary regions produce hairs with a variety of pigment patterns in which the distribution and relative amounts of black and yellow pigments do not necessarily conform to dominance relationships expected among agouti locus alleles as judged by their effects on the pigmentation of the dorsal pelage. We conclude that within genetically uniform integumental tissues, microenvironmental differences occur and are reflected as alterations in the metabolic pattern of differentiated cells.