An end-to-end software solution for the analysis of high-throughput single-cell migration data

The systematic study of single-cell migration requires the availability of software for assisting data inspection, quality control and analysis. This is especially important for high-throughput experiments, where multiple biological conditions are tested in parallel. Although the field of cell migration can count on different computational tools for cell segmentation and tracking, downstream data visualization, parameter extraction and statistical analysis are still left to the user and are currently not possible within a single tool. This article presents a completely new module for the open-source, cross-platform CellMissy software for cell migration data management. This module is the first tool to focus specifically on single-cell migration data downstream of image processing. It allows fast comparison across all tested conditions, providing automated data visualization, assisted data filtering and quality control, extraction of various commonly used cell migration parameters, and non-parametric statistical analysis. Importantly, the module enables parameters computation both at the trajectory-and at the step-level. Moreover, this single-cell analysis module is complemented by a new data import module that accommodates multiwell plate data obtained from high-throughput experiments, and is easily extensible through a plugin architecture. In conclusion, the end-to-end software solution presented here tackles a key bioinformatics challenge in the cell migration field, assisting researchers in their highthroughput data processing

Effect of overexpression of β- and γ-actin isoforms on actin cytoskeleton organization and migration of human colon cancer cells

Actins are eukaryotic proteins, which are involved in diverse cellular functions including muscle contraction, cell motility, adhesion and maintenance of cell shape. Cytoplasmic actin isoforms β and γ are ubiquitously expressed and essential for cell functioning. However, their unique contributions are not very well understood. The aim of this study was to determine the effect of β- and γ-actin overexpression on the migration capacity and actin cytoskeleton organization of human colon adenocarcinoma BE cells. In cells overexpressing β- or γ-actin, distinct cytoskeletal actin rearrangements were observed under the laser scanning confocal microscope. Overexpressed actins localized at the submembranous region of the cell body, especially near to the leading edge and on the tips of pseudopodia. The cells transfected with plasmids containing cDNA for β- or γ-actin were characterized by increased migration and invasion capacities. However, the migration velocity was statistically significantly higher only in the case of γ-actin overexpressing cells. In conclusion, the increased level of β- or γ-actin leads to actin cytoskeletal remodeling followed by an increase in migration and invasion capacities of human colon BE cells. These data suggest that expression of both actin isoforms has an impact on cancer cell motility, with the subtle predominance of γ-actin, and may influence invasiveness of human colon cancer

Combination of EGFR inhibitor lapatinib and MET inhibitor foretinib inhibits migration of triple negative breast cancer cell lines

Triple-negative breast cancer (TNBC) is the most challenging subtype to treat due to the lack of estrogen receptor, progesterone receptor, and HER2 expression, which excludes the usage of directed targeted therapy against them. Promising therapeutic targets are the hepatocyte growth factor receptor (MET) and epidermal growth factor receptor (EGFR), which expression is frequently elevated in TNBC. Inhibitors of these receptors used as monotherapy are often ineffective. Due to that, we studied the efficacy of combined therapy targeting MET and EGFR simultaneously. Two TNBC cell lines were treated with lapatinib (a dual EGFR and HER2 inhibitor), foretinib (a MET inhibitor), or a combination of the two. After the inhibitors treatment, we verified the cell viability (XTT assay), distribution of the cell cycle phases, the activation of signaling pathways (Western blotting), distribution of invadopodia, fluorescent gelatin digestion (immunofluorescence), and the invasion capacity of cells. A combination of foretinib and lapatinib effectively reduced the viability of examined cells, led to G2/M arrest and reduction of pAKT. There was also a decrease in number of invadopodia formed by cells, their ability to digest gelatin and reduction of cells migration/invasion capacity. Therapy targeting of both EGFR and MET receptors was much more effective against tested cells than monotherapy. We selected a combination of drugs that could be successfully used against this breast cancer subtype

Karakterisasi Pati Talas (Colocasia Esculenta (L.) Schott) Sebagai Alternatif Sumber Pati Industri Di Indonesia

Utilization of taro as the taro flour or taro starch will increase the economic value and the shelf life of products taro. Utilization of taro tubers during this simply as a snack, such as chips. Whereas in other countries namely the United States, Hawaii, Japan, Philippines, Brazil and Columbia have made a variety of taro tuber industrial commodities such as biscuits, bread, pasta and taro. Therefore, this study focused on the manufacture of starch and modified starch taro taro as one alternative source of starch industry in Indonesia. The purpose of this study is to get the characteristics of taro starch and modified starch and starch compared with markets such as corn starch, rice and cassava. In this study analysis, which includes characterization of starch. Characteristics of taro starch were tested consisted of moisture content, starch content, amylose and amylopectin content. The water content of taro starch produced ranged from 5.3 to 13.18 so still meet the quality standards of industrial starch is less than 14%. Starch content of tubers produced by 80% taro, taro flour by 75% while the starch content derived from the modification of taro only 65%. Low levels of starch modification due to the degradation that occurs in the process of modification. The resulting levels of amylose has an average value of 4.41%. Low levels of amylose due to the low starch content. Thus indirectly affect the levels of amylose. While the levels of amylopectin starch modification of taro only 60.88%. This is because the process of modification using acid hydrolysis led to a reduction of amylopectin. Starch is a starch modification process that has undergone physical or chemical treatment in a controlled manner so that changing one or more of its original properties, such as the initial temperature of gelatinization, during gelatinization characteristics, acidification and stirring. While hydrochloric acid hydrolysis of starch which aims to produce a more tenuous structure so that water is more volatile at the time of drying

Are non-muscle actin isoforms functionally equivalent?

Actin is highly conserved and it is the most widespread protein in eukaryotic cells. One of the most important features of actin, which allows it to have many different functions, is its ability to polymerize and interact with many other proteins. Actins are the major constituent of the actin cytoskeleton, which is an important system that is involved in various aspects of cell function, including cell motility, structure, integrity, regulation of signal transduction and transcription. Six mammal actin isoforms are highly conserved and share common functions. Two of them, β and γ non-muscle actin isoforms, which differ only by four amino acids located at the N-terminus of the polypeptide chain, are required for survival and proper cell functioning. We also summarized data about actbl2, which is suggested to be a newly discovered isoactin. Here, we review the current knowledge about tissue-specific expression of the non-muscle actin isoforms and possible functional differences between them. We also discuss molecular tools, which in recent years have allowed for a better understanding of the role of these proteins in cell functioning

Characterization of Melanoma Cell Lines Resistant to Vemurafenib and Evaluation of Their Responsiveness to EGFR- and MET-Inhibitor Treatment

Constitutively active mutated BRAF kinase occurs in more than 40% of patients suffering from melanoma. To block its activity, a specific inhibitor, vemurafenib, is applied as a therapy. Unfortunately, patients develop resistance to this drug rather quickly. Previously, we demonstrated that pairs of inhibitors directed against EGFR (epidermal growth factor receptor) and MET (hepatocyte growth factor receptor) trigger a synergistic cytotoxic effect in human melanoma cells, and decrease their invasive abilities. In this study, we aimed to generate and characterize melanoma cells resistant to vemurafenib treatment, and then to evaluate the effectiveness of a previously developed therapy in this model. We showed that melanoma cells resistant to the BRAF inhibitor are characterized by a lower proliferation rate and they acquire a spindle-like shape. Using Western Blot, we also noticed increased levels of EGFR, MET, and selected markers of cancer stem cells in generated cell lines. Resistant cells also exhibited increased invasive abilities and elevated proteolytic activity, observed using scratch wound assays and gelatin zymography. Moreover, combination therapy reduced their viability, as measured with a colorimetric cytotoxicity test, and decreased invasiveness. The obtained results validate the application of combination therapy directed against EGFR and MET in melanoma cells resistant to treatment with inhibitors of mutated BRAF

The Influence of Tumor Microenvironment on Immune Escape of Melanoma

The low efficiency of currently-used anti-cancer therapies poses a serious challenge, especially in the case of malignant melanoma, a cancer characterized by elevated invasiveness and relatively high mortality rate. The role of the tumor microenvironment in the progression of melanoma and its acquisition of resistance to treatment seems to be the main focus of recent studies. One of the factors that, in normal conditions, aids the organism in its fight against the cancer and, following the malignant transformation, adapts to facilitate the development of the tumor is the immune system. A variety of cell types, i.e., T and B lymphocytes, macrophages, and dendritic and natural killer cells, as well as neutrophils, support the growth and invasiveness of melanoma cells, utilizing a plethora of mechanisms, including secretion of pro-inflammatory molecules, induction of inhibitory receptors expression, or depletion of essential nutrients. This review provides a comprehensive summary of the processes regulated by tumor-associated cells that promote the immune escape of melanoma cells. The described mechanisms offer potential new targets for anti-cancer treatment and should be further studied to improve currently-employed therapies

Bleb-like protrusions in LS174T colon cancer cells with rounded morphology.

<p>Cells were plated onto coverslips. After fixation with 4% formaldehyde, cells were labeled to visualize filamentous actin (red) and ezrin (green). Merged image is shown on the right picture. Enlargements of the boxed, bleb-rich area are shown as insets. Scale bar: 10 μm</p