Morphology Of Cancer Cells Research Articles

BIOACTIVE COMPOUNDS OF INDONESIAN RED BETEL (PIPER CROCATUM) EXTRACT AND ITS INHIBITORY ACTIVITY IN MCF-7 CELL LINE

Piper crocatum, known as red betel, is a potential medicinal plant that grows in the tropics as an anti-cancer. Several previous studies of red betel showed the ability to inhibit various types of cancer cells. The research aims to analyze the bioactive compounds in red betel leaf extract origins from Indonesia on ethanol by GC-MS method, identify toxicity with MTT assay. Further, the morphology of MCF-7 breast cancer cells was determined to lead the ethanol extract of P. crocatum. The bioactive compounds of P. crocatum leaves have a function in cytotoxicity to cancer cells, such as phenol (1.36%), phytol (5.72%), and vitamin E (1.76%). Red betel leaves extract against MCF-7 cells has an IC50 = 6.68 μg mL-1 . The results showed that morphological changes occurred in MCF-7 breast cancer cells. Cell morphological changes were indicated by loss of contact with surrounding cells and decreased the number of cells after 24 h exposed to ethanol extract of P. crocatum. The ethanol extract of P. crocatum can inhibit MCF-7 breast cancer cells' growth and potential as an anti-cancer agent. However, further research is needed to confirm the mechanism of inhibition and expression of apoptotic genes.

Synthesis of silver nanoparticle (Ag NPs) using phytochemical rich medicinal plant Lonicera japonica for improve the cytotoxicity effect in cancer cells

Initially, the excellent anti-oxidant property of the Ag NPs was synthesized from Chinese medicinal plant of Lonicera japonica, and their spectroscopic evidences were confirmed by UV-spectrometer. The morphology, size and shape of the synthesized Ag NPs were effectively identified by scanning electron microscope and transmission electron microscope. Both the spectroscopic results were more evident, which UV-spectrometer peak was exhibited at 456 nm, scanning electron microscope and transmission electron microscope results were also clearly shown ball like spherical shape. The potential anti-cancer effect of the Ag NPs was revealed, the selected Chinese plant Lonicera japonica was influenced the A549 lung cancer cells through excessive ROS production. The IC50 concentration of 75 µg/mL was shown decreased viability in Ag NPs treated A549 lung cancer cells of MTT assay. Consecutively, the MTT assay result was proved by irregular shape and damaged morphology in A549 lung cancer cells by images of phase contrast microscope. Finally, the result was also clearly stated that the synthesized Ag NPs was very effective against A549 human lung cancer cells and it use for future drug discovery for inhibiting the lung cancer.

Study of new Zn(II)Prolinedithiocarbamate as a potential agent for breast cancer: Characterization and molecular docking

This study characterized the synthetic Zn(II)Prolinedithiocarbamate and determined its anti-cancer activity in vitro and in silico. Zn(II)Prolineditiocarbamate complex was synthesized, characterized, and determined by melting point, conductivity value, UV–Vis spectroscopy, FT-IR and XRD. Computational NMR spectrum analysis has been carried out. Zn(II)Prolinedithiocarbamate complex's binding to DNA was studied using an in vitro molecular docking anti-cancer activity assay. Molecular docking results showed the interaction of Zn(II)Prolinedithiocarbamate complex with DNA from MCF-7 strain cells. Cytotoxicity of Zn(II)Prolinethiocarbamate against the MCF-7 cell line showed changes in cancer cell morphology at an IC50 value of 360.10 g/mL. Zn(II)Prolinedithiocarbamate complex compounds can be a breakthrough in developing chemotherapy drugs that are potential and effective against the MCF-7 cell line.

Cancer-Cell Deep-Learning Classification by Integrating Quantitative-Phase Spatial and Temporal Fluctuations.

We present a new classification approach for live cells, integrating together the spatial and temporal fluctuation maps and the quantitative optical thickness map of the cell, as acquired by common-path quantitative-phase dynamic imaging and processed with a deep-learning framework. We demonstrate this approach by classifying between two types of cancer cell lines of different metastatic potential originating from the same patient. It is based on the fact that both the cancer-cell morphology and its mechanical properties, as indicated by the cell temporal and spatial fluctuations, change over the disease progression. We tested different fusion methods for inputting both the morphological optical thickness maps and the coinciding spatio-temporal fluctuation maps of the cells to the classifying network framework. We show that the proposed integrated triple-path deep-learning architecture improves over deep-learning classification that is based only on the cell morphological evaluation via its quantitative optical thickness map, demonstrating the benefit in the acquisition of the cells over time and in extracting their spatio-temporal fluctuation maps, to be used as an input to the classifying deep neural network.

Tuning the Nanotopography and Chemical Functionality of 3D Printed Scaffolds through Cellulose Nanocrystal Coatings.

In nature, cells exist in three-dimensional (3D) microenvironments with topography, stiffness, surface chemistry, and biological factors that strongly dictate their phenotype and behavior. The cellular microenvironment is an organized structure or scaffold that, together with the cells that live within it, make up living tissue. To mimic these systems and understand how the different properties of a scaffold, such as adhesion, proliferation, or function, influence cell behavior, we need to be able to fabricate cellular microenvironments with tunable properties. In this work, the nanotopography and functionality of scaffolds for cell culture were modified by coating 3D printed materials (DS3000 and poly(ethylene glycol)diacrylate, PEG-DA) with cellulose nanocrystals (CNCs). This general approach was demonstrated on a variety of structures designed to incorporate macro- and microscale features fabricated using photopolymerization and 3D printing techniques. Atomic force microscopy was used to characterize the CNC coatings and showed the ability to tune their density and in turn the surface nanoroughness from isolated nanoparticles to dense surface coverage. The ability to tune the density of CNCs on 3D printed structures could be leveraged to control the attachment and morphology of prostate cancer cells. It was also possible to introduce functionalization onto the surface of these scaffolds, either by directly coating them with CNCs grafted with the functionality of interest or with a more general approach of functionalizing the CNCs after coating using biotin-streptavidin coupling. The ability to carefully tune the nanostructure and functionalization of different 3D-printable materials is a step forward to creating in vitro scaffolds that mimic the nanoscale features of natural microenvironments, which are key to understanding their impact on cells and developing artificial tissues.

Evodiamine Inhibits Gastric Cancer Cell Proliferation via PTEN-Mediated EGF/PI3K Signaling Pathway.

Aims In this study, the pharmacological effects and potential molecular mechanisms of evodiamine in treating gastric cancer (GC) were investigated. Methods GC cells lines of AGS and BGC-823 were treated with evodiamine at various concentrations for different times (24, 48, and 72 h). Inhibition of the proliferation of AGS and BGC-823 cells was assessed using a CCK-8 assay. The morphology of gastric cancer cells was detected by high-content screening (HCS). The apoptosis-inducing effect of evodiamine on AGS and BGC-823 cells was detected by flow cytometric analysis. Cell migration and invasion were detected by Transwell assay. The relative mRNA and protein expression levels of PTEN-mediated EGF/PI3K signaling pathways were investigated via RT-qPCR or western blotting, respectively. Results Evodiamine substantially inhibited AGS and BGC-823 cells proliferation in a dose- and time-dependent manner. Flow cytometric analysis revealed that evodiamine could induce apoptosis of AGS and BGC-823 cells in a dose-dependent manner. In addition, evodiamine inhibited AGS and BGC-823 cell migration and invasion. Mechanistically, the results demonstrated that evodiamine promoted the relative mRNA and protein expression of PTEN and decreased expression of EGF, EGFR, PI3K, AKT, p-AKT, and mTOR. Most importantly, evodiamine could effectively increase the mRNA and protein expression of PTEN and decrease the protein expression of EGF/PI3K pathway, indicating that evodiamine downregulated EGF/PI3K through the activation of PTEN pathway. Conclusion Evodiamine inhibited the directional migration and invasion of GC cells by inhibiting PTEN-mediated EGF/PI3K signaling pathway. These findings revealed that evodiamine might serve as a potential candidate for the treatment or prevention of GC.

Phycocyanin inhibits pancreatic cancer metastasis via suppressing epithelial-mesenchymal transition and targeting Akt/β-catenin pathway.

According to our previous research, phycocyanin (PC) could inhibit pancreatic tumor growth obviously. However, little is known about the effects of phycocyanin on pancreatic tumor metastasis. This study was aimed to investigate whether phycocyanin inhibits pancreatic cancer cells' metastasis and the potential underlying mechanisms. Human pancreatic carcinoma cell lines PANC-1 and BxPC3 were treated with phycocyanin (5, 10, 20, and 40 μM) for 48 h or 72 h. BALB/c nude mice were inoculated intravenously with luciferase-PANC-1 cells to establish a tumor metastasis model. BALB/c nude mice were also inoculated subcutaneously with PANC-1 cells to establish a xenograft tumor model. Our results indicated that (1) phycocyanin inhibited migration, invasion, and movement of pancreatic cancer cells in vitro, and tumor metastasis in mouse xenograft tumor model; (2) phycocyanin significantly changed cell morphology and epithelial-mesenchymal transition (EMT) phenotype in pancreatic cancer cells; (3) phycocyanin decreased the phosphorylation level of Akt and the level of nuclear β-catenin. Moreover, administration of Akt activator SC79 counteracted the inhibitory effect of phycocyanin on the migration of pancreatic cancer cells, associated with the reversal of epithelial-mesenchymal transition and Akt/β-catenin signaling pathway. Taken together, our results suggest that phycocyanin inhibits pancreatic cancer metastasis via suppressing epithelial-mesenchymal transition and targeting the Akt/β-catenin pathway.

SHANK3 conformation regulates direct actin binding and crosstalk with Rap1 signaling

Actin-rich cellular protrusions direct versatile biological processes from cancer cell invasion to dendritic spine development. The stability, morphology, and specific biological functions of these protrusions are regulated by crosstalk between three main signaling axes: integrins, actin regulators, and small guanosine triphosphatases (GTPases). SHANK3 is a multifunctional scaffold protein, interacting with several actin-binding proteins and a well-established autism risk gene. Recently, SHANK3 was demonstrated to sequester integrin-activating small GTPases Rap1 and R-Ras to inhibit integrin activity via its Shank/ProSAP N-terminal (SPN) domain. Here, we demonstrate that, in addition to scaffolding actin regulators and actin-binding proteins, SHANK3 interacts directly with actin through its SPN domain. Molecular simulations and targeted mutagenesis of the SPN-ankyrin repeat region (ARR) interface reveal that actin binding is inhibited by an intramolecular closed conformation of SHANK3, where the adjacent ARR domain covers the actin-binding interface of the SPN domain. Actin and Rap1 compete with each other for binding to SHANK3, and mutation of SHANK3, resulting in reduced actin binding, augments inhibition of Rap1-mediated integrin activity. This dynamic crosstalk has functional implications for cell morphology and integrin activity in cancer cells. In addition, SHANK3-actin interaction regulates dendritic spine morphology in neurons and autism-linked phenotypes invivo.

Stereotactic Body Radiotherapy and Conventional Radiotherapy Induce Cytoskeleton Extension and Enlargement of Cell Morphology in Non-Small Cell Lung Cancer.

Stereotactic body radiotherapy (SBRT) is now widely used in cancer therapy. However, the biological effects of SBRT compared with conventional radiotherapy (CRT) are not clear. The cytoskeleton plays an important role in many biological processes and cellular life activities. The effects of SBRT or CRT on the morphology and cytoskeletal structure of non-small cell lung cancer (NSCLC) cells remain unknown. Based on the biologically equivalent dose (BED) formula, we designed SBRT and CRT fractionation regimens with the same BED. The morphology was captured during radiation, and rhodamine-phalloidin immunofluorescence was used to study the cytoskeleton. A lactate dehydrogenase assay kit was used to determine the cell membrane permeability, and western blot was used to detect the cytoskeleton protein expression levels. The morphology and cytoskeleton expanded after SBRT or CRT, with an increase in cell membrane permeability and stable cytoskeleton protein levels. Besides, different dose of SBRT (10,20,30 Gy) induce similar morphology and cytoskeleton enlargement. Our findings indicate that SBRT and CRT can induce cytoskeleton reorganization and the enlargement of cell morphology (at different rates) in NSCLC. The morphology and cytoskeleton enlargement after SBRT are dose independence.

Stereospecific inhibition of AMPK by (R)-crizotinib induced changes to the morphology and properties of cancer and cancer stem cell-like cells

Crizotinib is used in the clinic for treating patients with ALK- or ROS1-positive non-small-cell lung carcinoma. The objective of the present study was to determine if crizotinib enantiomers could induce changes to the properties of cancer and cancer stem cell (CSC)-like cells at a high concentration (∼ 3 μM). While (R)-crizotinib induced changes in morphologies or sizes of cells, (S)-crizotinib did not. Pretreatment with (R)-crizotinib suppressed the proliferation of cancer or CSC-like cells in vitro and tumor growth in vivo. In vivo administration of (R)-crizotinib inhibited the growth of tumors formed from CSC-like cells by 72%. %. Along with the morphological changes induced by (R)-crizotinib, the expression levels of CD44 (NCI-H23 and HCT-15), ALDH1 (NCI-H460), nanog (PC-3), and Oct-4A (CSC-like cells), which appear to be specific marker proteins, were greatly changed, suggesting that changes in cellular properties accompanied the morphological changes in the cells. The expression levels of Snail, Slug, and E-cadherin were also greatly altered by (R)-crizotinib. Among several signal transduction molecules examined, AMPK phosphorylation appeared to be selectively inhibited by (R)-crizotinib. BML-275 (an AMPK inhibitor) and AMPKα2 siRNA efficiently induced morphological changes to all types of cells examined, suggesting that (R)-crizotinib might cause losses of characteristics of cancer or CSCs via inhibition of AMPK. These results indicate that (R)-crizotinib might be an effective anticancer agent that can cause alteration in cancer cell properties.

Decreased prostasin expression is associated with aggressiveness of oral squamous cell carcinoma.

Prostasin is a glycosylphosphatidylinositol-anchored serine protease widely expressed in epithelial cells, with crucial epidermal barrier functions. Evidence has suggested prostasin may have served as a tumor suppressor in various cancers, but its role in oral squamous cell carcinoma (OSCC) remains unclear. Thus, herein, we conducted an immunohistochemical prostasin study in 119 resected OSCC cases. Prostasin expression was decreased in 63% (75/119) of cases. OSCC with decreased prostasin immunoreactivity (low prostasin cases) tended to show a higher histological grade (p = 0.0088) and a more infiltrative cancer cell morphology (p = 0.0024). We then explored the role of prostasin in the OSCC cell lines: SAS and HSC-4. SAS did not express detectable prostasin levels, whereas HSC-4 expressed low but distinct levels. Prostasin overexpression suppressed the proliferation and migration of both OSCC lines in vitro. Conversely, prostasin silencing significantly enhanced growth rates of HSC-4. Finally, we analyzed the impact of prostasin expression on the prognosis of patients with OSCC; decreased expression tended to correlate with shorter overall survival (p = 0.0291) after resection. This trend was supported by our analyses using a public database (Kaplan-Meier plotter) of head and neck squamous cell carcinomas. In conclusion, we showed decreased prostasin expression was associated with aggressive features and a poorer prognosis of OSCC.

Heterogeneous microenvironmental stiffness regulates pro-metastatic functions of breast cancer cells

Besides molecular and phenotypic variations observed in cancer cells, intratumoral heterogeneity also occurs in the tumor microenvironment. Correlative stiffness maps of different intratumor locations in breast tumor biopsies show that stiffness increases from core to periphery. However, how different local ECM stiffness regulates key functions of cancer cells in tumor progression remains unclear. Although increased tissue stiffness is an established driver of breast cancer progression, conclusions from 2D cultures do not correspond with newer data from cancer cells in 3D environments. Many past studies of breast cancer in 3D culture fail to recapitulate the stiffness of a real breast tumor or the various local stiffnesses present in a tumor microenvironment. In this study, we developed a series of collagen/alginate hybrid hydrogels with adjustable stiffness to match the core, middle, and peripheral zones of a breast tumor. We used this hydrogel system to investigate effects of different local stiffness on morphology, proliferation, and migration of breast cancer cells. RNA sequencing of cells in hydrogels with different stiffness revealed changes in multiple cellular processes underlying cancer progression, including angiogenesis and metabolism. We discovered that tumor cells in a soft environment enriched YAP1 and AP1 signaling related genes, whereas tumor cells in a stiff environment became more pro-angiogenic by upregulating fibronectin 1 (FN1) and matrix metalloproteinase 9 (MMP9) expression. This systematic study defines how the range of environmental stiffnesses present in a breast tumor regulates cancer cells, providing new insights into tumorigenesis and disease progression at the tumor-stroma interface. Statement of significanceApplied a well-defined hybrid hydrogel system to mimic the tumor microenvironment with heterogeneous local stiffness. Breast cancer cells tended to proliferate in soft core environment while migrate in stiff peripheral environment. Breast cancer cells shift from glycolysis to OXPHOS and fatty acid metabolism responding to stiff matrix microenvironment. The transcriptomic profile of breast cancer cells altered due to microenvironmental stiffness changes.

Antiproliferation Activity and Apoptotic Mechanism of Soursop (Annona muricata L.) Leaves Extract and Fractions on MCF7 Breast Cancer Cells.

IntroductionBreast cancer is the second most common cancer in women globally, and the incidence rate has increased annually. Traditional medicine is frequently used as a cancer treatment, and soursop or Annona muricata L (A. muricata) is a traditional medicinal plant that has been widely used as an anticancer treatment and requires more thorough study.MethodsIn this research, we prepared ethanol extract and three solvents, ie, ethyl acetate, n-hexane and water fractions of A. muricata leaves and assessed their antiproliferation and cytotoxic activity on MCF7 breast cancer cells compared with that on CV1 normal kidney cells; observation of cell morphology by stained with mixture of propidium iodide and 4ʹ,6-diamidino-2-phenylindole indicated that this treatment induced an ongoing process of apoptotic cell death in MCF7 cells. To clarify the cell death mechanism via apoptosis, we assessed the mRNA expression in the caspase cascade of caspase-9, caspase-3, and PARP-1, and anti-apoptotic, Bcl-2 which mediated cytotoxic activity of extracts and ethyl acetate fractions of A. muricata leaves against MCF7 cells.ResultsThe ethanol extract, ethyl acetate, n-hexane, and water fractions of A. muricata leaves had IC50 values of 5.3, 2.86, 3.08, and 48.31 µg/mL, respectively, in MCF7 cells but had no activity in CV1 cells. The high cytotoxic activity of A. muricata leaves was reflected by changes in the morphology of cancer cells that appeared after 6 h exposure to A. muricata leaf extract and ethyl acetate fraction; the membrane and nucleus of cells undergoing apoptosis were characterized by the rupture and loss of membranes and nuclei. The mechanism that mediates this cytotoxic activity in MCF7 cells was mediated through a decrease in the expression of Bcl-2 mRNA and an increase in caspase-9 and caspase-3 mRNA expression.ConclusionTherefore, the leaves of the medicinal plant A. muricata contained compounds that on extraction exerted a highly effective activity as an anticancer treatment for breast cancer via induced apoptotic cell death.

NEK5 activity regulates the mesenchymal and migratory phenotype in breast cancer cells.

Breast cancer remains a prominent global disease affecting women worldwide despite the emergence of novel therapeutic regimens. Metastasis is responsible for most cancer-related deaths, and acquisition of a mesenchymal and migratory cancer cell phenotypes contributes to this devastating disease. The utilization of kinase targets in drug discovery have revolutionized the field of cancer research but despite impressive advancements in kinase-targeting drugs, a large portion of the human kinome remains understudied in cancer. NEK5, a member of the Never-in-mitosis kinase family, is an example of such an understudied kinase. Here, we characterized the function of NEK5 in breast cancer. Stably overexpressing NEK5 cell lines (MCF7) and shRNA knockdown cell lines (MDA-MB-231, TU-BcX-4IC) were utilized. Cell morphology changes were evaluated using immunofluorescence and quantification of cytoskeletal components. Cell proliferation was assessed by Ki-67 staining and transwell migration assays tested cell migration capabilities. In vivo experiments with murine models were necessary to demonstrate NEK5 function in breast cancer tumor growth and metastasis. NEK5 activation altered breast cancer cell morphology and promoted cell migration independent of effects on cell proliferation. NEK5 overexpression or knockdown does not alter tumor growth kinetics but promotes or suppresses metastatic potential in a cell type-specific manner, respectively. While NEK5 activity modulated cytoskeletal changes and cell motility, NEK5 activity affected cell seeding capabilities but not metastatic colonization or proliferation in vivo. Here we characterized NEK5 function in breast cancer systems and we implicate NEK5 in regulating specific steps of metastatic progression.

Understanding the tumour immune microenvironment of stage III colorectal cancer using multiplexed imaging mass cytometry

Background/Aim: The tumour immune microenvironment is spatially heterogenous and plays a crucial role in cancer progression and response to therapy. Current histopathological examination cannot comprehensively assess the molecular heterogeneity in the tumour microenvironment. We aim to capture tissue morphology and single cell protein expression of cancer and immune cells within intact tissue sections of stage III colorectal tumours from 11 patients. Methods: Using Hyperion Image Mass Cytometry (IMC), we simultaneously profiled 16 protein markers from FFPE samples, including cancer (Ki-67, p53) and immune (CD3, CD4, CD8a, CD45RO, CD20, CD68, Granzyme B) cell markers. We integrated this with matched histopathological imaging and whole-exome-sequencing data. Results: We identified seven cancer, immune and stromal cell types and preserved their spatial organisation within the microenvironment. These cell types were validated on matched H&E histopathology. We found significant interactions of cancer cells with NK-cells and B-cells. Furthermore, we found that tumours with 17p deletion had significantly fewer NK-cells. Conclusion: Using IMC to acquire rich spatial data at single-cell resolution enabled us to assess the heterogeneity of tumour tissue by defining cell types, immune infiltration, and cancer-immune cell interaction within intact tissue sections. Integration of this data with H&E images could assist pathologists in cancer diagnosis.

Survival prediction based on the gene expression associated with cancer morphology and microenvironment in primary central nervous system lymphoma.

Dysregulation of cell morphology and cell-cell interaction results in cancer cell growth, migration, invasion, and metastasis. Besides, a balance between the extracellular matrix (ECM) and matrix metalloprotease (MMP) is required for cancer cell morphology and angiogenesis. Here, we determined gene signatures associated with the morphology and microenvironment of primary central nervous system lymphoma (PCNSL) to enable prognosis prediction. Next-generation sequencing (NGS) on 31 PCNSL samples revealed gene signatures as follows: ACTA2, ACTR10, CAPG, CORO1C, KRT17, and PALLD in cytoskeleton, CDH5, CLSTN1, ITGA10, ITGAX, ITGB7, ITGA8, FAT4, ITGAE, CDH10, ITGAM, ITGB6, and CDH18 in adhesion, COL8A2, FBN1, LAMB3, and LAMA2 in ECM, ADAM22, ADAM28, MMP11, and MMP24 in MMP. Prognosis prediction formulas with the gene expression values and the Cox regression model clearly divided survival curves of the subgroups in each status. Furthermore, collagen genes contributed to gene network formation in glasso, suggesting that the ECM balance controls the PCNSL microenvironment. Finally, the comprehensive balance of morphology and microenvironment enabled prognosis prediction by a combinatorial expression of 8 representative genes, including KRT17, CDH10, CDH18, COL8A2, ADAM22, ADAM28, MMP11, and MMP24. Besides, these genes could also diagnose PCNSL cell types with MTX resistances in vitro. These results would not only facilitate the understanding of biology of PCNSL but also consider targeting pathways for anti-cancer treatment in personalized precision medicine in PCNSL.

Protonectin peptides target lipids, act at the interface and selectively kill metastatic breast cancer cells while preserving morphological integrity

Despite the need for innovative compounds as antimicrobial and anticancer agents, natural sources of peptides remain underexplored. Protonectin (PTN), a cationic dodecapeptide of pharmacological interest, presents large hydrophobicity that is associated with the tendency to aggregate and supposedly influences bioactivity. A disaggregating role was assigned to PTN’ N-terminal fragment (PTN1-6), which enhances the bioactivity of PTN in a 1:1 mixture (PTN/PTN1-6). Spectroscopic techniques and model membranes (phospholipid bilayers and SDS micelles) revealed that environment-dependent aggregation is reduced for PTN/PTN1-6, but cytotoxicity of PTNs on MDA-MB-231 breast cancer showed the same CC50 values around 16 µM and on MCF-10A epithelial breast cells 6 to 5-fold higher values, revealing a selective interaction. Since PTN1-6 lacks activity on breast cells, its presence should differently affect PTN activity, suggesting that aggregation could modulate activity depending on the membrane characteristics. Indeed, increased partitioning and lytic activity of PTN/PTN1-6 were found in model membranes independently of charge density, but affected by the curvature tendency. PTN and PTN/PTN1-6 do not alter morphology and roughness of cancer cells, indicating a superficial interaction with membranes and consistent with results obtained in NMR experiments. Our results indicate that aggregation of PTNs depends on the membrane characteristics and modulates the activity of the peptides.

Cannabidiol Modulates Mitochondrial Redox and Dynamics in MCF7 Cancer Cells: A Study Using Fluorescence Lifetime Imaging Microscopy of NAD(P)H.

The cannabinoid, cannabidiol (CBD), is part of the plant's natural defense system that when given to animals has many useful medicinal properties, including activity against cancer cells, modulation of the immune system, and efficacy in epilepsy. Although there is no consensus on its precise mode of action as it affects many cellular targets, CBD does appear to influence mitochondrial function. This would suggest that there is a cross-kingdom ability to modulate stress resistance systems that enhance homeostasis. As NAD(P)H autofluorescence can be used as both a metabolic sensor and mitochondrial imaging modality, we assessed the potential of this technique to study the in vitro effects of CBD using 2-photon excitation and fluorescence lifetime imaging microscopy (2P-FLIM) of NAD(P)H against more traditional markers of mitochondrial morphology and cellular stress in MCF7 breast cancer cells. 2P-FLIM analysis revealed that the addition of CBD induced a dose-dependent decrease in bound NAD(P)H, with 20 µM treatments significantly decreased the contribution of bound NAD(P)H by 14.6% relative to the control (p < 0.001). CBD also increased mitochondrial concentrations of reactive oxygen species (ROS) (160 ± 53 vs. 97.6 ± 4.8%, 20 µM CBD vs. control, respectively, p < 0.001) and Ca2+ (187 ± 78 vs. 105 ± 10%, 20 µM CBD vs. the control, respectively, p < 0.001); this was associated with a significantly decreased mitochondrial branch length and increased fission. These are all suggestive of mitochondrial stress. Our results support the use of NAD(P)H autofluorescence as an investigative tool and provide further evidence that CBD can modulate mitochondrial function and morphology in a dose-dependent manner, with clear evidence of it inducing oxidative stress at higher concentrations. This continues to support emerging data in the literature and may provide further insight into its overall mode of action, not only in cancer, but potentially its function in the plant and why it can act as a medicine.

Organelle Topology Is a New Breast Cancer Cell Classifier

Breast cancer is a highly heterogenous disease, both phenotypically and genetically. Importantly a correlation between intra-tumoral heterogeneity, drug resistance and negative clinical outcome has been established. Previously, quantity or subcellular location of protein biomarkers have been compared to identify tumor tissue types. To-date, mass cytometry and single-cell sequencing studies have been used to assess spatial intra-tumor heterogeneity by identifying a range of phenotypic and genetic alterations across different regions of a single tumor. Recently, cell morphology has been indicated as a direct readout of the functional phenotypic state of an individual cancer cell. Here, we hypothesize that the spatial context of organelles within cancer cells, specifically their subcellular location and inter-organelle relationships (topology), can be used to inform breast cancer cell classification. Since numerous correlations between biologic behaviors and pathologic findings have been well-established, we propose that organelle topological heterogeneity reveals the long-term adaptation of organelles and cytoskeletal networks to match breast cancer cell type status. Thus, this study aims to investigate the heterogeneity of organelle topology and morphology in breast cancer cells to increase our basic understanding of breast cancer biology on a subcellular level. We have introduced a novel approach that quantifies, for the first time, the feature of organelles in breast cancer cells, removing the bias of visual interpretation to classify different cell lines based on organelle morphology and topology. This method was tested on three different organelle datasets: mitochondria, early endosomes and recycling endosomes in a panel of human breast cancer cells, including T47D (estrogen receptor-positive), MDAMB231, MDAMB436 and MDAMB468 (triple negative) and AU565 (HER2 positive), as well as non-cancerous mammary epithelial MCF10A cells. High resolution Airyscan microscopy was used to collect z-stacks across cells labeled with fluorescently labeled-transferrin and immunostained with anti-Tom20 and anti-EEA1 to label the recycling, early endosomal and mitochondria networks, respectively. Subsequently, 3D surface rendering of organelle objects was performed using IMARIS image analysis software. A morphometric evaluation of mitochondrial and endosomal compartments resulted in 34 topology and morphology parameters. Application of Random Forest machine learning based classification to 18 of these 34 parameters generated the highest accuracy in breast cancer cell classification. We systematically evaluated how different parameter combinations affected the machine learning-based cancer cell classification and discovered that topology parameters were crucial to achieve the highest accuracy. Based on organelle spatial distribution and their interaction with neighbor organelles (topology) a classification accuracy over 95% was achieved to distinguish between a variety of human breast cancer cell lines of differing subtype and aggressiveness. These findings lay the groundwork for using quantitative topological organelle features as an effective method to analyze and classify breast cancer cell phenotypes.

Cytotoxic impacts of escin via inducing apoptosis and morphological changes on human prostate cancer cells

Prostate cancer is one of the most prevalent cancer types in males with a percentages of 28% worldwide. This cancer type comprises approximately 37% of the cancer incidences in males in Turkey. Recent cancer investigations are focused on natural agents with good potent for cancer therapy. Escin is one of the most investigated agents of that kind but its effects on human prostate cancer cell’s morphology is not investigated in detail, yet. Thus, the aim of this study is to investigate the antiproliferative, cytotoxic and proapoptotic effects of escin, on prostate cancer cells Du-145. Cytotoxicity of escin on prostate cancer cells was intestigated by using sulforhodamine B (SRB) assay and viability percentages and IC50 value were detected from the elisa reader (BioTek Synergy HTX) results. For morphological changes, Du-145 cells treated with the IC50 value of escin were evaluated under a confocal microscope (Leica, TCS SP5 II, Germany). Apoptosis profiles of cells were investigated by flow cytometry. According to the SRB findings escin reduced the viability of prostate cancer cells in dose-dependent manner and the IC50 value was detected as 30.48 µM for 24 hours. On the confocal microscopy results it was confirmed that escin significantly changed the morphology of the treated cells as disintegrated and deformed nuclei, chromatin condensation, fragmentations in the cytoskeleton also shrinkage of prostate cells. Annexin-V technique indicated the apoptotic cell death trigered by escin in Du-145 cells. Based on the study results, it was concluded that escin changed the morphology of prostate cancer cells and induced apoptosis on prostate cancer cells.