Changes In Nuclear Morphology Research Articles

Co-Encapsulation of Simvastatin and Doxorubicin into pH-Sensitive Liposomes Enhances Antitumoral Activity in Breast Cancer Cell Lines.

Doxorubicin (DOX) is a potent chemotherapeutic drug used as the first line in breast cancer treatment; however, cardiotoxicity is the main drawback of the therapy. Preclinical studies evidenced that the association of simvastatin (SIM) with DOX leads to a better prognosis with reduced side effects and deaths. In this work, a novel pH-sensitive liposomal formulation capable of co-encapsulating DOX and SIM at different molar ratios was investigated for its potential in breast tumor treatment. Studies on physicochemical characterization of the liposomal formulations were carried out. The cytotoxic effects of DOX, SIM, and their combinations at different molar ratios (1:1; 1:2 and 2:1), free or co-encapsulated into pH-sensitive liposomes, were evaluated against three human breast cancer cell lines (MDA-MB-231, MCF-7, and SK-BR-3). Experimental protocols included cell viability, combination index, nuclear morphological changes, and migration capacity. The formulations showed a mean diameter of less than 200 nm, with a polydispersity index lower than 0.3. The encapsulation content was ~100% and ~70% for DOX and SIM, respectively. A more pronounced inhibitory effect on breast cancer cell lines was observed at a DOX:SIM molar ratio of 2:1 in both free and encapsulated drugs. Furthermore, the 2:1 ratio showed synergistic combination rates for all concentrations of cell inhibition analyzed (50, 75, and 90%). The results demonstrated the promising potential of the co-encapsulated liposome for breast tumor treatment.

Neuroprotective effects of hesperetin on H2O2-induced damage in neuroblastoma SH-SY5Y cells.

Oxidative stress is a fundamental neurodegenerative disease trigger that damages and decimates nerve cells. Neurodegenerative diseases are chronic central nervous system disorders that progress and result from neuronal degradation and loss. Recent studies have extensively focused on neurodegenerative disease treatment and prevention using dietary compounds. Heseperetin is an aglycone hesperidin form with various physiological activities, such as anti-inflammation, antioxidant, and antitumor. However, few studies have considered hesperetin's neuroprotective effects and mechanisms; thus, our study investigated this in hydrogen peroxide (H2O2)-treated SH-SY5Y cells. SH-SY5Y cells were treated with H2O2 (400 µM) in hesperetin absence or presence (10-40 µM) for 24 h. Three-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide assays detected cell viability, and 4',6-diamidino-2-phenylindole staining allowed us to observe nuclear morphology changes such as chromatin condensation and apoptotic nuclei. Reactive oxygen species (ROS) detection assays measured intracellular ROS production; Griess reaction assays assessed nitric oxide (NO) production. Western blotting and quantitative polymerase chain reactions quantified corresponding mRNA and proteins. Subsequent experiments utilized various non-toxic hesperetin concentrations, establishing that hesperetin notably decreased intracellular ROS and NO production in H2O2-treated SH-SY5Y cells (P < 0.05). Furthermore, hesperetin inhibited H2O2-induced inflammation-related gene expression, including interluekin-6, tumor necrosis factor-α, and nuclear factor kappa B (NF-κB) p65 activation. In addition, hesperetin inhibited NF-κB translocation into H2O2-treated SH-SY5Y cell nuclei and suppressed mitogen-activated protein kinase protein expression, an essential apoptotic cell death regulator. Various apoptosis hallmarks, including shrinkage and nuclear condensation in H2O2-treated cells, were suppressed dose-dependently. Additionally, hesperetin treatment down-regulated Bax/Bcl-2 expression ratios and activated AMP-activated protein kinase-mammalian target of rapamycin autophagy pathways. These results substantiate that hesperetin activates autophagy and inhibits apoptosis and inflammation. Hesperetin is a potentially potent dietary agent that reduces neurodegenerative disease onset, progression, and prevention.

PRR14 acts a novel oncogene activating the PI3K signal pathway in human cutaneous squamous cell carcinoma.

Proline rich protein 14 (PRR14) is considered as a new component of the nuclear fiber layer, it may be a key molecule in mediating nuclear morphological changes and functional changes in tumorigenesis. But, it's still unclear in human cutaneous squamous cell carcinoma (cSCC). In the study, the expression profiles of PRR14 in patients with cSCC were investigated by immunohistochemistry (IHC), also the PRR14 expression in cSCC tissues were detected using the methods of real-time quantitative PCR (RT-qPCR) and Western blot; cell counting kit-8 (CCK-8) assay, wound healing assay, matrigel-based transwell assay and Annexin V-FITC and PI double-staining with flow cytometry assay were used to investigate the biological functions of PRR14 in A431 and HSC-1 cSCC cells. Overexpression of PRR14 in cSCC patients was reported firstly in this study and its high expression was related to differentiation, thickness and tumor node metastasis (TNM) stage of cSCC. PRR14 inhibition with RNA interfering (RNAi) method resulted in the suppression of cell proliferation, migration and invasion but promotion the apoptosis of cSCC cells, and upregulation of the protein phosphorylation levels of mammalian target of rapamycin (mTOR), phosphoinositide 3-kinase (PI3K) and Akt. The study shows PRR14 maybe an activator of cSCC carcinogenesis through PI3K/Akt/mTOR signal pathway, and it also maybe a prognostic factor and new therapeutical target for cSCC treatment.

Assessing in vitro anti-cancer efficacy of sulfonated water-soluble phthalocyanine on breast and prostate cancer cells

Since cancer is one of the leading causes of death worldwide, the development of new anticancer drugs has become important. Phthalocyanines (Pcs) as anticancer agents have attracted particular attention in recent years. Especially water-soluble sulfonated phthalocyanines have been used consistently for both the diagnosis and treatment of many different types of cancer, such as small lung tumors. Motivated by these facts, since the anti-cancer activity of sulfonated water-soluble cobalt phthalocyanine in prostate and breast cancer without photoactivation is not known before, we aimed to investigate the in vitro anti-cancer effectiveness (cyto- and genotoxic effect) of alpha-substituted water-soluble cobalt phthalocyanine (WS-CoPc) complex on prostate and breast cancer cells for the first time. For this purpose ROS production, cell death, and DNA damage levels, changes in nuclear and cell morphology, and the expression of apoptosis and ROS-related genes were determined. WS-CoPc exhibited anti-cancer effects with cytotoxic and genotoxic features in both prostate and breast cancer cells also the anti-cancer efficacy is more prominent in prostate cancer cells. Therefore our results demonstrated that this Pc has high cytotoxic and genotoxic effects, especially in prostate cancer cells in vitro and further studies are needed to reveal the current anticancer potential of this Pc without photoactivation.

Nuclear envelope morphology change upon repetitive treatment with modified antisense oligonucleotides targeting Hutchinson-Gilford Progeria Syndrome

We present the influence of treating progeroid fibroblasts with two modified antisense oligonucleotides (ONs) on the nuclear envelope. Two modified ONs were designed to block ribosome binding during translation and spliceosome binding at the cryptic splice site. We analysed the changes in the nuclear morphology of progeria cell nuclei after repetitive transfection with modified ONs as a physical analysis tool for estimating alteration of the gene expression at the protein level. Confocal microscopy was used to image the nuclei, and the nuclear lobulations were quantified to study the changes in the morphology of the nuclear envelope upon treatment. PCR was used to identify the changes in the expression of lamin A and progerin after antisense treatment at the RNA level. We found a significant decrease in the number of nuclear envelope lobulations and a lower progerin expression in progeria cells after transfection with modified ONs.

Graph-based autoencoder integrates spatial transcriptomics with chromatin images and identifies joint biomarkers for Alzheimer’s disease

Tissue development and disease lead to changes in cellular organization, nuclear morphology, and gene expression, which can be jointly measured by spatial transcriptomic technologies. However, methods for jointly analyzing the different spatial data modalities in 3D are still lacking. We present a computational framework to integrate Spatial Transcriptomic data using over-parameterized graph-based Autoencoders with Chromatin Imaging data (STACI) to identify molecular and functional alterations in tissues. STACI incorporates multiple modalities in a single representation for downstream tasks, enables the prediction of spatial transcriptomic data from nuclear images in unseen tissue sections, and provides built-in batch correction of gene expression and tissue morphology through over-parameterization. We apply STACI to analyze the spatio-temporal progression of Alzheimer’s disease and identify the associated nuclear morphometric and coupled gene expression features. Collectively, we demonstrate the importance of characterizing disease progression by integrating multiple data modalities and its potential for the discovery of disease biomarkers.

Potential of the combination of Artemisia absinthium extract and cisplatin in inducing apoptosis cascades through the expression of p53, BAX, caspase 3 ratio, and caspase 9 in lung cancer cells (Calu-6)

Artemisia has received attention in treating cancer because of having bioactive components, antiproliferative and anti-inflammatory properties. The present study assessed the synergistic effect of Artemisia absinthium extract and cisplatin on Calu-6 human lung cancer cells. Separation and identification of extract components were assessed by TLC, HPLC-MS, and GC-MS. The cell viability, and antiproliferative activity of A. absinthium extract and cisplatin was measured separately and synergistically by MTT. Additionally, nuclear morphological changes were evaluated through DAPI staining. Annexin V/PI test and real-time PCR technique were respectively applied to determine apoptosis induction, and variations in expression level of P53, caspas-3 and 9, and BAX genes. The important components including costunolide, thujone, and artemisinin were identified in Artemisia absinthium extract. The results of MTT, extract, cisplatin, and their combination inhibited proliferation of Calu-6 cells in a dose-dependent manner. This combination significantly reduced the cell viability in comparison to control (p<0.001). DAPI staining results represented cell nuclear morphological changes including nuclear chromatin condensation, apoptotic body formation, and cell membrane shrinkage. Further, Annexin V/PI test and alterations in expression level of the apoptosis-involved genes revealed an increase in apoptosis in Calu-6 cells. The A. absinthium extract could enhanced cell sensitivity of cisplatin by its active compounds. The treatment with A. absinthium extract + cisplatin may reduce cisplatin side-effects in lung cancer cells through declining the dosage along with utilizing the potential of plant constituents. Finally, Artemisia can be suggested as an adjunctive compound with less side effects for lung cancer therapy.

Development of hydrazide hydrazone-tethered combretastatin-oxindole derivatives as antimitotic agents

A series of new combretastatin-oxindole derivatives via hydrazide hydrazone linker was synthesized by condensation using (E)-bis(phenyl) acrylohydrazides and substituted isatins as synthetic precursors. To determine the potential of these new derivatives, in vitro cytotoxic activity was evaluated using different cancer cell lines, particularly of lung, colorectal, breast and skin origin. A viability study also performed for the most potent compound 9o on HaCaT cells showed its non-toxic nature toward non-malignant cell line. Cell viability assay revealed that derivative 9o with trimethoxy substitution on ring A and N-benzyl substitution on isatin ring exhibited potent anti-cancer activity against lung cancer (A549) cell line with IC50 value 1.26±1.03 µM. Further, staining studies were performed using DAPI, AO/EB to determine induction of changes in nuclear morphology, DNA damage and apoptosis which was also affirmed by annexin V-FITC/PI assay, suggesting involvement of apoptotic mechanism in inhibition of cell growth. Additionally, inducing effect of compound 9o on ROS production indicates ROS-mediated cell death of unwanted cells as well as regulation of apoptosis. The antimitotic attribute was affirmed from cell cycle arrest at G2/M phase and tubulin polymerization inhibition property of compound 9o (IC50 0.17 µM). It is worth mentioning that the pattern of all results was alike standard colchicine. Furthermore, molecular modeling studies were also conducted to determine the binding interaction and ADMET profile of 9o on 3E22 protein.

A network pharmacology approach and experimental validation to investigate the anticancer mechanism and potential active targets of ethanol extract of Wei-Tong-Xin against colorectal cancer through induction of apoptosis via PI3K/AKT signaling pathway

Ethnopharmacological relevanceWei-Tong-Xin (WTX), derives from the Chinese herbal decoction (CHD) of Wan-Ying-Yuan in ancient China, has been shown to be effective therapeutic herbal decoction for treating gastrointestinal diseases. Present studies have demonstrated that WTX had potential to alleviate the symptoms of gastrointestinal inflammation, gastric ulcer and improve gastric motility. Aim of the studyThe study primarily focused on exploring the therapeutic effect and possible pharmacological mechanism of WTX on colorectal cancer (CRC) based on network pharmacology, in vitro and in vivo experiments. Materials and methodsFirstly, colorectal cancer and WTX associated with targets were searched from GeneCards database and TCM Systems Pharmacology Database and Analysis Platform (TCMSP) respectively. The protein–protein interaction (PPI) network also was constructed to screening key targets. In addition, the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were applied to predict the underlying biological function and mechanism involving in the anti-colorectal cancer effect of WTX. Next, CCK-8, colony formation and transwell assays were performed to verify the influence of proliferation and metastasizing ability of HCT116 cells after treated with WTX. Cell cycle, apoptosis and reactive oxygen species (ROS) were analysis by flow cytometry. Hoechst 33258 staining was conducted to observe nuclear morphology changes. Protein expression of apoptosis and PI3K/AKT signaling as well as mRNA expression of ferroptosis and apoptosis were determined by Western Blotting and RT-qPCR. The effects of WTX and LY294002 combination on the PI3K/Akt/mTOR signaling pathway were measured by Western Blotting. Finally, the xenograft tumor mouse model was established by subcutaneous injection of CT26 cells to measure tumors volume and weight. Hematoxylin and eosin (HE) staining and immunohistochemical analysis were used to observe the pathological changes and the protein expression in tumor tissues. ResultsThere were 286 potential treatment targets from 130 bioactive compounds in WTX, 1349 CRC-related targets were identified. Eleven core targets (TP53, AKT1, STAT3, JUN, TNF, HSP90AA1, IL-6, MAPK3, CASP3, EGFR, MYC) were found by PPI network analysis constructed of 142 common targets. The results of KEGG enrichment displayed PI3K/AKT signaling pathway as core pathway. After the treatment of WTX, the inhibitory of viability, metastases and cell cycle arrest at G2/M phase were observed in HCT116 cells. Moreover, WTX induced an increase in the expression of apoptosis proteins (Bak, cytochrome c, cleaved caspase-9/caspase-9 and cleaved caspase-3/caspase-3) and the levels of ROS and MDA, a decrease in the expression of PI3K/AKT signaling related proteins (PI3K, p-PI3K, p-AKT/AKT and p-mTOR/mTOR) and the level of SOD. WTX treatment significantly reduced the tumor weight, increased cleaved caspase-3 positive area and decreased that of ki67 in xenograft mouse model. ConclusionThrough a network pharmacology approach and in vitro experiments, we predicted and verified the effect of WTX on colorectal cancer cells mainly depended on the regulation of intrinsic apoptosis via PI3K/AKT signaling pathway, and further animal experiments proved that WTX has a good anti-colon cancer effect in vivo.

STEM-06. NUCLEAR STRUCTURE MAINTAINS GLIOBLASTOMA STEM CELLS

Abstract Glioblastoma stem cells (GSCs) contribute to therapeutic resistance, promote tumor angiogenesis, and evade anti-tumor immune responses. We and others have shown that GSCs display differential regulation of their epigenetic landscapes and metabolic profiles and contribute to the striking cellular heterogeneity and complex tumor microenvironments. Here, we find that GSCs undergo dramatic changes in chromatin organization and nuclear morphology upon differentiation. Leveraging unbiased RNA sequencing and protein profiling screens comparing GSCs and differentiated glioma cells, we identified nuclear membrane molecular regulation that defines nuclear size and lipid regulation. Genetic or pharmacologic targeting of nuclear envelope function reduced cancer stem cell proliferation, self-renewal, and in vivo tumor growth. Collectively, we define a novel role of the nuclear envelope in maintaining chromatin state and cancer stem cell self-renewing, informing the development of novel potential therapeutic paradigms for glioblastoma.

Redistribution of lamina-associated domains reshapes binding of pioneer factor FOXA2 in development of nonalcoholic fatty liver disease.

Nonalcoholic fatty liver disease (NAFLD) is highly prevalent in type 2 diabetes mellitus and the elderly, impacting 40% of individuals over 70. Regulation of heterochromatin at the nuclear lamina has been associated with aging and age-dependent metabolic changes. We previously showed that changes at the lamina in aged hepatocytes and laminopathy models lead to redistribution of lamina-associated domains (LADs), opening of repressed chromatin, and up-regulation of genes regulating lipid synthesis and storage, culminating in fatty liver. Here, we test the hypothesis that change in the expression of lamina-associated proteins and nuclear shape leads to redistribution of LADs, followed by altered binding of pioneer factor FOXA2 and by up-regulation of lipid synthesis and storage, culminating in steatosis in younger NAFLD patients (aged 21-51). Changes in nuclear morphology alter LAD partitioning and reduced lamin B1 signal correlate with increased FOXA2 binding before severe steatosis in young mice placed on a western diet. Nuclear shape is also changed in younger NAFLD patients. LADs are redistrubted and lamin B1 signal decreases similarly in mild and severe steatosis. In contrast, FOXA2 binding is similar in normal and NAFLD patients with moderate steatosis and is repositioned only in NAFLD patients with more severe lipid accumulation. Hence, changes at the nuclear lamina reshape FOXA2 binding with progression of the disease. Our results suggest a role for nuclear lamina in etiology of NAFLD, irrespective of aging, with potential for improved stratification of patients and novel treatments aimed at restoring nuclear lamina function.

Methanolic extract of Entada phaseoloides inhibits Colorectal carcinoma cells proliferation of HT-29 cell by Modulating mitochondria-mediated apoptotic pathway

Colorectal cancer (CRC) is among the leading three diseases with higher death rates worldwide, with an expected 2.2 million new cases continuously in 2030. The expanding utilization of common plant-inferred parts, investigating the counter proliferative impacts of phytochemicals is progressively picking up significance in planning anticancer medications. This study aimed to examine the effect of methanolic extracts of Entada phaseoloides (MEEP) on the apoptotic pathway in human colon carcinoma cells (HT-29 cells). MTT assay and live/dead staining with fluorescein diacetate/propidium iodide (FDA/PI) were utilized to quantify cell viability in cancer cells. This research facility's exploratory investigation showed the effects of colon malignant growth cells (HT-29) exposed to various portions (1.23, 3.70, 11.11, 33.33, and 100µg/mL) of MEEP. The pure compounds isolated from the extracts includes Oleic acid, Entadamide A, Entadamide A-beta-D-glucopyranoside, Phaseoloidin. The result showed that MEEP actuated the concealment of cell viability and apoptosis in HT-29 cells in a dose-dependent manner. This included characteristic changes in nuclear morphology, the breakdown of mitochondrial membrane potential (Δψm), up-regulation of pro-apoptotic BAX, and down-regulation of anti-apoptotic Bcl-2, which initiates the transformation of caspase-3 to cleaved caspase-3, thus actuating PARP promoting apoptosis. Furthermore, it was found that MEEP does not affect ROS production. Thus overall findings applies against proliferative impact through various signalling pathways, is an outstanding possibility for hostile to colon cancer therapy with the help of natural sources.

Bromoditerpenes from the Red Seaweed Sphaerococcus coronopifolius as Potential Cytotoxic Agents and Proteasome Inhibitors and Related Mechanisms of Action.

Seaweeds are a great source of compounds with cytotoxic properties with the potential to be used as anticancer agents. This study evaluated the cytotoxic and proteasome inhibitory activities of 12R-hydroxy-bromosphaerol, 12S-hydroxy-bromosphaerol, and bromosphaerol isolated from Sphaerococcus coronopifolius. The cytotoxicity was evaluated on malignant cell lines (A549, CACO-2, HCT-15, MCF-7, NCI-H226, PC-3, SH-SY5Y, and SK-MEL-28) using the MTT and LDH assays. The ability of compounds to stimulate the production of hydrogen peroxide (H2O2) and to induce mitochondrial dysfunction, the externalization of phosphatidylserine, Caspase-9 activity, and changes in nuclear morphology was also studied on MCF-7 cells. The ability to induce DNA damage was also studied on L929 fibroblasts. The proteasome inhibitory activity was estimated through molecular docking studies. The compounds exhibited IC50 values between 15.35 and 53.34 µM. 12R-hydroxy-bromosphaerol and 12S-hydroxy-bromosphaerol increased the H2O2 levels on MCF-7 cells, and bromosphaerol induced DNA damage on fibroblasts. All compounds promoted a depolarization of mitochondrial membrane potential, Caspase-9 activity, and nuclear condensation and fragmentation. The compounds have been shown to interact with the chymotrypsin-like catalytic site through molecular docking studies; however, only 12S-hydroxy-bromosphaerol evidenced interaction with ALA20 and SER169, key residues of the proteasome catalytic mechanism. Further studies should be outlined to deeply characterize and understand the potential of those bromoditerpenes for anticancer therapeutics.

Nuclear and Morphological Alterations in Erythrocytes, Antioxidant Enzymes, and Genetic Disparities Induced by Brackish Water in Mrigal Carp (Cirrhinus mrigala).

Salinization of aquatic ecosystem, abrupt climate change, and anthropogenic activities cause adverse impact on agricultural land/soil as well as the aquaculture industry. This experimental study was designed to evaluate different biomarkers of oxidative stress, antioxidant enzymes, and genotoxic potential of diverse salinities of brackish water on freshwater fish. A total of 84 fresh water mrigal carp (Cirrhinus mrigala) were randomly segregated and maintained in four groups (T0, T1, T2, and T3) in a glass aquarium under similar laboratory conditions at various salinity levels (0, 3, 5, and 7 parts per thousand) to determine the pathological influence of brackish water. All the fish in groups T1, T2, and T3 were exposed to various salinity levels of brackish water for a period of 90 days while the fish of group T0 served as the control group. The experimental fish reared in different groups T1, T2, and T3 displayed various physical and behavioral ailments. The results revealed significantly augmented quantity of different oxidative stress indicators including reactive oxygen species (ROS) and thiobarbituric acid reactive substance (TBARS) in different visceral tissues (kidneys, liver, and gills) of exposed fish. Different antioxidant enzymes such as reduced glutathione (GSH), peroxidase (POD), superoxide dismutase (SOD), and catalase (CAT) along with total proteins were remarkably reduced in the kidneys, gills, and liver tissues. Results showed significantly increased values of different nuclear abnormalities (erythrocyte with micronucleus, erythrocyte with condensed nucleus, and erythrocyte with lobed nucleus) and morphological changes (pear shaped erythrocyte, spindle-shaped erythrocytes, and spherocyte) in red blood cells of experimental fish. The results on genotoxic effects exhibited significantly increased DNA damage in isolated cells of liver, kidneys, and gills of exposed fish. The findings of our experimental research suggested that brackish water causes adverse toxicological impacts on different visceral tissues of fresh water fish at higher salinity level through disruption and disorder of physiological and biochemical markers.

Examination of Exopolysaccharides from Porphyridium cruentum for Estimation of Their Potential Antitumour Activity in Vitro

&#x0D; &#x0D; &#x0D; Globally, research is being conducted focusing on new biologically active substances and technologies for their application in various sectors, in the hope that their biological and medical applications will be widely used in the next decade.&#x0D; The introduction of therapeutic agents of biological origin isolated from certain species of bacteria and algae and their potential for cancer treatment is based on their lower toxicity and hence greater safety compared to synthetic pharmaceuticals. In this respect, the potential of microbial producers and microalgae is enormous. Among the biologically active components isolated from algae of the greatest importance for biology and medicine are complex polysaccharides, as well as some pigments.&#x0D; In the present work, the antitumour properties of two fractions of polysaccharides from the red microalgae Porphyridium cruentum were observed by determining their effect on cell viability of four different cell lines.&#x0D; New data from the study indicate that samples treated with P. cruentum polysaccharide have significant and dose-dependent antiproliferative and cytotoxic effects. Changes in nuclear morphology in MCF-7 tumour cells induced by polysaccharide treatment, including chromatin condensation, nucleus fragmentation, and apoptotic body formation, have been demonstrated. Polysaccharides show obvious pharmacological effects in the treatment of tumour cells without side effects.&#x0D; &#x0D; &#x0D;

H2O2 Induces Calcium and ERMES Complex-Dependent Mitochondrial Constriction and Division as Well as Mitochondrial Outer Membrane Remodeling in Aspergillus nidulans.

The dynamin-like protein DnmA and its receptor FisA are essential for H2O2-induced mitochondrial division in Aspergillus nidulans. Here, we show that in the absence of DnmA or FisA, mitochondria show few spontaneous transient constrictions, the frequency of which is extensively increased by H2O2 or the carbonyl cyanide m-chlorophenyl hydrazone (CCCP). While H2O2-induced constrictions are transient, CCCP induces a drastic and irreversible alteration of mitochondrial filaments. H2O2 induces a gradual mitochondrial depolarization, while CCCP-induced depolarization is abrupt. The calcium chelator BAPTA-AM prevents the formation of mitochondrial constrictions induced by either H2O2 or CCCP. H2O2 also induces major rearrangements of the mitochondrial outer membrane, which remain after constrictions dissipate, as well as changes in endoplasmic reticulum (ER) and nuclear morphology. Similar mitochondrial constriction, ER and nuclear morphology changes are detected during the early stages of asexual development. ER and ER-Mitochondria encounter structure (ERMES) complex—composed of proteins Mdm10, Mmm1, Mdm43 and Mdm12—are important for mitochondrial division in Saccharomyces cerevisiae. As the Mdm10 ortholog MdmB was found to be essential in A. nidulans, we evaluated its functions in ΔmdmB terminal mutants and ΔmdmB heterokaryons. ΔmdmB conidia produce a short germ tube that fails to grow further, in which inherited mitochondria become gigantic and round shaped, lacking clear contacts with the ER. In slow-growing ΔmdmB heterokaryotic mycelia, multiple hyphae contain very long mitochondria with high ROS levels, as occur in ΔdnmA and ΔfisA mutants. In this hyphae, H2O2 fails to induce mitochondrial constrictions but not outer mitochondrial membrane reshaping, indicating that these are two separate effects of H2O2. Our results indicate that H2O2 induces a generalized mitochondrial constriction response, prior to actual division, involving gradual depolarization; they also indicate that Ca2+ and the ERMES complex are critical for both mitochondrial constriction and division. This supports a view of mitochondrial dynamics as the result of a cascade of signaling events that can be initiated in vivo by H2O2.

Nesprin-1 LINC complexes recruit microtubule cytoskeleton proteins and drive pathology in Lmna-mutant striated muscle.

Mutations in LMNA, the gene encoding A-type lamins, cause laminopathies-diseases of striated muscle and other tissues. The aetiology of laminopathies has been attributed to perturbation of chromatin organization or structural weakening of the nuclear envelope (NE) such that the nucleus becomes more prone to mechanical damage. The latter model requires a conduit for force transmission to the nucleus. NE-associated Linker of Nucleoskeleton and Cytoskeleton (LINC) complexes are one such pathway. Using clustered regularly interspaced short palindromic repeats to disrupt the Nesprin-1 KASH (Klarsicht, ANC-1, Syne Homology) domain, we identified this LINC complex protein as the predominant NE anchor for microtubule cytoskeleton components, including nucleation activities and motor complexes, in mouse cardiomyocytes. Loss of Nesprin-1 LINC complexes resulted in loss of microtubule cytoskeleton proteins at the nucleus and changes in nuclear morphology and positioning in striated muscle cells, but with no overt physiological defects. Disrupting the KASH domain of Nesprin-1 suppresses Lmna-linked cardiac pathology, likely by reducing microtubule cytoskeleton activities at the nucleus. Nesprin-1 LINC complexes thus represent a potential therapeutic target for striated muscle laminopathies.

Responses of Prorocentrum cordatum (Ostenfeld) Dodge, 1975 (Dinoflagellata) to copper nanoparticles and copper ions effect.

In the present study, changes were determined in morphological, structural-functional, and fluorescent parameters of Prorocentrum cordatum with the addition of CuO nanoparticles (NPs) and copper ions (CuSO4). A stimulating effect of low Cu2+ concentrations (30μg L-1) on algal growth characteristics was observed. HigherCu2+ concentration of 60-600μg L-1 and CuO NPs concentration of 100-520μg L-1 inhibited algal growth. Ionic copper is more toxic to P. cordatum than NPs. After 72h of algae cultivation in the medium supplemented with CuSO4 and CuO NPs, EC50 values (calculated based on cell abundance) were of 60 and 300μg L-1 (in terms of copper ions), respectively. Reduction in algal growth rate is due to disruption in cell cycle, changes in nuclear morphology, chromatin dispersion, and DNA damage. The studied pollutants slightly affected the efficiency of P. cordatum photosynthetic apparatus. Addition of the pollutants resulted in an increased production of reactive oxygen species (ROS). At a concentration of Cu2+ of120μg L-1 and a concentration of CuO NPs 0-300μg L-1 of CuO NPs increase in ROS production is short-term with a decrease at later stages of the experiment. This is probably due to the activation of antioxidant mechanisms in cells and an increase in the concentration of carotenoids (peridinin) in cells. The high values of ROS production persisted throughout the experiment at sublethal copper concentrations (400-600μg L-1 of CuSO4 and 520μg L-1 of CuO NPs). Sublethal concentrations of pollutants caused restructuring of cell membranes in P. cordatum. Shedding of cell membranes (ecdysis) and formation of immobile stages (temporary or resting cysts) were recorded. The pronounced mechanical impact of NPs on the cell surface was observedsuch as-deformation and damage of a cell wall, its "wrinkling" and shrinkage, and adsorption of NP aggregates.

Apoptosis Induction Associated with Enhanced ER Stress Response and Up-Regulation of c-Jun/p38 MAPK Proteins in Human Cervical Cancer Cells by Colocasia esculenta var. aquatilis Hassk Extract

Colocasia esculenta var. Aquatilis Hassk, elephant ear (CF-EE) has been widely used as traditional food and medicine. It also shows other therapeutic properties, such as antimicrobial and anti-cancer activity. In this study, we aim to investigate the effect of CF-EE extract on apoptosis induction associated with ER stress in cervical cancer HeLa cells. Cell viability was determined by MTT assay. Assessments of nuclear morphological changes, mitochondrial membrane potential, and intracellular reactive oxygen species (ROS) production were conducted by hoeshst33342, JC-1, and DCFH-DA fluorescence staining, respectively. Sub-G1 DNA content was analyzed by flow cytometry, and protein expression was determined by Western blotting. The results demonstrate that CF-EE extract suppressed HeLa cell growth and induced nuclear condensation and apoptotic bodies. There was also a loss of mitochondrial membrane potential and increased apoptosis marker protein expression, including Bax, cleaved-caspase-7, and cleaved-PARP. In addition, the results show that CF-EE extract induced ROS, increased ER stress proteins (GRP78 and CHOP), enhanced p38 and c-Jun phosphorylation, and inhibited Akt expression in HeLa cells. In summary, CF-EE extract induced apoptotic cell death-associated ROS-induced ER stress and the MAPK/AKT signaling pathway. Therefore, CF-EE extract has anticancer therapeutic potential for cervical cancer treatment in the future.

FUCCItrack: An all-in-one software for single cell tracking and cell cycle analysis

Beyond the more conventional single-cell segmentation and tracking, single-cell cycle dynamics is gaining a growing interest in the field of cell biology. Thanks to sophisticated systems, such as the fluorescent ubiquitination-based cell cycle indicator (FUCCI), it is now possible to study cell proliferation, migration, changes in nuclear morphology and single cell cycle dynamics, quantitatively and in real time. In this work, we introduce FUCCItrack, an all-in-one, semi-automated software to segment, track and visualize FUCCI modified cell lines. A user-friendly complete graphical user interface is presented to record and quantitatively analyze both collective cell proliferation as well as single cell information, including migration and changes in nuclear or cell morphology as a function of cell cycle. To enable full control over the analysis, FUCCItrack also contains features for identification of errors and manual corrections.