Cell Blebbing Research Articles

Prolonged Intradialytic Hypoxemia Is Associated With Functional Disturbances in Erythrocytes

Prolonged Intradialytic Hypoxemia Is Associated With Functional Disturbances in Erythrocytes

Abstract 1054: Patient derived organoids demonstrate synergistic antitumor effect of senogenic and senolytic drug combination of Aurora kinase inhibitor and BCL2/xL inhibitor

Abstract The management of metastatic melanoma continues to be challenging, despite of evolving treatments such as immune checkpoint blockade and inhibitors of mutated BRAF. New therapies that work via immune and BRAF-independent mechanisms are urgently required for tumors that are unresponsive to currently available treatments. Here we hypothesized that combination of Aurora Kinase inhibitor (AURKAi) alisertib (senescence inducer) with a drug that selectively kills senescent cells, such as BCL2/xL inhibitor (BCLi) navitoclax (senolytic), will induce apoptosis in melanoma cells. Patient derived organoids (PDOs) were used to test anti- tumor effect of AURKAi and BCLi combination treatment. H&E staining of PDOs and their matched original tumors revealed similar histology. Fourteen out of ninety PDOs were highly sensitive to AURKAi and BCLi combination treatment exhibiting robust induction of cell death. NextGen DNA sequencing indicated that all sensitive PDOs retained wild type TP53, while resistant tumors had mutations in this gene. Knockout or knockdown of TP53 abrogated AURKAi and BCLi-induced cell death in melanoma cells, suggesting that transcription factor p53 encoded by TP53 is a key mediator of drug response. Addition of BCLi to AURKAi activated proapoptotic p53 targets BAX, NOXA and induced cleavage of PARP and Caspases 3, 7, and 8. Electron microscopy revealed morphological signs of apoptosis in combination-treated cells, including cell blebbing, apoptotic bodies, and nuclear fragmentation. This suggests that BCLi shifts cell fate decision in AURKAi-treated cells towards apoptosis. In conclusion, combination treatment with AURKAi and BCl2i induced p53-dependent apoptosis. Preclinical evaluation of this drug combination in PDO model demonstrated robust efficacy against melanomas with wild type p53. Since p53 mutations are relatively rare in melanoma, combination of AURKAi and BCLi presents promising therapeutic option for this deadly disease. Keywords: Apoptosis, melanoma, patient-derived organoids, aurora kinase, bcl-2 Citation Format: Vijaya Bharti, Vivian Weiss, Chengli Shen, Sheau Chiann Chen, Ashlyn Blevins, Rebecca L. Shattuck-Brandt, Ann Richmond, Anna E. Vilgelm. Patient derived organoids demonstrate synergistic antitumor effect of senogenic and senolytic drug combination of Aurora kinase inhibitor and BCL2/xL inhibitor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1054.

Colorado tick fever virus induces apoptosis in human endothelial cells to facilitate viral replication

Colorado tick fever virus (CTFV) belongs to the genus Coltivirus of the Reoviridae family, and it is the causative agent of Colorado tick fever. Symptoms of the infection are characterized by sudden biphasic fever, headache, and petechial rash, while severe forms of the disease can include meningoencephalitis, hemorrhagic fever, and death in children. However, the mechanisms underlying CTFV induced pathology and severe complications remain unknown. As CTFV is spread by tick bites and disseminates systemically via hematogenous routes, we performed in vitro analysis examining the interactions between endothelial cells (ECs) and CTFV. Our findings indicate that dermal microvascular ECs, HMEC-1, are susceptible and permissive to CTFV infection. To investigate the role of CTFV infection on endothelial barrier function, we assessed transendothelial electrical resistance (TEER) by xCELLigence and observed a dose-dependent decrease in cell index, indicating increased vascular permeability starting at approximately hour 18 (MOI=1) and hour 26 (MOI=0.1). Since CTFV induced cytopathic effect and increased vascular permeability in HMEC-1 cells, we hypothesized that CTFV causes apoptotic cell death. Our results showed that HMEC-1 cells infected with CTFV at 48 h caused a significant increase in Annexin V staining with reduced viability compared to uninfected cells suggesting CTFV induces apoptotic cell death in human ECs. Electron microscopy also was consistent with apoptotic features, including chromatin condensation and cell blebbing. Furthermore, CTFV induced caspase-3/7 activation at 24 and 48 h post-infection (hpi). The inhibition of caspase activity using Z-VAD-FMK reduced CTFV induced cell death and significantly reduced viral titer. These results indicated that CTFV can infect ECs, exerting direct adverse effects, leading to vascular permeability and cell death. Overall, our data suggest that caspase-mediated apoptosis is a critical mechanism by which CTFV induces disease in the host and enhances viral replication. Future studies will examine the viral and cellular determinants involved in CTFV induced apoptosis in human ECs.

A Screen of Natural Product Extracts Identifies Moenomycin as a Potent Antigonococcal Agent.

Increasing multidrug resistance in Neisseria gonorrheae is a growing public health crisis. Resistance to the last line therapies, cephalosporins and azithromycin, are of particular concern, fueling the need to discover new treatments. Here, we identified the phosphoglycolipid moenomycin from a screen of microbial natural products against drug-resistant N. gonorrheae as a potent antigonococcal agent. Moenomycin demonstrates excellent activity (MIC = 0.004-0.03 μg/mL) against a variety of multidrug-resistant N. gonorrheae. Importantly, moenomycin, thought to be a Gram-positive specific antibiotic, penetrates the Gram-negative gonococcal outer membrane. Moenomycin causes intracellular accumulation of peptidoglycan precursors, cell blebbing, and rupture of the cell envelope, all consistent with cell wall biosynthesis inhibition. Serial bacterial exposure to moenomycin for 14 days revealed slow development of resistance (MICDay14 = 0.03-0.06 μg/mL), unlike the clinically used drug azithromycin. Our results offer the potential utility of moenomycin as a lead for antigonococcal therapeutic candidates and warrant further investigation.

Establishing SW1353 Chondrocytes as a Cellular Model of Chondrolysis.

Osteoarthritis (OA) is the most common degenerative joint disease characterised by chondrocyte cell death. An in vitro model of chondrocyte cell death may facilitate drug discovery in OA management. In this study, the cytotoxicity and mode of cell death of SW1353 chondrocytes treated with 24 h of OA inducers, including interleukin-1β (IL-1β), hydrogen peroxide (H2O2) and monosodium iodoacetate (MIA), were investigated. The microscopic features, oxidative (isoprostane) and inflammatory markers (tumour necrosis factor-alpha; TNF-α) for control and treated cells were compared. Our results showed that 24 h of H2O2 and MIA caused oxidative stress and a concentration-dependent reduction of SW1353 cell viability without TNF-α level upregulation. H2O2 primarily induced chondrocyte apoptosis with the detection of blebbing formation, cell shrinkage and cellular debris. MIA induced S-phase arrest on chondrocytes with a reduced number of attached cells but without significant cell death. On the other hand, 24 h of IL-1β did not affect the cell morphology and viability of SW1353 cells, with a significant increase in intracellular TNF-α levels without inducing oxidative stress. In conclusion, each OA inducer exerts differential effects on SW1353 chondrocyte cell fate. IL-1β is suitable in the inflammatory study but not for chondrocyte cell death. H2O2 and MIA are suitable for inducing chondrocyte cell death and growth arrest, respectively.

Elastic wetting: Substrate-supported droplets confined by soft elastic membranes

While classical wetting is well captured by the famous Young's equation and classical bulge and blister models are readily available, there is limited understanding of a micro- or nano-scale droplet being covered by an ultrasoft elastic membrane. We call this phenomenon elastic wetting to feature the interplay between the liquid's surface tension and the membrane's elastic deformation. Examples of elastic wetting include cell blebs and 2D material bubbles, where the membrane thickness ranges from microns to sub-nanometers. In this work, we study the equilibrium of elastic wetting and solve for the profiles and the pressure-volume relations of the membrane-confined droplets. We show that in elastic wetting, the pressure across the membrane/droplet interface can be described by a simple superposition of the Young-Laplace equation and the nonlinear membrane equation. Furthermore, nonlinear elasticity, geometric nonlinearity, and surface tension, together with membrane-substrate adhesion, interweave at the contact line, leading to rich membrane-confined droplet configurations. Finally, we examine the effect of substrate compliance on elastic wetting and find that the rigid substrate assumption approximates well for most of the existing experiments in the literature. Our results provide fundamental mechanistic insights into the various phenomena of elastic wetting as well as viable means to extract physical parameters including the bubble pressure and the interface energies.

Synthesis, Cytotoxicity and Anti-Proliferative Activity Against AGS Cells of New 3(2H)-Pyridazinone Derivatives Endowed with a Piperazinyl Linker.

Novel twenty-three 3(2H)-pyridazinone derivatives were designed and synthesized based on the chemical requirements related to the anti-proliferative effects previously demonstrated within this scaffold. The introduction of a piperazinyl linker between the pyridazinone nucleus and the additional (un)substituted phenyl group led to some compounds endowed with a limited cytotoxicity against human gingival fibroblasts (HGFs) and good anti-proliferative effects against gastric adenocarcinoma cells (AGS) as evaluated by MTT and LDH assays, using doxorubicin as a positive control. Successive analyses revealed that the two most promising representative compounds (12 and 22) could exert their effects by inducing oxidative stress as demonstrated by the hydrogen peroxide release and the morphological changes (cell blebbing) revealed by light microscopy analysis after the haematoxylin-eosin staining. Moreover, to further assess the apoptotic process induced by compounds 12 and 22, Bax expression was measured by flow cytometry. These findings enlarged our knowledge of the structural requirements in this scaffold to display valuable biological effects against cancerous cell lines.

Optogenetic Control of Myocardin‐Related Transcription Factor A Subcellular Localization and Transcriptional Activity Steers Membrane Blebbing and Invasive Cancer Cell Motility

The myocardin-related transcription factor A (MRTF-A) controls the transcriptional activity of the serum response factor (SRF) in a tightly controlled actin-dependent manner. In turn, MRTF-A is crucial for many actin-dependent processes including adhesion, migration, and contractility and has emerged as a novel target for anti-tumor strategies. MRTF-A rapidly shuttles between cytoplasmic and nuclear compartment via dynamic actin interactions within its N-terminal RPEL domain. Here, optogenetics is used to spatiotemporally control MRTF-A nuclear localization by blue light using the light-oxygen-voltage-sensing domain 2-domain based system LEXY (light-inducible nuclear export system). It is found that light-regulated nuclear export of MRTF-A occurs within 10-20 min. Importantly, MRTF-A-LEXY shuttling is independent of perturbations of actin dynamics. Furthermore, light-regulation of MRTF-A-LEXY is reversible and repeatable for several cycles of illumination and its subcellular localization correlates with SRF transcriptional activity. As a consequence, optogenetic control of MRTF-A subcellular localization determines subsequent cytoskeletal dynamics such as non-apoptotic plasma membrane blebbing as well as invasive tumor-cell migration through 3D collagen matrix. This data demonstrates robust optogenetic regulation of MRTF as a powerful tool to control SRF-dependent transcription as well as cell motile behavior.

Combined treatment of cisplatin and 5-Fluorouracil Induces cell death in ARPE-19 cells through the endoplasmic reticulum stress pathway

Aim: Cisplatin and 5-Fluorouracil have been used for about 50 years as chemotherapy agents. Combination of these two agents is known as PF therapy and is widely used in the treatment of anal, esophageal cancer, head and neck cancer. One of the problems encountered with antineoplastic agents is the rapid and irreversible side effects on non-target cells and organs. It is common that occourence of ocular toxicity by chemotherapy. Materials and Methods: ARPE-19 cell viability and toxicity were determined by performing (MTT)-based colorimetric assays and IC50 values calculated by probit analyses options of SPSS 20 Software. Acridine orange/Ethidium bromide (AO/EB) fluorescent staining was performed to determine structural changes such as cell blebbing and chromatin condensation during the apoptosis process on cells. Images examined under a fluorescent microscope (Zeiss Axio Vert.A1) at 40X magnification with FITC filter for AO and Tex Red filter. And Real-time PCR was performed with ABI Step One Plus and Quant Studio 5 Real-Time PCR systems.Results: PF treatment within 24 hours and 48 hours caused ER stress due to toxic effect and this induced cell death through apoptosis via death receptor signaling and intrinsic pathway in ARPE-19 cells. Conclusion: PF-induced activation of the ER stress mechanism can be used as a novel therapeutic strategy for the prevention of side effects of non-target cells.

Membrane Homeostasis: The Role of Actin Cytoskeleton

The plasma membrane of a cell undergoes continuous deformations and turning-over of its constituents required for a diverse set of functions. And yet, the membrane retains its steady-state surface tension—without which these functions are adversely affected. In this review, we discuss how the establishment and maintenance of the homeostatic state of the membrane has a major contribution from the actin cytoskeleton. The regulation of tension at the global (or cellular) scale is extensively studied for the past few decades, leading to our understanding of how actin polymerization forces, myosin-II based contractile forces, and ezrin-mediated attachment to the membrane—each have a separate and sometimes multiple possible effects on membrane tension. Drawing examples from cell motility and blebbing cells, we highlight how the dynamics of the cytoskeleton decides if the steady-state tension has a uniform profile, front-rear gradients, or temporally varying tension profiles in single cells. Non-invasive studies open up new avenues especially allowing the investigations focusing on local regulation of tension—at the plasma membrane and inside cells.

Ethyl acetate fraction of Anethum graveolens seeds exerts an antiproliferative effect by inhibiting anti-apoptotic proteins in MCF-7 and PC-3 cells: An in vitro and molecular docking study

Background: Anethum graveolens seeds have therapeutic benefits, which may be a potential approach to the treatment of different cancers. Objectives: We investigated, the antiproliferative effect of ethyl acetate fraction of dill (EAFD), on MCF-7 and PC-3 cell lines and its two most active components, anethole and carvone by molecular docking analysis. Materials and Methods: In-vitro assays, like cell viability assay and measurement of reactive oxygen species, were performed besides performing Giemsa stain and other fluorescent stains; JC-1 dye, dual mixed stain ethidium bromide/acridine orange and 4,6-diamidino-2-phenylindole stain to study morphological characteristics, including molecular docking analysis. EAFD concentrations (0.2, 0.4, 0.6, 0.8, 1.0 mg/ml) were used. Results: The EAFD prominently inhibited the proliferation of MCF-7 cells and PC-3 cells by dose-dependent and time-dependent methods. Increased exposure of EAFD to MCF-7 and PC-3 cells increases the level of intracellular oxidative stress. Similarly, EAFD exposure confirms the morphological alternations such as cell shrinkage, membrane disruption, nuclear condensation, and blebbing in phase-contrast microscopy and even fluorescent microscopy stains can lead to mitochondrial membrane degradation, chromatin condensation, nuclear fragmentation. Analyses of docking results suggest that anethole and carvone bind to the hydrophobic patches of Bcl-2 and Bcl-xL through hydrophobic interactions. Conclusion: The EAFD may be antiproliferative activity and leading to pro-apoptotic cell death. Molecular docking analysis of Bcl-2 and Bcl-xL anti-apoptotic protein indicated that the antiproliferative activity and pro-apoptotic cell deaths by EAFD are possibly due to inhibition of these proteins.

Numerical Model for Formation and Evolution of the Bleb

The morphology of a bleb and its changes are critical to the amoeboid migration of a cell. By releasing bonds between the membrane and the cortex of a cell, the formation of a bleb can be observed experimentally, but the mechanism that affects the size and shape of a bleb during amoeboid migration requires further study. In this study, by adapting the governing equations and discrete equations of the two-dimensional fluid–solid coupling model recommended by Strychalski and Guy [2013 “A computational model of bleb formation,” Mathematical Medicine and Biology 30(2), 115–130], we overcome the defect that the bleb by traditional means is always too small compared with experimental results, and simulate the behaviors of cell blebs successfully. The effects of various parameters such as the number of broken bonds, the viscosity coefficient of the cortex, and the cell’s membrane modulus on the size and the shape of the bleb are investigated. Numerical results show that the model effectively simulates the formation and evolution of a bleb, thus, the contributions of several factors to bleb shape and size are successfully derived.

Alginate core–shell microcapsule reduces the DMSO addition-induced osmotic damage to cells by inhibiting cellular blebs

• Blebbing of cells encapsulated in alginate core–shell microcapsules is studied. • DMSO addition-induced blebs of cells in alginate microcapsules are small. • Relationship between DMSO-induced bleb size and cell viability is established. • Microencapsulation reduces the DMSO addition-induced osmotic damage to cells. In cryopreservation, the addition of cryoprotectant can change the intra- and extra-cellular osmotic pressure, affect the cell morphology, and induce blebs on the plasma membrane. In this study, the blebs of cells microencapsulated in the alginate microsphere induced by osmotic shock were studied, and the effects of microencapsulation on bleb size and cell viability were determined. Firstly, a coaxial co-flow focusing device was applied to generate cell-laden microcapsules using alginate hydrogel in this paper. Then, cellular blebs induced by DMSO with various concentrations under microencapsulation were compared with that when non-encapsulated, and the dynamic process of cellular bleb was investigated. Finally, the qualitative relationship between bleb size and cell viability in the presence of DMSO was built, and thus the effects of microencapsulation on bleb size and viability were evaluated. The results show that the bleb size is smaller and the cell viability is higher, and cell microencapsulation can significantly inhibit the excessively large blebs generated on the cell membrane and reduce the osmotic damage to cells when loading cryoprotectant and then to improve cell viability during cryopreservation. This work can provide insights for optimizing cryoprotectant-loading protocols, offer a new avenue to study cell blebbing, and advance future research on cryopreservation of rare cells and biomaterials.

Combined theoretical and experimental studies reveal the newly synthesized pyrimidinones as potential apoptotic agents

Reaction of acetophenone thiosemicarbazones (5–8) and (2-methyl) diethyl malonate in absolute ethanol under reflux conditions furnished the corresponding pyrimidinone analogs (9–12) in good to excellent yields. The resulting pyrimidines were characterised using spectral and analytical data analysis. The resulting products were studied for DNA interaction studies of by various techniques like UV–VIS, fluorescence spectroscopy, viscosity measurements, molecular docking and gel electrophoresis. To look into the various structural and molecular properties of the target compounds, theoretical DFT involving B3LYP/6-31G∗∗ level of theory was employed. Frontier molecular orbital analysis (FMO) was carried out which in turn helped us to determine and evaluate the descriptors like chemical hardness, potential, nucleophilicity and electrophilicity index, etc. Molecular docking and viscosity studies indicated that the synthesized pyrimidinones are minor groove binders of DNA with corresponding intrinsic binding constants of 3.41 × 103 M−1, 2.91 × 103 M−1, 3.24 × 103 M−1 and 3.04 × 103 M−1 for compounds (9–12) respectively indicating higher binding affinity of 9 towards DNA. Gel electrophoresis depicted that all new compounds display a dose dependent cleavage activity with pBR322 DNA and efficient nuclease activity was found at 2 μM. An in vitro MTT cytotoxicity assay revealed that the compounds (9–12) possess potential toxicity against the different human cancer cells. The AO/EB staining showed that compound 9 caused apoptosis as revealed by the notable apoptotic features like nuclear shrinkage, blebbing and chromatin condensation in HepG2 cells. Compound 9 also caused decrease in the level of apoptotic marker Bcl-XL in Western blotting indicating its prompt ability to cause apoptosis.

Recombinant human lactoferrin carrying humanized glycosylation exhibits antileukemia selective cytotoxicity, microfilament disruption, cell cycle arrest, and apoptosis activities.

Lactoferrin has gained extensive attention due to its ample biological properties. In this study, recombinant human lactoferrin carrying humanized glycosylation (rhLf-h-glycan) expressed in the yeast Pichia pastoris SuperMan5, which is genetically glycoengineered to efficiently produce functional humanized glycoproteins inclosing (Man)5(GlcNAc)2 Asn-linked glycans, was analyzed, inspecting its potential toxicity against cancer cells. The live-cell differential nuclear staining assay was used to quantify the rhLf-h-glycan cytotoxicity, which was examined in four human cell lines: acute lymphoblastic leukemia (ALL) CCRF-CEM, T-cell lymphoblastic lymphoma SUP-T1, cervical adenocarcinoma HeLa, and as control, non-cancerous Hs27 cells. The defined CC50 values of rhLf-h-glycan in CCRF-CEM, SUP-T1, HeLa, and Hs27 cells were 144.45 ± 4.44, 548.47 ± 64.41, 350 ± 14.82, and 3359.07 ± 164µg/mL, respectively. The rhLf-h-glycan exhibited a favorable selective cytotoxicity index (SCI), preferentially killing cancer cells: 23.25 for CCRF-CEM, 9.59 for HeLa, and 6.12 for SUP-T1, as compared with Hs27 cells. Also, rhLf-h-glycan showed significant antiproliferative activity (P < 0.0001) at 24, 48, and 72h of incubation on CCRF-CEM cells. Additionally, it was observed via fluorescent staining and confocal microscopy that rhLf-h-glycan elicited apoptosis-associated morphological changes, such as blebbing, nuclear fragmentation, chromatin condensation, and apoptotic bodies in ALL cells. Furthermore, rhLf-h-glycan-treated HeLa cells revealed shrinkage of the microfilament structures, generating a speckled/punctuated pattern and also caused PARP-1 cleavage, a hallmark of apoptosis. Moreover, in ALL cells, rhLf-h-glycan altered cell cycle progression inducing the G2/M phase arrest, and caused apoptotic DNA fragmentation. Overall, our findings revealed that rhLf-h-glycan has potential as an anticancer agent and therefore deserves further in vivo evaluation.

Design of antimicrobial and cytolytic peptides by computational analysis of bacterial, algal, and invertebrate proteomes.

The increase of antibiotic resistance in bacterial species has raised the need to search for novel antimicrobial molecules. Antimicrobial peptides are molecules that commonly display an amphipathic character. In this work, we developed a computational strategy to search for new peptide sequences within the proteome of any organism that includes in-house developed software and the use of artificial intelligence tools available online. Eleven peptides were selected after analyzing 63,343 proteins from the proteomes of bacteria, algae and invertebrates. Then, we validated the results by means of several assays which were carried out against five (5) pathogenic bacterial species and two (2) cancer cell lines. As a result, we found that ten of the peptides were antimicrobial, with minimum inhibitory concentration values between 4 and [Formula: see text]. Furthermore, two of the more active peptides were also cytotoxic to human red blood cells and cancer cells. In general, the antimicrobial peptides we discovered produced damage on the bacterial cell membrane that included membrane wrinkling, cell blebbing, and leakage of cytoplasmic material. Based on these results, we concluded that the computational approach proposed for finding sequences encrypted in proteins is appropriate for the discovery of selective and non-selective antimicrobial and anticancer peptides.

Holotomographic microscopy: A new approach to detect apoptotic cell features.

Holotomographic (HT) microscopy, combines two techniques, holography and tomography, and, in this way, it allows to quantitatively and noninvasively investigate cells and thin tissue slices, by obtaining three-dimensional (3D) images and by monitoring inner morphological changes. HT has indeed two significant advantages: it is label-free and low-energy light passes through the specimen with minimal perturbation. Using quantitative phase imaging with optical diffraction tomography, it can produce 3D images by measuring the refraction index (RI). Therefore, based on RI values, HT can provide structural and chemical cell information, such as dry mass values, morphological changes, or cellular membrane dynamics. In this study, suspended and adherent culture cells have been processed for HT analyses. Some of them have been treated with known apoptotic drugs or pro-oxidant agents and cell response has been investigated both by conventional microscopic approaches and by HT. The ultrastructural and fluorescence images have been compared to those obtained by HT and their congruence has been discussed, with particular attention to apoptotic cell death and on correlated plasma membrane changes. HT appears a valid approach to further characterize well-known apoptotic features such as cell blebbing, chromatin condensation, micronuclei, and apoptotic bodies. Taken together, our data demonstrate that HT appears suitable to highlight suspended or adherent cell behavior under different conditions. In particular, this technique appears an important new tool to distinguish healthy cells from the apoptotic ones, as well as to monitor outer and inner cell changes in a rapid way and with a noninvasive, label-free, approach.

Effect of the Organic Functionality on the Synthesis and Antimicrobial Activity of Silver Nanoparticles

In this paper, the effects of the organic reducing agent and 3-aminopropyltrimethoxysilane on the synthesis and properties of mono-, bi-, and trimetallic noble metal nanoparticles were considered; the antimicrobial activity of these nanomaterials was also evaluated. It was shown that 3-aminoptopyltrimethoxysilane-treated noble metal cations undergo rapid conversion into nanoparticles in the presence of three organic reducing agents, namely, 3-glycidoxypropyltrimethoxysilane (3-GPTMS), cyclohexanone and formaldehyde; the nanoparticles were formed on the order of one minute under microwave incubation. Bimetallic nanoparticles were formed by simultaneous or sequential reduction of metal cations; the formation of trimetallic nanoparticles containing gold, silver and palladium was demonstrated using a similar approach. The nanoparticles were characterized using UV-Visible light spectrophotometry, transmission electron microscopy and zeta potential measurements. All three nanoparticles exhibited a size [Formula: see text]10[Formula: see text]nm size. The nanoparticles showed antimicrobial activity against Acinetobacter baumannii. Scanning electron microscopy imaging showed an alteration in the size and shape of nanoparticle-treated bacterium, with bleb formation and cell wall disruption observed within 1[Formula: see text]h of incubation at the MBC values of the nanoparticles. Fluorescence spectrophotometric imaging of silver nanoparticle-Acinetobacter baumannii interactions suggested selective binding of silver nanoparticles to surface proteins. Our results showed the preparation of the novel silver nanoparticles with potent Anti-Acinetobacter baumannii activity, which can serve as an alternative to conventional antimicrobial agents.

Hemolysin BL from novel Bacillus toyonensis BV-17 induces antitumor activity both in vitro and in vivo

ABSTRACT The gut microbiota plays an important role in cancer development and immunotherapy. Bacterial toxins have enormous antitumor potential due to their cytotoxicity and ability to activate the immune system. Using 16S rRNA gene sequencing, we compared the gut microbiota composition of fecal samples from healthy individuals and patients with colorectal cancer (CRC) and observed that the genus Bacillus was common in the healthy donors but was absent in the CRC patients. Further, we isolated a novel Bacillus toyonensis BV-17 from the fecal samples of the healthy individuals. Our results showed that the supernatant of the Bacillus toyonensis BV-17 cultures could quickly kill various tumor cell lines within minutes in vitro, by causing cell membrane disruption, blebbing, and leakage of cytoplasmic content. Fast protein liquid chromatography (FPLC) and mass spectrometry analysis identified hemolysin BL (HBL) as the effector molecule, which exhibits a different cytotoxicity mechanism compared to previous studies. Intra-tumor injection of low dose HBL inhibited the growth of both treated and untreated tumors in mice. The outcomes of this pioneer study suggest that HBL exhibits antitumor activity and is a potential chemotherapeutic agent that could be engineered to target only tumor cells in future.

A Finite Element Method for a Fourth Order Surface Equation With Application to the Onset of Cell Blebbing

A variational problem for a fourth order parabolic surface partial differential equation is discussed. It contains nonlinear lower order terms, on which we only make abstract assumptions, and which need to be defined for specified problems. We derive a semi-discrete scheme based on the surface finite element method, show a-priori error estimates, and use the analytical results to prove well-posedness. Furthermore, we present a computational framework where specific problems can be conveniently implemented and, later on, altered with relative ease. It uses a domain specific language implemented in Python. The high level program control can also be done within the Python scripting environment. The computationally expensive step of evolving the solution over time is carried out by binding to an efficient C++ software back-end. The study is motivated by cell blebbing, which can be instrumental for cell migration. Starting with a force balance for the cell membrane, we derive a continuum model for some mechanical and geometrical aspects of the onset of blebbing in a form that fits into the abstract framework. It is flexible in that it allows for amending force contributions related to membrane tension or the presence of linker molecules between membrane and cell cortex. Cell membrane geometries given in terms of a parametrisation or obtained from image data can be accounted for by the software. The use of a domain specific language to describe the model makes is straightforward to add additional effects such as reaction-diffusion equations modelling some biochemistry on the cell membrane. Some numerical simulations illustrate the approach.