Changes In Cell Morphology Research Articles

Ripasudil as a Potential Therapeutic Agent in Treating Secondary Glaucoma in HTLV-1-Uveitis: An In Vitro Analysis.

Human T-cell leukemia virus type 1 (HTLV-1), a virus that affects 5-10 million people globally, causes several diseases, including adult T-cell leukemia-lymphoma and HTLV-1-associated uveitis (HU). HU is prevalent in Japan and often leads to secondary glaucoma, which is a serious complication. We investigated the efficacy of ripasudil, a Rho-associated coiled coil-forming protein kinase inhibitor, in alleviating changes in human trabecular meshwork cells (hTM cells) infected with HTLV-1. HTLV-1-infected hTM cells were modeled in vitro using MT-2 cells, followed by treatment with varying concentrations of ripasudil. We assessed changes in cell morphology, viability, and inflammatory cytokine levels, as well as NF-κB activation. The results showed that ripasudil treatment changed the cell morphology, reduced the distribution of F-actin and fibronectin, and decreased the levels of certain inflammatory cytokines, such as interleukin (IL)-6, IL-8, and IL-12. However, ripasudil did not significantly affect NF-κB activation or overall cell viability. These findings suggest that ripasudil has the potential to treat secondary glaucoma in patients with HU by modulating cytoskeletal organization and alleviating inflammation in HTLV-1-infected hTM cells. This study lays the foundation for further clinical studies exploring the effectiveness of ripasudil for the treatment of secondary glaucoma associated with HU.

A high-throughput gut-on-chip platform to study the epithelial responses to enterotoxins

Enterotoxins are a type of toxins that primarily affect the intestines. Understanding their harmful effects is essential for food safety and medical research. Current methods lack high-throughput, robust, and translatable models capable of characterizing toxin-specific epithelial damage. Pressing concerns regarding enterotoxin contamination of foods and emerging interest in clinical applications of enterotoxins emphasize the need for new platforms. Here, we demonstrate how Caco-2 tubules can be used to study the effect of enterotoxins on the human intestinal epithelium, reflecting toxins’ distinct pathogenic mechanisms. After exposure of the model to toxins nigericin, ochratoxin A, patulin and melittin, we observed dose-dependent reductions in barrier permeability as measured by TEER, which were detected with higher sensitivity than previous studies using conventional models. Combination of LDH release assays and DRAQ7 staining allowed comprehensive evaluation of toxin cytotoxicity, which was only observed after exposure to melittin and ochratoxin A. Furthermore, the study of actin cytoskeleton allowed to assess toxin-induced changes in cell morphology, which were only caused by nigericin. Altogether, our study highlights the potential of our Caco-2 tubular model in becoming a multi-parametric and high-throughput tool to bridge the gap between current enterotoxin research and translatable in vivo models of the human intestinal epithelium.

Analysis of changes in the action potential morphology of the mouse sinoatrial node true pacemaker cells during ontogenetic development in vitro and in silico.

Maturation of the mouse is accompanied by the increase in heart rate. However, the mechanisms underlying this process remain unclear. We performed an action potentials (APs) recordings in mouse sinoatrial node (SAN) true pacemaker cells and in silico analysis to clarify the mechanisms underlying pre-postnatal period heart rate changes. The APs of true pacemaker cells at different stages had similar configurations and dV/dtmax values. The cycle length, action potential duration (APD90), maximal diastolic potential (MDP), and AP amplitude decreased, meanwhile the velocity of diastolic depolarization (DDR) increased from E12.5 stage to adult. Using a pharmacological approach we found that in SAN true pacemaker cells ivabradine reduces the DDR and the cycle length significantly stronger in E12.5 than in newborn and adult mice, whereas the effects of Ni2+ and nifedipine were significantly stronger in adult mice. Computer simulations further suggested that the density of the hyperpolarization-activated pacemaker сurrent (If) decreased during development, whereas transmembrane and intracellular Ca2+ flows increased. The ontogenetic decrease in IK1 density from E12.5 to adult leads to depolarization of MDP to the voltage range in which calcium currents are activated, thereby shifting the balance from the "membrane-clock" to the "calcium-clock."

Systematic cryopreservation study of cardiac myoblasts in suspension.

H9c2 myoblasts are a cell line derived from embryonic rat heart tissue and demonstrate the ability to differentiate to cardiac myotubes upon reduction of the serum concentration (from 10% to 1%) and addition of all-trans retinoic acid in the growth medium. H9c2 cells are increasingly being used as an easy-to-culture proxy for some functions of cardiomyocytes. The cryobiology of cardiac cells including H9c2 myoblasts has not been studied as extensively as that of some cell types. Consequently, it is important to characterize the cryobiological response and systematically develop well-optimized cryopreservation protocols for H9c2 cells to have optimal and consistent viability and functionality after thaw for high quality studies with this cell type. In this work, an interrupted slow cooling protocol (graded freezing) was applied to characterize H9c2 response throughout the cooling profile. Important factors that affect the cell response were examined, and final protocols that provided the highest post-thaw viability are reported. One protocol uses the common cryoprotectant dimethyl sulfoxide combined with hydroxyethyl starch, which will be suitable for applications in which the presence of dimethyl sulfoxide is not an issue; and the other protocol uses glycerol as a substitute when there is a desire to avoid dimethyl sulfoxide. Both protocols achieved comparable post-thaw viabilities (higher than 80%) based on SYTO 13/GelRed flow cytometry results. H9c2 cells cryopreserved by either protocol showed ability to differentiate to cardiac myotubes comparable to fresh (unfrozen) H9c2 cells, and their differentiation to cardiac myotubes was confirmed with i) change in cell morphology, ii) expression of cardiac marker troponin I, and iii) increase in mitochondrial mass.

Comparison of pulsed and continuous electromagnetic field generated by WPT system on human dermal and neural cells

In recent decades, we have seen significant technical progress in the modern world, leading to the widespread use of telecommunications systems, electrical appliances, and wireless technologies. These devices generate electromagnetic radiation (EMR) and electromagnetic fields (EMF) most often in the extremely low frequency or radio-frequency range. Therefore, they were included in the group of environmental risk factors that affect the human body and health on a daily basis. In this study, we tested the effect of exposure EMF generated by a new prototype wireless charging system on four human cell lines (normal cell lines—HDFa, NHA; tumor cell lines—SH-SY5Y, T98G). We tested different operating parameters of the wireless power transfer (WPT) device (87–207 kHz, 1.01–1.05 kW, 1.3–1.7 mT) at different exposure times (pulsed 6 × 10 min; continuous 1 × 60 min). We observed the effect of EMF on cell morphology and cytoskeletal changes, cell viability and mitotic activity, cytotoxicity, genotoxicity, and oxidative stress. The results of our study did not show any negative effect of the generated EMF on either normal cells or tumor cell lines. However, in order to be able to estimate the risk, further population and epidemiological studies are needed, which would reveal the clinical consequences of EMF impact.

Nicotine reduces cell viability and induces oxidative stress in human gingival fibroblasts

BackgroundNicotine, as the main component of cigarettes, is known to interfere with the proliferation of human gingival fibroblasts (HGFs) and can trigger oxidative stress. This study aimed to analyze the impact of nicotine on viability, expression of the antioxidant Nrf2, levels of the product of oxidative stress malondialdehyde (MDA), and the migration capacity of HGFs. MethodsAn experimental laboratory study used fibroblasts isolated from healthy human gingiva. The cells were grouped into the non-treatment control group (NTC), the solvent control (SC), and the treatment groups, exposed to nicotine at various concentrations for twenty-four hours. Cell viability was assesed using the cell counting kit-8 (CCK-8), Nrf2 expression was examined using ELISA, MDA level was measured using an MDA kit, and migration capacity was assessed using a scratch assay. Statistical analysis used one-way Anova or Kruskal-Wallis test. A p-value of <0.05 was expressed statistically significant. ResultsThe Cell viability was substantially reduced in the nicotine group compared to the untreated group, accompanied by changes in cell morphology. In contrast, Nrf2 expression increased significantly (p=0.010) in the 5 mM nicotine group compared with the control group. The MDA levels were not significantly distinct across groups (p=0.056). Cell migration was delayed significantly in the 5 mM nicotine group at 72 hours after scratching compared to the control group. ConclusionNicotine decreased HGFs viability and increased Nrf2 expression significantly in a dose-dependent manner. Nicotine at 5 mM concentration did not alter MDA levels but delayed cell migration.

Hortensins, Type 1 Ribosome-Inactivating Proteins from Seeds of Red Mountain Spinach: Isolation, Characterization, and Their Effect on Glioblastoma Cells.

Ribosome inactivating proteins (RIPs) are specific N-β-glycosylases that are well-characterized in plants. Their enzymatic action is to damage ribosomes, thereby blocking protein translation. Recently, several research groups have been working on the screening for these toxins in edible plants to facilitate the use of RIPs as biotechnological tools and biopesticides and to overcome public prejudice. Here, four novel monomeric (type 1) RIPs have been isolated from the seeds of Atriplex hortensis L. var. rubra, which is commonly known as edible red mountain spinach. These enzymes, named hortensins 1, 2, 4, and 5, are able to release the β-fragment and, like many other RIPs, adenines from salmon sperm DNA, thus, acting as polynucleotide:adenosine glycosidases. Structurally, hortensins have a different molecular weight and are purified with different yields (hortensin 1, ~29.5 kDa, 0.28 mg per 100 g; hortensin 2, ~29 kDa, 0.29 mg per 100 g; hortensin 4, ~28.5 kDa, 0.71 mg per 100 g; and hortensin 5, ~30 kDa, 0.65 mg per 100 g); only hortensins 2 and 4 are glycosylated. Furthermore, the major isoforms (hortensins 4 and 5) are cytotoxic toward human continuous glioblastoma U87MG cell line. In addition, the morphological change in U87MG cells in the presence of these toxins is indicative of cell death triggered by the apoptotic pathway, as revealed by nuclear DNA fragmentation (TUNEL assay).

In vivo confocal microscopy findings after Descemet stripping only with and without topical ripasudil supplementation in Fuchs endothelial dystrophy

PurposeTo report corneal epithelial and corneal endothelial cell (CEC) changes following Descemet stripping only (DSO) with and without topical ripasudil using in vivo confocal microscopy (IVCM). MethodsProspective interventional case series of patients who underwent DSO for Fuchs endothelial dystrophy with or without postoperative topical ripasudil (4%, 6 times per day). Patients underwent IVCM (ConfoScan 3; NIDEK Technologies, Padova, Italy) at baseline, monthly until corneal clearance, and then every 6 months. En face images were obtained of the epithelium, anterior and posterior corneal stroma, and endothelium of the central and superior cornea. The epithelial and endothelial layer images were evaluated qualitatively. ResultsIVCM imaging was obtained from 34 patients, 26 (76%) of whom were supplemented with ripasudil and 8 (24%) of whom were not. Two eyes (6%) did not clear and required a keratoplasty. IVCM confirmed epithelial and endothelial cell morphology changes and reestablishment of a CEC mosaic. Hyperreflective nuclei, pleomorphism, multinucleated cells, and pseudoguttae were identified within the descemetorhexis. In patients on ripasudil, epithelial cells demonstrated transition from stratified squamous to spindle cell shape in regions of ripasudil-induced honeycomb edema. Migrating cells adopted an amoeboid shape and cytoplasm elongation when crossing to central stroma. Transient epithelial cell morphology changes and endothelial cell amoeboid shape changes appear to be uniquely associated with ripasudil. Rarely, figures were noted in both ripasudil-treated and non-ripasudil-treated corneas that resembled mitotic bodies. ConclusionsCECs display a range of adaptive mechanisms during healing to re-form the endothelial mosaic following DSO. Ripasudil administration appears to induce unique epithelial and endothelial cell changes.

EXPLORING THE THERAPEUTIC POTENTIAL OF AM114: A BORONIC CHALCONE DERIVATIVE INDUCE APOPTOSIS AND SUPPRESS PROINFLAMMATORY CYTOKINES AND CHEMOKINES IN INTERLEUKIN-1β STIMULATED HUMAN THP-1 DERIVED MACROPHAGES

Objective: Chalcones and their derivatives display a wide range of pharmacological activities. This study examined the effects of AM114, a boronic-chalcone derivative, on human THP-1-derived macrophages with and without interleukin-1β (IL-1β) stimulation. Methods: AM114 and Aspirin-treated THP-1-derived macrophages underwent activation with or without interleukin-1β. The IC50 concentrations of AM114 and Aspirin were determined through an MTT test. Apoptosis was measured using various techniques, including staining with acridine orange/Ethidium bromide, Hoechst 33342, and rhodamine 123 assays. Caspase-3 activity was measured using the spectrofluorimetric technique, while DNA fragmentation was assessed via agarose gel electrophoresis. Pro-inflammatory cytokines such as interleukin-6 (IL-6) and chemokines like interleukin-8 (IL-8) were measured using enzyme-linked immunosorbent assays. Results: AM114 and Aspirin showed dose-dependent cytotoxic effects on THP-1 macrophages. Induction of apoptosis was detected in AM114-treated THP-1 macrophages activated with IL-1β compared to macrophages without IL-1β. The gradation of dye uptake, membrane blebbing, increased caspase-3 activity, and DNA fragmentation ensures the induction of apoptosis, which indicates the cell's morphological changes, biochemical processes, and mitochondrial activity. Treating AM114 in IL-1β-activated THP-1 macrophages significantly reduced pro-inflammatory cytokines (IL-6) and chemokines (IL-8), suggesting its anti-cytokine potential in inflammatory diseases. Conclusion: The study results emphasize that AM114 could act as an anti-inflammatory agent by triggering apoptosis and reducing the release of cytokines and chemokines in inflammatory conditions. As a result, it may be used as a therapeutic option for inflammatory diseases.

Sh- Ambra1 inhibits IRS-1/PI3K/Akt signalling pathway to reduce autophagy in gestational diabetes.

Gestational diabetes mellitus (GDM) is the most common metabolic disease in pregnancy. However, studies of activating molecule of Beclin1-regulated autophagy (Ambra1) affecting the insulin substrate receptor 1/phosphatidylinositol 3 kinase/protein kinase B (IRS-1/PI3K/Akt) signalling pathway in GDM have not been reported. The aim of the study was to detect the difference of Ambra1 expression in the placenta of normal pregnant women and GDM patients. An in vitro model of gestational diabetes mellitus was established by inducing HTR8/Svneo cells from human chorionic trophoblast layer with high glucose. The changes of cell morphology were observed by inverted microscope, and the expression levels of Ambra1 gene and protein in model cells were detected. After this, Ambra1 gene was silenced by shRNA transfection, and PI3K inhibitor was added to detect changes in Ambra1, autophagy, and insulin (INS) signalling pathways. The protein expression levels of Ambra1, Bcl-2 interacting protein (Beclin-1), and microtubule-associated proteins 1A/1B light chain 3B (LC3-II) in the placentas of GDM pregnant women were higher than those of normal pregnant women. High glucose induces morphological changes in HTR8/Svneo cells and increases Ambra1 transcription and translation levels. sh-Ambra1 increased survival of HTR8/SvNEO-HG cells and inhibited Ambra1, Beclin1, and LC3-II transcription and translation levels. Also, sh-Ambra1 increased IRS-1/PI3K/Akt protein phosphorylation levels and inhibited the IRS-1/PI3K/Akt signalling pathway and its resulting autophagy. sh-Ambra1 increased IRS-1/PI3K/Akt protein phosphorylation levels to reduce autophagy in gestational diabetes.

Selenium protects against Pb-induced renal oxidative injury in weaning rats and human renal tubular epithelial cells through activating NRF2

BackgroundLead (Pb) poisoning posing a crucial health risk, especially among children, causing devastating damage not only to brain development, but also to kidney function. Thus, an urgent need persists to identify highly effective, safe, and low-toxicity drugs for the treatment of Pb poisoning. The present study focused on exploring the protective effects of Se on Pb-induced nephrotoxicity in weaning rats and human renal tubular epithelial cells, and investigated the possible mechanisms. MethodsForty weaning rats were randomly divided into four groups in vivo: control, Pb-exposed, Pb+Se and Se. Serum creatinine (Cr), urea nitrogen (BUN) and hematoxylin and eosin (H&E) staining were performed to evaluate renal function. The activities of antioxidant enzymes in the kidney tissue were determined. In vitro experiments were performed using human renal tubular epithelial cells (HK-2 cells). The cytotoxicity of Pb and Se was detected by 3-(4,5-dimethylthiazol-2yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. Inverted fluorescence microscope was used to investigate cell morphological changes and the fluorescence intensity of reactive oxygen species (ROS). The oxidative stress parameters were measured by a multi-detection reader. Nuclear factor-erythroid-2-related factor (NRF2) signaling pathways were measured by Western blot and reverse transcription polymerase chain reaction (RT-PCR) in HK-2 cells. ResultsWe found that Se alleviated Pb-induced kidney injury by relieving oxidative stress and reducing the inflammatory index. Se significantly increased the activity of the antioxidant enzymes glutathione (GSH), superoxide dismutase (SOD) and catalase (CAT), whereas it decreased the excessive release of malondialdehyde (MDA) in the kidneys of weaning rats and HK-2 cells. Additionally, Se enhanced the antioxidant defense systems via activating the NRF2 transcription factor, thereby promoting the to downstream expression of heme oxygenase 1. Furthermore, genes encoding glutamate-cysteine ligase synthetase catalytic (GCLC), glutamate-cysteine ligase synthetase modifier (GCLM) and NADPH quinone oxidoreductase 1 (NQO1), downstream targets of NRF2, formed a positive feedback loop with NRF2 during oxidative stress responses. The MTT assay results revealed a significant decrease in cell viability with Se treatment, and the cytoprotective role of Se was blocked upon knockdown of NRF2 by small interfering RNA (siRNA). MDA activity results also showed that NRF2 knockdown inhibited the NRF2-dependent transcriptional activity of Se. ConclusionsOur findings demonstrate that Se ameliorated Pb-induced nephrotoxicity by reducing oxidative stress both in vivo and in vitro. The molecular mechanism underlying Se’s action in Pb-induced kidney injury is related to the activation of the NRF2 transcription factor and the activity of antioxidant enzymes, ultimately suppressing ROS accumulation.

Multiscale transport and 4D time-lapse imaging in precision-cut liver slices (PCLS).

Monitoring cellular processes across different levels of complexity, from the cellular to the tissue scale, is important for understanding tissue structure and function. However, it is challenging to monitor and estimate these structural and dynamic interactions within three-dimensional (3D) tissue models. The aim of this study was to design a method for imaging, tracking, and quantifying 3D changes in cell morphology (shape and size) within liver tissue, specifically a precision-cut liver slice (PCLS). A PCLS is a 3D model of the liver that allows the study of the structure and function of liver cells in their native microenvironment. Here, we present a method for imaging liver tissue during anisosmotic exposure in a multispectral four-dimensional manner. Three metrics of tissue morphology were measured to quantify the effects of osmotic stress on liver tissue. We estimated the changes in the volume of whole precision cut liver slices, quantified the changes in nuclei position, and calculated the changes in volumetric responses of tissue-embedded cells. During equilibration with cell-membrane-permeating and non-permeating solutes, the whole tissue experiences shrinkage and expansion. As nuclei showed a change in position and directional displacement under osmotic stress, we demonstrate that nuclei could be used as a probe to measure local osmotic and mechanical stress. Moreover, we demonstrate that cells change their volume within tissue slices as a result of osmotic perturbation and that this change in volume is dependent on the position of the cell within the tissue and the duration of the exposure. The results of this study have implications for a better understanding of multiscale transport, mechanobiology, and triggered biological responses within complex biological structures.

Eugenol's Molecular Warfare against Human Leukemia K562 cells: In Vitro Insights to Chemotherapeutic Potentials

Background: The search for effective anticancer agents has led to an increased focus on natural compounds, given their role as the foundation for many chemotherapeutic drugs. Eugenol (EUG) and Bis-Eugenol (Bis-EUG), derived from clove oil, have shown promise due to their antioxidative, anti-inflammatory, pro-apoptotic, and anti-proliferative properties across various cancer cell lines. This study aims to elucidate the chemotherapeutic potential of EUG and Bis-EUG on human leukemia K562 cells, exploring their ability to modulate apoptosis and cell proliferation. Objective: To assess the in vitro anticancer effects of EUG and Bis-EUG on K562 cell lines through various assays measuring cell viability, apoptosis, gene expression, and nitric oxide release, with a focus on elucidating the mechanisms underlying their anticancer activity. Methods: The study employed an inter-collaborative approach, utilizing MTT assays to determine the IC50 values of EUG and Bis-EUG on K562 cells. Gene expression analysis of pro-apoptotic genes Caspase-3 and Caspase-9 was performed using RT-qPCR. Apoptotic changes were further analyzed through flow cytometry and Hoechst 33258 staining to observe morphological alterations in treated cells. The release of nitric oxide (NO) as an indicator of anti-proliferative activity was measured using a Griess assay. Statistical analysis was conducted using GraphPad Prism Software, Version 9, and Microsoft Excel, with a significance threshold of p ≤ 0.05. Results: The IC50 values for EUG and Bis-EUG were determined to be 16.7 µM and 0.14 mM, respectively. Gene expression profiling revealed a significant increase in Caspase-3 and Caspase-9 expression, indicating the activation of apoptotic pathways. Flow cytometry and Hoechst 33258 staining confirmed the induction of apoptosis, with noticeable changes in cell morphology and nuclear fragmentation. The NO release assay demonstrated a dose-dependent increase in NO levels, highlighting the compounds' anti-proliferative effects. Conclusion: EUG and Bis-EUG exhibit potent anticancer effects on leukemia K562 cells by inducing apoptosis and inhibiting cell proliferation. The significant increase in NO levels and the activation of caspases suggest that these compounds may serve as effective chemotherapeutic agents. Further in vivo studies are warranted to explore their potential in cancer treatment regimens.

Phillygenin Inhibits TGF-β1-induced Hepatic Stellate Cell Activation and Inflammation: Regulation of the Bax/Bcl-2 and Wnt/β-catenin Pathways.

Hepatic fibrosis (HF), a precursor to cirrhosis and hepatocellular carcinoma, is caused by abnormal proliferation of connective tissue and excessive accumulation of extracellular matrix in the liver. Notably, activation of hepatic stellate cells (HSCs) is a key link in the development of HF. Phillygenin (PHI, C21H24O6) is a lignan component extracted from the traditional Chinese medicine Forsythiae Fructus, which has various pharmacological activities such as anti-inflammatory, antioxidant and anti-tumour effects. However, whether PHI can directly inhibit HSC activation and ameliorate the mechanism of action of HF has not been fully elucidated. Therefore, the aim of the present study was to investigate the in vitro anti-HF effects of PHI and the underlying molecular mechanisms. Transforming growth factor-β1 (TGF-β1)-activated mouse HSCs (mHSCs) and human HSCs (LX-2 cells) were used as an in vitro model of HF and treated with different concentrations of PHI for 24h. Subsequently, cell morphological changes were observed under the microscope, cell viability was analyzed by MTT assay, cell cycle and apoptosis were detected by flow cytometry, and the mechanism of anti-fibrotic effect of PHI was explored by immunofluorescence, ELISA, RT-qPCR and western blot. The results showed that PHI suppressed the proliferation of TGF-β1-activated mHSCs and LX-2 cells, arrested the cell cycle at the G0/G1 phase, decreased the levels of α-SMA, Collagen I, TIMP1 and MMP2 genes and proteins, and promoted apoptosis in activated mHSCs and LX-2 cells. Besides, PHI reduced the expression of inflammatory factors in activated mHSCs and LX-2 cells, suggesting a potential anti-inflammatory effect. Mechanically, PHI inhibited TGF-β1-induced HSC activation and inflammation, at least in part through modulation of the Bax/Bcl-2 and Wnt/β-catenin pathways. Overall, PHI has significant anti-HF effects and may be a promising agent for the treatment of HF.

Involvement of GSTP1 in low dose radiation-induced apoptosis in GM12878 cells

BackgroundLow-dose ionizing radiation-induced protection and damage are of great significance among radiation workers. We aimed to study the role of glutathione S-transferase Pi (GSTP1) in low-dose ionizing radiation damage and clarify the impact of ionizing radiation on the biological activities of cells. ResultsIn this study, we collected peripheral blood samples from healthy adults and workers engaged in radiation and radiotherapy and detected the expression of GSTP1 by qPCR. We utilized γ-rays emitted from uranium tailings as a radiation source, with a dose rate of 14 μGy/h. GM12878 cells subjected to this radiation for 7, 14, 21, and 28 days received total doses of 2.4, 4.7, 7.1, and 9.4 mGy, respectively. Subsequent analyses, including flow cytometry, MTS, and other assays, were performed to assess the ionizing radiation’s effects on cellular biological functions. In peripheral blood samples collected from healthy adults and radiologic technologist working in a hospital, we observed a decreased expression of GSTP1 mRNA in radiation personnel compared to the healthy controls. In cultured GM12878 cells exposed to low-dose ionizing radiation from uranium tailings, we noted significant changes in cell morphology, suppression of proliferation, delay in cell cycle progression, and increased apoptosis. These effects were partially reversed by overexpression of GSTP1. Moreover, low-dose ionizing radiation increased GSTP1 gene methylation and downregulated GSTP1 expression. Furthermore, low-dose ionizing radiation affected the expression of GSTP1-related signaling molecules. ConclusionsThis study shows that low-dose ionizing radiation damages GM12878 cells and affects their proliferation, cell cycle progression, and apoptosis. In addition, GSTP1 plays a modulating role under low-dose ionizing radiation damage conditions. Low-dose ionizing radiation affects the expression of Nrf2, JNK, and other signaling molecules through GSTP1.

The pathogenicity of vancomycin-resistant Enterococcus faecalis to colon cancer cells

BackgroundThe aim of this study was to investigate the pathogenicity of vancomycin-resistant Enterococcus faecalis (VREs) to human colon cells in vitro.MethodsThree E. faecalis isolates (2 VREs and E. faecalis ATCC 29212) were cocultured with NCM460, HT-29 and HCT116 cells. Changes in cell morphology and bacterial adhesion were assessed at different time points. Interleukin-8 (IL-8) and vascular endothelial growth factor A (VEGFA) expression were measured via RT-qPCR and enzyme-linked immunosorbent assay (ELISA), respectively. Cell migration and human umbilical vein endothelial cells (HUVECs) tube formation assays were used for angiogenesis studies. The activity of PI3K/AKT/mTOR signaling pathway was measured by Western blotting.ResultsThe growth and adhesion of E. faecalis at a multiplicity of infection (MOI) of 1:1 were greater than those at a MOI of 100:1(p < 0.05). Compared to E. faecalis ATCC 29212, VREs showed less invasive effect on NCM460 and HT-29 cells. E. faecalis promoted angiogenesis by secreting IL-8 and VEGFA in colon cells, and the cells infected with VREs produced more than those infected with the standard strain (p < 0.05). Additionally, the PI3K/AKT/mTOR signaling pathway was activated in E. faecalis infected cells, with VREs demonstrating a greater activation compared to E. faecalis ATCC 29212 (p < 0.05).ConclusionVREs contribute to the occurrence and development of CRC by promoting angiogenesis and activating the PI3K/AKT/mTOR signaling pathway.

Ionic and nanoparticulate silver alleviate the toxicity of inorganic mercury in marine microalga Chaetoceros muelleri.

Toxicological effects of silver nanoparticles (SNPs) in different organisms have been studied; however, interactions of SNPs with other environmental pollutants such as mercury are poorly understood. Herein, bioassay tests were performed according to ΟECD 201 guideline to assess the toxic effects induced by mercury ions (mercury chloride, MCl) on the marine microalga Chaetoceros muelleri in the presence of SNPs or silver ions (silver nitrate, SN). Acute toxicity tests displayed that the presence of SNPs or SN (0.01mg L-1) significantly reduced the toxicity of MCl (0.001, 0.01, 0.1, 1, 10, and 100mg L-1) and increased the IC50 of MCl from 0.072 ± 0.014 to 0.381 ± 0.029 and 0.676 ± 0.034mg L-1, respectively. In the presence of SN or SNPs, the mercury-reducing effect on algal population growth significantly decreased. Considering the increase of IC50, the mercury toxicity decreased approximately 5.44 and 9.66 times in the presence of SNPs or SN, respectively. The chlorophyll a and c contents decreased at all exposures; however, the decrease by MCl-SNPs and MCl-SN was significantly less than MCl except at 1mg L-1. The lowering effect of MCl-SN on chlorophyll contents was less than MCl and MCl-SNPs. MCl exposure induced significant raises in total protein content (TPC) at concentrations < 0.01mg L-1, with a maximum of ~ 70.83% attained at 100mg L-1. The effects of MCl-SNPs and MCl-SN on TPC were significantly less than MCl. Total lipid content (TLC) at all MCl concentrations was higher than the control, while at coexposure to MCl-SN, TLC did not change until 0.01mg L-1 compared with the control. The effects of MCl-SN and MCL-SNPs on TPC and TLC were in line with toxicity results, and were significantly less than those of MCl individually, confirming their antagonistic effects on MCl. The morphological changes of algal cells and mercury content of the cell wall at MCl-SN and MCl-SNPs were mitigated compared with MCl exposure. These findings highlight the mitigatory impacts of silver species on mercury toxicity, emphasizing the need for better realizing the mixture toxicity effects of pollutants in the water ecosystem.

Morphological Changes of 3T3 Cells under Simulated Microgravity.

Cells are sensitive to changes in gravity, especially the cytoskeletal structures that determine cell morphology. The aim of this study was to assess the effects of simulated microgravity (SMG) on 3T3 cell morphology, as demonstrated by a characterization of the morphology of cells and nuclei, alterations of microfilaments and microtubules, and changes in cycle progression. 3T3 cells underwent induced SMG for 72 h with Gravite®, while the control group was under 1G. Fluorescent staining was applied to estimate the morphology of cells and nuclei and the cytoskeleton distribution of 3T3 cells. Cell cycle progression was assessed by using the cell cycle app of the Cytell microscope, and Western blot was conducted to determine the expression of the major structural proteins and main cell cycle regulators. The results show that SMG led to decreased nuclear intensity, nuclear area, and nuclear shape and increased cell diameter in 3T3 cells. The 3T3 cells in the SMG group appeared to have a flat form and diminished microvillus formation, while cells in the control group displayed an apical shape and abundant microvilli. The 3T3 cells under SMG exhibited microtubule distribution surrounding the nucleus, compared to the perinuclear accumulation in control cells. Irregular forms of the contractile ring and polar spindle were observed in 3T3 cells under SMG. The changes in cytoskeleton structure were caused by alterations in the expression of major cytoskeletal proteins, including β-actin and α-tubulin 3. Moreover, SMG induced 3T3 cells into the arrest phase by reducing main cell cycle related genes, which also affected the formation of cytoskeleton structures such as microfilaments and microtubules. These results reveal that SMG generated morphological changes in 3T3 cells by remodeling the cytoskeleton structure and downregulating major structural proteins and cell cycle regulators.

Synthesis of Lawsonia inermis-encased silver–copper bimetallic nanoparticles with antioxidant, antibacterial, and cytotoxic activity

Abstract The extract of the medicinal plant Lawsonia inermis, known as henna, was employed to synthesize silver–copper bimetallic nanoparticles (Ag–Cu NPs) in a unique, efficient, and cost-effective method. The shape, size, and structural features of synthesized Ag–Cu NPs were determined by ultra–visible spectroscopy, Fourier transform infrared spectroscopy, X-ray diffractometer, field emission scanning electron microscopy, transmission electron microscopy, and energy dispersive X-ray spectroscopy methods. The rod-shaped Ag–Cu nanoparticles, averaging 41.66 ± 17.18 nm in size, synthesized from L. inermis, exhibited potent antioxidant activity by inhibiting 2,2-diphenyl-1-picrylhydrazyl and 2,2′-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) free radicals. They also displayed significant antibacterial effects against Pseudomonas aeruginosa (28 mm), Staphylococcus aureus (27 mm), Bacillus cereus (26 mm), and Escherichia coli (24 mm). Additionally, these nanoparticles induced notable morphological changes in cancer cells and demonstrated promising cytotoxicity against MDA-MB-231 tumor cells (IC50 = 37.40 µg·mL−1). However, they exhibited biotoxicity in Artemia nauplii, resulting in mortality rates ranging from 3.0% to 32.5%. The LC50 and LC90 values recorded for a 48-h exposure were 1.51 mg·L−1 and 2.59 mg·L−1, respectively. These findings highlight the potential application of L. inermis-derived Ag–Cu NPs in pharmacology and bio-nanomedicine.

Krüppel-like factor 4 modulates the miR-101/COL10A1 axis to inhibit renal fibrosis after AKI by regulating epithelial–mesenchymal transition

Acute kidney injury (AKI) can progress to renal fibrosis and chronic kidney disease (CKD), which reduces quality of life and increases the economic burden on patients. However, the molecular mechanisms underlying renal fibrosis following AKI remain unclear. This study tested the hypothesis that the Krüppel-like factor 4 (KLF4)/miR-101/Collagen alpha-1X (COL10A1) axis could inhibit epithelial-mesenchymal transition (EMT) and renal fibrosis after AKI in a mouse model of ischemia-reperfusion (I/R)-induced renal fibrosis and HK-2 cells by gene silencing, overexpression, immunofluorescence, immunohistochemistry, real-time quantitative PCR, Western blotting, dual-luciferase reporter assay, fluorescence in situ hybridization (FISH) and ELISA. Compared with the Sham group, I/R induced renal tubular and glomerular injury and fibrosis, and increased the levels of BUN, serum Scr and neutrophil gelatinase-associated lipocalin (NGAL), Col10a1 and Vimentin expression, but decreased E-cadherin expression in the kidney tissues of mice at 42 days post-I/R. Similarly, hypoxia promoted fibroblastic morphological changes in HK-2 cells and enhanced NGAL, COL10A1, Vimentin, and α-SMA expression, but reduced E-cadherin expression in HK-2 cells. These pathological changes were significantly mitigated in COL10A1-silenced renal tissues and HK-2 cells. KLF4 induces miR-101 transcription. More importantly, hypoxia upregulated Vimentin and COL10A1 expression, but decreased miR-101, KLF4, and E-cadherin expression in HK-2 cells. These hypoxic effects were significantly mitigated or abrogated by KLF4 over-expression in the HK-2 cells. Our data indicate that KLF4 up-regulates miR-101 expression, leading to the downregulation of COL10A1 expression, inhibition of EMT and renal fibrosis during the pathogenic process of I/R-related renal fibrosis.