Lactate Dehydrogenase Leakage Assay Research Articles

Cytoprotective and Anti-inflammatory Effects of Derma Genie™-H002 against Paraphenylenediamine-induced Cytotoxicity and Inflammation in RAW 264.7 Cells

Purpose: This study aimed to establish an <i>in vitro</i> model to examine paraphenylenediamine (PPD)-induced cytotoxicity and inflammation within RAW 264.7 cells. Additionally, we investigated the cytoprotective and anti-inflammatory effects of Derma Genie™-H002 derived from extracts of five medicinal herbs. Methods: Various biochemical analyses, including water-soluble tetrazolium salt, crystal violet staining, and lactate dehydrogenase leakage assays, were employed to evaluate the impact of Derma Genie™-H002 on PPD-induced cytotoxicity and inflammation. Furthermore, apoptosis- and inflammation-related gene expression was analyzed using reverse transcription-polymerase chain reaction and Western blotting. Results: Derma Genie™-H002 exhibited no significant cytotoxicity in RAW 264.7 cells, whereas PPD induced notable cytotoxicity at concentrations >25 μM. Cotreatment with PPD and Derma Genie™-H002 not only improved cell proliferation rates but also alleviated cell viability in PPD-induced RAW 264.7 cells. Furthermore, findings demonstrated that Derma Genie™-H002 mitigates the cytotoxicity of PPD-induced RAW 264.7 cells by reducing the expression of the proapoptotic protein Bax and increasing the expression of the antiapoptotic protein Bcl-2. Moreover, cotreatment with PPD and Derma Genie™-H002 downregulated the mRNA expression of inflammasome-related genes, inflammatory cytokines, and allergic contact dermatitis-related genes, all elevated by PPD. The increased expression of nuclear factor-κB protein induced by PPD was also reduced upon cotreatment with Derma Genie™-H002, providing compelling evidence of its anti-inflammatory efficacy in the PPD-induced inflammation model. Conclusion: This study demonstrated that Derma Genie™-H002 effectively alleviates PPD-induced cytotoxicity and inflammation in RAW 264.7 cells. Therefore, the potential application of Derma Genie™-H002 as a naturally derived functional anti-inflammatory material has been validated.

Tributyltin chloride induces chondrocyte damage through the activation of NLRP3‑mediated inflammation and pyroptosis.

Tributyltin chloride (TBTC) is known to have effects and mechanisms in various diseases; however, whether TBTC is detrimental to joints and causes osteoarthritis (OA), as well as its underlying mechanism, has not yet been fully elucidated. The present study explored the effects of TBTC on rat chondrocytes, as well as on mouse OA. The toxicity of TBTC toward rat chondrocytes was detected using a lactate dehydrogenase (LDH) leakage assay and cell viability was evaluated using the Cell Counting Kit‑8 assay. The results showed that TBTC decreased the viability of rat chondrocytes and increased the LDH leakage rate in a concentration‑dependent manner. Moreover, compared with in the control group, TBTC increased the expression levels of interleukin (IL)‑1β, IL‑18, matrix metalloproteinase (MMP)‑1, MMP‑13, NLR family pyrin domain containing 3 (NLRP3), caspase‑1, PYD and CARD domain containing, and gasdermin D in chondrocytes. Furthermore, knockdown of NLRP3 reversed the TBTC‑induced increases in LDH leakage and NLRP3 inflammasome‑associated protein levels. In vivo, TBTC exacerbated cartilage tissue damage in mice from the OA group, as evidenced by the attenuation of safranin O staining. In conclusion, TBTC may aggravate OA in mice by promoting chondrocyte damage and inducing pyroptosis through the activation of NLRP3 and caspase‑1 signaling. The present study demonstrated that TBTC can cause significant damage to the articular cartilage; therefore, TBTC contamination should be strictly monitored.

Effect of Silica in the Manufacture of Autologous Serum Eye Drops on Corneal Stromal Cells.

The aim of this study was to evaluate the effect of serum clot activator, silica (SiO 2 ), which may be used for making autologous serum eye drops, on human corneal fibroblasts. Cultured human corneal fibroblasts were exposed to 10%, 20%, and 30% silica for 1, 6, and 24 hours; methyl thiazolyl tetrazolium-based colorimetric assay was performed to determine the survival rate of fibroblasts and lactate dehydrogenase leakage assay to assess the cytotoxicity. The apoptotic response was evaluated by flow cytometric analysis and fluorescence staining with Annexin V and propidium iodide. Cellular morphology was evaluated by inverted phase-contrast light microscopy and electron microscopy. The survival rate of human corneal fibroblasts and cytotoxicity showed both dose-dependent and time-dependent responses. The fluorescent micrograph and flow cytometry showed that as the exposure time increased, more cells underwent apoptosis or necrosis after treatment with 30% silica. When observed with light and electron microscopy, the number of corneal fibroblasts decreased and they were more detached from the dish. In addition, damaged corneal fibroblasts showed degenerative changes after exposure to 30% silica. Silica showed dose-dependent and time-dependent toxicity in human corneal fibroblasts. It is safer to keep the blood in tubes without a clot activator when manufacturing autologous serum eye drops to prevent possible corneal cytotoxicity.

Biocompatibility Evaluation of TiO2, Fe3O4, and TiO2/Fe3O4 Nanomaterials: Insights into Potential Toxic Effects in Erythrocytes and HepG2 Cells.

Nanomaterials such as titanium dioxide and magnetite are increasingly used in several fields, such as water remediation and agriculture. However, this has raised environmental concerns due to potential exposure to organisms like humans. Nanomaterials can cause adverse interactions depending on physicochemical characteristics, like size, morphology, and composition, when interacting with living beings. To ensure safe use and prevent the risk of exposure to nanomaterials, their biocompatibility must be assessed. In vitro cell cultures are beneficial for assessing nanomaterial-cell interactions due to their easy handling. The present study evaluated the biocompatibility of TiO2, Fe3O4, and TiO2/Fe3O4 nanomaterials thermally treated at 350 °C and 450 °C in erythrocytes and HepG2 cells. According to the hemolysis experiments, non-thermally treated NMs are toxic (>5% hemolysis), but their thermally treated counterparts do not present toxicity (<2%). This behavior indicates that the toxicity derives from some precursor (solvent or surfactant) used in the synthesis of the nanomaterials. All the thermally treated nanomaterials did not show hemolytic activity under different conditions, such as low-light exposure or the absence of blood plasma proteins. In contrast, non-thermally treated nanomaterials showed a high hemolytic behavior, which was reduced after the purification (washing and thermal treatment) of nanomaterials, indicating the presence of surfactant residue used during synthesis. An MTS cell viability assay shows that calcined nanomaterials do not reduce cell viability (>11%) during 24 h of exposure. On the other hand, a lactate dehydrogenase leakage assay resulted in a higher variability, indicating that several nanomaterials did not cause an increase in cell death as compared to the control. However, a holotomographic microscopy analysis reveals a high accumulation of nanomaterials in the cell structure at a low concentration (10 µg mL-1), altering cell morphology, which could lead to cell membrane damage and cell viability reduction.

IN VITRO ANTIOXIDANT AND ANTICANCER ACTIVITY OF THE FRUIT EXTRACT OF RIVINA HUMILIS L. (RED PIGEON BERRY)

The purpose of this study was to evaluate the cytotoxic and antioxidant properties of a hydromethanolic extract of Rivina humilis L fruits extract (HMERH) on human cancer cell lines and various in vitro models. HMERH concentrations ranging from 5 to 400mg/ml were utilized in several assay methods such as hydroxyl radicals, DPPH, superoxide radical scavenging activity, metal chelating, and nitric oxide radical test, ABTS, and reducing power assay. All antioxidant assays employed ascorbic acid as the standard. Its short-term cytotoxicity on EAC and DLA cells was determined using the trypan blue dye exclusion method, as well as the lactate dehydrogenase (LDH) leakage assay and in vitro cytotoxicity on MEF-L929, DU-145, and PC-3 cells using the MTT assay. The total antioxidant potential was also determined using the phosphomolybdenum test. HMERH demonstrated considerable cytotoxicity against all cancer cell lines examined. The findings of the free radical scavenging activities demonstrated a concentration-dependent and antiradical activity caused by the reduction of ABTS, DPPH, NO, OH, and SO radicals to non-radical form. The findings reveal that antioxidants and cytotoxic agents have potent antioxidant and cytotoxic effects on cancer cell lines. Hence, this fruit extract has the potential to be a natural antioxidant and anticancer drug source, which could be valuable in the development of new anticancer therapies.

Exploration of the ameliorative effect of dietary polyphenol on Bisphenol-A prompted DNA damage by in vitro and in silico approaches

Bisphenol A (BPA), an infamous xenoestrogen, can cause the inimical environmental and health implications via generation of extremely reactive oxygen species (ROS) and/or by mimicking the endocrine activity. The intention of this study was to understand the concentration range of BPA between 0.02-200 µg mL−1 that may induce DNA damage and cytotoxicity, individually and/or through amelioration by gallic acid (GA) (10–50 µg mL−1) in HepG2 cell line by Comet, Lactate Dehydrogenase (LDH) leakage and MTT assay. The results revealed that BPA was not found genotoxic (0.02–0.2 µg mL−1) and cytotoxic at low dose exposure (0.02 µg mL−1). However, BPA at a concentration of 2-200 µg mL−1, induced significant DNA damage and cytotoxicity (p<0.05) in a dose-relying manner in HepG2 cell line. However, DNA damage and cytotoxicity were inhibited when GA (10–50 g mL-1) and BPA (200 g mL-1) were added together. To investigate the mechanism of toxicity and the binding potential of BPA/BPA-Q to DNA and GA, an in silico molecular docking approach and molecular dynamics simulation were performed. The obtained results revealed strong binding affinity between BPA/BPA-Q with DNA which induces DNA damage.

Peimine Inhibits MCF-7 Breast Cancer Cell Growth by Modulating Inflammasome Activation: Critical Roles of MAPK and NF-κB Signaling.

Peimine (PM) is a bioactive compound obtained from Fritillaria. It has been documented that PM exhibits potent antitumor properties against multiple cancers. However, the antitumor properties of PM in breast cancer and its associated mechanisms have not been clarified. Proliferation and apoptosis of MCF-7 and MCF-10A cells were detected by CCK8, colony formation, and flow cytometry assays. Cytotoxicity was measured by Lactate dehydrogenase (LDH) leakage assay. The level of IL-1β and IL-18 were detected with ELISA kits. Western blotting and real-time Polymerase Chain Reaction were performed to analyze the expression of proteins and genes related to the NLRP3 inflammasome pathway and Endoplasmic reticulum stress. The doses of PM (5, 10, and 20 μM) inhibited cell viability significantly, apoptotic induction, and inflammasome activation in breast cancer cells in vitro. Inflammasome components were decreased, including the apoptosisassociated speck like protein containing a CARD (ASC) and NOD-like receptor pyrindomain-containing protein3 (NLRP3), as well as the inhibition of caspase-1 and interleukin-1β activation. Moreover, inflammasome inhibitors suppressed cell growth and induced apoptosis, implying that PM suppresses the growth of breast cancer cells through regulating inflammasome. Mechanistically, PM inhibited the activity of inflammasome by alleviating endoplasmic reticulum (ER) stress and by down-regulating the expression of multiple proteins in transcription factor nuclear factor- κB (NF-κB) and mitogen-activated protein kinases (MAPKs) signaling pathways. These findings show that PM suppresses the growth of breast cancer cells by inhibiting inflammasome activation, to a certain extent, by primarily acting on the MAPK/NF-κB pathway's inactivation-dependent mechanisms.

Neuroprotective effect of a novel brain-derived peptide, HIBDAP, against oxygen-glucose deprivation through inhibition of apoptosis in PC12 cells.

The effect of a novel brain-derived peptide,hypoxic-ischemic brain damage associated peptide (HIBDAP), on apoptosis after oxygen-glucose deprivation (OGD) in PC12 cells was investigated. The HIBDAP sequence (HSQFIGYPITLFVEKER) was coupled with the carrier peptide of the transactivator of transcription (TAT) sequence (YGRKKRRQRRR). FITC-labelled TAT-HIBDAP was observed by fluorescence microscopy. After TAT-HIBDAP treatment and OGD treatment, the PC12 cell apoptosis rate was analysed using lactate dehydrogenase (LDH) leakage and Annexin V-fluorescein isothiocyanate (FITC) assays. Mitochondrial membrane potential (ΔΨm) was examined by fluorescence microscopy. Protein expression of apoptotic factors was examined by Western blotting. FITC-labelled TAT-HIBDAP entered the PC12 cell nucleus. Compared with the OGD group, TAT-HIBDAP at low concentrations (1 µM, 5 µM, 10 µM) significantly reduced the apoptosis rate of PC12 cells (except at 20 µM); 5 µM TAT-HIBDAP had the most obvious effect. There were remarkable increases in ΔΨm at different concentrations (1 µM, 5 µM, 10 µM, 20 µM) of TAT-HIBDAP pretreatment, and 5 µM TAT-HIBDAP also had the most obvious effect. TAT-HIBDAP reversed the increased ratio of Bax/Bcl-2 and activation of Caspase-3 induced by OGD. TAT-HIBDAP is resistant to OGD-induced PC12 cell apoptosis by regulating the Bax/Bcl-2/Caspase-3 pathway, which may provide a novel therapeutic strategy for neonatal HIBD.

Enclathration of Mn(II)(H2O)6 guests and unusual Cu⋯O bonding contacts in supramolecular assemblies of Mn(II) Co-crystal hydrate and Cu(II) Pyridinedicarboxylate: Antiproliferative evaluation and theoretical studies

Two new coordination compounds of Mn(II) and Cu(II) involving 2,6-pyridinedicarboxylate viz. [Mn3(2,6-PDC)4(H2O)4][Mn(H2O)6]·8H2O (1) and [Cu(2,6-PDC)(H2O)(4-CNpy)]·H2O (2) (2,6-PDC = 2,6-pyridinedicarboxylate, 4-CNpy = 4-cyanopyridine) have been synthesized at room temperature. Compound 1 crystallizes as a co-crystal hydrate of Mn(II); whereas compound 2 is a mononuclear compound of Cu(II). Crystal structure analysis of 1 unfolds the presence of Mn(II)(H2O)6 guests within the supramolecular host cavities formed by anionic complex moieties and lattice water molecules. The lattice water molecules of the compounds 1 and 2 are involved in the formation of (H2O)2 cores which are enclathrated in the supramolecular host cavities. Furthermore, the unconventional anion⋯π, antiparallel CN⋯CN, CO⋯CO, CR⋯CR (CR: chelate ring) and Cu⋯O bonding interactions observed in compound 2 play crucial role to the stability of the crystal structure. Theoretical calculations confirm that the H-bonding network in 1 and anion-π, unconventional Cu···O bonds, antiparallel CO···CO along with CR···CR interactions in 2 play crucial roles in the solid state stability of the compounds. In vitro anticancer activities of the compounds considering cell viability and apoptosis assays against Dalton's lymphoma (DL) cancer cell lines reveal that the compounds induce considerable cytotoxicity in DL cells with negligible cytotoxicity in normal PBMC cells. Lactate dehydrogenase (LDH) leakage assay also unfolds the release of LDH from DL cells to extracellular space suggesting the irreversible cell death with cell membrane damage. The molecular docking simulations of the compounds further demonstrate the structure activity relationships involving the active sites of some important antiapoptotic proteins.

The effect of steviol on differentiated rat PC-12 cells induced by MPP+

Background: The 1-methyl-4-phenylpyridinium ion (MPP+) is a neurotoxin. It inhibits mitochondrial complex I by interfering with oxidative phosphorylation in mitochondria. That causes ATP depletion and eventual cell apoptosis. Objective: This study investigates steviol's effect, a major metabolite of stevia obtained from Stevia rebaudiana, on MPP+-induced apoptosis of differentiated rat adrenal pheochromocytoma (PC-12) cells. Methods: The PC12 cells were differentiated into neuronal cells by adding nerve growth factor and then treated with MPP+ and steviol in different concentrations; twenty-four hours later, two in vitro toxicity assays. 3-(4,5-Dimethylthiazol-2-Yl)-2,5-Diphenyltetrazolium Bromide (MTT) and Lactate Dehydrogenase Leakage assays (LDH) were conducted to detect the cells’ viability. Results: The present study demonstrated that steviol caused statistically significant cell damage and inhibited the proliferation of the differentiated rat PC-12 cell line in a dose-dependent manner. Furthermore, the results from MTT and LDH assays complemented the outcome of our experiments that steviol acted synergistically in the presence of MPP+ to increase cell death in differentiated rat PC-12 cells. Conclusion: The data obtained from MTT and LDH assays indicated that steviol acted synergistically in the presence of MPP+ free radicals, increasing cytotoxicity. Both assays revealed that steviol aggravated the cytotoxicity produced by MPP+ in differentiated rat PC-12 cells. However, as the MTT assay does not correspond to mitochondrial dysfunction, it may give a poor estimate of cytotoxicity of steviol as compared to the LDH assay that quantifies the damage to the cell membrane, especially when there are small-scale variations in cellular reduction capacity.

Trimethylamine oxide induces pyroptosis of vascular endothelial cells through ALDH2/ROS/NLRP3/GSDMD pathway.

Trimethylamine oxide (TMAO) is a metabolite of intestinal flora and is known to promote the progression of atherosclerotic plaques. However, how TMAO works, including its effect on vascular endothelial cells, is not fully understood. This study aims to explore the biological role of TMAO in human umbilical vein endothelial cells (HUVECs) and the underlying mechanism. Cell pyroptosis and the loss of plasma membrane integrity were induced under TMAO stimulation in HUVECs. The plasma membrane integrity of the cells was measured by Hoechst 33342/propidium iodide (PI) staining and lactate dehydrogenase leakage assay, and the changes in cell morphology were observed by atomic force microscope. The expression of proteins related to pyroptosis was determined by Western blotting or immunofluorescence. Mitochondrial acetaldehyde dehydrogenase 2 (ALDH2) activity in HUVECs was measured by the ALDH2 activity assay kit, and the level of reactive oxygen species (ROS) was detected by fluorescent probe DCFH-DA. TMAO induced pyroptotic cell death, manifesting by the presence of propidium iodide-positive cells, the leakage of lactate dehydrogenase, the production of N-terminal gasdermin D (GSDMD-N), and the formation of plasma membrane pores. Moreover, TMAO induced elevated expression of inflammasome components, nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3 (NLRP3), apoptosis-associated speck-like protein containing a caspase activation and recruitment domain (ASC), and caspase-1 in cells. TMAO significantly inhibited ALDH2 activity and increased intracellular ROS production. However, the activation of ALDH2 by pharmacological manipulation attenuated TMAO-induced inflammasome activation and GSDMD-N production. TMAO induces pyroptosis of vascular endothelial cells through the ALDH2/ROS/NLRP3/GSDMD signaling pathway, which may be a potential therapeutic target for improving the treatment of atherosclerosis.

Amelioration of dextran sulfate sodium-induced colitis by autoinducer-2-deficient Lactiplantibacillus plantarum is mediated by anti-inflammatory effects and alleviation of dysbiosis of the gut microbiota.

Lactic acid bacteria (LAB) attenuate dextran sulfate sodium (DSS)-induced colitis in mice by restoring gut flora homeostasis and modulating the immune response. Because synchronous behavior can be controlled by autoinducer-2 (AI-2)/LuxS-mediated quorum sensing, the Caco-2 cell model and DSS-induced model in C57BL/6 mice were used to explore the unknown effects of these communications involving AI-2 among various intestinal symbiotic species. The results of the cell viability and lactate dehydrogenase leakage assays indicated that the tested strains (the wild-type strains and AI-2-deficient mutants) were characterized by equal cytoprotection from hydrogen peroxide-induced injury independently of AI-2. The results of the assays of multiple indicators and proinflammatory cytokines characteristic for the symptoms of colitis in mice showed that oral administration of AI-2-deficient mutants for 7 days was more effective in ameliorating inflammation than the treatment with the wild-type strains. The treatment with AI-2-deficient mutants enriched potential probiotics (e.g., Lactobacillaceae) and controlled the proliferation of potentially harmful bacteria (e.g., Helicobacteraceae) to achieve the transformation of intestinal flora. These mutants regulated short-chain fatty acids and the intestinal epithelial barrier, thereby promoting the maintenance of relatively favorable intestinal homeostasis. These results demonstrated that the AI-2-deficient mutants provided a more pronounced ameliorative effect on colitis in a mouse model, suggesting that the background of the LAB effect is associated with the alterations in colonic flora induced by AI-2.

Fatty Acid Modification of the Anticancer Peptide LVTX-9 to Enhance Its Cytotoxicity against Malignant Melanoma Cells.

Spider venom is a valuable resource for the development of novel anticancer drugs. In this study, we focused on novel linear amphipathic α-helical anticancer peptide LVTX-9, which was derived from the cDNA library of the venom gland of the spider Lycosa vittata. The cytotoxicity of LVTX-9 against murine melanoma cells in the range of 1.56–200 μM was tested and found to be significantly lower than those of most anticancer peptides reported. Its IC50 was determined to be 59.2 ± 19.8 μM in a serum or 76.3 ± 12.7 μM in serum-free medium. Fatty acid modification is a promising strategy for improving peptide performance. Therefore, to enhance the cytotoxic activity of LVTX-9, fatty acid modification of this peptide was performed, and five different carbon chain length lipopeptides named LVTX-9-C12-C20 were produced. Among them, the lipopeptide LVTX-9-C18 showed the highest cytotoxic activity in relation to B16-F10 cells, whether in a serum or serum-free medium. Most importantly, the cytotoxic activity of LVTX-9-C18 was improved by about 12.9 times in a serum medium or 19.3 times in a serum-free medium compared to that of LVTX-9. Subsequently, assays including scanning electron microscopy, trypan blue staining, lactate dehydrogenase leakage assay, and hemolytic activity could indicate that the potential direct cell membrane disruption is the main mechanism of LVTX-9-C18 to induce cancer cell death. Furthermore, the LVTX-9-C18 also showed strong cytotoxicity in relation to 3D B16-F10 spheroids, which indicates it might be a promising lead for developing anticancer drugs.

Acacia catechu seed extract provokes cytotoxicity via apoptosis by intrinsic pathway in HepG2 cells.

Acacia catechu Willd (Fabaceae) is a thorny tree widely distributed in India and commonly used as traditional Ayurvedic medicine for various ailments. The current study evaluates the cytotoxic potentials of A. catechu ethanolic seed extract (ACSE) in HepG2 cells, a human hepatocellular carcinoma cell line. The HepG2 cells were treated with 0.1, 0.3, 1, 3, 10, 30, 100, 300 and 1000 μg/ml of ACSE and the cytotoxic effect was evaluated by MTT and lactate dehydrogenase (LDH) leakage assays. The IC50 of ACSE was found at 77.04 μg/ml and therefore, further studies were carried out with the concentrations of 35 and 70 μg/ml. The intracellular reactive oxygen species (ROS) generation and apoptosis-related morphological changes were evaluated. Gene expressions of Bax, Bcl-2, cytochrome C (Cyt-c), caspases-9 and 3 were analyzed by qPCR. The ACSE treatments caused LDH leakage was associated with an increased ROS generation. The increased ROS generation was associated with the downregulation of intracellular antioxidant enzyme superoxide dismutase and reduced glutathione content. AO/EB and PI staining also confirmed chromatin condensation and apoptosis. The flow cytometric analysis showed an accumulation of HepG2 cells at sub G0/G1 (apoptotic) phase upon ACSE treatments. The ACSE induced cytotoxicity and oxidative stress were related to increased apoptotic marker gene expressions such as Bax, Cyt-c, caspase-9 and 3, and decreased anti-apoptotic marker Bcl-2. The current finding suggests that ACSE has apoptosis-inducing potential via the mitochondrial pathway in HepG2 cells.

Cytotoxicities and wound healing effects of contact lens multipurpose solution on human corneal epithelial cell

ABSTRACT Clinical Relevance Contact lens multipurpose solutions (MPSs) contain several components that have the potential to cause corneal epithelial cell toxicity. Evaluating the components and the toxic effect of MPS should be considered for effective eye care. Background The cytotoxic and wound healing effects of five commercially available MPSs on human corneal epithelial cells (HCECs) are is investigated. Methods The following commercially available MPSs were used: Queen’s PLURISOL®, Frenz®, Boston SIMPLUS®, DL+PLUS EYE® (DL), and NEW YORK DEFINE® (NY). The proliferation of HCECs exposed to each MPS for 1, 6, and 24 h and the cytotoxicity of these solutions were analyzed using methyl thiazolyl tetrazolium-based colorimetric and lactate dehydrogenase leakage assays, respectively. The cellular morphology was evaluated by inverted phase-contrast and electron microscopy. A scratch-wound assay was performed to measure wound widths 24 h after confluent HCEC monolayers were scratch-wounded. Results The tested MPS had a time-dependent inhibitory effect on HCEC proliferation and cytotoxicity, significantly at 24 h after exposure (p< 0.05 in all MPSs). HCECs exposed to MPS detached from the bottom of the culture dishes, showed degenerative changes such as loss of microvilli, cytoplasmic vacuole formation and nuclear condensation, and decreased wound healing, compared to the controls (p< 0.001 in Boston, DL and NY). Among the tested MPS, DL and NY were more cytotoxic and showed less wound healing. Conclusion MPS has a toxic effect on HCECs, which is dependent on the concentration of the disinfecting component. Since the components that constitute the MPS are absorbed and retained in the lens, cautious scrutiny of the concentration and attention to lens cleaning are warranted to mitigate the related cytotoxicity.

Flusilazole Induced Cytotoxicity and Inhibition of Neuronal Growth in Differentiated SH-SY5Y Neuroblastoma Cells by All-Trans-Retinoic Acid (Atra)

Flusilazole (FLUS) is a broad-spectrum organosilicon triazole fungicide used for protecting economically important cereals and orchard fruits. Considering the exposure route of pesticides, pesticide contamination of food is inevitable. Furthermore, excessive exposure to pesticides causes health problems in both target and non-target organisms. It was aimed to evaluate the effects of the triazole fungicide FLUS on cytotoxicity and neurite extension in differentiated SH-SY5Y neuroblastoma cells. The SH-SY5Y cells were differentiated into mature neurons using 10-µM all-trans-retinoic acid (RA) treatment for 7 days. Then the differentiated SH-SY5Y cells were treated with 50, 100 and 200 μM FLUS for 24 h. Afterwards, cell viability assays were performed including crystal violet, neutral red cell viability, and lactate dehydrogenase leakage assays. The morphological examinations were performed and neurite lenghts of the cells were measured in all experimental groups. FLUS treatment induced cytotoxicity in SH-SY5Y cells differentiated with RA. Significant decreases in cell viability percentages were observed. Furthermore, neurite lengths were negatively affected by the treatment of FLUS at the highest concentration. FLUS is a fungicide widely used in agriculture to protect crops from fungal diseases. However, the intensive use of these compounds causes a potential risk to human and environmental health. According to the results of the study, it can be concluded that high concentrations of FLUS cause neurotoxicity by causing neural cell death and adverse effects on neurite outgrowth in differentiated SH-SY5Y cells. FLUS exposure can cause neuronal degeneration in mammals.

The Impact of the Combined Administration of 1MeTIQ and MK-801 on Cell Viability, Oxidative Stress Markers, and Glutamate Release in the Rat Hippocampus

MK-801, as an N-methyl-D-aspartate (NMDA) receptor inhibitor, causes elevation in glutamate release, which may lead to an increase in excitotoxicity, oxidative stress and, consequently, cell death. 1-Methyl-1,2,3,4-tetrahydroisoquinoline (1MeTIQ) shows antioxidant activity. The aim of the present study was to evaluate the effect of combined treatment with 1MeTIQ and MK-801 on cell viability, antioxidant enzyme activity, and glutamate release in the rat hippocampus. Cytotoxicity was measured using lactate dehydrogenase leakage assay (LDH) and the methyl tetrazolium (MTT) assay; antioxidant enzyme activity (glutathione peroxidase (GPx), glutathione reductase (GR), superoxide dismutase (SOD), and catalase (CAT)) were measured by ELISA kits. The release of glutamate in the rat hippocampus was measured using in vivo microdialysis methodology. An in vitro study showed that MK-801 induced cell death in a concentration-dependent manner and that 1MeTIQ partially reduced this adverse effect of MK-801. An ex vivo study indicated that MK-801 produced an increase in antioxidant enzyme activity (GPx, GR, and SOD), whereas coadministration of MK-801 and 1MeTIQ restored the activity of these enzymes to the control level. An in vivo microdialysis study demonstrated that combined treatment with both drugs decreased the release of glutamate in the rat hippocampus. The above results revealed that 1MeTIQ shows limited neuroprotective activity under conditions of glutamate-induced neurotoxicity.

CoNi alloy nanoparticles for cancer theranostics: synthesis, physical characterization, in vitro and in vivo studies

Nanomaterials are attracting increasing interest in many biomedical fields, including the fight against cancer. In this context, we successfully synthesized CoNi alloy nanoparticles (NPs) by a simple polyol process. The magnetic characteristics of the products were measured by vibration sample magnometry, which revealed that the samples have soft ferromagnetic behavior. The microstructure and morphology were inspected by X-ray diffraction and scanning electron microscopy, respectively. Human cancer cells derived from the breast (MCF7) and oral cavity (C152) and normal cells derived from human umbilical vein endothelial cells (HUVECs) were treated with increasing concentrations of CoNi NPs, and their cytotoxic effect was measured via MTT and lactate dehydrogenase (LDH) leakage assays. We found that treatments by using 12.5 to 400 µg/mL of Co0.5Ni0.5, Co0.6Ni0.4, and Co0.4Ni0.6 NPs were associated with significant concentration-dependent toxicity toward such cell lines and profoundly enhanced LDH leakage following 48 h of exposure (P < 0.05 compared with untreated cells). Besides, a NP dose of 6.25 µg/mL did not affect the survival of HUVECs while leading to marked cell death in MCF7 and C152 cells. In vivo experiments in rats were done to investigate the biochemical and histopathological changes over three weeks, following intraperitoneal administration of Co0.5Ni0.5, Co0.6Ni0.4, and Co0.4Ni0.6 NPs (100 mg/kg). As compared with the controls, the exposure to NPs caused significant elevations in aspartate aminotransferase, alanine aminotransferase, blood urea nitrogen, serum creatinine, serum catalase activity, serum superoxide dismutase, and liver malondialdehyde levels. Also, rats treated with Co0.6Ni0.4 NPs showed more severe histopathological changes of the liver and kidney. Our findings represent an essential step toward developing theranostic nanoplatforms for selective cancer treatment.

Photo induced mechanistic activity of GO/Zn(Cu)O nanocomposite against infectious pathogens: Potential application in wound healing

Treating infection causing microorganisms is one of the major challenges in wound healing. These may gain resistance due to the overuse of conventional antibiotics. A promising technique is antimicrobial photodynamic therapy (aPDT) used to selectively cause damage to infectious pathogenic cells via generation of reactive oxygen species (ROS). We report on biocompatable nanomaterials that can serve as potential photosensitizers for aPDT. GO/Zn(Cu)O nanocomposite was synthesized by co-precipitation method. Graphene Oxide (GO) is known for its high surface to volume ratio, excellent surface functionality and enhanced antimicrobial property. ZnO nanoparticle induces the generation of reactive oxygen species (ROS) under light irradiation and it leads to recombination of electron-hole pair. Nanocomposites of GO and Cu doped ZnO increases visible light absorption and enhances the photocatalytic property. It generates more ROS and increases the bacterial inhibition. GO/Zn(Cu)O nanocomposite was tested against Staphylococcus aureus (S. aureus), Enterococcus faecium (E. faecium), Escherichia coli (E. coli), Salmonella typhi (S. typhi), Shigella flexneri (S. flexneri) and Pseudomonas aeruginosa (P. aeruginosa) by well diffusion method, growth curve, colony count, biofilm formation under both dark and visible light condition. Reactive Oxygen Species assay (ROS), Lactate dehydrogenase leakage (LDH) assay, Protein estimation assay and membrane integrity study proves the mechanism of inhibition of bacteria. Inhibition kinetics shows the sensitivity between bacteria and GO/Zn(Cu)O nanocomposite.

Energetically significant cooperative π-stacked ternary assemblies in Ni(II) phenanthroline compounds involving discrete water clusters: Anticancer activities and theoretical studies

Two new coordination compounds viz. [Ni(phen)2(NO3)](NO3)·4H2O (1) and [Ni(phen)2(H2O)Cl]Cl·4H2O (2) (phen = 1,10-phenanthroline) have been synthesized and characterized using elemental analysis, TGA, PXRD, electronic, FT-IR spectroscopy and single crystal X-ray diffraction techniques. Crystal structure analysis reveals the formation of discrete water and anion-water clusters that contribute to the layered assemblies of the compounds. Moreover, the enclathration of (H2O)4 cluster in the layered assembly of 2 also provides rigidity to the crystal structure. Unconventional anion-π interaction is also observed in compound 1 involving the π-system of the chelate ring of coordinated phen. Detailed structural investigation reveals the presence of biologically relevant cooperative π-stacked (π–π)1/(π–π)2/(π–π)1 ternary assemblies in both the compounds. Interestingly, DFT calculations reveal that (π-π)2 binding mode in 2 is energetically significant than in compound 1 which is further analyzed by NCI plot index computational tool. The compounds have been screened for anticancer activities considering cell cytotoxicity and apoptosis in Dalton's lymphoma (DL) malignant cancer cell line. The complexes significantly inhibit cell viability by inducing apoptotic cell death in cancer cell lines with negligible cytotoxicity in normal cells. DNA ladder assay results in the degradation of genomic DNA into small inter-nucleosomal fragments (damage DNA) which is a biochemical hallmark of DL cells undergoing apoptosis. Lactate dehydrogenase (LDH) leakage assay also reveals the release of the intracellular enzyme LDH from DL cells indicating the irreversible cell death involving cell membrane damage. The molecular docking simulations reveal that the compounds interact and accommodated well with the active site of anti-apoptotic BCL proteins. Structure activity relationship (SAR) analysis based on pharmacophore modelling reveal that the molecular features of the compounds play important roles in the biological activities.