HEp-2 Cells Research Articles

Synthesis and characterization of Pluronic F-127-coated titanium dioxide nanoparticles synthesized from extracts of Atractylodes macrocephala leaf for antioxidant, antimicrobial, and anticancer properties

Abstract Globally, nanotechnology is generating significant interest because of its promise in a wide range of industries. The most commonly used nanoparticles are titanium dioxide nanoparticles (PF-127 coated TiO2 NPs), which can be formulated with physical, chemical, and environmental factors. The establishment of an economical and environmentally beneficial method for its fabrication is due to increasing concerns about human health impacts. In this exploration, green Pluronic F-127 (PF-127) coated TiO2 NPs using leaf extracts of Atractylodes macrocephala have been formulated and studied through various methods. PF-127 coated TiO2 NPs were 60 nm large and a polygonal rutile-type crystalline structure was observed. Moreover, the NPs’ antimicrobial capacity against several pathogens was investigated. The cytotoxicity of the NPs against HEp-2, KB, and Vero cell lines was assessed using the MTT test. Increased antimicrobial potential of PF-127 coated TiO2 NPs against several pathogens was noted. Furthermore, NPs displayed remarkable antioxidant activity, which increased with concentration. The NPs exhibited significant cytotoxic effects against HEp-2 and KB cell lines but failed to demonstrate toxicity against Vero cells. This is indicative of their cytotoxic potential against cancer cell lines and non-toxic nature towards healthy cells. This indicates that PF-127 coated TiO2 NPs possess beneficial antimicrobial and antitumor properties.

Some new insights into the biological activities of carboxymethylated polysaccharides from Lasiodiplodia theobromae

BackgroundCarboxymethylated Lasiodiplodan (LaEPS-C), Lasiodiplodia theobromae β-glucan exopolysaccharide derivative, has a well-known range of biological activities. Compared to LaEPS-C, its fractions, Linear (LLaEPS-C) and Branched (BLaEPS-C), have biological potentialities scarcely described in the literature. So, in this study, we investigate the immunomodulatory, antiviral, antiproliferative, and anticoagulant activities of LLaEPS-C and BLaEPS-C and compare them to the LaEPS-C.MethodsLaEPS was obtained from L. theobromae MMBJ. After carboxymethylation, LaEPS-C structural characteristics were confirmed by Elementary Composition Analysis by Energy Dispersive X-Ray Detector (EDS), Fourier Transform Infrared (FTIR), and Nuclear Magnetic Resonance (NMR). The immunomodulatory activity on cytokine secretion was evaluated in human monocyte-derived macrophage cultures. The antiviral activity was evaluated by Hep-2 cell viability in the presence or absence of hRSV (human respiratory syncytial virus). In vitro antiproliferative activity was tested by sulforhodamine B assay. The anticoagulant activity was determined by APTT (Activated Partial Thromboplastin Time) and PT (Prothrombin Time).ResultsLaEPS-C showed low macrophage cell viability only at 100 µg/mL (52.84 ± 24.06, 48 h), and LLaEPS-C presented no effect. Conversely, BLaEPS-C showed cytotoxicity from 25 to 100 µg/mL (44.36 ± 20.16, 40.64 ± 25.55, 33.87 ± 25.16; 48 h). LaEPS-C and LLaEPS-C showed anti-inflammatory activity. LaEPS-C presented this at 100 µg/mL (36.75 ± 5.53, 48 h) for IL-10, and LLaEPS-C reduces TNF-α cytokine productions at 100 µg/mL (18.27 ± 5.80, 48 h). LLaEPS-C showed an anti-hRSV activity (0.7 µg/ml) plus a low cytotoxic activity for Hep-2 cells (1.4 µg/ml). LaEPS-C presented an antiproliferative activity for NCI-ADR/RES (GI50 65.3 µg/mL). A better PT was achieved for LLaEPS-C at 5.0 µg/mL (11.85 ± 0.87s).ConclusionsThese findings demonstrated that carboxymethylation effectively improves the biological potential of the LaEPS-C and their fractions. From those polysaccharides tested, LLaEPS provided the best results with low toxicity for anti-inflammatory, antiviral, and anticoagulant activities.

Respiratory syncytial virus co-opts hypoxia-inducible factor-1α-mediated glycolysis to favor the production of infectious virus.

Respiratory syncytial virus (RSV) is the leading etiological agent of lower respiratory tract illness. However, efficacious vaccines or antiviral drugs for treating RSV infections are currently not available. Indeed, RSV depends on host cells to provide energy needed to produce progeny virions. Glycolysis is a series of oxidative reactions used to metabolize glucose and provide energy to host cells. Therefore, glycolysis may be helpful for RSV infection. In this study, we show that RSV increases glycolysis by inducing the stabilization, transcription, translation, and activation of hypoxia-inducible factor (HIF)-1α in infected cells, which is important for the production of progeny RSV virions. This study contributes to understanding the molecular mechanism by which HIF-1α-mediated glycolysis controls RSV infection and reveals an effective target for the development of highly efficient anti-RSV drugs.

Five-Year Analysis of Anti-Nuclear Antibody (ANA) and Extractable Nuclear Antigen Antibody (anti-ENA) Immunoblot Test Results

Autoimmune disorders may cause systemic disease in which multiple organs are affected or may also be localized as single-organ involvement. Various ancillary immunological tests used in cases where systemic involvement is observed, and differential diagnosis becomes difficult or non-specific symptoms are observed. For this purpose, investigation of antinuclear antibodies (ANA) and antibodies against extractable nuclear antigens (ENA) in patient serums using various methods is a guide for clinicians. In this study, we aimed to evaluate the effectiveness of ANA and ENA antibody tests by examining the test results, the patients’ demographic data, and the clinics that ordered the tests. In the study, a retrospective review of the results of 37,791 ANA tests studied on serum samples sent to the medical microbiology laboratory of our hospital from various clinics between January 2018 and December 2022 is presented. ANA tests were resulted semi-quantitatively by evaluating the slides prepared using diluted serum samples at a ratio of 1:100 and HEp-2 cells and monkey liver tissue under a fluorescent microscope with the indirect immunofluorescence (IIF) antibody method. Of the patients who requested ANA test, 11,604 (30.7%) were male and 26,187 (69.3%) were female. The mean ages were 43.46 (0-94) and 46.94 (0-96), respectively. Of the samples tested, 18.8% (7,107/37,791) were evaluated as ANA positive. The ANA positivity rate was significantly higher in women (24.8%) than in men (19.7%) (p<0.001). ANA positive samples were re-tested using the anti-ENA immunoblot method. Of the 7,107 ANA-positive patients, 3,809 (53.6%) were found to be anti-ENA positive for at least one specific antibody, while 3,298 (46.4%) were negative. In the anti-ENA profile, antibodies against 11 different antigen types were screened and the most frequently detected antibody was anti-SS-A (22.7%), and this was followed by anti-centromere (12.8%), anti-RNP (10.2%), and anti-dsDNA (9.7%) antibodies. Immunoblot-based anti-ENA tests provide specific information for the diagnosis of autoimmune diseases. In our study, the fact that anti-ENA tests were negative in a significant part of ANA positive patients indicates the inadequacy of the immunoblot test panel and reveals the need for immunoblot tests with wider antibody diversity or any alternative methods.

Effects of ceramide C2 application on human laryngeal carcinoma cells: a cell culture study.

In the present study, we investigated the effects of Ceramide C2 application on human laryngeal carcinoma cells. Human larynx epidermoid carcinoma HEp-2 (ATCC® CCL-23™) cells were purchased from the American Type Culture Collection (ATCC, USA). Human larynx epidermoid carcinoma HEp-2 cells were cultured in complete Dulbecco's Modified Eagle's Medium (DMEM) supplemented with fetal bovine serum (FBS) (10%) and penicillin/streptomycin (1%) in a CO2 (5%) incubator under standard cell culture conditions. Ceramide C2 was prepared, and further dilutions ranging from 3.13 to 100 μM were prepared in a fresh culture medium. Cells on 96 well plates were exposed to the prepared concentrations of ceramide C2 for 24 and 48 hours. Cytotoxicity evaluation was performed by MTT. Apoptosis profiles of HEp-2 cells were detected by annexin-V analysis. The activated caspases 3/7 on HEp-2 cells after ceramide C2 exposure were evaluated with flow cytometric analysis. The morphological changes on HEp-2 cells caused by ceramide C2 were evaluated by staining with phalloidine and acridine orange via confocal microscopy. For the Wound Healing Assay, HEp-2 cells were cultured in 6 well-plates until they became confluent. MTT cytotoxicity test findings revealed that the viability of human laryngeal carcinoma cells decreased with the increased application of ceramide C2 for 24 hours compared to untreated (control) cells. The highest growth inhibition by ceramide C2 for short-term application for 24 hours was detected at the highest concentration of ceramide C2 (100 µM). Annexin-V findings showed that 98.97 of HEp-2 cells were alive, and 1.63% were detected as early apoptosis for the control group. The results showed that ceramide C2 triggered apoptosis on HEp-2 cells with a percentage of total apoptotic cells of 61,40 compared to untreated HEp-2 cells. Cysteine proteases (caspases) 3/7 activation percentages of HEp-2 cells exposed to ceramide C2 for 24 hours were compared to control cells, and the morphology of HEp-2 cells was changed with clear apoptotic signs that underlined the cytotoxicity and pro-apoptotic activity of ceramide C2. Scratch Assay assessed the migration capability of HEp-2 cells before and after the exposure to ceramide C2. It showed that ceramide C2 reduced human laryngeal carcinoma cells' migration capability and proliferation for 24 hours. Based on all study findings, it can be considered that short-chain ceramide C2 exerted cytotoxicity on human laryngeal carcinoma cells in a dose and time-dependent manner and reduced the viability via inducing caspase-dependent apoptosis. The overall effect might be derived from the elevated intracellular ceramide levels by the exogenous application of ceramide C2. Consequently, it was concluded that ceramide C2 has good potential to cause cytotoxicity and apoptosis in human laryngeal carcinoma cells and, after deeper in vitro and in vivo investigations, can be a good candidate for designing anti-cancer drugs with high efficiency.

Longitudinal study of patients with discrepant results in CLIFT and a solid-phase dsDNA antibody assay: does a gold standard dsDNA assay exist?

ObjectiveAntidouble-stranded DNA (dsDNA) antibodies are essential for diagnosis and follow-up of systemic lupus erythematous (SLE). To ensure the best diagnostic approach, most healthcare laboratories opt for a combination of highly...

Intermittent Hypoxia Promotes TAM-Induced Glycolysis in Laryngeal Cancer Cells via Regulation of HK1 Expression through Activation of ZBTB10.

Obstructive sleep apnea (OSA), characterized by intermittent hypoxia (IH), may increase the risk of cancer development and a poor cancer prognosis. TAMs of the M2 phenotype, together with the intermittent hypoxic environment within the tumor, drive tumor aggressiveness. However, the mechanism of TAMs in IH remains unclear. In our study, IH induced the recruitment of macrophages, and IH-induced M2-like TAMs promoted glycolysis in laryngeal cancer cells through hexokinase 1. The hexokinase inhibitor 2-deoxy-D-glucose and HK1 shRNA were applied to verify this finding, confirming that M2-like TAMs enhanced glycolysis in laryngeal cancer cells through HK1 under intermittent hypoxic conditions. Comprehensive RNA-seq analysis disclosed a marked elevation in the expression levels of the transcription factor ZBTB10, while evaluation of a laryngeal cancer patient tissue microarray demonstrated a positive correlation between ZBTB10 and HK1 expression in laryngeal carcinoma. Knockdown of ZBTB10 decreased HK1 expression, and overexpression of ZBTB10 increased HK1 expression in both laryngeal cancer cells and 293T cells. The luciferase reporter assay and Chromatin immunoprecipitation assay confirmed that ZBTB10 directly bound to the promoter region of HK1 and regulated the transcriptional activity of HK1. Finally, the CLEC3B level of the M2 supernatant is significantly higher in the IH group and showed a protumor effect on Hep2 cells. As ZBTB10-mediated regulation of HK1 affects glycolysis in laryngeal cancer, our findings may provide new potential therapeutic targets for laryngeal cancer.

Anti-nuclear valosin-containing protein-like autoantibody is associated with calcinosis and higher risk of cancer in systemic sclerosis.

Systemic sclerosis (SSc)-specific autoantibodies allow the diagnosis and predict the prognosis of SSc patients with different clinical characteristics. The aim of this study was to describe new SSc-related autoantibodies by a novel protein immunoprecipitation (IP) assay. Serum samples and clinical data were collected from 307 SSc patients. Antinuclear autoantibodies were tested in all patients by indirect immunofluorescence (IIF) on HEp-2 cells. SSc-specific autoantibodies were evaluated with a commercial immunoblot and chemiluminescence immunoassay, and traditional RNA-IP. Patients negative for all these autoantibodies (n = 51) were further tested with a non-radioactive protein IP assay. Protein bands detected on SDS-PAGE were then analysed by mass spectrometry (MS) and confirmed by western blot (WB). Additional 56 patients with nucleolar pattern by IIF were tested by protein IP-WB. Five patients who underwent protein IP testing showed a 110-115kDa molecular weight band on SDS-PAGE and a homogeneous nucleolar pattern by IIF. MS identified the bands as nuclear valosin-containing protein-like (NVL). An additional positive patient was detected by IP-WB. As compared with the remaining 101 negative patients, anti-NVL positive patients showed a greater prevalence of calcinosis (100% vs 18.9%, P < 0.001), and cancer (66.7% vs 8.9%, P = 0.002), with a particular association with synchronous cancer (OR = 16.3; P = 0.024). We identified NVL as a new autoantibody target by a novel protein IP assay in SSc patients with a homogeneous nucleolar IIF pattern, testing negative for all known SSc-specific autoantibodies by commercial assays and RNA IP. Anti-NVL identifies a new clinical phenotype, characterized by calcinosis and cancer.

MORPHOLOGICAL CHARACTERISTICS AND CYTOTOXIC ACTIVITY OF THE BLACK SEA STRAIN STREPTOMYCES SP. LIM 10

Aim. The aim of the work was to study the morphological characteristics and cytotoxic activity of the strain of actinobacteria Streptomyces sp. Lim 10 isolated from natural fouling of shell rock in the Odesa Bay of the Black Sea. Materials and methods. The object of the study was a strain of marine actinobacteria Streptomyces sp. Lim 10. The morphology and growth patterns were studied on the media of Gauze 2, Czapek, Eshbi, TSB and on the media of ISP (ISP-1–ISP-7). SG medium was used to accumulate secondary metabolites of actinobacteria. Extraction of exometabolites from the culture liquid was carried out with ethyl acetate followed by dissolution in dimethyl sulfoxide (DMSO). The cytotoxic activity of exometabolites of actinobacteria strain Streptomyces sp. Lim 10 was studied on the model of cultures cells human - rhabdomyosarcoma (RD,) and of the laryngeal adenocarcinoma (Hep-2). The cytotoxic effect of exometabolites of the actinobacteria strain Streptomyces sp. Lim 10 was determined after 24 hours of incubation by microscopy. The degree of destruction of the monolayer was assessed by the number of viable cells in terms of optical density, which was measured using a spectrophotometer at a wavelength of 630 nm. The exometabolites of actinobacteria strain Streptomyces sp. Lim 10 were identified using LC-ESIMS. Results. Study of the morphological properties of the Black Sea strain of actinobacteria Lim 10 showed that it is characterized by pleomorphism. А high cytotoxic activity of exometabolites of the marine strain of actinobacteria Streptomyces sp. Lim 10 was established. The level of cytotoxic effect of metabolites of the strain of actinobacteria Streptomyces sp. Lim 10 on Hep-2 cell culture depended on their concentration. RD cell culture was more sensitive to the cytotoxic effect of exometabolites of Streptomyces sp. Lim 10 – at all tested concentrations of exometabolites (from 2.5 to 500.0 μg/ml), the number of viable cells in the RD monolayer decreased to 51.3 ± 3.0 – 63.7 ± 5.4%, compared to the control. This can be at least partially explained by action of apoptosis-inducing agent staurosporine, which is produced by strain Streptomyces sp. Lim 10 along with itsstructural homologs. Conclusion. Strain Streptomyces sp. Lim 10 is a promising producer of antitumor compounds and can be recommended for further more in-depth studies.

Anticancer and antimicrobial evaluation of extract from brown algae Hormophysa cuneiformis.

We investigated the antimicrobial and anticancer properties of an ethanol crude extract of Red Sea brown alga (Hormophysa cuneiformis) from Egypt. Extraction was achieved by mixing 100 g of sample powder with absolute ethanol, incubating at 37 °C overnight in a shaking incubator, and then collecting the extract. The extract's antimicrobial activity was tested using a well diffusion assay against the tested pathogens (Escherichia coli, Bacillus subtilis, Staphylococcus aureus, and Candida albicans) in comparison to commercial antibiotics. Anticancer activity was assessed using MTT assay on MCF-7, HepG-2, and HEP-2 cell lines. The anticancer mechanism of action against the HepG-2 cell line was investigated using cell cycle analysis, Annexin V, and antioxidant enzymes, in addition to transmission electron microscopy. GC-MS phytoconstituent profile of the extract was dominant with fatty acids. A broad antimicrobial effect against all the pathogenic isolates of E. coli, S. aureus, B. subtitles, and C. albicans was demonstrated, especially at the high concentration in comparison to commercial antibiotics. The extract could inhibit the growth of the tested cell lines. We observed the most significant effect on HepG-2 cells, and the concentration of the extract played a role in the level of inhibition (IC50 of 44.6 ± 0.6 µg/ml). The extract had negligible effects on Vero normal cell lines at the lower concentration, with slight toxicity (90.8% viability) at the highest concentration (500 µg/ml). At this same concentration, the extract caused 80-92% inhibition of the cancer cell lines. The extract appears to have demonstrated promising effects on cancer cells. It induces programmed cell death (apoptosis), arrests the cell cycle, and affects the oxidative/antioxidant balance within the cells, potentially leading to the suppression or elimination of cancer cells. These findings are encouraging and may have implications for cancer treatment or further research in this area. More action of extract was seen against bacteria than fungi, with a wide antibacterial impact against all of the tested isolates, notably at the high concentration in comparison to conventional antibiotics. According to the findings, H. cuneiformis may be a valuable source of chemicals that are both antimicrobial and anticancer.

Dual anticancer activity of Aspergillus nidulans pigment and Ionizing γ-Radiation on human larynx carcinoma cell line

BackgroundFungi are a readily available source of naturally generated colored compounds. These compounds might be used as radiosensitizers for treating cancer cells.MethodsAspergillus nidulans was examined for its color-producing ability in Potato dextrose agar (PDA) broth medium. The pigment was characterized by Ultraviolet (UV) spectrophotometer and Gas Chromatography Mass Spectrometry (GC/MS). Pigment extracts from A. nidulans were studied for their cytotoxic effects on the growth of human larynx carcinoma cell line (HEp-2) with or without exposure to γ-radiation at three different doses (5, 10, and 15 Gy). A. nidulans pigment cytotoxic activity was tested against normal Vero cells. Cell apoptosis was studied using flow cytometry. Gene expression of P53, Caspase 3 and Bcl-2 were quantified.ResultsUltraviolet spectrum and GC/MS revealed the ability of Aspergillus nidulans to produce Rhodopin pigment. HEp-2 cells treated with A. nidulans pigment only give IC50 about 208 µg/ml. In contrast, when treated with the pigment +10 Gy γ-radiation, it give about 115 µg/ml. However, for normal cells, lower cytotoxic activity was detected. Treatment with pigment (208 g/mL) caused about 50% ± 1.0 total apoptosis level and gene expression of P53: 2.3 fold and Caspase 3: 1.84 fold in respect to untreated HEp-2), while Bcl-2 was decreased (Bcl-2: 0.63 fold in respect to untreated HEp-2). Furthermore, treated with pigment (115 µg/mL) + 10Gy caused about 47.41% ± 1.7 total apoptosis level and P53: 2.53 fold and Caspase 3: 2.0 fold in respect to untreated HEp-2, while Bcl-2 was downregulated (Bcl-2: 0.61 fold in respect to untreated HEp-2).ConclusionThis study concluded that the anti-cancer activity of Aspergillus nidulans pigment was enhanced by ionizing radiation at 10 Gy, as well as its low cytotoxic activity against normal Vero cells.

Effect on cell survival and cytoophidium assembly of the adRP-10-related IMPDH1 missense mutation Asp226Asn.

Introduction: Inosine monophosphate dehydrogenase 1 (IMPDH1) is a critical enzyme in the retina, essential for the correct functioning of photoreceptor cells. Mutations in IMPDH1 have been linked to autosomal dominant retinitis pigmentosa subtype 10 (adRP-10), a genetic eye disorder. Some of these mutations such as the Asp226Asn (D226N) lead to the assembly of large filamentous structures termed cytoophidia. D226N also gives IMPDH1 resistance to feedback inhibition by GDP/GTP. This study aims to emulate the adRP-10 condition with a long-term expression of IMPDH1-D226N in vitro and explore cytoophidium assembly and cell survival. We also assessed whether the introduction of an additional mutation (Y12C) to disrupt the cytoophidium has an attenuating effect on the toxicity caused by the D226N mutation. Results: Expression of IMPDH1-D226N in HEp-2 cells resulted in cytoophidium assembly in ∼70% of the cells, but the presence of the Y12C mutation disrupted the filaments. Long-term cell survival was significantly affected by the presence of the D226N mutation, with a decrease of ∼40% in the cells expressing IMPDH1-D226N when compared to IMPDH1-WT; however, survival was significantly recovered in IMPDH1-Y12C/D226N, with only a ∼10% decrease when compared to IMPDH1-WT. On the other hand, the IMPDH1 expression level in the D226N-positive cells was <30% of that of the IMPDH1-WT-positive cells and only slightly higher in the Y12C/D226N, suggesting that although cell survival in Y12C/D226N was recovered, higher expression levels of the mutated IMPDH1 were not tolerated by the cells in the long term. Conclusion: The IMPDH1-D226N effect on photoreceptor cell survival may be the result of a sum of problems: nucleotide unbalance plus a toxic long-life cytoophidium, supported by the observation that by introducing Y12C in IMPDH1 the cytoophidium was disrupted and cell survival significantly recovered, but not the sensibility to GDP/GTP regulation since higher expression levels of IMPDH1-D226N were not tolerated.

Design, synthesis, and antitumor screening of new thiazole, thiazolopyrimidine, and thiazolotriazine derivatives as potent inhibitors of VEGFR-2.

New thiazole, thiazolopyrimidine, and thiazolotriazine derivatives 3-12 and 14a-f were synthesized. The newly synthesized analogs were tested for in vitro antitumor activity against HepG2, HCT-116, MCF-7, HeP-2, and Hela cancer cells. Results indicated that compound 5 displayed the highest potency toward the tested cancer cells. Compound 11b possessed enhanced effectiveness over MCF-7, HepG2, HCT-116, and Hela cancer cells. In addition, compounds 4 and 6 showed promising activity toward HCT-116, MCF-7, and Hela cancer cells and eminent activity against HepG2 and HeP-2 cells. Moreover, compounds 3-6 and 11b were tested for their capability to inhibit vascular endothelial growth factor receptor-2(VEGFR-2) activity. The obtained results showed that compound 5 displayed significant inhibitory activity against VEGFR-2 (half-maximal inhibitory concentration[IC50 ] = 0.044 μM) comparable to sunitinib (IC50 = 0.100 μM). Also, the synthesized compounds 3-6 and 11b were subjected to in vitro cytotoxicity tests over WI38 and WISH normal cells. It was found that the five tested compounds displayed significantly lower cytotoxicity than doxorubicin toward normal cell lines. Cell cycle analysis proved that compound 5 induces cell cycle arrest in the S phase for HCT-116 and Hela cancer cell linesand in the G2/M phase for theMCF-7 cancer cell line. Moreover, compound 5 induced cancer cell death through apoptosis accompanied by a high ratio of BAX/BCL-2 in the screened cancer cells. Furthermore, docking results revealedthat compound 5 showedthe essential interactionbonds with VEGFR-2, which agreed with in vitro enzyme assay results. In silico studies showed that most of the analyzed compounds complied with the requirements of good oral bioavailability with minimal toxicity threats in humans.

Poly(2-n-propyl-2-oxazoline)/tannic acid nanocapsules with in-situ synthesized gold nanorods for cancer theranostics

The concept of hollow polymeric capsules fabricated with thermoresponsive polymers has revealed immense possibilities in the field of nanomedicine due to their tunable temperature and pH responsiveness caused by lower critical solution temperature (LCST) behavior. Herein, we report a new methodology to in-situ synthesize gold nanorods (AuNRs) within the pores of mesoporous silica nanoparticles (NPs) by trapping gold seed crystals and growing them into NRs in the presence of a weak reducing agent. The mesoporous silica NPs with incorporated NRs, i.e., modified silica NPs (SiO2-NRs) were used as sacrificial template for the fabrication of hollow polymeric multilayer capsules via layer-by-layer (LbL) assembly using poly(2-n-propyl-2-oxazoline) (PnPrOx) and tannic acid (TA) as layer components. The NRs of 25 ± 5 nm in size were uniformly distributed in the interior structure of PnPrOx/TA nanocapsules. The photo-thermal conversion efficiency of the PnPrOx/TA nanocapsules was found to be 47.5%. The release of encapsulated doxorubicin was found to be 55%, which increased to 68% when the incubation time was increased from 1 to 2 h. An on-demand burst release of 56% of the drug occurred within 2 min of NIR light exposure, and it reached 72% within 30 min, which inflicted major damage to cancer cells. Notably, the capsule suspension temperature increase from room temperature to 47 °C under laser light exposure, which is not only playing a key role in rupturing the capsules to enhance the drug release but also induces a hyperthermia effect in the tumor environment. Further, the in-vitro cytotoxicity experiments with human epithelial HEp-2 cells and fibrosarcoma HT1080 cells revealed an enhanced anti-cancer activity due to a synergistic chemo-photothermal effect. Hence, the proposed nanocapsule system can provide a unique, cheap, and efficient way of fabricating cancer theranostics for in-vivo applications.

Fabrication of BiCuOS nanoflowers acting as nanoarrays on photonic nanoparticles for chemo-photothermal therapy

In recent years, 3D flower shaped nanostructures with an ability to improve internal photo reflections and absorptions is gaining popularity for the development of photo-thermal heat generating materials for cancer therapy. The use of bimetallic oxysulfide materials as photo-thermal agent is especially interesting as it is not only resulting in heat generation by absorbing laser light in the biological window but also participates in reactive oxygen species generation in tumor environment. In this work, we have reported a novel flower shaped BiCuOS nanostructures made of CuS base structures with vertical nanoarrays of bismuth oxide nanoplates for synergistic chemo-photothermal cancer therapy. Notably, 2D nanoplates (∼150 ± 50 nm in length and thickness of about ∼25 nm) could be seen grown in the form of nanoflowers (NFs) of 800 ± 100 nm in size on CuS base structures. The electronic structure, density of states, bond angle, bond length and thermal stability of formed NFs were investigated by density functional theory and molecular dynamic studies. The photo-thermal conversion efficiency was increased as a function of NF concentration. The percentage of doxorubicin released was estimated to be only ∼4% even after one day of incubation at pH 7.4 which increased to 38 and 45% at pH 5.5 without and with near infra-red light exposure, respectively. The cell viability experiments with fibroblast cells and red blood cells exhibited excellent biocompatibility of BiCuOS NFs. The synthesized BiCuOS NFs showed excellent anticancer activity against Hep-2 cells. Hence, the BiCuOS NFs reported here have huge potential for use as novel dual performing chemo-and-photothermal agent for advanced cancer therapy.

Age-differential CD13 and interferon expression in airway epithelia affect SARS-CoV-2 infection - Effects of vitamin D

Young age and high vitamin D plasma levels have been associated with lower SARS-CoV-2 infection risk and favourable disease outcomes. This study investigated mechanisms associated with differential responses to SARS-CoV-2 across age groups and effects of vitamin D. Nasal epithelia were collected from healthy children and adults and cultured for four weeks at the air-liquid interface with and without vitamin D. Gene expression and DNA methylation were investigated. Surface protein expression was confirmed by immunofluorescence while vitamin D receptor recruitment to the DNA was analysed through chromatin immunoprecipitation. HEp-2 cells were used for protein co-immunoprecipitation and luciferase reporter assays. Compared to children, airway epithelia from adults show higher viral RNA recovery following infection. This was associated with higher ANPEP/CD13, reduced type I interferon expression, and differential DNA methylation. In cells from adults, exposure to vitamin D reduced TTLL-12 expression, a negative regulator of the interferon response. This was mediated by vitamin D receptor recruitment to TTLL12, where it instructs DNA methylation through DNA methyltransferase 1. This study links age-dependent differential expression of CD13 and type I interferon to variable infection of upper airway epithelia. Furthermore, it provides molecular evidence for vitamin D reducing viral replication by inhibiting TTLL-12.

Cytotoxic Copper(II) Complexes Based on 2,2'-Bipyridine/1,10- Phenanthroline and 5-(4-Chlorophenyl)-1H-tetrazole: Synthesis and Structures

Five coordination compounds [Cu2(Bipy)2L4]·C2H5OH (Iа, Ib), [Cu2(Dmbipy)2L4] (II),[Cu2(Phen)2L4]·H2O (IIIa), [Cu2(Dmphen)2L4] (IVa), and [Cu2(Phendione’)2L4]·2C2H5OH·2H2O (V) aresynthesized from 5-(4-chlorophenyl)-1H-tetrazole (HL), where Bipy is 2,2'-bipyridine, Dmbipy is 4,4'-dimethyl-2,2'-bipyridine, Phen is 1,10-phenanthroline, Dmphen is 4,7-dimethyl-1,10-phenanthroline, andPhendione’ is 6-ethoxy-6-hydroxy-1,10-phenanthrolin-5-one. The crystal structures of the complexes aredetermined by X-ray diffraction (XRD) of single crystals (CIF files CCDC nos. 2225368 (Ia), 2225369 (Ib),2225370 (II), 2225372 (IIIa), 2225373 (IVa), and 2225371 (V)). The compounds are binuclear due to thebridging function of the tetrazolate anion, and the coordination number of copper is five in all synthesizedcomplexes. The cytotoxic activity of the complexes against the Hep2 and HepG2 cancer cell lines and noncanceroushuman fibroblasts MRC-5 is studied. The complexes exhibit pronounced cytotoxic properties, andcompound V has the maximum selectivity index with respect to the cancer cells.

MiR-5189-3p Suppresses cell Proliferation, Invasion and Migration Through Targeting EIF5A2 in Laryngeal Squamous Cell Carcinoma.

A growing body of evidence suggests that miR-5189-3p plays a critical role in multiple diseases. This study aimed to investigate the function of miR-5189-3p in laryngeal squamous cell carcinoma (LSCC) and explore its underlying mechanisms. qRT-PCR was designed to determine the expression levels of miR-5189-3p and eukaryotic translation initiation factor 5A2 (EIF5A2), while CCK-8 assay was performed to measure the effects of miR-5189-3p on cell proliferation. Transwell assay was performed to evaluate cell invasion as well as migration, and wound healing assay was applied to demonstrate cell migratory ability. Target gene prediction and luciferase reporter assay were developed to screen the possible target gene of miR-5189-3p, and Western blot was designed to measure EIF5A2 protein expression. MiR-5189-3p was down-regulated in LSCC tissues and cell lines. Up-regulation of miR-5189-3p notably inhibited cell proliferation, invasion, and migration in HEP2 and FADU cells. EIF5A2 was the potential downstream gene of miR-5189-3p, and overexpression of miR-5189-3p apparently reduced EIF5A2 expression. Moreover, reintroduction of EIF5A2 rescued the tumor suppressive effects of miR-5189-3p. MiR-5189-3p functions as a tumor inhibitor in LSCC progression via directly regulating EIF5A2 and may be a potential therapeutic target for LSCC.

Motility provides specific adhesion patterns and improves Listeria monocytogenes invasion into human HEp-2 cells.

Listeria monocytogenes is motile at 22°C and non-motile at 37°C. In contrast, expression of L. monocytogenes virulence factors is low at 22°C and up-regulated at 37°C. Here, we studied a character of L. monocytogenes near surface swimming (NSS) motility and its effects on adhesion patterns and invasion into epithelial cells. L. monocytogenes and its saprophytic counterpart L. innocua both grown at 22°C showed similar NSS characteristics including individual velocities, trajectory lengths, residence times, and an asymmetric distribution of velocity directions. Similar NSS patterns correlated with similar adhesion patterns. Motile bacteria, including both pathogenic and saprophytic species, showed a preference for adhering to the periphery of epithelial HEp-2 cells. In contrast, non-motile bacteria were evenly distributed across the cell surface, including areas over the nucleus. However, the uneven distribution of motile bacteria did not enhance the invasion into HEp-2 cells unless virulence factor production was up-regulated by the transient shift of the culture to 37°C. Motile L. monocytogenes grown overnight at 22°C and then shifted to 37°C for 2 h expressed invasion factors at the same level and invaded human cells up to five times more efficiently comparatively with non-motile bacteria grown overnight at 37°C. Taken together, obtained results demonstrated that (i) NSS motility and correspondent peripheral location over the cell surface did not depend on L. monocytogenes virulence traits; (ii) motility improved L. monocytogenes invasion into human HEp-2 cells within a few hours after the transition from the ambient temperature to the human body temperature.

The Synthesis of BODIPY-TKI Conjugates and Investigation of Their Ability to Target the Epidermal Growth Factor Receptor

A near-IR BODIPY was covalently conjugated via its isothiocyanate groups to one or two Erlotinib molecules, a known tyrosine kinase inhibitor (TKI), via triethylene glycol spacers, to produce two novel BODIPY-monoTKI and BODIPY-diTKI conjugates. The ability of these conjugates to target the intracellular domain of the epidermal growth factor receptor (EGFR) was investigated using molecular modeling, surface plasma resonance (SPR), EGFR kinase binding assay, time-dependent cellular uptake, and fluorescence microscopy. While both the BODIPY-monoTKI and the BODIPY-diTKI conjugates were shown to bind to the EGFR kinase by SPR and accumulated more efficiently within human HEp2 cells that over-express EGFR than BODIPY alone, only the BODIPY-monoTKI exhibited kinase inhibition activity. This is due to the high hydrophobic character and aggregation behavior of the BODIPY-diTKI in aqueous solutions, as shown by fluorescence quenching. Furthermore, the competition of the two Erlotinibs in the diTKI conjugate for the active site of the kinase, as suggested by computational modeling, might lead to a decrease in binding relative to the monoTKI conjugate. Nevertheless, the efficient cellular uptake and intracellular localization of both conjugates with no observed cytotoxicity suggest that both could be used as near-IR fluorescent markers for cells that over-express EGFR.