Toxicity of commercially available engineered nanoparticles to Caco-2 and SW480 human intestinal epithelial cells

The effects of ingestion of engineered nanoparticles (NPs), especially via drinking water, are unknown. Using NPs spiked into synthetic water and cell culture media, we investigated cell death, oxidative stress, and inflammatory effects of silver (Ag), titanium dioxide (TiO₂), and zinc oxide (ZnO) NPs on human intestinal Caco-2 and SW480 cells. ZnO NPs were cytotoxic to both cell lines, while Ag and TiO₂ NPs were toxic only at 100 mg/L to Caco-2 and SW480, respectively. ZnO NPs led to significant cell death in synthetic freshwaters with 1 % phosphate-buffered saline in both cell lines, while Ag and TiO₂ NPs in buffered water led to cell death in SW480 cells. NP exposures did not yield significant increased reactive oxygen species generation but all NP exposures led to increased IL-8 cytokine generation in both cell lines. These results indicate cell stress and cell death from NP exposures, with a varied response based on NP composition.     

Evaluation of 5-HT7 Receptor Trafficking on In Vivo and In Vitro Model of Lipopolysaccharide (LPS)-Induced Inflammatory Cell Injury in Rats and LPS-Treated A549 Cells

This study aimed to investigate the effects of the 5-HT7 receptor agonist (LP44) and antagonist (SB269970) on LPS-induced in vivo tissue damage and cell culture by molecular methods. This study was conducted in two steps. For in vivo studies, 24 female rats were divided into four groups. Group I: healthy; II (2nd h): LPS 5 mg/kg administered intraperitoneally (i.p.); III (4th h): LPS 5 mg/kg administered i.p.; IV (8th h): LPS 5 mg/kg administered i.p. For in vitro studies, we used the A549 cell line. Groups: I control (healthy) (2–4 h); II LPS: 1 µg/ml E. Coli O55:B5 strain (2–4 h); III agonist (LP44) 10^?9 M (2–4 h); IV antagonist (SB269970) 10^?9 M (2–4 h); V LPS+agonist 10^?9 M (LP44 1 µg/ml) (2–4 h); VI LPS+antagonist 10^?9 M (2–4 h). In molecular analyses, we determined increased TNF-α, IL-1β, NF-κB, and 5-HT7 mRNA expressions in rat lung tissues and increased TNF-α, iNOS, and 5-HT7 mRNA expressions in the A549 cell line. In in vitro parameters, LP44 agonist administration-related decrease was observed. Our study showed that lung 5-HT7 receptor expression is increased in LPS-induced endotoxemia. All this data suggest that 5-HT7 receptor overexpression is an important protective mechanism during LPS-induced sepsis-related cell damage.     

Effects of Boron Supplementation on Peripartum Dairy Cows’ Health

In the present study, toxicity of commercial zinc oxide nanoparticles (ZnO NPs) was studied on the bacterium Pseudomonas sp., human promyelocytic leukemia (HL-60) cells, and peripheral blood mononuclear cells (PBMC). The toxicity was assessed by measuring growth, cell viability, and protein expression in bacterial cell. The bacterial growth and viability decreased with increasing concentrations of ZnO NP. Three major proteins, ribosomal protein L1 and L9 along with alkyl hydroperoxides reductase, were upregulated by 1.5-, 1.7-, and 2.0-fold, respectively, after ZnO NP exposure. The results indicated oxidative stress as the leading cause of toxic effect in bacteria. In HL-60 cells, cytotoxic and genotoxic effects along with antioxidant enzyme activity and reactive oxygen species (ROS) generation were studied upon ZnO NP treatment. ZnO NP exhibited dose-dependent increase in cell death after 24-h exposure. The DNA-damaging potential of ZnO NP in HL-60 cells was maximum at 0.05 mg/L concentration. Comet assay showed 70–80% increase in tail DNA at 0.025 to 0.05 mg/L ZnO NP concentration. A significant increase of 1.6-, 1.4-, and 2.0-fold in ROS level was observed after 12 h. Genotoxic potential of ZnO NPs was also demonstrated in PBMC through DNA fragmentation. Thus, ZnO NP, besides being an essential element having antibacterial activity, also showed toxicity towards human cells (HL-60 and PBMC).     

The effect of nanoparticle size on endocytosis dynamics depends on membrane–nanoparticle interaction

Molecular simulation studies on the interaction between nanoparticles (NPs) and cell membranes have been limited by small NP size of several nanometres. In this work, by using a simplified lipid model, we study the endocytosis of large NPs with a size being enlarged to 37.5 nm. It is found that the effect of NP size on endocytosis dynamics depends on the membrane–NP interaction. As the interaction strength between NP and lipid changes, different wrapping modes are observed. For the system with weak membrane–NP attraction, the wrapping process is controlled by the membrane bending, and thus large size of NPs (within the range of NP size we studied) would promote the wrapping dynamics. While for the case with strong membrane–NP adhesion, the wrapping process is dominated by lipid diffusion and small NPs show a larger wrapping rate. In this wrapping mode, the membrane–NP adhesion drives small NPs to move towards the membrane as the wrapping process proceeds. For relatively larger NPs, however, the membrane moves towards the NPs instead. We also find that for the second wrapping mode, the rapid wrapping rate, especially with the hydrophobic ligands on the hydrophilic NP would impose significant perturbations on membrane stability, and consequently, membrane pores may be induced during the process of NP endocytosis.     

Resolvin D2 Relieving Radicular Pain is Associated with Regulation of Inflammatory Mediators,Akt/GSK-3β Signal Pathway and GPR18

Neuroinflammation induced by protruded nucleus pulposus (NP) has been shown to play a significant role in facilitation of radicular pain. Resolvin D2 (RvD2), a novel member of resolvin family, exhibits potent anti-inflammatory, pro-resolving and antinociceptive effects. But the effect of RvD2 in radicular pain remains unknown. The radicular pain rat models were induced by application of NP to L5 dorsal root ganglion. Each animal received intrathecal injections of vehicle or RvD2 (10 ng µl^?1 or 100 ng µl^?1). Mechanical thresholds were determined by measuring the paw withdrawal threshold for 7 days. The expressions of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and transforming growth factor-β1 (TGF-β1) in ipsilateral lumbar segment of rat spinal dorsal horns were measured by using ELISA and real time-PCR. Western blot was used to measure the expressions of phosphorylated Akt (p-Akt) and phosphorylated glycogen synthase kinase 3 beta (p-GSK-3β). The expressions and distributions of RvD2 receptor, G-protein-coupled receptor 18 (GPR18), were also explored in the spinal cord of rats by using double-label immunofluorescence. RvD2 treatment caused significant reductions in the intensity of mechanical hypersensitivity and spinal expressions of TNF-α and IL-6. Meanwhile, RvD2 increased the expressions of TGF-β1 and regulated Akt/GSK-3β signaling. Furthermore, immunofluorescence showed that GPR18 colocalized with neurons and astrocytes in spinal cord. The results suggested that RvD2 might attenuate mechanical allodynia via regulating the expressions of inflammatory mediators and activation of Akt/GSK-3β signal pathway. RvD2 might offer a hopeful method for radicular pain therapy.     

Interaction of iron nanoparticles with nervous system: an in vitro study

Nanoparticles (NPs) are one of the interesting and widely studying issues mainly because of their particular physico-chemical features and broad applications in the field of biomedical sciences, such as diagnosis and drug delivery. In this study, the interaction of iron nanoparticles (Fe–NPs) with Tau protein and PC12 cell, as potential nervous system models, was investigated with a range of techniques including dynamic light scattering, intrinsic fluorescence spectroscopy, circular dichroism, [(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium-bromid] assay, and acridine orange/ethidium bromide (AO/EB) dual staining method. An inverse correlation between Stern and Volmer constant (K_SV) and temperature indicated a probable static quenching mechanism occurred between Tau protein and Fe–NPs. The number of binding site (n = 0.86) showed that there is almost one binding site of Fe–NP per protein. The negative values of ?H (?53.21 kJ/mol) and T?S (?42.44 kJ/mol) revealed that Fe–NPs interacts with Tau protein with dominate role of hydrogen bonds and van der Waals interactions and this interaction was spontaneous (?G = ?10.77 kJ/mol). Also, Fe–NPs stabilized the random coil structure of Tau protein. Moreover, Fe–NPs reduced PC12 cells viability by fragmentation of DNA in an apoptotic manner. In conclusion, induced conformational changes of Tau protein and cytotoxicity of PC12 cells by Fe–NP were revealed to be in a concentration and time-dependent manner.     

Synthesis of silver nanoparticles using plant derived 4-N-methyl benzoic acid and evaluation of antimicrobial,antioxidant and antitumor activity

The present study is to investigate the antitumor, antioxidant and antibacterial potential of silver nanoparticles (Ag NPs) synthesized from a phenolic derivative 4-N-methyl benzoic acid, isolated from a medicinal plant (Memecylon umbellatum Burm F). The Bio-inspired nanoparticles (NPs) were analyzed by using UV–vis spectroscopy, FTIR, HRTEM, Zeta potential and XRD techniques. The UV–vis spectroscopy study at the band of 430 nm confirmed the nanoparticles formation. HRTEM report showed that the AgNPs synthesized were in the size range 7–23 nm. The harvested nanoparticles were subjected to anti-bacterial assay and a dose dependent inhibitory action was observed against the tested human pathogens. Among the tested bacteria, Acinetobacter baumannii was found to be highly sensitive to AgNPs (diameter of zone of inhibition was 31 mm). Further, the silver nanoparticles exhibited a good anti-tumor activity against the breast cancer cell line (MCF 7) with an IC₅₀ value of 42.19 µg/mL. As the present study confirmed a good antibacterial, antioxidant and antitumor activity in the nanoparticles synthesized using 4-N-methyl benzoic acid derived from a medicinal plant, the product can be further tested to formulate a good lead compound for biomedical applications.     

Rice leaf extract synthesized silver nanoparticles: An in vitro fungicidal evaluation against Rhizoctonia solani,the causative agent of sheath blight disease in rice

Silver nanoparticles (Ag NP) were synthesized using rice leaf extract and optimized synthetic conditions were found to be 0.4 % leaf extract, 0.6 mM AgNO₃ and 30 min of autoclaving. Produced NP were characterized using UV–vis, DLS, zeta potential, XRD, TEM and FTIR. Ag NP formation was established from UV–vis spectra and NP showed zeta potential value of −27.4 mV. NP were spherical, polydisperse and average size was 16.5 ± 6.2 nm. Antifungal activity of Ag NP was assessed by poisoned food technique and resazurin broth dilution against mycelium and sclerotia of fungus R. solani, the causative agent of sheath blight disease in rice. Results confirmed effective hyphal growth inhibition and % growth inhibition was dose dependent (2.5–10 μg/mL). Ag NP showed enhanced mycelial inhibition (81.7–96.7 %) at 10 μg/mL. MIC values of Ag NP were in the range of 5–10 and 15–20 μg/mL towards fungal mycelium and sclerotia, respectively. Ag NP treatment (20 μg/mL) completely inhibited the disease incidence at 20 μg/mL. Ag NP treatment (10 μg/mL) caused 1.3 and 1.5 times enhancement in seedling vigor index. Hence, Ag NP can be utilized towards management and control of various fungal diseases of crops.     

Biogenic antimicrobial silver nanoparticles produced by fungi

Aspergillus tubingensis and Bionectria ochroleuca showed excellent extracellular ability to synthesize silver nanoparticles (Ag NP), spherical in shape and 35?±?10 nm in size. Ag NP were characterized by transmission electron microscopy, X-ray diffraction analysis, and photon correlation spectroscopy for particle size and zeta potential. Proteins present in the fungal filtrate and in Ag NP dispersion were analyzed by electrophoresis (sodium dodecyl sulfate polyacrylamide gel electrophoresis). Ag NP showed pronounced antifungal activity against Candida sp, frequently occurring in hospital infections, with minimal inhibitory concentration in the range of 0.11–1.75 μg/mL. Regarding antibacterial activity, nanoparticles produced by A. tubingensis were more effective compared to the other fungus, inhibiting 98.0 % of Pseudomonas. aeruginosa growth at 0.28 μg/mL. A. tubingensis synthesized Ag NP with surprisingly high and positive surface potential, differing greatly from all known fungi. These data open the possibility of obtaining biogenic Ag NP with positive surface potential and new applications.     

Cupric oxide nanoparticles‐enhanced chemiluminescence method for measurement of β‐lactam antibiotics

A simple, sensitive cupric oxide nanoparticles (CuO NPs) enhanced chemiluminescence (CL) method was developed for the measurement of β‐lactam antibiotics, including amoxicillin and cefazolin sodium. The method was based on suppression of the CuO NPs–luminol–H₂O₂ CL reaction by β‐lactam antibiotics. Experimental parameters that influenced the inhibitory effect of the antibiotic drugs on the CL system, such as NaOH (mol/L), luminol (µmol/L), H₂O₂ (mol/L) and CuO NPs (mg/L) concentrations, were optimized. Calibration graphs were linear and had dynamic ranges of 1.0 × 10^–6 to 8.0 × 10^–6 mol/L and 3.0 × 10^–5 to 5.0 × 10^–3 mol/L for amoxicillin and cefazolin sodium, respectively, with corresponding detection limits of 7.9 × 10^–7 mol/L and 1.8 × 10^–5 mol/L. The relative standard deviations of five replicate measurements of 5.0 × 10^–6 amoxicillin and 5 × 10^–4 cefazolin sodium were 5.43 and 5.01%, respectively. The synthesized CuO NPs were characterized by X‐ray diffraction (XRD) and transmission electronmicroscopy (TEM). The developed approach was exploited successfully to measure antibiotics in pharmaceutical preparations. Copyright © 2014 John Wiley & Sons, Ltd.     

Influence of SiO<Subscript>2</Subscript> Spacer Layer Thickness on Performance of Plasmonic Textured Silicon Solar Cell

We investigated the effect of SiO₂ spacer layer thickness between the textured silicon surface and silver nanoparticles (Ag NPs) on solar cell performance using quantum efficiency analysis. Separation of Ag NPs from high index silicon with SiO₂ layer led to modified absorption and scattering cross-sections due to graded refractive index medium. The forward scattering from Ag NPs is very sensitive to SiO₂ layer thickness in plasmonic silicon cell performance due to the evanescent character of generated near-fields around the NPs. With the optimized ～30–40 nm SiO₂ spacer layer, we observed an enhancement of solar cell efficiency from ～8.7 to ～10 %, which is due to the photocurrent enhancement in the off-resonance surface plasmon region. We also estimated minority carrier diffusion lengths (L _eff) from internal quantum efficiency data, which are also sensitive to SiO₂ spacer layer thickness. We observed that the L _eff values are enhanced from ～356 to ～420 μm after placing Ag NPs on ～40 nm spacer layer due to improved forward (angular) scattering of light from the Ag NPs into silicon.     

Biophysical and computational comparison on the binding affinity of three important nutrients to β-lactoglobulin: folic acid,ascorbic acid and vitamin K3

Small globular protein, β-lactoglobulin (βLG), which has significant affinity toward many drugs, is the most abundant whey protein in milk. In this study, the interaction of βLG with three important nutrients, ascorbic acid (ASC), folic acid (FOL), and vitamin K3 (VK3) was investigated by spectroscopic methods (UV–visible and fluorescence) along with molecular docking technique. The results of fluorescence measurements showed that studied nutrients strongly quenched βLG fluorescence in static (FOL and ACS) or static–dynamic combined quenching (VK3) mode. The values of binding constants (K_βLG-ASC ~ 4.34 × 10⁴ M^?1, K_βLG-FOL ~ 1.67 × 10⁴ M^?1and K_βLG-VK3 ~ 13.49 × 10⁴ M^?1 at 310 K) suggested that VK3 and FOL had stronger binding affinity toward βLG than ASC. Thermodynamic analysis indicated that hydrophobic interactions are the major forces in the stability of FOL–βLG complex with enthalpy- and entropy-driving mode while, hydrogen bonds and van der Waals interactions play a major role for βLG–ASC and βLG–VK3 associations. The results of 3D fluorescence FT-IR and UV–Visible measurements indicated that the binding of above nutrients to βLG may induce conformational and micro-environmental changes of protein. Also, there is a reciprocal complement between spectroscopic techniques and molecular docking modeling. The docking results indicate that the ASC, FOL, and VK3 bind to residues located in the subdomain B of βLG. Finally, this report suggests that βLG could be used as an effective carrier of above nutrients in functional foods.     

Evaluation of in vitro efficacy of docetaxel-loaded calcium carbonate aragonite nanoparticles (DTX-CaCO<Subscript>3</Subscript>NP) on 4T1 mouse breast cancer cell line

Cockle shell-derived calcium carbonate nanoparticles have shown promising potentials as slow drug-releasing compounds in cancer chemotherapy. In this study, we evaluated the in vitro efficacy of docetaxel (DTX)-loaded CaCO₃NP on 4T1 cell line. This was achieved by evaluating the following: cytotoxicity using MTT assay, fluorescence imaging, apoptosis with Annexin V assay, cell cycle analysis, scanning (SEM) and transmission electron microscopy (TEM), and scratch assay. Based on the results, DTX-CaCO₃NP with a DTX concentration of 0.5 μg/mL and above had comparable cytotoxic effects with free DTX at 24 h, while all concentrations had similar cytotoxic effect on 4T1 cells at 48 and 72 h. Fluorescence and apoptosis assay showed a higher (p < 0.05) number of apoptotic cells in both free DTX and DTX-CaCO₃NP groups. Cell cycle analysis showed cycle arrest at subG0 and G2/M phases in both treatment groups. SEM showed presence of cellular blebbing, while TEM showed nuclear fragmentation, apoptosis, and vacuolation in the treatment groups. Scratch assay showed lower (p < 0.05) closure in both free DTX and DTX-CaCO₃NP groups. The results from this study showed that DTX-CaCO₃NP has similar anticancer effects on 4T1 cells as free DTX, and since it has a slow release rate, it is a more preferred substitute for free DTX.     

Mn3O4 nanoparticles: Synthesis,characterization and their antimicrobial and anticancer activity against A549 and MCF-7 cell lines

Due to their inexpensive and eco-friendly nature, and existence of manganese in various oxidation states and their natural abundance have attained significant attention for the formation of Mn₃O₄ nanoparticles (Mn₃O₄ NPs). Herein, we report the preparation of Mn₃O₄ nanoparticles using manganese nitrate as a precursor material by utilization of a precipitation technique. The as-prepared Mn₃O₄ nanoparticles (Mn₃O₄ NPs) were characterized by using X-ray powder diffraction (XRD), UV–Visible spectroscopy (UV–Vis), High-Resolution Transmission electron microscopy (HRTEM), Field emission scanning electron microscopy (FESEM), Thermal gravimetric analysis (TGA) and Fourier-transform infrared spectroscopy (FT-IR). The antimicrobial properties of the as-synthesized Mn₃O₄ nanoparticles were investigated against numerous bacterial and fungal strains including S. aureus, E. coli, B. subtilis, P. aeruginosa, A. flavus and C. albicans. The Mn₃O₄ NPs inhibited the growth of S. aureus with a minimum inhibitory concentration (MIC) of 40 μg/ml and C. albicans with a MIC of 15 μg/ml. Furthermore, the Mn₃O₄ NPs anti-cancer activity was examined using MTT essay against A549 lung and MCF-7 breast cancer cell lines. The Mn₃O₄ NPs revealed significant activity against the examined cancer cell lines A549 and MCF-7. The IC₅₀ values of Mn₃O₄ NPs with A549 cell line was found at concentration of 98 µg/mL and MCF-7 cell line was found at concentration of 25 µg/mL.     

SKI knockdown suppresses apoptosis and extracellular matrix degradation of nucleus pulposus cells via inhibition of the Wnt/β-catenin pathway and ameliorates disc degeneration

This study aimed to determine the effects of SKI on interleukin (IL)-1β-induced apoptosis of nucleus pulposus (NP) cells, intervertebral disc degeneration (IDD), and the Wnt signaling pathway. NP tissue specimens of different Pfirrmann grades (II–V) were collected from patients with different grades of IDD. Real-time polymerase chain reaction and western blotting were used to compare SKI mRNA and protein expression in NP tissues from patients. Using the IL-1β-induced IDD model, NP cells were infected with lentivirus-coated si-SKI to downregulate the expression of SKI and treated with LiCl to evaluate the involvement of the Wnt/β-catenin signaling pathway. Western blotting, immunofluorescence, and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining were used to detect NP cell apoptosis, extracellular matrix (ECM) metabolism, and related protein expression changes in the Wnt/β-catenin signaling pathway. To investigate the role of SKI in vivo, a rat IDD model was established by needle puncture of the intervertebral disc. Rats were injected with lentivirus-coated si-SKI and evaluated by magnetic resonance imaging (MRI), and hematoxylin and eosin (HE) and safranin O staining. SKI expression positively correlated with the severity of human IDD. In the IL-1β-induced NP cell degeneration model, SKI expression increased significantly and reached a peak at 24 h. SKI knockdown protected against IL-1β-induced NP cell apoptosis and ECM degradation. LiCl treatment reversed the protective effects of si-SKI on NP cells. Furthermore, lentivirus-coated si-SKI injection partially reversed the NP tissue damage in the IDD model in vivo. SKI knockdown reduced NP cell apoptosis and ECM degradation by inhibiting the Wnt/β-catenin signaling pathway, ultimately protecting against IDD. Therefore, SKI may be an effective target for IDD treatment.

     

Effects of inorganic nanoparticles on viability and catabolic activities of Agrobacterium sp. PH-08 during biodegradation of dibenzofuran

This study investigated the cytotoxicity, genotoxicity, and growth inhibition effects of four different inorganic nanoparticles (NPs) such as aluminum (nAl), iron (nFe), nickel (nNi), and zinc (nZn) on a dibenzofuran (DF) degrading bacterium Agrobacterium sp. PH-08. NP (0–1,000 mg L^?1) -treated bacterial cells were assessed for cytotoxicity, genotoxicity, growth and biodegradation activities at biochemical and molecular levels. In an aqueous system, the bacterial cells treated with nAl, nZn and nNi at 500 mg L^?1 showed significant reduction in cell viability (30–93.6 %, p < 0.05), while nFe had no significant inhibition on bacterial cell viability. In the presence of nAl, nZn and nNi, the cells exhibited elevated levels of reactive oxygen species (ROS), DNA damage and cell death. Furthermore, NP exposure showed significant (p < 0.05) impairment in DF and catechol biodegradation activities. The reduction in DF biodegradation was ranged about 71.7–91.6 % with single NPs treatments while reached up to 96.3 % with a mixture of NPs. Molecular and biochemical investigations also clearly revealed that NP exposure drastically affected the catechol-2,3-dioxygenase activities and its gene (c23o) expression. However, no significant inhibition was observed in nFe treatment. The bacterial extracellular polymeric materials and by-products from DF degradation can be assumed as key factors in diminishing the toxic effects of NPs, especially for nFe. This study clearly demonstrates the impact of single and mixed NPs on the microbial catabolism of xenobiotic-degrading bacteria at biochemical and molecular levels. This is the first study on estimating the impact of mixed NPs on microbial biodegradation.     

Retention of titanium dioxide nanoparticles in biological activated carbon filters for drinking water and the impact on ammonia reduction

Given the increasing discoveries related to the eco-toxicity of titanium dioxide (TiO₂) nanoparticles (NPs) in different ecosystems and with respect to public health, it is important to understand their potential effects in drinking water treatment (DWT). The effects of TiO₂ NPs on ammonia reduction, ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) in biological activated carbon (BAC) filters for drinking water were investigated in static and dynamic states. In the static state, both the nitrification potential and AOB were significantly inhibited by 100 μg L^?1 TiO₂ NPs after 12 h (p < 0.05), and the threshold decreased to 10 μg L^?1 with prolonged exposure (36 h, p < 0.05). However, AOA were not considerably affected in any of the tested conditions (p > 0.05). In the dynamic state, different amounts of TiO₂ NP pulses were injected into three pilot-scale BAC filters. The decay of TiO₂ NPs in the BAC filters was very slow. Both titanium quantification and scanning electron microscope analysis confirmed the retention of TiO₂ NPs in the BAC filters after 134 days of operation. Furthermore, the TiO₂ NP pulses considerably reduced the performance of ammonia reduction. This study identified the retention of TiO₂ NPs in BAC filters and the negative effect on the ammonia reduction, suggesting a potential threat to DWT by TiO₂ NPs.     

Selenium (Na<Subscript>2</Subscript>SeO<Subscript>3</Subscript>) Upregulates Expression of Immune Genes and Blood–Testis Barrier Constituent Proteins of Bovine Sertoli Cell In Vitro

Sertoli cells were isolated from newborn calves and cultured in a medium supplemented with 0, 0.25, 0.50, 0.75, and 1.00 mg/L of sodium selenite to study their immune stimulatory effect, influence on cell’s viability, and expression of blood–testis barrier proteins (occludin, connexin-43, zonula occluden, E-cadherin) using quantitative PCR and western blot analyses. Results showed that medium supplemented with 0.50 mg/L of selenium significantly (P?<?0.05) promoted cell viability, upregulated toll-like receptor gene (TLR4), anti-inflammatory cytokines (IL-4, IL-10, TGFβ1), and expressions of blood–testis barrier proteins, and modulated expressions of pro-inflammatory cytokines (TNF-α, IL-1β, IFN-γ). Sertoli cells grown in culture medium supplemented with 0.25 mg/L of selenium significantly upregulated TLR4, IL-4, IL-10, TGFβ1, and blood–testis barrier proteins compared to the control group. Sodium selenite supplementation at 0.75 and 1.00 mg/L levels was cytotoxic and temporarily downregulated the expression of blood–testis barrier protein within 24 h after culture; however, commencing from 72 h post culture, increased cell viability and upregulation of expression of blood–testis barrier proteins were observed. In conclusion, the results of this study showed that selenium supplementation in the culture medium up to 0.50 mg/L concentration upregulates immune genes and blood–testis barrier constituent proteins of bovine Sertoli cells.