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.     

The Effect of Antimicrobial Peptide Temporin-Ra on Cell Viability and Gene Expression of Pro-inflammatory Factors in A549 Cell Line

Antimicrobial peptide Temporin-Ra was isolated from the skin secretions of marsh frog Rana ridibunda. Temporin-Ra was chemically synthesized and purified using RP-HPLC technique. The cytotoxicity of peptide on lung airway epithelial cell line (A549) and peripheral blood mononuclear cells (PBMC) was studied by MTT assay. Furthermore, the effect of Temporin-Ra on the expression of pro-inflammatory factors such as IL-1β and IL-8 in A549 cell line was evaluated at peptide concentrations of 15, 30 and 50 μg/mL for 6, 12 and 24 h using semi-quantitative RT-PCR and real-time PCR methods. The result of our experiments revealed that Temporin-Ra decreased cell viability about 3–13 % in A549 cells and 4–6 % in PBMC cells. Moreover, Temporin-Ra induced the mRNA expression of IL-1β and IL-8 genes in a dose- and time-dependent manner. According to our results, maximum IL-8 mRNA expression was observed after a 24-h treatment of cancer cells with 50 μg/mL peptide with approximately 18-fold increase in expression. The least expression level of IL-1β was observed after 6-h of incubation in the presence of 15 μg/mL peptide with 2.5-fold increase in expression whereas the most expression level was obtained following 24 h-treatment with 50 μg/mL peptide with 26-fold increase in mRNA expression. Eventually, the present study highlights the role of Temporin-Ra as a potent antimicrobial peptide in the activation and maintenance of inflammatory processes.     

Investigation of the antibacterial activity of a short cationic peptide against multidrug-resistant Klebsiella pneumoniae and Salmonella typhimurium strains and its cytotoxicity on eukaryotic cells

With the growing microbial resistance to conventional antimicrobial agents, the development of novel and alternative therapeutic strategies are vital. During recent years novel peptide antibiotics with broad spectrum activity against many Gram-positive and Gram-negative bacteria have been developed. In this study, antibacterial activity of CM11 peptide (WKLFKKILKVL-NH2), a short cecropin–melittin hybrid peptide, is evaluated against antibiotic-resistant strains of Klebsiella pneumoniae and Salmonella typhimurium as two important pathogenic bacteria. To appraise the antibacterial activity, minimal inhibitory concentration (MIC), minimal bactericidal concentration (MBC) and bactericidal killing assay were utilized with different concentrations (2–128 mg/L) of peptide. To evaluate cytotoxic effect of peptide, viability of RAJI, Hela, SP2/0, CHO, LNCAP cell lines and primary murine macrophage cells were also investigated with MTT assay in different concentrations (3–24 and 0.5–16 mg/L, respectively). MICs of K. pneumoniae and S. typhimurium isolates were in range of 8–16 and 4–16 mg/L, respectively. In bactericidal killing assay no colonies were observed at 2X MIC for K. pneumoniae and S. typhimurium isolates after 80–90 min, respectively. Despite the fact that CM11 reveals no significant cytotoxicity on RAJI, Hela, SP2/0, and CHO cell lines beneath 6 mg/L at first 24 and 48 h, the viability of LNCAP cells are about 50 % at 3 mg/L, which indicates strong cytotoxicity of the peptide. In addition, macrophage toxicity by MTT assay showed that LD₅₀ of CM11 peptide is 12 μM (16 mg/L) after 48 h while in this concentration after 24 h macrophage viability was about 70 %.     

Low-Dose Homocystine Enhances Proliferation and Migration of Bv2 Microglia Cells

Homocysteine (Hcy) is a non-essential amino acid that is derived from the breakdown of dietary methionine. Hyperhomocysteinemia (HHcy) is an independent risk factor for a variety of chronic diseases, especially neurodegenerative conditions. To better understand the role of HHcy in the pathogenesis of neurodegenerative disorders, we investigated the effect of Hcy on the proliferation and activation of microglia Bv2 cells. Cells were treated with six different Hcy concentrations: 0, 50, 100, 300, 500, and 1000 µM for different time periods (8, 12, 16, 24, and 48 h). The morphology of Bv2 cells was observed, and cell activity and proliferation were detected. Cell migration and secretion of pro-inflammatory cytokines were detected by the scratch wound assay, the transwell assay, and ELISA, respectively. The effect of Hcy on Bv2 proliferation occurred earlier (<24 h, especially 16 h) after treatment with concentrations between 100 and 300 μM, and there was no cytotoxicity to Bv2 cells. Meanwhile, functional assays suggested that Hcy not only promoted Bv2 secretion of the pro-inflammatory cytokines IL-1β, TNF-α, and IL-6, but also enhanced Bv2 migration and invasion, with 100 μM being the most effective concentration. In summary, Bv2 proliferation and activation can be promoted by short-term treatment with low-dose Hcy.     

Boron Affects Immune Function Through Modulation of Splenic T Lymphocyte Subsets,Cytokine Secretion,and Lymphocyte Proliferation and Apoptosis in Rats

This study demonstrated the mechanisms of boron effects in a rat model and provided a scientific basis for the rational of boron use. These findings were achieved by investigating the effects of boron (10, 20, 40, 80, 160, 320, and 640 mg/L in drinking water or 1.5, 3, 6, 12, 24, 48, and 96 mg/kg BW) on rat serum immunoglobulins (IgGs), splenic cytokines, lymphocyte subsets, as well as on lymphocyte proliferation and apoptosis. Addition of 20 (3) and 40 (6) mg/L (mg/kg BW) of boron to drinking water significantly increased rat serum IgG concentrations, splenic IFN-γ and IL-4 expression as well as the number of splenic CD3⁺, CD4⁺ and proliferating cell nuclear antigen (PCNA)⁺ cells. Supplementation of drinking water with 40 mg/L (6 mg/kg BW) boron also markedly increased splenic IL-2 expression and the CD4⁺/CD8⁺ cell ratio and reduced splenic CD8⁺ cell number. Supplementation with 80 mg/L (12 mg/kg BW) boron significantly increased CD3⁺ and PCNA⁺ cell numbers (P < 0.05) and decreased the IL-10 expression in the spleen. Addition of 320 (48) and 640 (96) mg/L (mg/kg BW) boron markedly reduced the serum IgG concentrations; splenic IL-2 and IL-10 expression; the number of CD3⁺, CD4⁺ and PCNA⁺ cells; and increased the number of splenic CD8⁺ and caspase-3⁺ cells and promoted caspase-3 expression in CD3⁺ cells. In conclusion, these findings suggest that the supplementation of rat drinking water with 20(3) and 40(6) mg/L (mg/kg BW) boron can markedly enhance humoral and cellular immune functions, while boron concentrations above 320 mg/L (48 mg/kg BW) can have an inhibitory effect or even toxicity on immune functions. These results exhibit a U-shaped response characteristic of low and high doses of boron supplementation on immune function and imply that proper boron supplementation in food for humans and animals could be used as an immunity regulator.     

Simultaneous application of cisplatin and static magnetic field enhances oxidative stress in HeLa cell line

In this study, we reported the effects of simultaneous application of static magnetic field (SMF) and cisplatin as an anticancer drug on the oxidative stress in human cervical cancer (HeLa) cell line and normal skin fibroblast cells (Hu02). The cells were exposed to different SMF intensities (7, 10, and 15 mT) for 24 and 48 h. IC₅₀ concentrations of cisplatin were obtained by MTT assay. The cytotoxic effects of combined treatment were studied by measuring the intracellular reactive oxygen species content using flow cytometric method and estimation of membrane lipid peroxidation by spectrophotometry. Statistical analysis was assessed using one-way repeated measures analysis of variance (ANOVA) followed by Tukey’s test. Based on the obtained results, the highest and lowest death rate, respectively, in HeLa and Hu02 cell lines was observed at the intensity of 10 mT. Also, we found that membrane lipid peroxidation in cancer cells is higher than that of normal counterparts. SMF potently sensitized human cervical cancer cells to cisplatin through reactive oxygen species (ROS) accumulation while it had small effects on normal cells. The combination of both treatments for 48 h led to a marked decrease in the viability percentage of HeLa cells by about 89% compared to untreated cells. This study suggests that conjugation of both physical and chemical treatments could increase the oxidative stress in HeLa cell line and among three optional intensities of SMF, the intensity of 10 mT led to the higher damage to cancer cells in lower doses of drug.     

Total Glucosides of Peony Attenuates 2,4,6-Trinitrobenzene Sulfonic Acid/Ethanol-Induced Colitis in Rats Through Adjustment of Th1/Th2 Cytokines Polarization

The present study is to investigate effects of total glucosides of peony (TGP) on 2,4,6-trinitrobenzene sulfonic acid (TNBS)/ethanol-induced colitis in rats and to explore potential clinical use of TGP for treatment of inflammatory bowel disease. Sixty Sprague–Dawley rats were randomly grouped into normal controls, model controls, sulfasalazine (SASP) controls (100 mg/kg/day), and low, medium, and high-dose TGP groups (25, 50, and 100 mg/kg/day, respectively). 24 h following colonic instillation of TNBS, TGP, and SASP were given by gastric gavage three times a day for 7 days. Disease activity index (DAI), colon macroscopic damage index (CMDI), histopathological score (HPS), and myeloperoxidase (MPO) activity were evaluated. Levels of serum TNF-α, IL-1β, and IL-10 were measured by ELISA, and expression of TNF-α, IL-1β, and IL-10 mRNA and protein in colonic tissues was detected by RT-PCR and western blot, respectively. Compared with rats in the model controls, TGP (50 or 100 mg/kg/day)-treated rats with TNBS/ethanol-induced colitis showed significant improvements of DAI, CMDI, HPS, and MPO activity. Moreover, administration of TGP (50 or 100 mg/kg/day) decreased the up-regulated levels of serum TNF-α and IL-1β, and expression of TNF-α and IL-1β mRNA and protein in colonic tissues, and increased the serum IL-10 and colonic IL-10 mRNA and protein level. And there was no significant difference compared with administration of SASP (P > 0.05). TGP attenuates TNBS/ethanol-induced colitis in rats and its efficacy is similar to SASP, the potential mechanism might be related to the adjustment of Th1/Th2 cytokines polarization by decreasing pro-inflammatory cytokine TNF-α and IL-1β, and increasing anti-inflammatory cytokine IL-10.     

Ginkgolides and bilobalide protect BV2 microglia cells against OGD/reoxygenation injury by inhibiting TLR2/4 signaling pathways

Ginkgolide and bilobalide are major trilactone constituent of Ginkgo biloba leaves and have been shown to exert powerful neuroprotective properties. The aims of this study were to observe the inhibitory effects of ginkgolide and bilobalide on the activation of microglial cells induced by oxygen–glucose deprivation and reoxygenation (OGD/R) and the specific mechanisms by which these effects are mediated. For detecting whether ginkgolide and bilobalide increased cell viability in a dose-dependent manner, BV2 cells were subjected to oxygen–glucose deprivation for 4 h followed by 3 h reoxygenation with various concentrations of drugs (6.25, 12.5, 25, 50, and 100 μg/ml). The extent of apoptosis effect of OGD/R with or without ginkgolide and bilobalide treatment were also measured by Annexin V-FITC/PI staining. Similarly, the levels of pro-inflammatory cytokines TNF-α, IL-1β, IL-6, IL-8, and IL-10 were detected using a specific Bio-Plex Pro? Reagent Kit. The effects of ginkgolide and bilobalide on protein levels of TLR2/4, MyD88, p-TAK1, p-IKKβ, p-IkBα, NF-κB p65, Bcl-2, Bax, Bak, RIP3, cleaved-Caspase-3, cleaved PARP-1 and cellular localization of NF-κB p65 were evaluated by Western blot and double-labeled immunofluorescence staining, respectively. OGD/R significantly decreased the cell viability and increased the release of IL-1β, IL-6, IL-8, IL-10, TNF-α in BV2 microglia cells; these effects were suppressed by ginkgolide and bilobalide. Meanwhile, ginkgolide and bilobalide also attenuated the OGD/R-induced increases in TLR2, TLR4, MyD88, Bak, RIP3 levels and reversed cleaved caspase-3/caspase-3, Bax/Bcl-2 and cleaved PARP-1/PARP-1 ratio. Furthermore, ginkgolide and bilobalide also downregulated p-TAK1, p-IkBα, and p-IKKβ and inhibited the OGD/R-induced transfer of NF-κB p65 from cytoplasm to nucleus in BV2 microglia cells. The results showed that ginkgolide and bilobalide can inhibit OGD/R-induced production of inflammatory factors in BV2 microglia cells by regulating the TLRs/MyD88/NF-κB signaling pathways and attenuating inflammatory response. The possible mechanism of anti-inflammatory and neuroprotective effects of ginkgolides results from the synergistic reaction among each monomer constituents.     

Enteroaggregative Escherichia coli infection induces IL-8 production via activation of mitogen-activated protein kinases and the transcription factors NF-κB and AP-1 in INT-407 cells

Multiple mucosal immune factors, such as TNF-α and IL-1β, are thought to be key mediators involved in inflammatory bowel disease. We evaluated the role of the pro-inflammatory cytokine TNF-α on nitric oxide synthase (NOS) expression in indomethacin-induced jejunoileitis in rats. Jejunoileitis was induced in rats with subcutaneous injections of indomethacin (7.5 mg/kg) 24 h apart for two consecutive days, and animals were randomized into four groups. Group 1 received only indomethacin. Group 2 was treated with a daily dose of phosphodiesterase (PDE) inhibitor (theophylline or pentoxifylline) by oral gavage for 2 days before and 4 days after indomethacin. Group 3 received a single dose of anti-TNF-α monoclonal antibody (TNF-Ab, IP) 30 min before indomethacin. Group 4 was treated with 1 h hyperbaric oxygenation (HBO₂) for 5 days after indomethacin. Rats were sacrificed at 12 h or 4 days after final indomethacin injection. PDE inhibitor, TNF-Ab, or HBO₂ treatment significantly decreased indomethacin-induced ulceration, myeloperoxidase activity, and disease activity index. Although indomethacin significantly increased serum TNF-α and nitrate/nitrite (NOx) concentrations above control values at 12 h, inducible NOS (iNOS) expression was detected only at day 4. Serum IL-1β levels did not change at 12 h but increased 4-fold after 4 days. Indomethacin had no effect on constitutive NOS. Treatment with PDE inhibitor, TNF-Ab, or HBO₂ significantly reduced serum/tissue TNF-α, IL-1β, NOx, and iNOS expression. Our data show TNF-α plays an early pro-inflammatory role in indomethacin-induced jejunoileitis. Additionally, down-regulation of NOx by PDE inhibitors, TNF-Ab, or HBO₂ suggests that TNF-α modulates iNOS expression.     

Effect of <Emphasis Type="Italic">Lactobacillus reuteri</Emphasis> on Cell Viability and PGE<Subscript>2</Subscript> Production in Human Gingival Fibroblasts

Emerging evidence suggests that probiotic therapy can play a role in the prevention and management of oral inflammatory diseases through immunomodulation and down-regulation of the inflammatory cascade. The aim of this in vitro study was to investigate the viability of human gingival fibroblasts (HGF) and its production of prostaglandin E₂ (PGE₂), when exposed to supernatants of two mixed Lactobacillus reuteri strains (ATCC PTA 5289 and DSM 17938). The experiments were conducted in the presence and absence of the pro-inflammatory cytokine IL-1β. L. reuteri strains were grown and the bacterial supernatant was collected. The cell-free supernatant was diluted to concentrations equivalent to the ones produced by 0.5 to 5.0 × 10⁷ CFU/mL bacteria. Cell viability was assessed with the MTT colorimetric assay and the amount of PGE₂ in the cell culture medium was determined using the monoclonal enzyme immune assay kits. Our findings showed that none of the L. reuteri supernatants were cytotoxic or affected the viability of HGF. The most concentrated bacterial supernatant stimulated the production of PGE₂ by the gingival cells in a significant way in the presence of IL-1β (p < 0.05), suggesting that bacterial products secreted from L. reuteri might play a role in the resolution of inflammation in HGF. Thus, our findings justify further investigations on the influence of probiotic bacteria on gingival inflammatory reactions.     

Anti-Inflammatory Effects of Ginsenoside-Rh2 Inhibits LPS-Induced Activation of Microglia and Overproduction of Inflammatory Mediators Via Modulation of TGF-β1/Smad Pathway

Microglia activation plays an important role in neuroinflammation and contributes to several neurological disorders. Hence, inhibition of both microglia activation and pro-inflammatory cytokines may lead to the effective treatment of neurodegenerative diseases. In this study, we found that GRh2 inhibited the inflammatory response to lipopolysaccharide (LPS) and prevented the LPS-induced neurotoxicity in microglia cells. GRh2 significantly decreased the generation of nitric oxide production, and tumor necrosis factor-α, interleukin (IL)-6, IL-1β, cyclooxygenase-2 and inducible nitric oxide synthase in LPS-induced activated microglia cells. Furthermore, GRh2 (20 and 50 μM) significantly increased TGF-β1 expression and reduced the expression of Smad. These results suggest that GRh2 effectively inhibits microglia activation and production of pro-inflammatory cytokines via modulating the TGF-β1/Smad pathway.     

Sorafenib Inhibition of Hepatic Stellate Cell Proliferation in Tumor Microenvironment of Hepatocellular Carcinoma: A Study of the Sorafenib Mechanisms

We investigated the mechanism and effects of sorafenib on hepatic stellate cell (HSC) viability and in the liver tumor microenvironment. The expression of α-smooth muscle actin (α-SMA) was measured immunocytochemically in the LX2 cells treated with differing concentrations of sorafenib. Changes in the platelet-derived growth factor (PDGF)-BB and tumor growth factor (TGF)-β1 concentrations were detected in the LX2 supernatant using an enzyme-linked immunosorbent assay (ELISA). Expressions of the extracellular signal-regulated kinase 1 (ERK₁), ERK₂, and Akt signaling pathways were measured using a western blot assay. The LX2 cells were cocultured with HepG2 cells for 24 h to observe their effects on HepG2 cell invasive ability. (1) After treatment with various concentrations of sorafenib for 12, 24, 36, or 48 h, MTT assay showed that the viability of the treated LX2 cells was lower than in the controls. (2) As sorafenib concentration and time of exposure increased, α-SMA expression became weaker in the treated cells. (3) The PDGF-BB and TGF-β1 concentrations decreased with higher concentration, and longer exposures under the same sorafenib concentration. (4) The ERK₁, ERK₂, and Akt expressions were identical between the treated and the control groups, but their phosphorylated expression decreased with increased concentrations of sorafenib. (5) The invasive ability of the HepG2 cells induced by the LX2 gradually decreased as sorafenib concentrations increased. Sorafenib suppressed α-SMA expression, inhibited PDGF-dependent signaling pathways in HSCs, downregulated the PDGF-BB and TGF-β1 expression in the HSCs supernatant, and restrained viability of the HSCs, resulting in suppressed proliferation and invasion in the HepG2 cells.     

Gastrodin Ameliorates Depressive-Like Behaviors and Up-Regulates the Expression of BDNF in the Hippocampus and Hippocampal-Derived Astrocyte of Rats

Gastrodin (GAS), a main constituent of a Chinese herbal medicine Tian ma, has been shown to be effective in treating various mood disorders. The purpose of the present study was to assess the effects of GAS on alleviating depressive-like behaviors in a rat model of chronic unpredictable stress (CUS) and regulating the expression of BDNF in the hippocampus and hippocampal-derived astrocyte from Sprague–Dawley (SD) rats. Following CUS, rats were intraperitoneally administered gastrodin (50, 100, or 200 mg/kg daily) or vehicle for 2 weeks. Rats were then experienced sucrose preference test and forced swim test. The expressions of GFAP and BDNF in the hippocampus were evaluated. In addition, hippocampal astrocytes were isolated from neonatal SD rats and exposed to different concentrations of GAS (sham, 5, 10, 20, 50 and 100 μg/mL) for 48 and 72 h before the cell viability and the levels of pERK1/2 and BDNF were analyzed. Furthermore, the cell viability was also tested after exposure to serum-free condition that contain different concentrations of GAS for 48 and 72 h. GAS administration (100 and 200 mg/kg daily) reversed depressive-like behaviors in rats exposed to CUS paradigm and restored the expression of GFAP and BDNF in the hippocampus. Moreover, in vitro experiments revealed that GAS did not increase the cell viability of astrocytes but protected it from 72 h’s serum-free damage at the dosage 20 μg/mL. Increased levels of ERK1/2 phosphorylation and BDNF protein were also observed after GAS (20 μg/mL) treatment for 72 h. These results indicate that gastrodin possesses antidepressant effect. The changes of the astrocyte activation and the level of BDNF may play a critical role in the pharmacological action of GAS.     

Protective effects of calcitonin on IL-1 stimulated chondrocytes by regulating MMPs/TIMP-1 ratio via suppression of p50-NF-κB pathway

The aim of this study was to investigate the effects and underlying mechanisms of calcitonin (CT) on interleukin 1 beta (IL-1β) stimulated human chondrocytes. IL-1β (5 ng/mL) was added into chondrocytes to establish osteoarthritis (OA) model in vitro. Different concentrations of CT (0.1, 0.5, 1, 5, 10 and 50 nM) were used for treating IL-1β stimulated chondrocytes. Cell viability of chondrocytes was measured by cell counting kit-8 (CCK8) method. Western blotting was performed to evaluate the expression of matrix metalloproteinases (MMP-13), tissue inhibitor of metalloproteinases 1 (TIMP-1), p50 and p38. CT inhibited MMP-13 expression and promoted TIMP-1 expression in the IL-1β stimulated human chondrocytes. The CT-mediated alteration of MMP-13/TIMP-1 ratio was partially attributed to the inactivation of the p50- nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway by suppressing p50 in IL-1β stimulated chondrocytes. CT might play a protective role in IL-1β stimulated OA model via p50-NF-κB pathway.

Abbreviations: CT: calcitonin; IL-1β: interleukin-1β; MMP-13: matrix metalloproteinases-13; TIMP-1: tissue inhibitors of metalloproteinases-1.     

黄曲霉素对裸鼹鼠肺泡上皮Ⅱ型细胞基因表达的影响

目的:观察不同浓度黄曲霉素对裸鼹鼠肺泡上皮Ⅱ型细胞(AEC II)相关因子m RNA表达的影响,探讨裸鼹鼠抵御肺癌的分子机制。方法:通过酶消化法分离裸鼹鼠肺组织细胞并用免疫黏附法进行纯化得到裸鼹鼠肺泡上皮Ⅱ型细胞进行原代培养,40小时后,实验组分别给予不同浓度(0.25、0.5、1.0、2.0和4.0 mg/L)的黄曲霉素处理,溶剂对照组给予DMSO(0.4 m L/L)处理。黄曲霉素作用48小时后收集细胞,用实时定量PCR技术检测裸鼹鼠AECII细胞中的SP-C基因及TNF-α、IL-1、IL-6、IL-8、IL-12等炎症因子的m RNA表达变化。结果:原代分离的裸鼹鼠AEC II细胞纯度70%,活性90%,可用于体外实验。定量PCR结果显示黄曲霉素使裸鼹鼠AEC II细胞SP-C表达水平显著下降,TNF-α、IL-1、IL-6、IL-8、IL-12等炎症因子的表达水平在黄曲霉低于2 mg/L浓度的条件下没有显著变化。结论:裸鼹鼠AEC II细胞在黄曲霉素导致细胞受损的情况下,仍然保持较低水平的炎症因子表达,这可能是其抵抗肺癌的机制之一。     

Hydrogen sulfide delays nicotinamide-induced premature senescence via upregulation of SIRT1 in human umbilical vein endothelial cells

The present study was designed to investigate the effect of hydrogen sulfide on cellular senescence of human umbilical vascular endothelial cells (HUVECs CC-2517) and its underlying mechanism. The premature senescence-like phenotype HUVECs (the fourth passage) was induced by treatment with nicotinamide (NAM, an inhibitor of SIRT1, 5 mmol/L, 12 h). Cells were cultured with sodium hydrosulfide (NaHS, 12.5, 25, 50 and 100 μmol/L) for 48 h in premature senescence-like phenotype HUVECs. The fourth passage of HUVECs was considered as young group. Senescence-associated (SA)-β-galactosidase activities were detected to evaluate cell senescence, and the expression of SA heterochromatin foci (SAHF) was visualized by DAPI DNA staining. The mRNA and protein levels of SIRT1 were detected using RT-PCR and western blotting analysis, respectively. The results showed that β-galactosidase positive cells and the formation of SAHF were markedly increased after treatment with NAM (5 mmol/L) for 12 h. We also found that NaHS (12.5 μmol/L) had no effect on the percentage of SA β-gal positive cells and the expression of SAHF, and the hallmarks decreased at the concentration of 25 and 50 μmol/L, reaching the minimum at 50 μmol/L, while the percentage of SA β-gal positive cells and the expression of SAHF increased at the concentration of 100 μmol/L. Furthermore, we found that both on protein and mRNA levels of SIRT1 in the Y+N+S50 group was significantly increased compared with that in Y+N group. In conclusion, NaHS delays senescence of HUVECs induced by NAM via upregulation of SIRT1 expression.     

Evaluating cytotoxic effect of nanoliposomes encapsulated with umbelliprenin on 4T1 cell line

Cytotoxicity of umbelliprenin has been found in various cancer cell lines such as, prostate, breast, CLL, and skin. Encapsulating chemotherapeutic agents with nanoliposomes have been resulted in improved cytotoxicity effects than their free forms. However, whether nanoliposomal form of umbelliprenin could have higher cytotoxic effect than free umbelliprenin is not clarified yet. After synthesizing umbelliprenin, different concentrations (3, 6, 12, 25, 50, 100, 200 μg/ml) applied on the mouse mammary carcinoma cell line (4T1) for 24, 48, and 72 h at 37°C. Afterwards, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay was performed to analyze cytotoxicity. MTT assay results showed that IC₅₀ of umbelliprenin in dimethyl sulfoxide (DMSO) (30.92, 30.64, and 62.23 for 24, 48, 72 h incubation, respectively) decreased (5.8, 5.0, 3.5 for 24, 48, 72 h incubation, respectively) when encapsulated with nanoliposomes. Nanoliposomal umbelliprenin cytotoxicity affected cell viability in concentration and time-dependent manner. Our study recommended nanoliposomal umbelliprenin as the most effective chemotherapeutic agent against the mouse mammary carcinoma cell line viability. Future in vivo studies and clinical trials are needed.