[Effects of pre-electroacupuncture on blood pressure and cardiac function in high-salt-induced hypertension rats].

OBJECTIVE: To observe the effects of pre-electroacupuncture at "Taichong"(LR3), "Neiguan"(PC6) and "Waiguan"(TE5) on blood pressure and cardiac function of high-salt-induced hypertension rats, so as to explore the possible mechanism of pre-electroacupuncture in improving hypertension. METHODS: Twenty-four SD rats were randomly divided into control group, high-salt group and pre-electroacupuncture group, with 8 rats in each group. The hypertension model was established by feeding high-salt diet for 7 weeks. In the pre-electroacupuncture group, rats received electroacupuncture intervention at bilate-ral LR3, PC6 and TE5 (2 Hz/15 Hz, 2 mA) for 30 min, once a day, from the first day of modeling, for a total of 7 weeks. The blood pressure of rats was monitored by caudal artery noninvasive blood pressure measurement technique before and at the 1st, 3rd, 5th and 7th week of modeling. At the 8th week of the experiment, left ventricular catheterization was performed and biological signal acquisition system was used to detect left ventricular hemodynamics indexes and analyze left ventricular function, the car-diac mass ratio was measured to evaluate the degree of myocardial hypertrophy. The mRNA expressions of atrial natriuretic peptide(ANP), myosin heavy chain 7(MYH7), α-smooth muscle actin(α-SMA), interleukin(IL)-1ß, and IL-6 of myocardial tissues were detected by quantitative real-time PCR. Sirius red staining was used to observe the degree of myocardial fibrosis. RESULTS: Compared with the control group, systolic blood pressure, diastolic blood pressure, mean arterial pressure, left ventricular end-diastolic pressure (LVEDP), cardiac mass ratio，and the mRNA expressions of ANP, MYH7, α-SMA, IL-1ß, and IL-6, and sirius red staining area of myocardium were all significantly increased(P<0.01,P<0.05)，maximal rate of rise and descent of left ventricular pressure(LVP±dP/dtmax) were decreased (P<0.05,P<0.01) in the high-salt group. Compared with the high-salt group, rats in the pre-electroacupuncture group had lower systolic blood pressure, diastolic blood pressure, mean arterial pressure, LVEDP，cardiac mass ratio，higher LVP±dP/dtmax,down-regulated mRNA expressions of ANP, MYH7, α-SMA, IL-1ß, IL-6, and smaller area of sirius red staining(P<0.05, P<0.01). CONCLUSION: Pre-electroacupuncture tends to lower blood pressure, improve cardiac function and reduce myocardial fibrosis in high-salt-induced hypertension rats, which may be associated with inhibiting inflammatory response.

The Effect of Nursing Management of Patients Undergoing Interventional Therapy for Liver Cancer Compared with Standard Care on Patient-Reported Outcomes.

BACKGROUND: To investigate the efficacy of individualized symptom management based on patients' self-reports during interventional therapy (IT) for liver cancer. METHODS: Patients with liver cancer who recieved IT from April to August 2019 were assigned to either the intervention (n=70) or control group (n=70). The control group received routine nursing care and the intervention group received a nursing management program. The severity of specific symptoms, as measured by the Karnofsky Performance Scale (KPS), and satisfaction with nursing care, were analyzed. RESULTS: Compared to the control group, patients given individualized management experienced significantly less severe pain, nausea, anxiety, and fatigue (p < .05). The scores for KPS and satisfaction with care were both significantly improved in the intervention group than in the control group (p < .05). CONCLUSION: This high-quality nursing management program predicated on patients' self-reports is worthy of clinical application and popular adoption.

Single-Cell Isotope Dilution Analysis with LA-ICP-MS: A New Approach for Quantification of Nanoparticles in Single Cells.

Laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) is an emerging method for the analysis of metal nanoparticles (NPs) in single cells. However, two main obstacles, low analytical throughput and lack of commercial reference materials, need to be overcome. In this work, we demonstrated the principles of a new approach termed "single-cell isotope dilution analysis" (SCIDA) to remove the two obstacles. For a proof of concept, macrophage cells were chosen as a model to study the uptake of silver NPs (AgNPs) at a single-cell level. Single cells exposed to AgNPs were placed in an array by a microfluidic technique; each cell in the array was precisely dispensed with a known picoliter droplet of an enriched isotope solution with a commercial inkjet printer; accurate quantification of AgNPs in single cells was done by using isotope dilution LA-ICP-MS. The average Ag mass of 1100 single cells, 396 ± 219 fg Ag per cell, was in good accord with the average of the population of cells determined by solution ICP-MS analysis. The detection limit was 0.2 fg Ag per cell. The SCIDA approach is expected to be widely applied for the study of cell-NP interactions and biological effects of NPs at the single-cell level.

Antitumor Activity and Mechanism Study of Riluzole and Its Derivatives.

To explore novel antitumor agents with high efficiency and low toxicity, riluzole alkyl derivatives (4a-4i) were synthesized. Their anti-proliferative activities against HeLa, HepG2, SP2/0, and MCF-7 cancer cell lines were assessed by the CCK-8 assay and compared with human normal liver (LO2) cells. Most of them showed potent cytotoxic effects against four human tumor cell lines and low toxic to LO2 cells. In particular, 2-(N-ethylamine)-6-trifluoromethoxy- benzothiazole (4a) showed a IC50 value of 7.76 µmol/L in HeLa cells and was found to be nontoxic to LO2 cells up to 65 µmol/L. Furthermore, flow cytometry indicated that 4a could induce remarkable early apoptosis and G2/M cell cycle arrest in HeLa cells. It also impaired the migration ability of HeLa cells in wound healing assays. Western blot results demonstrated that 4a suppressed Bcl-2 protein expression but increased the level of Bax in HeLa cells, and elevated the Bax/Bcl-2 expression ratio. These new findings suggest that 4a exhibited beneficially anti-cervical cancer effect on HeLa cells by inducing HeLa cell apoptosis.

NLRP3 in human glioma is correlated with increased WHO grade, and regulates cellular proliferation, apoptosis and metastasis via epithelial-mesenchymal transition and the PTEN/AKT signaling pathway.

Glioma is the most prevalent and fatal primary tumor of the central nervous system in adults, while the development of effective therapeutic strategies in clinical practice remain a challenge. Nucleotide-binding domain leucine-rich family pyrin-containing 3 (NLRP3) has been reported to be associated with tumorigenesis and progression; however, its expression and function in human glioma remain unclear. The present study was designed to explore the biological role and potential mechanism of NLRP3 in human glioma. The results demonstrated that overexpression of NLRP3, apoptosis-associated speck-like protein containing a caspase-recruitment domain (ASC), caspase­1 and interleukin (IL)­1ß protein in human glioma tissues were significantly correlated with higher World Health Organization grades. The in vitro biological experiments demonstrated that NLRP3 downregulation significantly inhibited the proliferation, migration and invasion, and promoted the apoptosis of SHG44 and A172 glioma cell lines. Furthermore, western blot assays revealed that the downregulation of NLRP3 significantly reduced the expression of ASC, caspase­1 and IL­1ß protein. Furthermore, NLRP3 knockdown caused the inhibition of epithelial-mesenchymal transition (EMT), and inhibited the phosphorylation of AKT serine/threonine kinase (AKT) and phosphorylation of phosphatase and tensin homolog (PTEN). Consistently, the upregulation of NLRP3 significantly increased the expression of ASC, caspase­1, IL­1ß and phosphorylated-PTEN, promoted proliferation, migration, invasion and EMT, inhibited apoptosis, and activated the AKT signaling pathway. The data of the present study indicate that NLRP3 affects human glioma progression and metastasis through multiple pathways, including EMT and PTEN/AKT signaling pathway regulation, enhanced inflammasome activation, and undefined inflammasome-independent mechanisms. Understanding the biological effects of NLRP3 in human glioma and the underlying mechanisms may offer novel insights for the development of glioma clinical therapeutic strategies.

AP-1 confers resistance to anti-cancer therapy by activating XIAP.

The underlying cause of treatment failure in many cancer patients is intrinsic and acquired resistance to chemotherapy. Recently, histone deacetylase (HDAC) inhibitors have developed into a promising cancer treatment. However, resistance mechanism induced by HDAC inhibitors remains largely unknown. Here we report that a HDAC inhibitor, JNJ-2648158 induced transcription of XIAP by activating AP-1 expression, which conferring resistance to chemotherapeutics. Our results showed that high expression of c-Fos caused by HDAC inhibitor promoted AP-1 formation during acquired resistance towards chemo-drugs, indicating an extremely poor clinical outcome in breast cancers and liver cancers. Our study reveals a novel regulatory mechanism towards chemo-drug resistance, and suggests that XIAP may serve as a potential therapeutic target in those chemo-resistant cancer cells.

Empirically derived dietary patterns and ovarian cancer risk: a meta-analysis.

Dietary patterns, which reflect overall diet and possible nutrient and food interactions, have been reported to be related to ovarian cancer (OC) risk. However, studies on the relationship between dietary patterns and OC risk have been inconsistent. Thus, we carried out a systematic meta-analysis to assess the relationship between dietary patterns and the risk of OC. Relevant studies are identified by searching the Medline and Embase electronic databases up to December 2016. The Cochrane Q statistic and the I statistical were used to evaluate heterogeneity. A total of 22 studies fulfilled the inclusion criteria and were included in this meta-analysis. There was evidence of a decreased risk for OC in the highest versus the lowest categories of healthy dietary pattern [odds ratio (OR)=0.86; 95% confidence interval (CI): 0.74-0.99; P=0.04]. An increased risk of OC was shown for the highest versus the lowest category of a western-style dietary pattern (OR=1.19; 95% CI: 1.01-1.41; P=0.04). No significant association with OC risk was observed in the highest versus the lowest category of a heavy drinking pattern (OR=0.89; 95% CI: 0.67-1.19; P=0.42). The results of this meta-analysis suggest that a healthy dietary pattern is associated with reduced risk for OC and a western-style dietary pattern is associated with an increased risk of OC. Further studies are needed to confirm our results.

Immunization with alkyl hydroperoxide reductase subunit C reduces Fusobacterium nucleatum load in the intestinal tract.

Fusobacterium nucleatum (Fn) is an important tumour-associated bacterium in colorectal cancer (CRC). The antioxidant protein alkyl hydroperoxide reductase subunit C (AhpC) can induce strong antibacterial immune response during various pathogen infections. Our study aimed to evaluate the efficacy of Fn-AhpC as a candidate vaccine. In this work, by western blot analysis, we showed that Fn-AhpC recombinant protein could be recognized specifically by antibodies present in the sera of CRC patients; using the mouse Fn-infection model, we observed that systemic prophylactic immunization with AhpC/alum conferred significant protection against infection in 77.3% of mice. In addition, we measured the anti-AhpC antibody level in the sera of CRC patients and found that there was no obvious increase of anti-AhpC antibodies in the early-stage CRC group. Furthermore, we treated Fn with the sera from both immunized mice and CRC patients and found that sera with high anti-AhpC antibodies titre could inhibit Fn growth. In conclusion, our findings support the use of AhpC as a potential vaccine candidate against inhabitation or infection of Fn in the intestinal tract, which could provide a practical strategy for the prevention of CRC associated with Fn infection.

Epidermal growth factor and tumor necrosis factor α cooperatively promote the motility of hepatocellular carcinoma cell lines via synergistic induction of fibronectin by NF-κB/p65.

BACKGROUND: The interaction between hepatocellular carcinoma (HCC) cells and their microenvironment plays a fundamental role in tumor metastasis. The HCC microenvironment is rich in epidermal growth factor (EGF) and tumor necrosis factor α (TNFα), which may cooperatively, rather than individually, interact with tumor cells to influence their biological behavior. METHODS: Immunohistochemistry was performed to study the expression of EGF and TNFα in HCCs. Western blotting, immunofluorescence, qRT-PCR, wound healing scratch and invasion assay, and chromatin immunoprecipitation assays were used to study the combined roles of EGF and TNFα in the motility of HCC cells in vitro. RESULTS: We demonstrated that both EGF and TNFα were highly expressed in HCCs, and HCCs with higher expression of both EGF and TNFα were more frequently rated as high-grade tumors. In vitro, EGF and TNFα cooperatively promoted the motility of HCC cells mainly via synergistic induction of an extracellular matrix glycoprotein fibronectin (FN). Mechanistically, EGF and TNFα jointly increased the nuclear translocation and PKC mediated phosphorylation of NF-κB/p65 which could bind to the -356bp to -259bp fragment of the FN promoter, leading to a markedly increased activity of the FN promoter in HCC cells. CONCLUSIONS: HCCs with higher expression of both EGF and TNFα were more frequently rated as high-grade tumors. EGF and TNFα cooperatively promoted the motility of HCC cells mainly through NF-κB/p65 mediated synergistic induction of FN in vitro. GENERAL SIGNIFICANCE: These findings highlight the crosstalk between EGF and TNFα in promoting HCC, and provide potential targets for HCC prevention and treatment.

The TGF-ß-induced up-regulation of NKG2DLs requires AKT/GSK-3ß-mediated stabilization of SP1.

Natural killer (NK) cells play an important role in preventing cancer development. NK group 2 member D (NKG2D) is an activating receptor expressed in the membrane of NK cells. Tumour cells expressing NKG2DL become susceptible to an immune-dependent rejection mainly mediated by NK cells. The paradoxical roles of transforming growth factor beta (TGF-ß) in regulation of NKG2DL are presented in many studies, but the mechanism is unclear. In this study, we showed that TGF-ß up-regulated the expression of NKG2DLs in both PC3 and HepG2 cells. The up-regulation of NKG2DLs was characterized by increasing the expression of UL16-binding proteins (ULBPs) 1 and 2. TGF-ß treatment also increased the expression of transcription factor SP1. Knockdown of SP1 significantly attenuated TGF-ß-induced up-regulation of NKG2DLs in PC3 and HepG2 cells, suggesting that SP1 plays a key role in TGF-ß-induced up-regulation of NKG2DLs. TGF-ß treatment rapidly increased SP1 protein expression while not mRNA level. It might be due to that TGF-ß can elevate SP1 stability by activating PI3K/AKT signalling pathway, subsequently inhibiting GSK-3ß activity and decreasing the association between SP1 and GSK-3ß. Knockdown of GSK-3ß further verified our findings. Taken together, these results revealed that AKT/GSK-3ß-mediated stabilization of SP1 is required for TGF-ß induced up-regulation of NKG2DLs. Our study provided valuable evidence for exploring the tumour immune modulation function of TGF-ß.

Histone deacetylase inhibitors deplete myeloid-derived suppressor cells induced by 4T1 mammary tumors in vivo and in vitro.

Myeloid-derived suppressor cells (MDSC) have been identified as a population of immature myeloid cells that suppress anti-tumor immunity. MDSC are increased in tumor-bearing hosts; thus, depletion of MDSC may enhance anti-tumor immunity. Histone deacetylase inhibitors (HDACi) are chemical agents that are primarily used against hematologic malignancies. The ability of these agents to modulate anticancer immunity has recently been extensively studied. However, the effect of HDACi on MDSC has remained largely unexplored. In the present study, we provide the first demonstration that HDACi treatment decreases MDSC accumulation in the spleen, blood and tumor bed but increases the proportion of T cells (particularly the frequency of IFN-γ- or perforin-producing CD8+ T cells) in BALB/C mice with 4T1 mammary tumors. In addition, HDACi exposure of bone marrow (BM) cells significantly eliminated the MDSC population induced by GM-CSF or the tumor burden in vitro, which was further demonstrated as functionally important to relieve the inhibitory effect of MDSC-enriched BM cells on T cell proliferation. Mechanistically, HDACi increased the apoptosis of Gr-1+ cells (almost MDSC) compared with that of Gr-1- cells, which was abrogated by the ROS scavenger N-acetylcysteine, suggesting that the HDACi-induced increase in MDSC apoptosis due to increased intracellular ROS might partially account for the observed depletion of MDSC. These findings suggest that the elimination of MDSC using an HDACi may contribute to the overall anti-tumor properties of these agents, highlighting a novel property of HDACi as potent MDSC-targeting agents, which may be used to enhance the efficacy of immunotherapeutic regimens.

Nodal signaling modulates the expression of Oct-4 via nuclear translocation of ß-catenin in lung and prostate cancer cells.

Nodal is a member of transforming growth factor beta (TGF-ß) superfamily. Nodal promotes the self-renewal of human cancer stem cells (CSCs) and triggers carcinogenesis of human cancers via an autocrine manner through Smad2/3 pathway. In our study, generation of Nodal-overexpressed cancer cells was constructed, and the effect of Nodal on the stem cell marker Oct-4 was evaluated by overexpression or blocked Nodal/ALKs signaling pathway in non-small cell lung cancer cells A549 and prostate cancer cells PC3. Functionally, Nodal also increased the proliferation via the ß-catenin nuclear translocation. This increase was attributed to GSK-3ß dephosphorylating, and activin receptor-like kinase 4/7 (ALK4/7) played a major role in human cancer cells. Our study provides a positive understanding of Nodal function in cancer cells and suggests a potential novel target for clinical therapeutic research.

Evaluation of antibody level against Fusobacterium nucleatum in the serological diagnosis of colorectal cancer.

Fusobacterium nucleatum (F. nucleatum, Fn) is associated with the colorectal cancer (CRC). Fn-infection could induce significant levels of serum Fn-specific antibodies in human and mice. The objective of this study was to identify Fn-infection that elicit a humoral response in patients with CRC and evaluate the diagnostic performance of serum anti-Fn antibodies. In this work, we showed the mean absorbance value of anti-Fn-IgA and -IgG in the CRC group were significantly higher than those in the benign colon disease group and healthy control group (P < 0.001). The sensitivity and specificity of ELISA for the detection of anti-Fn-IgA were 36.43% and 92.71% based on the optimal cut-off. The combination of anti-Fn-IgA and carcino-embryonic antigen (CEA) was better for diagnosing CRC (Sen: 53.10%, Spe: 96.41%; AUC = 0.848). Furthermore, combining anti-Fn-IgA with CEA and carbohydrate antigen 19-9 (CA19-9) (Sen: 40.00%, Spe: 94.22%; AUC = 0.743) had the better ability to classify CRC patients with stages I-II. These results suggested that Fn-infection elicited high level of serum anti-Fn antibodies in CRC patients, and serum anti-Fn-IgA level may be a potential diagnosing biomarker for CRC. Serum anti-Fn-IgA in combination with CEA and CA19-9 increases the sensitivity of detecting early CRC.

[Inhibition of Paeoniflorin on TNF-α-induced TNF-α Receptor Type I /Nuclear Factor-κB Signal Transduction in Endothelial Cells].

OBJECTIVE: To study the inhibitory effect of paeoniflorin (PAE) on TNF-α-induced TNF receptor type I (TNFR1)-mediated signaling pathway in mouse renal arterial endothelial cells (AECs) and to explore its underlying molecular mechanisms. METHODS: Mouse AECs were cultured in vitro and then they were treated by different concentrations PAE or TNF-α for various time periods. Expression levels of intercellular cell adhesion molecule-1 (ICAM-1) were detected in the normal group (cultured by serum-free culture media), the TNF-α group (cultured by 2-h serum-free culture media plus 6-h TNF-α 30 ng/mL), the low dose PAE group (cultured by 2-h PAE 0.8 µmo/L plus 6-h TNF-α 30 ng/mL), the middle dose PAE group (cultured by 2-h PAE 8 µmol/L plus 6-h TNF-α 30 ng/mL), the high dose PAE group (cultured by 2-h PAE 80 µmol/L plus 6-h TNF-α 30 ng/mL) with Western blot analysis. Nuclear translocation of transcription factor NF-κB (NE-κB) was detected in the normal group (cultured by serum-free culture media), the TNF-α group (cultured by 2-h serum-free culture media plus 45-mm TNF-α 30 ng/mL), and the high dose PAE group (cultured by 2-h PAE 80 µmol/L plus 45-min TNF-α 30 ng/mL) by immunofluorescent staining. Expression levels of the phosphorylation of extracellular signal-regulated (protein) kinase (ph-ERK) and p38 (ph- p38) were detected in the normal group (cultured by serum-free culture media) and the high dose PAE group (2-h PAE 80 µmol/L culture) by Western blot. NF-κB inhibitor-α (IκBα) protein expressions were detected in the normal group (cultured by serum-free culture media), the TNF-α group (cultured by 2-h serum-free culture media plus 30-min TNF-α 30 ng/mL), the high dose PAE group (cultured by 2-h PAE 80 µmol/L plus 30-min TNF-α 30 ng/mL), the p38 inhibitor group (SB group, pretreatment with SB238025 25 µmol/L for 30 min, then treated by PAE 80 µmol/L for 2 h, and finally treated by TNF-α 30 ng/mL for 30 min), the ERK inhibitor group (PD group, treated by PD98059 50 µmol/L for 30 min, then treated by PAE 80 µmol/L for 2 h, and finally treated by TNF-α 30 ng/mL for 30 min) by Western blot. RESULTS: Compared with the normal group, ICAM-1 protein expression levels obviously increased (P < 0.01). Compared with the TNFα group, ICAM-1 protein expression levels were obviously inhibited in the high dose PAE group (P < 0.05). Protein expression levels of ph-p38 and ph-ERK were obviously higher in the hIgh dose PAE group (P < 0.05). Compared with the normal group, IκBα protein expression levels obviously decreased in the TNF-α group (P < 0.01). Compared with the TNFα group, TNF-α-induced IκBα degradation could be significantly inhibited in the high dose PAE group (P < 0.01); the inhibition of PAE on IκBα degradation could be significantly inhibited in the SB group (P < 0.05). NF-κB/p65 signal was mainly located in cytoplasm in the normal group. NF-κB/p65 was translocated from cytoplasm to nucleus after stimulated by 45 min TNF-α in the TNF-α group, while it could be significantly inhibited in the high dose PAE group. CONCLUSIONS: PAE inhibited TNF-α-induced expression of lCAM-1. Its action might be associated with inhibiting TNFR1/NF-κB signaling pathway. p38 participated and mediated these actions.

Expression and significance of Nodal in human cancers: a meta-analysis.

Cancer is a global and growing problem. Nodal, which has been showed to be involved in occurrence and development of cancers, is an important embryonic morphogen. The aim of this study was to evaluate the significance of Nodal expression in human cancers based on the published related articles. Online databases were searched to retrieve relevant articles published between 2000 and 2015. The odds ratio (OR) with its 95% confident intervals (CI) were employed to calculate the strength of significance. Finally, a total of 11 articles were screened out, including 801 cancer patients and 372 healthy controls. Nine kinds of cancers were contained, and Nodal was detected in 56.7% of all participants (665/1173). Overall, our result found that Nodal was highly expressed in cancer patients than that in healthy controls, indicating that Nodal expression was significantly associated with cancers progression (OR=21.72, 95% CI=9.94-47.46, P<0.00001). Subgroup analysis showed that Nodal expression was significantly corrected with high WHO grade of human cancers (III+IV versus I+II: OR=2.46, 95% CI=1.63-3.71, P<0.00001). This significant relationship was also found in tumor size, differentiation degree, not observed in gender, age and lymphatic metastasis status of patients with all studied cancers in this meta-analysis. In conclusion, our results demonstrated that Nodal might be implicated in cancer progression, suggesting that it was a potential biomarker and therapeutic target for cancers.

A comparative study on inhibition of total astragalus saponins and astragaloside IV on TNFR1-mediated signaling pathways in arterial endothelial cells.

BACKGROUND: Both total astragalus saponins (AST) and it's main component astragaloside IV (ASIV) have been used in China as cardiovascular protective medicines. However, the anti-inflammatory activities that are beneficial for cardiovascular health have never been compared directly and the molecular mechanisms remain unresolved. This study was conducted to compare the inhibitory effects of these drugs on TNFα-induced cell responses, related signaling pathways, and the underlying mechanisms in mouse arterial endothelial cells. METHODOLOGY/PRINCIPAL FINDINGS: Real-time qRT-PCR was performed to determine the expression of cell adhesion molecule (CAM) genes. Immunofluorescent staining was used to detect the nuclear translocation of transcription factor NF-κB-p65. Western Blot analysis was used to identify TNFα-induced NF-κB-p65 phosphorylation, IκBα degradation, and caspase-3 cleavage. Cell surface proteins were isolated and TNFα receptor-1(TNFR1) expression was determined. The results suggest that both AST and ASIV attenuate TNFα-induced up-regulation of CAMs mRNA and upstream nuclear translocation and phosphorylation of NF-κB-p65. However, TNFR1-mediated IκBα degradation, cleavage of caspase-3 and apoptosis were inhibited only by AST. These differences in the actions of AST and ASIV could be explained by the presence of other components in AST, such as ASII and ASIII, which also had an inhibitory effect on TNFR1-induced IκBα degradation. Moreover, AST, but not ASIV, was able to reduce TNFR1 protein level on the cell surface. Furthermore, mechanistic investigation demonstrated that TNFR1-mediated IκBα degradation was reversed by the use of TAPI-0, an inhibitor of TNFα converting enzyme (TACE), suggesting the involvement of TACE in the modulation of surface TNFR1 level by AST. CONCLUSION: ASIV was not a better inhibitor than AST, at least on the inhibition of TNFα-induced inflammatory responses and TNFR1-mediated signaling pathways in AECs. The inhibitory effect of AST was caused by the reduction of cell surface TNFR1 level, and TACE could be involved in this action.

[Comparative analysis for the cytotoxicity and genotoxicity of multi-walled carbon nanotubes with different lengths and surface modifications in A549 cells].

OBJECTIVE: To compare the cytotoxicity and DNA strand breakage induced by multi-walled carbon nanotubes (MWCNTs) with different lengths and different surface modifications in human alveolar type II cells (A549 cells). METHODS: Two different lengths (5-15 µm, 350-700 nm) of MWCNTs and three different kinds of surface modified MWCNTs (COOH-MWCNTs, NH2-MWCNTs, and Tau-MWCNTs) were used in the experiments. The short MWCNTs were used as pristine MWCNTs to compare with the 3 surface modified MWCNTs. The cytotoxicity was determined by cell counting kit-8 (CCK-8) assay at the concentrations of 2, 8, and 32 mg/L at hours 12, 24, 36, and 48 respectively. Single cell gel electrophoresis (SCGE) assay was performed to evaluate DNA strand breakage in A549 cells after 24 h treatment of 8 mg/L of each tested material. RESULTS: Long multi-walled carbon nanotubes (Long-MWCNTs) and short multi-walled carbon nanotubes (Short-MWCNTs) showed a dose-dependent cytotoxicity within the exposure time 12-48 h. Especially, Long-MWCNTs showed greater cytotoxicity than Short-MWCNTs from 24 to 48 h at the same concentration. The relative cell viability of the 3 surface modified MWCNTs was higher than that of the pristine MWCNTs at h 12 at the concentration of 32 mg/L [COOH-MWCNTs (86.55±1.80)%, NH2-MWCNTs (84.67±1.32)%, Tau-MWCNTs (80.15±3.53)% and Pristine-MWCNTs (71.44±5.58)%], at h 24 at the concentration of 8 mg/L [COOH-MWCNTs (96.74±1.00)%, NH2-MWCNTs (96.74±3.35)%, Tau-MWCNTs (106.39±3.83)% and Pristine-MWCNTs (91.02±2.53)%], at h 24 at the concentration of 32 mg/L [COOH-MWCNTs (80.88±2.67)%, NH2-MWCNTs (82.90±3.25)%, Tau-MWCNTs (82.55±3.32)% and Pristine-MWCNTs (76.08±4.27)%] and at h 36 at the concentration of 8 mg/L [COOH-MWCNTs (96.87±1.05)%, NH2-MWCNTs (96.66±4.76)%, Tau-MWCNTs (100.23± 2.84)% and Pristine-MWCNTs (89.61±3.78)%], and the differences were statistically significant (P<0.05). Compared with the Pristine-MWCNTs, the relative cell viability of the 3 surface modified MWCNTs didn't demonstrate a statistically significant difference (P>0.05) at other observation time and exposure concentrations. The DNA strand breakage of the 3 surface modified MWCNTs: the Olive tail moment of COOH-MWCNTs was 1.56±0.22, the Olive tail moment of NH2-MWCNTs 2.25±1.62 and the Olive tail moment of Tau-MWCNTs 2.23±0.94; the tail DNA% of COOH-MWCNTs was (3.96± 0.60)%, the tail DNA% of NH2-MWCNTs (6.16±4.68)% and the tail DNA% of Tau-MWCNTs (6.05±2.31)%, which were lower than that of the pristine MWCNTs (P<0.05), whose Olive tail moment was 3.00±0.64 and tail DNA% (8.23±2.27)%. Moreover, the COOH-MWCNTs induced the lowest DNA damage among the three modified MWCNTs. CONCLUSION: Long-MWCNTs compared with Short-MWCNTs demonstrated a greater cytotoxicity and lower DNA strand breakage damage. The surface modifications of MWCNTs can reduce the cytotoxicity and DNA strand breakage in A549 cells.

[The role of lipid raft in TNFR1-mediated signal transduction in osteoblasts].

AIM: To investigate the role of membrane cholesterol in TNFR1-mediated signal transduction in osteoblastic MC3T3 cells. METHODS: MCD binds cholesterol specifically and was commonly used to deplete cholesterol from cell plasma membrane. MC3T3 cells were serum-starved for 22 h, treated with MCD (10 g/L) for 60 min followed by TNF-α (10 µg/L) for 0, 5, 10, 15 or 30 min, or TNF-α plus CHX (10 mg/L) for 4 h to induce apoptosis, then TNFR1-mediated IκBα degradation, phosphorylation of AKT, ERK or p38, and processing of caspase-3 were analyzed by using SDS-PAGE/Western blotting method. RESULTS: MC3T3 cell membrane cholesterol level was reduced to 35% within 60 min by MCD (10 g/L). Reduction of MC3T3 cell surface cholesterol dramatically inhibited TNFR1-mediated AKT phosphorylation, while did not affect the degradation of IκBα, activation of ERK or p38, and processing of caspase-3 induced by TNF-α. CONCLUSION: Cholesterol depletion can destruct lipid rafts; therefore our results suggest that lipid raft is essential for TNFR1-mediated AKT phosphorylation, but is dispensable for TNFR1-mediated degradation of IκBα, activation of ERK or p38 and processing of caspase-3.

[Cytotoxicity and its mechanism of zinc oxide nanoparticles on human leukemic monocyte lymphoma cell line U937].

OBJECTIVE: To investigate the cytotoxicity and its mechanism of ZnO nanoparticles on human leukemic monocyte lymphoma cell line U937. METHODS: Four different size ZnO (10, 30, 60, 500 nm) were carefully characterized. The survival rate and viability were measured by trypan blue assay and MTT assay for each size ZnO particles at different concentrations (12, 120, 240, 600, 1200 µmol/L). The zinc probe, Fluozin-3, was used to detect the intracellular free zinc. Transmission electron microscopy was adopted to observe the cellular ultrastructure and the uptake of ZnO. RESULTS: All four kinds of ZnO were rod shape, with a purity of > 99.9 wt%, and they were classified as zincite phase crystal and their surface areas were in accordance with the sizes. The viability (ZnO-n10: (97 ± 19)%, (91 ± 4)%, (24 ± 4)%, (15 ± 2)%; ZnO-n30: (111 ± 4)%, (81 ± 3)%, (24 ± 2)%, (27 ± 8)%; ZnO-n60: (105 ± 11)%, (73 ± 20)%, (43 ± 11)%, (28 ± 14)%; ZnO-µm: (88 ± 16)%, (62 ± 7)%, (22 ± 4)%, (13 ± 5)%) of cells exposed to ZnO decreased with the increasing of the concentration of ZnO from 12 to 600 µmol/L (r values were 0.965, 0.979, 0.998, 0.992, and the t values were 19.8, 25.3, 76.3, 40.9, respectively, P < 0.05). The liability (ZnO-n10: (98 ± 1)%, (67 ± 2)%, (59 ± 7)%, (13 ± 13)%, (5 ± 4)%; ZnO-n30: (98 ± 1)%, (97 ± 2)%, (50 ± 3)%, (20 ± 14)%, (7 ± 2)%; ZnO-n60: (97 ± 2)%, (88 ± 5)%, (48 ± 10)%, (12 ± 5)%, (4 ± 1)%; ZnO-µm: (96 ± 1)%, (76 ± 3)%, (58 ± 3)%, (19 ± 5)%, (20 ± 10)%) of cells exposed to ZnO decreased with the increasing of the concentration of ZnO from 12 to 600 µmol/L (r valued at 0.982, 0.956, 0.972, 0.980, and the t valued at 19.3, 12.1, 15.6, 18.5, respectively, P < 0.05). The increase of the zinc concentration showed by the zinc fluorescence probe was 121 ± 11, which was similar to the fluorescence of cells treated with ZnAc(2) (132 ± 14, F = 0.6, P > 0.05) at the Zn-equivalent concentration. There was no statistic difference for the percents of high zinc content cells in total cells exposed to ZnO-n30 (87.6 ± 2.6)% and these exposed to ZnAc(2) (86.9 ± 3.2)% (F = 1.5, P > 0.05). CONCLUSION: ZnO nanoparticles are highly cytotoxic to U937 cells and the solubilization of ZnO is the main toxicological mechanism.

[The role of Angelica polysaccharides in inducing effector molecule release by peritoneal macrophages].

AIM: To observe the effect of Angelica polysaccharides on effector molecule production by peritoneal macrophages. METHODS: Macrophages were isolated from the peritoneal cavity of BALB/c mice and the primary culture was performed. MTT colorimetry and spectrophotometry were used to examine the effects of Angelica polysaccharides on the releases of effector molecules, such as nitric oxide(NO), tumor necrosis factor-alpha(TNF-alpha), and reactive oxygen species(ROS) as well as inducible nitric oxide synthase (iNOS) and lysozyme(LSZ) activity by peritoneal macrophages. RESULTS: Angelica polysaccharides could promote the releases of NO, TNF-alpha and ROS from macrophages and improved iNOS and LSZ activities in macrophages. However, Angelica polysaccharides had no direct cytotoxicity to tumor cells, but the cultural supernatant of macrophages cocultured with Angelica polysaccharides could kill L929 cells. CONCLUSION: Angelica polysaccharides can promote the releases of NO, TNF-alpha and ROS by macrophages. Angelica polysaccharides may indirectly play the role of anti-tumors through increased TNF-alpha production by macrophages.