[Caspase-1/-11 participates in LPS-induced sepsis-associated acute kidney injury by cleaving GSDMD].

The present study was aimed to investigate whether Gasdermin D (GSDMD)-mediated pyroptosis participated in lipopolysaccharide (LPS)-induced sepsis-associated acute kidney injury (AKI), and to explore the role of caspase-1 and caspase-11 pyroptosis pathways in this process. The mice were divided into four groups: wild type (WT), WT-LPS, GSDMD knockout (KO) and KO-LPS. The sepsis-associated AKI was induced by intraperitoneal injection of LPS (40 mg/kg). Blood samples were taken to determine the concentration of creatinine and urea nitrogen. The pathological changes of renal tissue were observed via HE staining. Western blot was used to investigate the expression of pyroptosis-associated proteins. The results showed that the concentrations of serum creatinine and urea nitrogen in the WT-LPS group were significantly increased, compared with those in the WT group (P < 0.01); whereas serum creatinine and urea nitrogen in the KO-LPS group were significantly decreased, compared with those in the WT-LPS group (P < 0.01). HE staining results showed that LPS-induced renal tubular dilatation was mitigated in GSDMD KO mice. Western blot results showed that LPS up-regulated the protein expression levels of interleukin-1ß (IL-1ß), GSDMD and GSDMD-N in WT mice. GSDMD KO significantly down-regulated the protein levels of IL-1ß, caspase-11, pro-caspase-1, caspase-1(p22) induced by LPS. These results suggest that GSDMD-mediated pyroptosis is involved in LPS-induced sepsis-associated AKI. Caspase-1 and caspase-11 may be involved in GSDMD cleavage.

Berberine Reverses Breast Cancer Multidrug Resistance Based on Fluorescence Pharmacokinetics In Vitro and In Vivo.

Exploring the mechanism through which berberine (Ber) reverses the multidrug resistance (MDR) of breast cancer is of great importance. Herein, we used the methyl thiazolyl tetrazolium assay to determine the drug resistance and cytotoxicity of Ber and doxorubicin (DOX) alone or in combination on the breast cancer cell line MCF-7/DOXFluc. The results showed that Ber could synergistically enhance the inhibitory effect of DOX on tumor cell proliferation in vitro, and the optimal combination ratio was Ber/DOX = 2:1. Using a luciferase reporter assay system combined with the bioluminescence imaging technology, the efflux kinetics of d-luciferin potassium salt in MCF-7/DOXFluc cells treated with Ber in vivo was investigated. The results showed that Ber could significantly reduce the efflux of d-luciferin potassium salt in MCF-7/DOXFluc cells. In addition, western blot and immunohistochemistry experiments showed that the expression of P-glycoprotein (P-gp/ABCB1) and multidrug resistance protein 1 (MRP1/ABCC1) in MCF-7/DOXFluc cells was downregulated upon Ber treatment. Finally, high-performance liquid chromatography was used to investigate the effect of Ber on DOX tissue distribution in vivo, and the results showed that the uptake of DOX in tumor tissues increased significantly when combined with Ber (P < 0.05). Thus, the results illustrated that Ber can reverse MDR by inhibiting the efflux function of ATP-binding cassette transporters and downregulating their expression levels.

Low level of microplastic contamination in wild fish from an urban estuary.

Microplastic accumulation in estuarine environments is considered the dominant input of land-based plastics into the oceans. In this study, the level of microplastic contamination was evaluated in 26 species of wild fish from the Pearl River Estuary, South China. Results showed that microplastics abundance ranged from 0.17 items individual-1 (Boleophthalmus pectinirostris & Acanthogobius flavimanus) to 1.33 items individual-1 (Plectorhynchus cinctus) among different species. The distribution of microplastic abundance in the gills and gastrointestinal tracts was not significantly different. Microplastics in gills are strongly related to the filtration area of gills in 15 fish species. Fibers were the dominant shapes accounting for 93.45% of the total shapes. The majority of microplastics were <3 mm in size. The most common polymer composition was polyethylene terephthalate (38.2%) and the most common color was black (30.36%). The findings of this study provide baseline data for microplastic contamination in wild fish from an urban estuary.

Microplastics in oysters Saccostrea cucullata along the Pearl River Estuary, China.

As a transitional zone between riverine and marine environments, an estuary plays an important role for the sources, accumulation and transport of microplastics. Although estuarine environments are hotspots of microplastic pollution, the correlation between microplastic pollution and aquatic organisms is less known. Here we investigated microplastic pollution in wild oysters Saccostrea cucullata from 11 sampling sites along the Pearl River Estuary in South China. The microplastic abundances in oysters ranged from 1.4 to 7.0 items per individual or from 1.5 to 7.2 items per gram tissue wet weight, which were positively related to those in surrounding waters. The oysters near urban areas contained significantly more microplastics than those near rural areas. Fibers accounted for 69.4% of the total microplastics in oysters. Microplastic sizes varied from 20 to 5000 µm and 83.9% of which were less than 100 µm. Light color microplastics were significantly more common than dark color ones. Based on the results, oysters are recommended as a biomonitor for the microplastic pollution in estuaries.

Telekin suppresses human hepatocellular carcinoma cells in vitro by inducing G2/M phase arrest via the p38 MAPK signaling pathway.

AIM: Telekin, isolated from the Chinese herb Carpesium divaricatum, has shown anti-proliferation effects against various cancer cells, including hepatocellular carcinoma cells. In this study, we investigated the anti-proliferation mechanisms of telekin in human hepatocellular carcinoma HepG2 cells in vitro. METHODS: HepG2 cells were treated with telekin. Cell viability was evaluated using MTT assay. Flow cytometry was used to measure cell cycle profiles, ROS level and apoptosis. The protein expression levels were analyzed with Western blotting. RESULTS: Telekin (3.75-30 µmol/L) dose-dependently inhibited the viability of HepG2 cells and induced l apoptosis. Furthermore, the treatment induced cell cycle arrest at G2/M phase, accompanied by significantly increased the phosphorylation of Cdc25A and Cdc2, and decreased Cyclin B1 level. Moreover, the treatment significantly stimulated ROS production, and increased the phosphorylation of p38 and MAPKAPK-2 in the cells. Pretreatment with the antioxidant NAC (2.5, 5, and 10 mmol/L), or the p38 MAPK inhibitor SB203580 (2.5 and 5 µmol/L) dose-dependently attenuated these telekin-induced effects in the cells. CONCLUSION: Telekin suppresses hepatocellular carcinoma cells in vitro by inducing G2/M phase arrest via activating the p38 MAPK pathway.

[Residues and health risk assessment of HCHs, DDTs and heavy metals in water and Tilapias from fish ponds of Guangdong].

Concentrations of copper, lead, cadmium, arsenic, hexachlorcyclohexane (HCHs) and dichlorodiphenyltrichloroethane (DDTs) in the water and the fish samples collected separately from fish pond, markets and supermarkets in four cities of Guangdong Province were measured by using GC-ECD, flame atomic absorption spectrometry and atomic fluorescence spectrometry. Health risk assessments associated with Cu, Pd, Cd, As, HCHs and DDTs were conducted based on the model of health risk assessment recommended by the US EPA. The results showed that the concentration ranges of Cu, Pd, Cd, As, HCHs and DDTs in water samples were nd-0.101 mg x L(-1), nd-0.097 mg x L(-1), nd-0.003 27 mg x L(-1), 0.0121-0.08127 mg x L(-1), 2.63-37.18 ng x L(-1) and 2.05-12.21 ng x L(-1), respectively. The health risk assessment indicates that the carcinogenic and non-carcinogenic risks of Cu, Pd, Cd, HCHs and DDTs in Tilapias both lower than the highest acceptable level of risk set by ICRP, but As cancer risk value slightly exceeded the upper limit of the acceptable risk levels in city population.

Fluopsin C induces oncosis of human breast adenocarcinoma cells.

AIM: Fluopsin C, an antibiotic isolated from Pseudomonas jinanesis, has shown antitumor effects on several cancer cell lines. In the current study, the oncotic cell death induced by fluopsin C was investigated in human breast adenocarcinoma cells in vitro. METHODS: Human breast adenocarcinoma cell lines MCF-7 and MD-MBA-231 were used. The cytotoxicity was evaluated using MTT assay. Time-lapse microscopy and transmission electron microscopy were used to observe the morphological changes. Cell membrane integrity was assessed with propidium iodide (PI) uptake and lactate dehydrogenase (LDH) assay. Flow cytometry was used to measure reactive oxygen species (ROS) level and mitochondrial membrane potential (Δψm). A multimode microplate reader was used to analyze the intracellular ATP level. The changes in cytoskeletal system were investigated with Western blotting and immunostaining. RESULTS: Fluopsin C (0.5-8 µmol/L) reduced the cell viability in dose- and time-dependent manners. Its IC50 values in MCF-7 and MD-MBA-231 cells at 24 h were 0.9 and 1.03 µmol/L, respectively. Fluopsin C (2 µmol/L) induced oncosis in both the breast adenocarcinoma cells characterized by membrane blebbing and swelling, which was blocked by pretreatment with the pan-caspase inhibitor Z-VAD-fmk. In MCF-7 cells, fluopsin C caused PI uptake into the cells, significantly increased LDH release, induced cytoskeletal system degradation and ROS accumulation, decreased the intracellular ATP level and Δψm. Noticeably, fluopsin C exerted comparable cytotoxicity against the normal human hepatocytes (HL7702) and human mammary epithelial cells with the IC50 values at 24 h of 2.7 and 2.4 µmol/L, respectively. CONCLUSION: Oncotic cell death was involved in the anticancer effects of fluopsin C on human breast adenocarcinoma cells in vitro. The hepatoxicity of fluopsin C should not be ignored.

1-Oxoeudesm-11(13)-eno-12,8a-lactone induces G2/M arrest and apoptosis of human glioblastoma cells in vitro.

AIM: To investigate the effects of 1-oxoeudesm-11(13)eno-12,8a-lactone (OEL), a novel eudesmane-type sesquiterpene isolated from Aster himalaicus, on the cell cycle and apoptosis in human glioblastoma cells in vitro. METHODS: Human malignant glioblastoma cell lines U87 and A172 were used. The cytotoxicity of OEL was examined using the MTT assay. Cell apoptosis was assessed with DAPI staining and flow cytometry. DNA damage was determined by measuring the phosphorylation of H2AX using immunofluorescence staining and Western blotting. Cell cycle profiles were measured with flow cytometry. The mRNA expression of p53 and p21Waf1/Cip1 was investigated using real-time PCR. The protein expression of γ-H2AX, caspase-9, caspase-3, p53, p21Waf1/Cip1, cyclin B1, and cdc2 was analyzed with Western blotting. RESULTS: Treatment of the malignant glioblastoma cells with OEL inhibited the cell growth in dose- and time-dependent manners (the values of IC(50) at 48 and 72 h were 29.5 and 16.99 µmol/L, respectively, in U87 cells; 7.2 and 9.5 µmol/L, respectively, in A172 cells). OEL (10-30 µmol/L) induced apoptosis and G(2)/M phase arrest in both U87 and A172 cells. OEL induced the phosphorylation of cdc2, a G(2)/M phase cyclin-dependent kinase, and decreased the expression of cyclin B1 required for progression through the G(2)/M phase in U87 cells. The compound remarkably increased the phosphorylation of H2AX in U87 cells. Moreover, OEL increased the mRNA and protein levels of p53 and its target gene p21(Waf1/Cip1) in U87 cells. The compound also induced p53 phosphorylation. Pretreatment with PFT-α, a specific inhibitor of p53 transcriptional activity, could partially reverse the inhibition of OEL on the viability of U87 and A172 cells. CONCLUSION: OEL suppresses the growth of human glioblastoma cells in vitro via inducing DNA damage, p53-mediated cell cycle arrest and apoptosis, thus warrants further studies as a lead compound of anti-glioblastoma drug.

[A dynamic finite element analysis of stress distribution in bone tissue surrounding solely or splinted implant-borne fixed partial denture].

OBJECTIVE: To study the distribution patterns of stresses induced in bone tissue surrounding solely and splinted implants under dynamic loads. METHODS: Three dimensional finite-element models were created of two 765 sections of the mandible with solely or splinted implants embedded in. Vertical and oblique dynamic loads were applied in a circle of mastication (0.875 s). The stress distribution was analyzed to study the biomechanical behavior of bone tissue surrounding solely or splinted implants. RESULTS: As loading on the solely implant 5, the maximum von Mises value in the surrounding bone tissue under oblique loads at 0.300 s was 4.2 times as much as that under vertical loads at 0.150 s. Meanwhile, as coincidently loading on the splinted implants, the maximum von Mises value at 0.300 s was 1.2 times as much as that at 0.150 s. As loading on the solely implant 5, the maximum stress value was 48.393 MPa at 0.300 s. As separately loading on the splinted implant 5, the maximum stress value of the whole model was 9.541 MPa in the same loading course, and the maximum stress was located at the distal cervical of the indirectly loaded implant 7. When loading on the pontic, the stress in bone tissue surrounding implant 7 was more than that of implant 5. CONCLUSIONS: Stress in the bone-interface of the splinted implants is evenly distributed at the cervical level, which may also reduce disadvantages from oblique loads.

Sesquiterpenoids from Carpesium divaricatum and their cytotoxic activity.

In the course of searching for cytotoxic terpenoids from medicinal plants in China, two new eudesmane sesquiterpenoids, 5α-hydroxy-13-methoxy-7αH,11αH-eudesm-4(15)-en-12,8ß-lactone (1) and 1ß-hydroxy-7αH,11αH-eudesm-4(15)-en-12,8ß-lactone (2), along with fourteen known sesquiterpenoids were isolated from the whole plant of Carpesium divaricatum. The structures of new compounds were determined using spectroscopic methods, including IR, HRESIMS, and 1D and 2D NMR spectroscopy. The cytotoxicity of selected sesquiterpene lactones against human oral epidermoid carcinoma (KB), human breast cancer (MCF-7) and human hepatoma (HepG-2) cells was also evaluated by MTT method.

Regulation of the human vitamin C transporters expressed in COS-1 cells by protein kinase C [corrected].

Protein kinase C (PKC) regulation of l-ascorbic acid transport mediated by the Na+/ascorbic acid transporters, hSVCT1 and hSVCT2, expressed in COS-1 cells was studied using recombinant carboxyl-terminal V5 epitope-tagged forms of the transporters. The PKC activator phorbol 12-myristate 13-acetate (PMA) caused a time-dependent and concentration-dependent decrease (40-60%) in ascorbic acid transport activity. Effects of PMA were not observed with the inactive phorbol ester 4 alpha-phorbol and were reversed by treatment of the cells with the PKC-specific inhibitor Ro-31-8220. Kinetically, the reduction in hSVCT1 and hSVCT2 activity arose from a decrease in maximal velocity with no change in the apparent affinity. Western blot and confocal microscopy analyses indicated that the total pool of hSVCT1 or hSVCT2 proteins expressed in the transfected COS-1 cells remained unaffected by PMA treatment. For hSVCT1 the decrease in L-ascorbic acid correlated with a redistribution of the transporter from the cell surface to intracellular membranes. However, for hSVCT2 there was no apparent change in transporter distribution, suggesting that the PKC-dependent modulation of L-ascorbic acid transport mediated by hSVCT2 was the result of reduced catalytic transport efficiency.