[Effects of MSP on Cell Cycle and EMT of Non-Small Cell Lung Cancer PC14].

OBJECTIVE: To study the effect of macrophage stimulating protein (MSP) on the cell cycle of non-small cell lung cancer PC14 cells without expression of recepteur d'originenanta (RON) and MSP,and analyse its effect on PC14's epithelial mesenchymal transition (EMT) capacity. METHODS: Vitro culture PC14 (blank control),PC14-Mst1-pEGFP-N1 (stablely expressed MSP) and PC14-pEGFP-N1. Cell cycles were detected by flow cytometry and the gaps between cells during growth were measured by transmission electron microscope (TEM); RT-PCR and Western blot were used to figure out the shifts of EMT related gene expression in PC14-Mst1-pEGFP-N1 cells. RESULTS: Compared with the PC14 group and PC14-pEGFP-N1 group,PC14-Mst1-pEGFP-N1 population of G1/G0 phase were significantly increased while S and G2/M phase were significantly reduced;The gaps between PC14-Mst1-pEGFP-N1 cells decreased; RT-PCR and Western blot showed that mRNA and protein levels of E-cadherin of PC14-Mst1-pEGFP-N1 were significantly higher than that of PC14,but mRNA and protein levels of Vimentin were significantly lower. CONCLUSION: MSP may affect the cell cycle of PC14 and inhibit its EMT procedure by regulating the expression of related proteins including E-cadherin and Vimentin when RON was not expressed.

[The Correlation Between MicroRNAs in Serum and the Extent of Liver Injury].

OBJECTIVES: To investigate the correlation between the absolute quantification of the microRNAs (miR-122, miR-451, miR-92a, miR-192) in serum during acute liver injury and the extent of liver injury on rat models of CCl4 induced acute liver injury and mice models of acetaminophen (APAP) induced acute liver injury. Furthermore, to investigate the correlation between the absolute quantification of microRNAs in serum and the drug induced liver injury pathological scoring system (DILI-PSS). METHODS: The acute liver injury model in rat by CCl4 (1.5 mL/kg), and the acute liver injury model in mice by APAP (160 mg/kg) were established. The serum at different time points on both models were collected respectively. The absolute quantification of microRNAs in serum were detected by using MiRbayTM SV miRNA Assay kit. Meanwhile, the pathological sections of liver tissue of the mice at each time point were collected to analyze the correlation between microRNAs and the degree of liver injury. RESULTS: In CCl4-induced rat acute liver injury model and APAP induced mouse acute liver injury, miR-122 and miR-192 appeared to be rising significantly, which remained the highest level at 24 h after treatment, and declined to the normal level after 72 h. In CCl4-induced rat acute liver injury model, the change of miR-92a was fluctuated and had no apparent rules, miR-451 declined gradually, but not obviously. In mice acute liver injury model induced by APAP, miR-92a and miR-451 in the progress of liver injury declined gradually, reached the lowest point at 48 h, and then recovered. The result of correlation analysis indicated that miR-122 and miR-192 presented a good positive correlation with the DILI-PSS ( r=0.741 3, P<0.05; r=0.788 3, P<0.01). CONCLUSIONS: The absolute quantification of miR-122 and miR-192 in serum has the highest level in 24 h, then decrease in 72 h, in both drug-induced and chemical liver injury. In addition, both the two microRNAs have good correlation with DILI-PSS in APAP-induced liver injury models.

[Effects of Msp on the Proliferation, Migration and Invasion of Human Non-small Cell Lung Cancer Cells].

OBJECTIVES: To determine the effects of macrophage stimulating protein (Msp) on the proliferation, migration and invasion of human non-small cell lung cancer cells PC14. METHODS: The eukaryotic expression vector for st1was constructed and transfected into Msp(-)and RON(-)human non-small cell lung cancer cells PC14. The expression of st1mRNA in PC14 cells was observed by RT-PCR. The expression levels of Msp protein in PC14, PC14-st1-pEGFP-N1 and PC14-pEGFP-N1 groups as well as the expression of RON in PC14 and SKBR-3 cells were detected by Western blot. RAW264.7 (mouse monocyte macrophage) and SKBR-3 cells were cultured in the supernatant of cells(PC14, PC14-st1-pEGFP-N1and PC14-pEGFP-N1 groups)and tested with Transwell microporous membrane, through which the biologic activity of Msp was evaluated by calculating the cell number migrated. The proliferation of PC14 was measured by MTT assay. The capabilities of PC14 to migrate and invade were measured by Transwell chamber and Matrigel invasion tests, respectively. RESULTS: The expressions of mRNA and protein of Mst1 in PC14 were stable after transfection with Mst1. Msp (PC14-st1 -pEGFP-N1 group) promoted the migration of RON (+) cells (SKBR-3 and RAW264.7). Compared with PC14 and PC14-pEGFP-N1 groups, the proliferation, migration and invasion of PC14 cells in PC14-st1 -pEGFP-N1 group were inhibited significantly. CONCLUSIONS: Msp can promote the migration of RON (+) cancer cells in paracrine secretion manner and inhibit the proliferation, migration and invasion of human non-small cell lung cancer cells PC14 in an unknown way.

[Monitoring urine metabolic profiles of rat inflammation model with capillary electrophoresis].

OBJECTIVE: To determine the comprehensive metabolic profiling of inflammation with capillary zone electrophoresis (CZE) in rat urine. METHODS: CZE-based metabolomics method was used to acquire urine metabolom data, with Computer Aided Similarity Evaluation System aligning peaks and analyzing urine profiling. Principle component analysis (PCA) was performed to compare and classify the phenotypes of the urine CZE spectra. RESULTS: The PCA revealed different phenotypes of metabolites between rats with and without inflammation, which were independent from gender, individual diversity, day-night diversity and diurnal variation. CONCLUSION: CZE-based metabolomics method could be used as a potential tool for urinary profiling for disease diagnosis and drug studies.

Effect of acupuncture on rats with acute gouty arthritis inflammation: a metabonomic method for profiling of both urine and plasma metabolic perturbation.

Acute gouty arthritis is a common inflammation model with multiple pathogenic mechanisms seen in clinical practice, for which acupuncture may potentially be an alternative therapy. To investigate the effect of acupuncture on acute gouty arthritis and search for its mechanism, a metabonomic method was developed in this investigation. Acute gouty arthritis model rats were induced by monosodium urate (MSU) crystals. The urine and plasma samples were collected at several time points and the endogenous metabolites were analyzed by an ultra-performance liquid chromatography coupled with a mass spectrometry (UPLC-MS). Data were analyzed using principal components analysis (PCA) and partial least squares (PLS) analysis to compare metabolic profiles of MSU crystal-induced acute gouty arthritis rats with MSU crystal-induced acute gouty arthritis, treated with acupuncture rats. The results showed that acupuncture could restore the metabolite network that disturbed by MSU administration. Our study indicates that UPLC-MS-based metabonomics can be used as a potential tool for the investigation of biological effect of acupuncture on acute gouty arthritis.

A multiple-dose pharmacokinetics of polyethylene glycol recombinant human interleukin-6 (PEG-rhIL-6) in rats.

Radiation therapy has been widely applied in cancer treatment. However, it often causes thrombocytopenia (deficiency of white blood cells) as an adverse effect. Recombinant human interleukin-6 (rhIL-6) has been found to be a very effective way against this thrombocytopenia, but IL-6 has low stability in blood, which reduces its efficacy. To increases the stability and half-life of rhIL-6, it was modified by polyethylene glycol (PEG). The pharmacokinetics and the tissue distribution of PEG-rhIL-6 labeled with (125)I were examined after subcutaneous injection in rats. The pharmacokinetic pattern of PEG-rhIL-6 was defined with linear-kinetics, and we fitted a one-compartment model with half-lives of 10.44-11.37 h (absorption, t(1/2Ka)) and 19.77-21.53 h (elimination, t(1/2Ke)), and peak concentrations at 20.51-21.96 h (t(peak)) in rats. Half-lives and t(peak) of PEG-rhIL-6 were longer than those of rhIL-6 previously reported. In the present study, for deposition of PEG-rhIL-6 in rats, the tissue distribution examination showed that blood was the major organ involved, rather than liver. However, as to the elimination of PEG-rhIL-6, the major organ was the kidney. The excretion fraction of the injection dose recovered from urine was 23.32% at 192 h after subcutaneous administration. Less than 6% of PEG-rhIL-6 was eliminated via the feces at 192 h. These results indicate that PEG-rhIL-6 is a good candidate drug formulation for patients with cancer.

[Metabonomic study of the urine of rat acute gouty arthritis model].

UNLABELLED: [ OBJECTIVE: To analysis the metabonome characteristics of the urine of MSU crystal-induced acute gouty arthritis rats by the method of metabolomics. METHODS: Based on the method of metabolomics, which applies LC/MS as data acquisition platform, incorporating with the means of stoechiometry such as principal component analysis, we analyzed the metabonome difference between the urine of acute gouty arthritis rats and that of normal rats. RESULTS: Compare with control group, the metabolism status of acute gouty arthritis model group deviated. After that, with the time lapsed, it retrieved gradually to the incipient metabolism status. The variation of metabolism locus of rats measured by the methods of metabolomics properly reflects the genesis, development, and recuperation course of acute gouty arthritis. CONCLUSION: The whole metabolism status of rat model is able to be presented finely with the method of metabolomics. The metabolomics study could offer a satisfactory research approach to acute gouty arthritis.

[Pharmacokinetics of PEG recombinant human interleukin-6 in rats].

OBJECTIVE: To study the pharmacokinetics and tissue distribution of PEG-rhIL-6 in rats after a single dose administration. METHODS: Pharmacokinetics and distribution of PEG-rhIL-6 in rats were studied by 125I isotope tracing method. Pharmacokinetic analysis was performed using 3P97 computer software. RESULTS: PEG-rhIL-6 declined in one-compartment model with half-lives of 10.44-11.37 h for t1/2 Ka, 19.77-21.53 h for t1/2 Ke and 20.51-21.96 h for T(pcak), respectively. PEG-rhIL-6 was mainly distributed in blood and excreted via urine. CONCLUSION: The half-lives of PEG-rhIL-6 are prolonged after being modified by PEG.

[Effects of leptin on development of mouse preimplantation embryos in vitro].

OBJECTIVE: To investigate the effects of leptin on development of mouse preimplantation embryos in vitro. METHODS: (1) Groups of 2-cell stage embryos randomly selected were placed in drops of CZB medium with or without recombinant leptin (10, 50, 100 and 500 ng/ml) and were cultured to the hatched blastocyst stage, and then embryo-transfer was carried out and the implantation rate was observed and determined. (2) Groups of 2-cell stage embryos randomly selected were placed in drops of leptin free CZB medium and cultured to morula stage. Then we changed the medium, randomized the embryos into CZB medium with or without recombinant leptin (10, 50, 100 and 500 ng/ml), and cultured them to the hatched blastocyst stage. RESULTS: Addition of leptin to embryo culture media promoted the development from 2-cell stage embryo to morula, blastocyst and hatched blastocyst and improved the implantation rate. Leptin at 0, 10, 50 ng/ml concentrations caused a dose-dependent promotion, nevertheless Leptin at even higher concentrations (100 and 500 ng/ml) only brought on the same promotion as what the Leptin at concentration of 50 ng/ml did (P>0.05). Addition of leptin to embryo culture media seemed to have no effects on the in vitro development from morula to blastocyst and hatched blastocyst. CONCLUSION: Leptin plays an important role in the development of preimplantation embryo. It can promote embryo development and embryo implantation.