Preparation and functional characteristics of protein from Ginkgo endophytic Pseudomonas R6 and Ginkgo seed.

Ginkgo seed protein (GSP) has excellent processing characteristics and antioxidant properties. In this study, Gingko endophytic protein (GEP) was synthesized by Ginkgo endophytic Pseudomonas R6. SDS-PAGE analysis indicated that the molecular weights of GSP and GEP were mainly distributed at 17 KDa and 48 KDa, respectively. FTIR showed that GEP and GSP exhibited characteristic absorption in the amide I, II, and III bands, and absorption in amide A and B indicated the presence of hydrogen bonding. HPLC analysis showed that both proteins had 17 amino acids, but their relative abundance was different, with GSP having the highest Ser content (74.713 mg/g) and GEP having the highest Val content (35.905 mg/g). Stomata were observed on the surface of both proteins by SEM, and there were lamellar and some spherical structures on GEP, while the opposite was observed on GSP. GEP had superior solubility, OHC, FC and EC, while GSP showed good WHC. Both proteins exhibited antioxidant activities, with GSP exhibiting stronger hydroxyl radical scavenging ability than GEP, with IC50 of 0.46 mg/mL and 1.54 mg/mL, respectively. This work demonstrates the antioxidant potential of GEP as an alternative to GSP in the food industry.

Characterization of lipopeptide produced by Bacillus altitudinis Q7 and inhibitory effect on Alternaria alternata.

This study identified the antifungal metabolites produced by Bacillus altitudinis Q7 against Alternaria alternata and investigated the antifungal activity and antifungal action. Lipopeptide, the important secondary metabolites were identified by Fourier transform infrared (FTIR) and liquid chromatography-mass spectrometry as lichenysin. The antifungal activity of lipopeptide on A. alternata was determined by microdilution technique, and its minimum inhibitory concentration was 1.2 mg/ml. Stability test showed that lipopeptide had excellent temperature and pH resistance. To investigate whether lichenysin acted on the cell membrane and changed its permeability, the ultra-violet absorption of protein and nucleic acid were measured using a colorimetric method. The antifungal metabolites produced by B. altitudinis Q7 was lichenysin, which showed stable antifungal activity in the extreme environments. Lichenysin could inhibit A. alternata by altering the permeability of cell membrane, leading to the outflow of proteins and nucleic acids from the cytoplasm. This research suggests the lipopeptide from B. altitudinis Q7 is a potential biological control agent against A. alternata.

Identification of subtypes of hepatocellular carcinoma and screening of prognostic molecular diagnostic markers based on cell adhesion molecule related genes.

Cell adhesion molecules can predict liver hepatocellular carcinoma (LIHC) metastasis and determine prognosis, while the mechanism of the role of cell adhesion molecules in LIHC needs to be further explored. LIHC-related expression data were sourced from The Cancer Genome Atlas (TCGA) and the gene expression omnibus (GEO) databases, and genes related to cell adhesion were sourced from the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. First, the TCGA-LIHC dataset was clustered by the nonnegative matrix factorization (NMF) algorithm to find different subtypes of LIHC. Then the difference of prognosis and immune microenvironment between patients of different subtypes was evaluated. In addition, a prognostic risk model was obtained by least shrinkage and selection operator (LASSO) and Cox analysis, while a nomogram was drawn. Furthermore, functional enrichment analysis between high and low risk groups was conducted. Finally, the expressions of model genes were explored by quantitative real-time polymerase chain reaction (qRT-PCR). The 371 LIHC patients were classified into four subtypes by NMF clustering, and survival analysis revealed that disease-free survival (DFS) of these four subtypes were clearly different. Cancer-related pathways and immune microenvironment among these four subtypes were dysregulated. Moreover, 58 common differentially expressed genes (DEGs) between four subtypes were identified and were mainly associated with PPAR signaling pathway and amino acid metabolism. Furthermore, a prognostic model consisting of IGSF11, CD8A, ALCAM, CLDN6, JAM2, ITGB7, SDC3, CNTNAP1, and MPZ was built. A nomogram consisting of pathologic T and riskScore was built, and the calibration curve illustrated that the nomogram could better forecast LIHC prognosis. Gene Set Enrichment Analysis (GSEA) demonstrated that DEGs between high and low risk groups were mainly involved in cell cycle. Finally, the qRT-PCR illustrated the expressions of nine model genes between normal and LIHC tissue. A prognostic model consisting of IGSF11, CD8A, ALCAM, CLDN6, JAM2, ITGB7, SDC3, CNTNAP1, and MPZ was obtained, which provides an important reference for the molecular diagnosis of patient prognosis.

Biosynthesis of selenoproteins by Saccharomyces cerevisiae and characterization of its antioxidant activities.

The study investigated biosynthesis of selenoproteins by Saccharomyces. cerevisiae using inorganic selenium. Selenium supplement via two stages was carried out during fermentation and the physicochemical characteristics of selenoproteins and its antioxidant activities were examined through in vitro assessment procedures. After fermentation, dry cells weight (7.47 g/L) and selenium content (3079.60 µg/kg) in the yeast were achieved when fermentation time points at the 6th hour and the 9th hour were chosen to supplement 30% and 70% of 30 µg/mL Na2SeO3 respectively. A maximal yield of selenium content in selenoproteins reached 1013.07 µg/g under optimized culture conditions and was 133-fold higher than the control. One new band with molecular weight of 26.76 KDa appeared in conjugated selenoproteins of sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Surface structure of selenoproteins and the control was different by Scanning electron microscopy images. Infrared spectrometry analysis demonstrated that groups of HSe, SeO and C-Se-O involved in selenoproteins were important pieces of evidence showing presence of Se embedded in the protein molecule. Selenoproteins showed strong antioxidant activities on DPPH·, OH and ·O2-, which was much higher than the control proteins. Therefore, the study provided an efficient selenium-enriched culture method of inorganic selenite to organic selenium and basis for selenoproteins applications.

[Dynamic evolution of MMP-2 gene expression and its enzymatic activities in experimental liver fibrosis].

OBJECTIVES: To explore the dynamic changes and interactions between MMP-2 and TIMP-2 during experimental liver fibrosis. METHODS: Wistar rats were randomly allocated into a normal group and a model group. To induce liver fibrosis, rats were injected intraperitoneally with dimethylnitrosamine (DMN) three consecutive times in the first week, then two consecutive times per week, totally for 6 weeks. In the normal control group, rats were injected with saline by the same method as the model group. Animals were sacrificed 1, 4, 10, 17, 28, 42, 56 days after starting DMN injections. Conventional histological examinations of the livers were performed with hematoxylin and eosin and Masson staining. The fibrosis was classified into 0 to 4 stages. Hydroxyproline content was determined after liver tissues were hydrolyzed in HCl at 160 degree C for 2 hrs and then measured with spectrometry at 560 nm wavelength. mRNA levels of MMP-2 and TIMP-2 were determined by semi-quantitive RT-PCR. Gelatinase activity of MMP-2 was examined by zymography using gelatin substrate. RESULTS: In the model group the hepatic MMP-2 mRNA expression started to increase 10 days after DMN administration and remained at a much higher level than in the normal group throughout the study period, while TIMP-2 mRNA expression started to be lower than in the normal group 17 days after DMN administration and reached the lowest level on the 28th day. Then it rapidly rebounded and remained higher than that in the normal group from the 42nd day to the end of the study period. TIMP-2/MMP-2 began to be lower by several days than that of the normal group after DMN administration through the remaining study period. Zymography showed that the enzymatic activities of both latent MMP-2 and active MMP-2 were increased during the process of liver fibrosis. CONCLUSION: In liver fibrosis, MMP-2 expression increases, while TIMP-2 expression relatively decreases. The enzymatic activities of MMP-2 increase as the liver fibrosis develops.

[The effect of angiotensin II type 1 receptor blocker valsartan in preventing hepatic fibrosis induced by dimethylnitrosamine in rats].

OBJECTIVE: To observe the effects of angiotensin II type 1 receptor blocker valsartan in preventing hepatic fibrosis induced by dimethylnitrosamine in rats. METHODS: Except rats in the control group, all were given intraperitoneal injections of 1% dimethylnitrosamine (DMN 1 ml/kg, two or three consecutive days/a week for 6 weeks). From the first day of the intraperitoneal injection, rats in treatment groups were given valsartan for 8 weeks by gastric gavage. Liver tissue and blood samples of all rats were examined at 56 days (8 weeks). AngII levels were determined by radioimmunoassay. Hepatic mRNA levels of Collagen type I (Col I) and tissue inhibitor of metalloproteinase1 (TIMP1) were evaluated by reverse-transcription polymerase chain reaction (RT-PCR). RESULTS: Valsartan significantly attenuated the degree of liver fibrosis and decreased the hepatic AngII content compared with DMN treated rats (P<0.01). mRNA levels of Col I and TIMP1 were upregulated in DMN treated rats compared with normal rats. Valsartan downregulated the elevation of Col I and TIMP1 mRNA levels (P<0.01). CONCLUSION: Hepatic AngII content of the model group was increased, the local tissue RAS was activated in DMN induced liver fibrosis. Valsartan can retard the progression of hepatic fibrosis and may provide an effective new strategy for anti-liver fibrosis therapy.

[Dynamic evolution of MMP-13, TIMP-1, type I and III collagen and their interaction in experimental liver fibrosis].

OBJECTIVE: To obtain a detailed pattern of the dynamic evolution and interactions among MMP-13, TIMP-1, type I and III collagen during experimental liver fibrosis. METHODS: Wistar rats were randomly allocated into a normal group, and a model group. To induce liver fibrosis, rats were intraperitoneally injected with dimethylnitrosamine (DMN) three consecutive times in the first week, then two consecutive times per week, totally for 6 weeks. In the normal control group, rats were treated with saline by the same means. Animals were sacrificed 1, 4, 10, 17, 28, 42, 56 days after starting DMN injections. Conventional histological examinations were performed after hematoxylin and eosin, and Masson stain. Fibrosis stages were classified into 0 to 4. Hydroxyproline contents were determined after liver tissues were hydrolyzed in HCl at 160 degrees C for 2 h and then measured with spectrometry at 560 nm wavelength. mRNA levels of MMP-13, TIMP-1, type I and III collagen were determined by semi-quantitive RT-PCR. RESULTS: In the model group, hepatic type I pro-collagen mRNA expression started to increase on the 10th day after DMN administration (t = 2.85, P < 0.05), type III started to increase on the 28th day (t = 4.16, P< 0.01), and TIMP-1 mRNA expression started to increase on the 4th day (t = 2.60, P < 0.05). They all remained much higher than in the normal group throughout the remaining study period. Hepatic MMP-13 mRNA expression started to increase on the 17th day after DMN administration and remained at a higher level than in the normal group until he 28th day (t = 4.08, P < 0.01), then gradually returned to normal level at the end of the study period. CONCLUSION: Although hepatic MMP-13 expression transiently increased during liver fibrosis, enhanced expression of TIMP-1 from the early periods of liver fibrosis inhibited the collagen degrading ability of MMP-13, therefore, over-expressed collagen accumulated in the liver. Thus, it is hypothesized that TIMPs play a pivotal role in liver fibrosis.