ObjectiveTo analysis the death causes of patients with acute traumatic spinal cord injury(ATSCI) during the initial four months, and determine the clinical predictors of mortality. MethodsMultiple factors that could contribute to mortality were evaluated in the retrospective study. Results3.3% of ATSCI patients died during the initial four months. The death causes were multiple factors inculding arrest of heart and respiration resulting from excessive stimulus to trachea and bronchus, interventions of surgical procedures, pneumonia, multiple organ failure, jam of sputum, and inhalation of food, etc. ConclusionsThe leading causes were the arrest of heart and respiration resulting from excessive stimulus to trachea and bronchus, interventions of surgical procedures and pneumonia. The predictors of mortality include nurological level at C5 and above, motor index of upper extremities less than 6, copious sputum, disability of cough, low blood oxygen pressure especially with progressive decrease, and neurological deterioration, etc.

Objective To obtain optimum scheme on reaction system for randomly amplified polymorphic DNA(RAPD) of Pseudomonas aeruginosa. Methods The optimum reaction systems of RAPD were obtained by single factor design and response surface design(RSD). Results The multiple optimum reaction systems caused by effect of interaction on multifactor were observed in single factor design, and the concentrations of key components of the optimum reaction system with good stability obtained by response surface design were 2.0 mmol/L of Mg 2+ , 0.5 ?mol/L of primer and 0.5 ng/?l of template. Conclusion To optimize the reaction system of RAPD, RSD is simpler, more scientific, and more reasonable than single factor design.

Objective:To obtain optimum template for randomly amplified polymorphic DNA(RAPD). Methods:Four different methods(rod winding, precipitation, boiling and lysis)for extraction of template DNA were used.The length and purity of template DNA and its RAPD profile were observed separately by agarose gel electrophoresis, spectrophotometric scan assays and RAPD reaction.Results:The template DNA (>23 kb) with high purity and the same RAPD profile with 2-4 kb DNA fragments were obtained by both rod winding and precipitation method. However,the template DNA (4 kb and 2 kb,respectively) with break and dispersion and low purity was extracted by method of boiling and lysis, and 600-2 000 bp DNA fragments were seen in the similar RAPD profile.Conclusions: Template DNA extracted by rod winding or precipitation method was optimized for RAPD.

The separation of anionic compounds by strong anion-exchange capillary electrochromatography (SAX-CEC) was carried out. It was found that the analytes could be absorbed onto the stationary phase, and this would lessen the retention factors (k) of the analytes, thus the column separation capability decreased. For the acidic compounds, k increased with increase of applied voltage. And the change of the applied voltage could provide different separation selectivity for the solutes. The separation with different eluent was studied. It showed that the logarithm of the capacity factor linearly decreased with increase of the logarithm of the ionic strength. The different retention behavior of the anionic compounds in SAX-CEC and CE was also studied

The incidence rate of primary liver cancer continues to increase around the world, with a younger age of onset and poorer prognosis. As the most classic regulator of cell polarity and density, mechanical signal transduction, cell proliferation, and organ development, the Hippo pathway can promote the development and progression of various cancers including primary liver cancer. YAP, a classic nuclear effector of the Hippo pathway, is significantly upregulated in primary liver cancer and promotes the development of drug resistance. This article aims to investigate the association of the dysregulation of the Hippo signaling pathway with the development and progression of primary liver cancer and analyzes the mechanism of action of the Hippo signaling pathway in the drug resistance of primary liver cancer as an early event of the development of primary liver cancer, which is of great significance for exploring new treatment strategies for primary liver cancer.

ObjectiveTo investigate the effect of the Yap1 gene and tanshinone ⅡA on the proliferation, migration, and invasion abilities of Huh-7 hepatoma cells. MethodsA total of 10 pairs of human hepatocellular carcinoma (HCC) samples and adjacent tissue samples were collected in Zhongnan Hospital of Wuhan University from June 1 to December 1, 2019. Quantitative real-time PCR and Western blotting were used to measure the expression of the Yap1 gene and phenotype-related molecules. MTT cell proliferation detection reagent was used to measure the inhibition rate of cell proliferation after the treatment with different concentrations of tanshinone ⅡA. Western blotting was used to measure the changes in the expression of apoptosis-and migration-related markers after different interventions. Flow cytometry and Transwell assay were used to measure apoptosis and cell migration and invasion abilities. The data of 375 cases of liver cancer and 50 cases of relatively normal liver tissue samples were downloaded from The Cancer Genome Atlas, including clinicopathological information. The t-test was used for comparison of continuous data between two groups, and the chi-square test was used for comparison of categorical data between two groups. ResultsIn 8 of the 10 pairs of HCC samples and adjacent tissue samples, HCC samples had significantly higher expression of Yap1 than the adjacent tissue samples. Compared with the normal human liver epithelial cells L02, the Huh-7 and HCCL-M3 hepatoma cells had a significant increase in the expression of Yap1. The silencing efficiency of si-Yap1-3 transfection reached 87.004% at the protein level. MTT results showed that tanshinone ⅡA effectively inhibited the proliferation of Huh-7 cells, with a half inhibitory concentration of 8.683 μmol/L. After the cells were treated with si-Yap1-3 and tanshinone ⅡA, there was an increase in the expression of the downstream marker for proliferation and migration E-cadherin and a reduction in the expression of vimentin, and the results of Transwell assay showed that compared with the si-NC group, the tanshinone ⅡA+si-Yap1-3 group had significant reductions in the migration and invasion abilities of Huh-7 cells (migration: 43.19±2.88 vs 132.20±10.03, t=8.527, P=0.001; invasion: 53.95±4.20 vs 179.10±11.11, t=4.484, P=0.011). The group treated with si-Yap1-3 and tanshinone ⅡA had an increase in the expression of the apoptosis-related marker Bax and a reduction in the expression of Bcl-2, as well as a significantly higher early apoptosis rate than the si-NC group (2598% vs 9.21%, χ2=4.078, P＜0.05). ConclusionOncogene Yap1 silencing combined with tanshinone ⅡA can promote the apoptosis of Huh-7 hepatoma cells and inhibit their migration and invasion, which can provide certain guiding significance for clinical medication.

The human serum proteome is closely associated with the state of the body. Endogenous peptides derived from proteolytic enzymes cleaving on serum proteins are widely studied due to their potential application in disease-specific marker discovery. However, the reproducibility of peptidome analysis of endogenous peptides is significantly influenced by the proteolytic enzymes within body fluids, thereby limiting the clinical use of the endogenous peptides. We comprehensively investigated the N and C terminus of endogenous peptides using peptidomics. The cleavage site patterns of the N and C terminus and adjacent sites from all the identified endogenous peptides were highly conserved under different sample preparation conditions, including long-term incubation at 37°C and pretreatment with repeated freeze-thaw cycles. Furthermore, a distinguishable cleavage site pattern was obtained when a different disease serum was analyzed. The conserved cleavage site pattern derived from proteolytic enzymes holds potential in highly specific disease diagnosis.
Carcinoma, Hepatocellular ; blood ; diagnosis ; Chromatography, High Pressure Liquid ; Chromatography, Reverse-Phase ; Humans ; Liver Neoplasms ; blood ; diagnosis ; Mass Spectrometry ; Nanotechnology ; Peptide Hydrolases ; metabolism ; Peptides ; blood ; Protein Structure, Tertiary ; Proteome ; analysis ; Proteomics ; Silicon Dioxide ; chemistry ; Time Factors

Objectives: To investigate the expression of phosphoserine aminotransferase 1 (PSAT1) in pancreatic cancer tissues, and its potential role in pancreatic cancer. Methods: The expression of PSAT1 in 98 human pancreatic cancer tissues, which were collected from the People＇s Hospital of Guizhou, between July 2013 to July 2017, and the corresponding adjacent normal tissues was analyzed by immunohistochemical staining. Additionally, the relationship between the expression of PSAT1 and the clinicopathological parame-ters, overall survival (OS), and disease-free survival (DFS) of patients with pancreatic cancer was evaluated. The human pancreatic can-cer cell lines, BxPC-3 and SW1990, were transfected with PSAT1-siRNA, to investigate the effect of PSAT1 knockdown on cell prolifera-tion, migration, and invasion. Additionally, we performed Western blot to assess the expression of PI3K/Akt/mTOR-related proteins in PSAT1-knockdown cells. Results: The percentages of PSAT1-positive cells in pancreatic cancer and adjacent non-tumor tissues were 69.4% (68/98) and 5.0% (5/98), respectively, indicating a significantly higher expression of PSAT1 in pancreatic cancer tissues com-pared to adjacent non-tumor tissues (P<0.05). The increased expression of PSAT1 in pancreatic cancer tissues correlated with lymph node metastasis and TNM stage. Kaplan-Meier analysis suggested that a high expression of PSAT1 correlated with a poor OS and DFS compared to a low expression of PSAT1 (P<0.05). Cox regression analysis showed that the expression of PSAT1 is an independent prog-nostic marker for OS and DFS in pancreatic cancer patients (P<0.05, all). Transient transfection of BxPC-3 and SW1990 cells with PSAT1-siRNA markedly reduced the cell proliferation, migration, and invasion abilities of these cells compared to transfection with NC-siRNA (P<0.05). Knockdown of PSAT1 in pancreatic cancer cells also inhibited the expression of p-Akt and p-mTOR (P<0.05). Conclusions: The expression of PSAT1 increases in human pancreatic cancer tissues and cell lines. Additionally, PSAT1 regulates cell proliferation and in-vasion through the PI3K/Akt/mTOR pathway.

Objective To investigate the mechanism and protective effect of Humanin(HN)on rotenone(Rot)-induced toxic damage for dopamine neurons.Methods The Rot-poisened PC12 cell model was constructed,and the control group,the Rot poisening group,the HN pretreated Rot poisening group,and the HN treatment group were set up.ELISA was used to detect the content of HN inside and outside of Rot-infected cells,CCK-8 assay was used to detect cell viability,and ATP detection kit was used to detect the intracellular ATP content.Dichloro-dihydro-fluorescein diacetate(DCFH-DA)assay was used to detect the level of reactive oxygen species(ROS)in cells.Western blotting was performed to detect the expression level of mitochondrial autophagy regulatory proteins Pink1,Parkin,p62,LC3,mitochondrial biogenesis regulatory protein PGC1α,division/fusion regulatory proteins OPA1,MFN2,DRP1,p-DRP1 and antioxidant stress regulatory proteins Keap1 and Nrf2.HBAD-mcherry-EGFP-LC3 adenovirus transfected cells was used to observed the number of autophagosomes and autophagolysosomes.Results The results showed that the intracellular concentration of HN in PC12 in the Rot poisening group was significantly higher than that in the control group(P＜0.05);Compared with the control group,the Rot poisening group had significantly decreased activity of PC12 cells,decreased ATP content and increased production of ROS.After the poisen of Rot in PC12 cells,the expression of Pink1 and p-Parkin,the ratio of LC3Ⅱ/LC3Ⅰ and the expression of p-DRP1 in mitochondrial fusion protein was increased,while the expression of p62,the expression of mitochondrial biogenesis protein PGC1 α,mitochondrial fusion proteins MFN2 and OPA1,and antioxidant stress proteins Keap1 and Nrf2 were decreased(all P＜0.05).The number of autophagosomes and autophagolysosomes in PC12 cells in the Rot poisening group was higher than that in the control group(P＜0.05),and HN pretreatment(20 μmol/L)could significantly improve the changes mentioned above caused by Rot poisening(P＜0.05).Conclusion HN ameliorates Rot-induced toxic damage for dopamine neurons by inhibiting mitophagy and mitochondrial division and promoting mitochondrial biogenesis and fusion,and anti-oxidative stress.

OBJECTIVETo investigate the spiral ganglion degeneration and the expression of EFR3A in the cochlea of the deaf mice induced by co-administration of kanamycin and furosemide.

METHODSEight weeks old C57BL/6J mice were administered with a single dose of kanamycin followed by furosemide, then fluorescent immunohistochemistry staining and transmission electron microscopy were applied to observe the SGNs' degeneration process and extent characteristics at 1, 5, 15, 30 and 60 days following treatment. We detected the expression of EFR3A during the degeneration of SGNs via fluorescent immunohistochemistry and western blotting.

RESULTSCo-administration of kanamycin and furosemide quickly induced cochlear hair cell death in mice, and then caused progressive degeneration of SGNs. Our results showed that the abnormal morphology of SGNs occurredat day 5 following administration, and the number of SGNs began to decrease at day 15. Compared to the control group, it was found the remarkable increase of the EFR3A protein at the fifth day after co-administration, then decreased to the nearly normal at 15 days following treatment, and no further significant changes thereafter.

CONCLUSIONThe changes of the EFR3A protein expression in the spiral ganglion of the cochlea in mice are coincidence with the time of the SGNs degeneration to happen, which imply that EFR3A may play an important role in the occurrence of the SGNs' degeneration in the cochlea in mice following hair cells loss.

Animals ; Cochlea ; metabolism ; Furosemide ; Hair Cells, Auditory ; Kanamycin ; Membrane Proteins ; metabolism ; Mice ; Mice, Inbred C57BL ; Saccharomyces cerevisiae Proteins ; metabolism ; Spiral Ganglion ; metabolism ; pathology