BACKGROUND:Silk fibroin, as an inorganic mineralization template, can induce hydroxyapatite crystal growth, and combined with nano-hydroxyapatite can simulate the inorganic and organic components of natural bone, which is expected to become the most ideal bone graft material. OBJECTIVE:To prepare the silk fibroin/nano-hydroxyapatite composite material and investigate its treatment outcomes in spinal fusion. METHODS:Silk fibroin/nano-hydroxyapatite composite was synthesized by the co precipitation method with silk fibroin and calcium phosphate as raw materials, to simulate the structure and composition of the bone tissue. The crystal phase composition and microstructure of the composite scaffold were analyzed by X-ray diffraction and transmission electron microscope. Osteoblasts were seeded onto the composite, and the cel adhesion and proliferation were observed under inverted microscope. The lumbar posterolateral spinal fusion models were established in the New Zealand rabbits, fol owed by treated with autologous bone graft (control group) and composite (composite group), respectively. The gross, radiological and histological observations of bone fusion were compared between groups. RESULTS AND CONCLUSION:Silk fibroin/nano-hydroxyapatite composite appeared to be fascicular under electron microscope, the length was 200-500 nm and width was 20-30 nm. And the hydroxyapatite was about 200 nm in length and 50 nm in width. X-ray diffraction showed that the bottom of diffraction peak was wide, and the peak was not sharp. Transmission electron microscope found that cel s adhered wel onto the composite scaffold at 2 days. Scanning electron microscope showed that the polygonal, oval or conical cel s covered most of the composite scaffold holes, presented obvious mitotic phase at 5 days. The third generation of MC3T3-E cel s tended to rise at first 3 days, and then decreased. The fusion site of L5-6 transverse process was strong, and non-bony fusion occurred. At the same time, numerous new bones were visible in the composite group. Hematoxylin-eosin staining showed a large number of cel aggregation, abundant osteoblasts surrounding cartilage, and the bone tissues were in a regular arrangement in the composite group. Moreover, irregular trabecular bone with medul ary cavity was found in the composite material. These results suggest that the silk fibroin/nano-hydroxyapatite composite with the similar structure and composition of natural bone can achieve satisfactory fusion effect in the rabbit lumbar posterolateral fusion.

BACKGROUND:Although the traditional surgical treatment can improve the symptoms of patients with senile osteoporotic vertebral fracture, the treatment easily produces bone graft fusion failure and pseudoarticulation formation and affects clinical effects. OBJECTIVE:To investigate biomechanical properties of anterior cervical pedicle screw and the effects on osteoporotic vertebral stability. METHODS:A total of 16 fresh cadaver cervical specimens contained 64 motion segments (C3-4, C4-5, C5-6 and C6-7). The 64 segments by the way of implantation were randomly divided into ordinary anterior locking screw fixation group and lower anterior vertebral pedicle screw system group (32 segments per group). The mechanical properties were determined on the biomechanical testing machine for each group. RESULTS AND CONCLUSION:(1) Biomechanics:Compared with the ordinary anterior locking screw fixation group, the maximum pul-out strength, screw path length, postoperative vertebral column height, the maximum surface strain, strain maximum and the range of maximum values were increased in the lower anterior vertebral pedicle screw system group (P<0.05). (2) Results suggest that compared with the ordinary anterior locking screw fixation group, lower cervical anterior pedicle screw required larger extraction force and was more stable for osteoporotic vertebrae.

Objective To assess the association between NOD2 gene single nucleotide polymorphisms (SNPs) and leprosy in Chinese Yi population.Methods Whole blood samples were obtained from 300 patients with leprosy and 300 healthy human controls of Yi nationality in Sichuan province.Genomic DNA was extracted,and a SNaPshot assay was performed to determine the genotypes of four single nucleotide polymorphisms (SNPs) of the NOD2 gene,including rs9302752,rs7194886,rs8057341 and rs3135499.Chi-square test was conducted to compare allele frequency,and Hardy-Weinberg equilibrium was tested.Results The genotype distribution of all the four SNPs was consistent with Hardy-Weinberg equilibrium (all P ＞ 0.05).Significant differences were observed between the patients with leprosy and healthy controls in both genotype distribution and allele frequency of the SNP rs3135499 (both P ＜ 0.01),but not in those of the other three SNPs (all P ＞ 0.05).Conclusion The SNP rs3135499 of the NOD2 gene may be associated with the development of leprosy in Chinese Yi population.

Objective Detailed descriptions of the double-labeling immunofluorescence staining for fluorescence microscopy provides an ideal sample.Methods Rat liver frozen sections were used fixative were 95％ alcohol,95％ formaldehyde and acetone,frozen sections,with anti-CSE,Ki-67 polyclonal antibody and incubated with FITC,Cy3 fluorescence-labeled secondary fluorescently labeled secondary antibody staining,observed under a fluorescence microscope.Results Acetone fixed group visible in the proliferative phase (S phase) cells showed a red fluorescent nucleus,cytoplasmic green fluorescence.Conclusion The impact of double labeling immunofluorescence the effect of sample links and many factors,including the two most important factors are 2 and coordination with the primary antibody and select the appropriate fixative.

Background Ultraviolet irradiation promotes cellular apoptosis by affecting the mitochondrial transmembrane potential,including human lens epithelial cells (LECs).Gene associated with retinoid-interferoninduced mortality-19 (GRIM-19),a cell death regulatory protein,is essential for the assembly and function of mitochondrial complex Ⅰ.However,whether LECs apoptosis induced by ultraviolet irradiation is related to GRIM-19 is still unclear.Objective The purpose of this study was to investigate the relationship between the apoptosis of human LECs caused by ultraviolet with GRIM-19 expression in vitro.Methods Human LEC line(SRA01/04)was cultured in α-MEM containing 10％ fetal bovine serum.The cells were exposed to ultraviolet ray at doses of 0,30,60,90,120 or 150 mJ/cm2 when cell growth reached the logarithmic phase and 80％ confluency.The rate of apoptosis of the cells was assayed using flow cytometry,and the level of expression and relative amount of GRIM-19 protein (GRIM-19/β-actin) were detected by Western blot.The relationship between apoptosis and the GRIM-19/β-actin value among the different treatment groups was compared using One-way ANOVA,and the correlation of LECs apoptosis rate and GRIM-19 expression level was assessed by Pearson linear analysis.Results A significant difference was found in the apoptosis rate among the different treatment groups(F=149.32,P＜0.01).Compared with the 0 mJ/cm2 ultraviolet irradiation group,the apoptosis rate of LECs was significantly increased in the 60,90,120 and 150 mJ/cm2 ultraviolet irradiation groups (q =17.02,-25.20,-29.41,-8.61,P ＜ 0.01).The expression of the GRIM-19 protein in the LECs suspension was enhanced by ultraviolet irradiation at 60,90,120 and 150 mJ/cm2.The relative expression of the GRIM-19 protein (GRIM-19/β-actin) was significantly different among the various groups (F=6.87,P＜0.05),and the GRIM-19/β-actin values in the 60,90,120,150 mJ/cm2 ultraviolet irradiation groups were elevated in comparison with the un-irradiated group(2.01±0.76,2.98± 1.80,3.97± 1.61,2.42± 1.28 vs.0.56±0.23),which showed statistically significant differences (q =4.12,-5.04,-7.09,-3.85,P ＜ 0.01).In addition,a positive correlation was seen between the rate of apoptosis and the expression of the GRIM-19 protein(r=0.71,P＜0.01).Conclusions GRIM-19 is expressed in normal human LECs.The apoptosis of human LECs accompanies the up-regulation of GRIM-19.The expression of GRIM-19 in LECs increases with ultraviolet irradiation in a doseindependent manner.

Background Epithelial-myofibroblast transition (EMT) of human lens epithelial cells (LECs) induced by transforming growth factor-β2 (TGF-β2) is the main mechanism in the pathogenesis of posterior capsular opacification(PCO).Seeking an effective drug capable of inhibiting this process is important for the prevention and treatment of PCO.Objective The purpose of this study was to investigate the inhibitory effect of rapamycin (RAPA)on the proliferation of human LECs and TGF-β2-induced EMT.Methods Human LEC strain(SRA01/04)was cultured in DMEM with high glucose and 10％ fetal bovine serum.The cells were consequently cultured in serumfree DMEM with 5 mg/L TGF-β2,TGF-β2+10 mg/L RAPA,TGF-β2 + 100 mg/L RAPA,TGF-β2 + 1000 mg/L RAPA or TGF-β2 +10 000 mg/L RAPA for 72 hours,and SRA01/04 cultured in serum-free DMEM were used as control.The proliferation rate(A490)of SRA01/04 in the different groups was detected using the MTT assay and the rate of inhibition of RAPA was calculated.The expressions of the α-smooth muscle actin(α-SMA) and E-cadherin(E-cad)mRNA and protein were detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot,respectively.The changes in the expression of α-SMA and E-cad in SRA01/04 were evaluated by Western blot 24,48 and 72 hours after TGF-β2 +400 mg/L RAPA treatment.Results The A490 value of SRA01/04 was 0.680±0.020,0.550±0.013,0.480±0.014,0.400±0.011 and 0.200±0.019 in the control group,TGF-β2 group,TGF-β2 + 10 mg/LRAPA group,TGF-β2 + 100 mg/L RAPA group,TGF-β2 + 1000 mg/L RAPA and TGF-β2 + 10 000 mg/L RAPA group,respectively,showing a gradually declining trend in SRA01/04 rate of proliferation with increasing RAPA concentrations (F =101.920,P =0.000).RT-PCR and Western blot assay showed that the relative expression levels of α-SMA mRNA (α-SMA mRNA/β-actin mRNA)and protein (α-SMA/β-actin)in the cells were significantly increased in the TGF-β2 treatment group.However,with exposure to RAPA,the relative expression levels of α-SMA mRNA and protein were significantly lowered with increasing RAPA concentrations,but the expression levels of E-cad mRNA and protein were raised (α-SMA mRNA:F =294.660,P =0.000 ; α-SMA protein:F =346.950,P =0.000 ; E-cad mRNA:F =264.250,P =0.000 ; E-cad protein:F =317.327,P =0.000).In addition,after exposure to 400 mg/L RAPA,the expression levels of α-SMA protein gradually reduced and those of E-cad protein gradually increased with increasing treatment durations,showing significant differences among the different time points (α-SMA:F =693.864,P =0.000 ;E-cad:F=369.286,P =0.000).Conclusions RAPA can inhibit the proliferation of SRA01/04 in vitro and arrest EMT of SRA01/04 induced by TGF-β2 in a dose-and time-dependent manner.

OBJECTIVETo analyse the polymorphism of squalene synthase gene and reveal the influence of squalene synthase (SQS) gene polymorphism on the catalytic efficiency of its encode enzyme in Glycyrrhiza uralensi.

METHODThe total RNA was extracted. PCR was used to amplify the coding sequences of squalene synthase gene, which were sequenced and analysed. The expression vectors containing different SQS gene sequences, including SQS1C, SQS1F, SQS2A, SQS2B, were constructed and transformed into Escherichia coli BL21. The fusion protein was induced to express by IPTG, then was isolated, purified and used to carry out the enzymatic reaction in vitro. GC-MS was used to analyse the production.

RESULTThere were three kinds of gene polymorphism existing in SQS1 gene of G. uralensis, including single nucleotide polymorphism (SNPs), insertion/deletion length polymorphism (InDels) and level of amino acid, the proportion of conservative replace of SQS1 was 53.94%, and there were 2 mutational sites in structural domains. The proportion of conservative replace of SQS2 was 60%, and there was 1 mutational site in structural domains. The production squalene could be detected by GC-MS in all the 4 kinds of enzymatic reactions. The capacity of accumulating squalene of SQS1F was higher than other SQS genes.

CONCLUSIONThe polymorphism of SQS gene was quite abundant in G. uralensis, which maybe the molecular foundation of the formation of high-quality liquorice.

Amino Acid Substitution ; Biocatalysis ; Cloning, Molecular ; DNA, Complementary ; chemistry ; genetics ; Electrophoresis, Polyacrylamide Gel ; Escherichia coli ; genetics ; Farnesyl-Diphosphate Farnesyltransferase ; genetics ; metabolism ; Gas Chromatography-Mass Spectrometry ; Glycyrrhiza uralensis ; enzymology ; genetics ; INDEL Mutation ; Isoenzymes ; genetics ; metabolism ; Molecular Sequence Data ; Plant Proteins ; genetics ; metabolism ; Polymorphism, Genetic ; Polymorphism, Single Nucleotide ; Recombinant Proteins ; metabolism ; Sequence Analysis, DNA ; Squalene ; metabolism

BACKGROUND: The present study aimed to determine the short-term and long-term outcomes of critically ill patients with acute respiratory insufficiency who had received sedation or no sedation. METHODS: The data of 91 patients who had received mechanical ventilation in the first 24 hours between November 2008 and October 2009 were retrospectively analyzed. These patients were divided into two groups: a sedation group (n=28) and a non-sedation group (n=63). The patients were also grouped in two groups: deep sedation group and daily interruption and /or light sedation group. RESULTS: Overall, the 91 patients who had received ventilation ≥48 hours were analyzed. Multivariate analysis demonstrated two independent risk factors for in-hospital death: sequential organ failure assessment score (P=0.019, RR 1.355, 95％CI 1.051–1.747, B=0.304, SE=0.130, Wald=50483) and sedation (P=0.041, RR 5.015, 95％CI 1.072–23.459, B=1.612, SE=0.787, Wald=4.195). Compared with the patients who had received no sedation, those who had received sedation had a longer duration of ventilation, a longer stay in intensive care unit and hospital, and an increased in-hospital mortality rate. The Kaplan-Meier method showed that patients who had received sedation had a lower 60-month survival rate than those who had received no sedation (76.7％ vs. 88.9％, Log-rank test=3.630, P=0.057). Compared with the patients who had received deep sedation, those who had received daily interruption or light sedation showed a decreased in-hospital mortality rate (57.1％ vs. 9.5％, P=0.008). The 60-month survival of the patients who had received deep sedation was significantly lower than that of those who had daily interruption or light sedation (38.1％vs. 90.5％, Log-rank test=6.783, P=0.009). CONCLUSIONS: Sedation was associated with in-hospital death. The patients who had received sedation had a longer duration of ventilation, a longer stay in intensive care unit and in hospital, and an increased in-hospital mortality rate compared with the patients who did not receive sedation. Compared with daily interruption or light sedation, deep sedation increased the in-hospital mortality and decreased the 60-month survival for patients who had received sedation.

BACKGROUND:Esophagectomy is a very important method for the treatment of resectable esophageal cancer, which carries a high rate of morbidity and mortality. This study was undertaken to assess the predictive score proposed by Ferguson et al for pulmonary complications after esophagectomy for patients with cancer. METHODS:The data of patients who admitted to the intensive care unit after transthoracic esophagectomy at Cancer Hospital of Chinese Academy of Medical Sciences and Peking Union Medical College between September 2008 and October 2010 were retrospectively reviewed. RESULTS:Two hundred and seventeen patients were analyzed and 129 (59.4％) of them had postoperative pulmonary complications. Risk scores varied from 0 to 12 in all patients. The risk scores of patients with postoperative pulmonary complications were higher than those of patients without postoperative pulmonary complications (7.27±2.50 vs. 6.82±2.67;P=0.203). There was no significant difference in the incidence of postoperative pulmonary complications as well as in the increase of risk scores (χ2=5.477,P=0.242). The area under the curve of predictive score was 0.539±0.040 (95％CI 0.461 to 0.618;P=0.324) in predicting the risk of pulmonary complications in patients after esophagectomy. CONCLUSION:In this study, the predictive power of the risk score proposed by Ferguson et al was poor in discriminating whether there were postoperative pulmonary complications after esophagectomy for cancer patients.

BACKGROUND:Readmission to intensive care unit (ICU) after discharge to ward has been reported to be associated with increased hospital mortality and longer length of stay (LOS). The objective of this study was to investigate whether ICU readmission are preventable in critical y il cancer patients. METHODS:Data of patients who readmitted to intensive care unit (ICU) at National Cancer Center/Cancer Hospital of Chinese Academy of Medical Sciences (CAMS) and Peking Union Medical College (PUMC) between January 2013 and November 2016 were retrospectively collected and reviewed. RESULTS:A total of 39 patients were included in the final analysis, and the overall readmission rate between 2013 and 2016 was 1.32％ (39/2,961). Of 39 patients, 32 (82.1％) patients were judged as unpreventable and 7 (17.9％) patients were preventable. There were no significant differences in duration of mechanical ventilation, ICU LOS, hospital LOS, ICU mortality and in-hospital mortality between patients who were unpreventable and preventable. For 24 early readmission patients, 7 (29.2％) patients were preventable and 17 (70.8％) patients were unpreventable. Patients who were late readmission were all unpreventable. There was a trend that patients who were preventable had longer 1-year survival compared with patients who were unpreventable (100％ vs. 66.8％, log rank=1.668, P=0.196). CONCLUSION:Most readmission patients were unpreventable, and all preventable readmissions occurred in early period after discharge to ward. There were no significant differences in short term outcomes and 1-year survival in critically ill cancer patients whose readmissions were preventable or not.