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Objective To explore the feasibility of silk fibron(SF)-phlorotannins(PT) double layer membranes as barrier membrane for guided bone regeneration(GBR) by exploring the mechanical property, biocompatibility, and the effect on osteogenic differentiation of SF-PT double layer membranes. Methods The SF solution treated with different degumming time was prepared into SF bilayer by solution cast technique, ice template method and lyophilization. The tensile strength of the membranes under hydration were measured via mechanical universal machine(n=5). The microstructures of the composite membranes with the highest tensile strength under hydration condition were observed by scanning electron microscopy. Then the rough surfaces of SF double layer membranes was loaded with PT to prepare SF-PT double layer membranes, Fourier transformed infrared spectroscopy was carried out to characterize the interaction between PT and SF double layer membranes,and the bone marrow mesenchymal stem cells of human alveolar origins( h ABMSCs) adhesion was studied by SEM. CCK-8 assay and live dead staining were used to evaluated the biocompatibility of SF-PT double layer membranes. qRT-PCR,ALP staining and alizarin red stain were carried out to measure the osteogenic differentiation of h ABMSCs in different groups. Results The SF membranes prepared from SF solution with a degumming time of 30 min has the highest tensile strength,reaching( 3. 293 ± 0. 122 8) MPa. The smooth surface of SF double layer membrane was dense and smooth,while the rough surface was longitudinally ordered loose porous structure. PT was loaded onto the rough surface of the SF double layer membranes by hydrogen bond,hydrophobic interaction and other forces. There was no significant difference in the results of CCK-8 and live dead staining between blank group,SF group and PT-SF group. Compared with blank group and SF group,PT-SF double layer membranes promoted osteogenic differentiation of h ABMSCs. Conclusion Thus,the PT-SF double layer membranes expressed desired mechanical,biological properties and can promote the osteogenic differentiation of h ABMSCs to a certain extent. These results indicate that PT-SF double layer membranes is successfully fabricated,providing a solid foundation for future in vivo experiments.

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Objective To investigate the effects of interleukin(IL)-10 on the proliferation of HaCaT cells and CaCl2induced expression of differentiation markers and its possible molecular mechanisms. Methods HaCaT cells were treated with various concentrations of IL-10(0, 3, 10, 30 ng/ml) for different time(0, 24, 48, 72 h), cell proliferation was measured using MTS, and cell cycle was determined by flow cytometry. HaCaT cells were pretreated with IL-10(final concentration 10 ng/ml) for 1 h, then incubated with or without CaCl2(final concentration 1.2 mmol/L) for 24, 48, 72 h, Western blot was performed to detect the effect of IL-10 on the expression of HaCaT keratinocyte differentiation markers. After pretreatment of HaCaT cells with PD98059, an inhibitor of mitogen-activated kinase-ERK1/2, and LY294002, an inhibitor of phosphatidylinositol kinase-serine/threonine kinase(PI3K-AKT), the total RNA and proteins were extracted separately, real time quantitative polymerase chain reaction(RT-qPCR) and Western blot were used to examine the influence of IL-10 on the expression of differentiation markers(Keratin1, Keratin5, Involucrin). Results MTS results revealed that IL-10(30 ng/ml and lower doses) did not alter the proliferation of HaCaT cells in 72 h. Flow cytometry analysis demonstrated that IL-10 had no significant influence on cell cycle progression. The results of Western blot showed that IL-10 upregulated the expression of differentiation markers Involucrin, while there was no significant effect on Keratin1 and Keratin5. Mechanism research analysis demonstrated that IL-10 could activate ERK1/2 and AKT, increase their phosphorylation levels; RT-qPCR and Western blot results showed that PD98059 and LY294002 partially blocked IL-10 induced Involucrin expression. Conclusion At a particular concentration range, IL-10 has little effect on HaCaT proliferation, but it partially upregulates the expression of differentiation marker Involucrinviathe MAPKs-ERK1/2 and PI3K-AKT pathways.

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Objective To investigate the effect of berberine(BBR) on the proliferation and autophagy of mesangial cells in high glucose(HG) environment and the specific molecular mechanism. Methods Mesangium cells at exponential growth stage were divided into the following groups: normal group, high glucose group, high glucose + BBR treatment group(30, 60 and 90 μmol/L), high glucose + BBR(90 μmol/L) + AMPK inhibitor Compound C group(CC group); the number of mesangial cells was calculated by high content cell imager. The expressions of type Ⅳ collagen(Col-Ⅳ), fibronectin(FN) and microtubule-associated protein 1 light chain 3B(LC3B) in mesangial cells were detected by immunofluorescence assay. The protein expression levels of LC3B, Beclin-1, p62, Col-Ⅳ, FN and silencing regulatory factor 1(SIRT1)/adenylate activated protein kinase(AMPK) signaling pathway were detected by Western blot. Results Compared with the normal group, high content cell imaging showed abnormal proliferation of mesangial cells in the hyperglycemic group. The results of immunofluorescence and Western blot showed that the expression levels of Col-Ⅳ and FN deposited in mesangial extracellular matrix increased in the high glucose group. The results of Western blot showed that the protein expressions of SIRT1, p-AMPK, LC3B and Beclin-1 decreased, while the protein expressions of p-p65 and p62 increased. BBR inhibited the abnormal proliferation of mesangial cells induced by high glucose. BBR could reduce the expression levels of Col-Ⅳ and FN deposited in mesangial extracellular matrix. BBR could increase the expressions of SIRT1, p-AMPK, LC3B and Beclin-1 proteins in mesangial cells, while decrease the expressions of p-p65 and p62 proteins. CC group weakened the inhibition of mesangial cell proliferation and autophagy by high dose BBR. Conclusion Berberine can effectively inhibit the proliferation of mesangial cells induced by high glucose and increase the level of autophagy, which may be related to SIRT1/AMPK signaling pathway.

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Objective To investigate the reversal effect of cinobufagin(CINO) combined with 5-fluorouracil(5-FU) on human colorectal cancer(CRC) drug-resistant cell line HCT15/5-FU, and to clarify the regulatory role of hypoxia-inducible factor-1α(HIF-1α)/vascular endothelial growth factor(VEGF) pathway in reversing chemoresistance of colorectal cancer. Methods MTT assay was used to detect the changes of drug resistance and drug resistance index, flow cytometry was used to evaluate the apoptosis of cells, scratch test and Transwell assay were used to detect the changes of cell migration and invasion ability. Western blot was used to detect the expressions of epithelial-mesenchymal transition(EMT) related proteins and HIF-1α/VEGF pathway-related proteins. Results Compared with HCT15 cells, the resistance index of HCT15/5-FU was about 8.720. CINO combined with 5-FU could significantly enhance the drug sensitivity of HCT15/5-FU cells, reduce drug resistance index, up-regulate the level of apoptosis, and inhibit cell migration and invasion in a dose-dependent manner. Western blot results showed that CINO combined with 5-FU could inhibit the activity of EMT and HIF-1α/VEGF pathway. Conclusion CINO can reverse 5-FU resistance of colorectal cancerin vitro, and its mechanism may be related to the regulation of the HIF-1α/VEGF pathway and the inhibition of EMT and angiogenesis.

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Objective To investigate whether Mitochondria-targeted antioxidant peptide SS-31 can inhibit the ozone(O3)-induced mice lung airway hyperresponsiveness and mucus hypersecretion. Methods Eight-week C57BL/6 mice were randomized into four groups, including phosphate buffer saline(PBS)+Air group, SS-31+Air group, PBS+O3group and SS-31+O3group. C57BL/6 mice were injected intraperitoneally with SS-31(10 mg/kg) one hour before ozone exposure, and then single-exposed to ozone at a concentration of 5.01×10-6mol/m3for 3 hours. After 24 hours, airway hyperresponsiveness(AHR) and bronchoalveolar lavage fluid(BALF) cells numbers were measured. Lung tissue schiff periodic acid shiff(PAS) staining, malondialdehyde(MDA), inflammatory factors(interleukin, IL)-1β, IL-6, IL-18 and monocyte chemoattractant protein-1(MCP-1)) and mucin factor(MUC5B)were detected, and the protein expression levels of NOD-like receptor thermal protein domain associated protein 3(NLRP3), pro-Caspase 1/Caspase 1(p20), Gasdermin D(GSDMD) and Cleaved GSDMD were determined by Western blot. Results O3exposure caused both mice lung airway hyperresponsiveness and mucus hypersecretion. However, SS-31 could inhibit the O3-induced airway hyperresponsiveness and mucus secretion, reduce the levels of oxidative stress and inflammatory factor mRNA expression, and downregulate the protein expression level of NLRP3 and the activated forms of Caspase 1 and GSDMD. Conclusion SS-31 could suppress O3-induced mice airway hyperresponsiveness and mucus hypersecretion by inhibiting the NLRP3/Caspase 1/GSDMD signaling pathway.

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Objective To observe the therapeutic effect of electroacupuncture on rabbit knee osteoarthritis(KOA) model and to explore the mechanism of electroacupuncture in the treatment of KOA in rabbits. Methods Eighteen healthy New Zealand rabbits were randomly divided into a blank control group, a model control group and an electroacupuncture group, with 6 rabbits in each group. The rabbits were kept normally in the blank control group, while the model control and electroacupuncture groups were fixed in a modified Videman left hind limb extension and braking position for 6 weeks to create a KOA model,and electroacupuncture was given to the electroacupuncture group 1 week after completion of the modeling for 4 weeks,five times a week for 20 min each time. The functional score of the left knee joint of rabbits was evaluated by Lequesne's functional algorithm index; serum and joint fluid interleukin( IL)-1β and IL-6 levels were measured by enzyme-linked immunosorbent( ELISA) assay; serum and joint fluid nitric oxide( NO) levels were measured by Griess; morphological changes in knee cartilage were observed by hematoxylin-eosin( HE) staining,and the degree of cartilage degeneration was evaluated by Mankin' s scale; p-P38MAPK positive cell expression in knee cartilage was observed by immunofluorescence technique. Results Compared with the blank control group,the Lequesne score increased in the model control group( P<0. 05); the levels of IL-1β,IL-6,and NO in serum and joint fluid increased( P<0. 05); the cartilage surface was less smooth and the Mankin score increased( P<0. 05); and the immunofluorescence staining positive cells increased( P<0. 05). Compared with the model control group,the Lequesne score was reduced in the electroacupuncture group( P<0. 05); the levels of IL-1β,IL-6,and NO in serum and joint fluid were reduced( P<0. 05); the cartilage surface was smoother,and the Mankin score was reduced( P<0. 05); and the immunofluorescence staining positive cells were reduced( P<0. 05). Conclusion Electroacupuncture can effectively improve knee joint behavior in rabbit KOA model,and the mechanism may be to delay the onset of KOA by decreasing the expression level of inflammatory factors and affecting the expression of MAPK signaling pathway.

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Objective To investigate the mechanism of AMD3100 reversing the resistance of human hepatocellular carcinoma cells to Sorafenib by regulating carbonic anhydrase Ⅸ(CA9) and chemokine receptor 4(CXCR 4).Methods The Sorafenib resistant cell lines Huh7/Sor and HepG2/Sor were established from human hepatocellular carcinoma cells Huh7 and HepG2.The effects of Sorafenib alone or in combination with AMD3100 on the proliferation of Huh7,HepG2,Huh7/Sor, HepG2/Sor cells were detected.The difference of invasive ability between Sorafenib resistant cells and non-resistant cells, and the effect of AMD3100 on the invasive ability of hepatocellular carcinoma cells were observed.And the regulation effect of Sorafenib alone or in combination with AMD3100 on the expression of CA9 and CXCR4 proteins in Huh7,HepG2,Huh7/Sor and HepG2/Sor cells was detected.Results Compared with the Control group, AMD3100(50 μmol/L) increased the inhibitory effect of Sorafenib on the proliferation of Huh7/Sor and HepG2/Sor cells(P<0.05).Compared with Huh7 cells, the invasive ability of Sorafenib resistant Huh7/Sor cells was significantly enhanced(P<0.05),and AMD 3100(50 μmol/L) decreased the invasive ability of Huh7 and Huh7/Sor cells(P<0.05).Compared with Huh7 and HepG2 cells, the protein expression of CA9 and CXCR4 in Huh7/Sor and HepG2/Sor cells increased(P<0.05).AMD3100(50 μmol/L) down-regulated the protein expression of CA9 and CXCR4 in Huh7,HepG2,Huh7/Sor and HepG2/Sor cells(P<0.05).Conclusion AMD3100 can reduce the resistance of human hepatocellular carcinoma cells to Sorafenib by down-regulating the expression of CA9 and CXCR4,and enhance the inhibition of Sorafenib on the proliferation and invasion of human hepatocellular carcinoma cells.

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Objective To explore the correlation between epidermal growth factor receptor(EGFR) mutation and brain metastasis in lung adenocarcinoma. Methods Comparisons of brain metastasis between lung adenocarcinoma patients with EGFR exon 19 deletion(19 Del) and a single point mutation of L858R in exon 21(21 L858R) at initial diagnosis and after EGFR-TKIs targeted therapy were analyzed. The CCK-8 assay was used to detect and calculate the semi-inhibitory concentration(IC50) of gefitinib against EGFR-mutated lung adenoma cell lines PC9 and H3255. Results Of the 410 patients with lung adenocarcinoma, a total of 153(37.3%) cases had brain metastasis. Age, lymph node metastasis and 21 L858R mutation were high-risk factors for brain metastasis of lung adenocarcinoma. Among newly diagnosed 48 EGFR-mutated lung adenocarcinoma patients with brain metastasis, the number of 19 Del and 21 L858R were 14 and 34, respectively(P=0.006). There were 17 patients with 19 Del and 20 patients with 21 L858R were observed to complicated by brain metastasis after receiving EGFR-TKIs targeted therapy. The median time of brain metastasis after EGFR-TKIs treatment were 15 months(8.50, 25.00) and 7 months(4.25, 12.25), respectively(P=0.013). The IC50of PC9 and H3255 to gefitinib were(0.037±0.008)and(0.150±0.040)μmol/L, respectively(P=0.007). Conclusion Age younger than or equal to 60 years, lymph node N2-3 stage, and EGFR 21 L858R mutation are high-risk factors for brain metastases in patients with lung adenocarcinoma, and the latter still has a shorter time to brain metastases than 19 Del mutations after treatment with EGFR-TKIs, which may be related to drug sensitivity.

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Objective To investigate the effect of18F-deoxythymidine nucleoside(18F-FLT) positron emission computed tomography(PET/CT) imaging to evaluate the radiosensitization effect of Olaparib on breast cancer model in nude mice. Methods According to the random number table method, twenty-four BALB/C nude mice MCF-7 breast cancer models were established and divided into four groups with 6 mice in each group, namely the control group, radiotherapy group, Olaparib group and Olaparib + radiotherapy group.18F-FLT micro PET/CT imaging was performed on nude mice before and 48 h after treatment, respectively. The changes of maximum standardized uptake value(SUVmax), total proliferation volume(TPV) and tumor volume before and after tumor treatment in four groups were compared. The tumors were extracted and weighed to observe the changes of tumor weight, and the expression of Ki-67 and PCNA was analyzed by immunohistochemistry staining. The correlation of tumor SUVmaxwith Ki-67 and PCNA was analyzed. Results Before treatment, there were no significant differences in SUVmax, TPV and tumor volume among the 4 groups(F=0.041, 0.061, 0.045,P>0.05). 48 h after treatment, SUVmaxin the control and Olaparib groups increased significantly(t=-12.111,P<0.001;t=-3.001,P=0.03), SUVmaxwas reduced in the radiotherapy and Olaparib + radiotherapy groups(t=5.829,P<0.01;t=4.448,P<0.01), while SUVmax, TPV and tumor volume of tumors in the Olaparib + radiotherapy group were lower than those in the radiotherapy group(t=3.388, 5.884, 5.990,P<0.01).Tumor weight was significantly lower in the Olaparib + radiotherapy group than in the other three groups(F=44.405,P<0.001). Immunohistochemical staining showed that Ki-67 and PCNA were the least expressed in the Olaparib + radiotherapy group than in the other three groups(F=16.289,39.645,P<0.001). SUVmaxwas positively correlated with Ki-67 and PCNA expression(r=0.920,0.918,P<0.01). Conclusion 18F-FLT Micro PET/CT imaging can evaluate the radiosensitizing effect of Olaparib on nude mouse breast cancer model.

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Objective To investigate the effects of miR-130a-3p targeting Sirtuin7(Sirt7) on the migration, invasion and epithelial-mesenchymal transformation(EMT) of human trophoblast cells HTR-8/SVneo under hypoxic condition and its mechanism. Methods HTR-8/SVneo cells were studied as subjects, miR-130a-3p inhibitor and its negative control inhibitor-NC were transfected into the cells or combined with Sirt7 inhibitor 97491 intervention, treated with hypoxia(1% O2) for 48 h. qRT-PCR was used to detect the expression levels of miR-130a-3p and Sirt7 mRNA. Transwell assay was used to detect the ability of cells migration and invasion. The protein expression levels of Sirt7, HIF-1α, MMP-2, MMP-9, E-cadherin, N-cadherin and Vimentin were detected by Western blot. The targeting relationship between miR-130a-3p and Sirt7 was predicted and verified by TargetScan Human 7.1 online software and dual luciferase reporter assay. Results Hypoxia up-regulated the expression of miR-130a-3p in HTR-8/SVneo cells, down-regulated the expression of Sirt7 mRNA and protein. The low expression of miR-130a-3p increased the expression of Sirt7 mRNA, promoted the migration and invasion ability of cells, down-regulated the expression of HIF-1α and E-cadherin protein, and up-regulated the expression of Sirt7, MMP-2, MMP-9, N-cadherin and Vimentin protein. However, Combined treatment with Sirt7 inhibitor 97491 reversed the promotion effect of miR-130a-3p low expression on the migration, invasion and EMT of hypoxic cells. Conclusion miR-130a-3p is highly expressed in hypoxia-induced trophoblast cells. Inhibiting the expression of miR-130a-3p promotes the ability of migration, invasion and EMT of HTR-8/SVneo cells under hypoxic condition, which may be achieved by targeting Sirt7 to regulate HIF-1α.

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Objective To investigate the role of Vitamin D in the pathogenesis of inflammatory bowel disease(IBD) by studying the changes of inflammation related genes in peripheral blood mononuclear cells(PBMC). Methods 24 sample sizes were downloaded from GSE50012 in the GEO database, and they were divided into two groups: patients taking Vitamin D and patients not taking Vitamin D, with 12 patients in each group. Find out the differential expression of mRNA in PBMC of two groups of patients. At the same time, we conducted GO/KEGG and GSEA enrichment analysis on the differentially expressed genes, searched for possible pathways or molecular functions, conducted molecular interaction network analysis on the differential genes, and searched for key genes(Hub genes). A total of 128 patients with ulcerative colitis(UC) and 62 patients with Crohn′s disease(CD) in the outpatient department of our hospital were collected and divided into 4 groups by whether to take Vitamin D or not. Candidate key gene expression in PBMC in each group was measured to evaluate the effect of Vitamin D on inflammatory bowel disease. Results The GEO database screened a total of 128 differential expression genes(DEGs), of which 53 were highly expressed and 75 were low expressed. GO analysis showed that DEGs were mainly involved in cytokine activity, chemokine receptor binding, regulating inflammatory reactions, participating in immune response processes, mediating lymphocyte immunity, and other processes. KEGG analysis showed that cytokine receptor interaction, inflammatory bowel disease IgA mediated signaling pathways such as intestinal immune networks; GSEA analysis showed that the differentially expressed molecules were mainly involved in the Vitamin D receptor pathway and the KEGG chemokine signaling pathway. Protein interaction network analysis screened seven key genes, IL-6, MMP9, CXCL9, CXCL10, TREM1, IL1R2, and C1QB. After adding Vitamin D, HLA-DMA, HLA-DMB, IL-6, MMP9, CXCL9, and CXCL10 showed low expression, while NOD2 showed high expression. Conclusion Vitamin D may play a protective role in IBD by influencing PBMC inflammation related genes, thereby inhibiting inflammation and regulating immune function, providing ideas for the basic and clinical research of late IBD.

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Objective To investigate the expression and molecular mechanism of dickkopf 1(DKK1) in tongue squamous cell carcinoma(TSCC) by bioinformatics method and molecular biology experiments. Methods The patients information wasdownload from TCGA-TSCC database,the differentially expressed genes between the cancer and normal tissues were screened by Network Analysed site,the key genes and clinical prognosis were identified through Kaplan-Meier analysis and Lasson regression,the functions and pathways of differentially expressed genes were gained by GO and KEGG database,the expression of DKK1 mRNA and protein in TSCC as well as its relationship with clinicopathological features were analyzed by UALCAN database and immunohistochemistry. Western blot assay was conducted to detect the protein expression of DKK1 in TSCC cells,and siRNA was used to konck down the expression of DKK1 protein in Cal27 cells. Results The three key genes DKK1,CYP19A1 and IRX4,which were highly expressed in tongue squamous cell carcinoma and the survival rate of TSCC patients with high expression group was poor,were screened through Network Analysed,Kaplan-Meier analysis and Lasson regression method. UALCAN database showed that the mRNA level of DKK1 in TSCC tissues was higher than that in normal tissues,and its high expression was significantly correlated with clinical stage,histological grade and lymph node metastasis of TSCC patients. The immunohistochemistry assay suggested that the positive rate of DKK1 protein in clinical stage Ⅲ + Ⅳ TSCC tissues was significantly higher than that in stage Ⅰ + Ⅱ TSCC tissues. In addition,the expression level of DKK1 protein in TSCC tissues was significantly higher than that in adjacent tissues. Western blot assay also showed that the protein expression of DKK1 in TSCC cell Cal27 was much higher than normal oral epithelial cell HOEC. When knock down the protein expression of DKK1 in Cal27,the expression of β-catenin、pp65 和 p65 werealso reduced. Conclusion DKK1 is highly expressed in tongue squamous cell carcinoma tissues and cells and plays an important role. It may be a new target for early diagnosis and drug treatment of TSCC.

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Objective To investigate the regulatory effect and mechanism of specific antagonist of acid-sensitive ion channel 3(APETx2) on visceral sensitivity in mice with post-infectious irritable bowel syndrome(PI-IBS). Methods The PI-IBS model was established by National Institutes of Health(NIH) mice infected with Trichinella spiralis. Gastrointestinal transport function was assessed by measuring the time to first black stool and the number of fecal pellets collected for 6 hours; abdominal wall withdrawal reflex(AWR) was used to assess visceral sensitivity; the expression of calcitonin gene-related peptide(CGRP) in the colon tissue was detected by immunohistochemistry; the expression of brain-derived neurotrophic factor(BDNF) and CGRP mRNA in the colon tissues was detected by quantitative real time polymerase chain reaction(qRT-PCR). The expression levels of acid sensing ion channel 3(ASIC3), CGRP, and transient receptor potential vanilloid 1(TRPV1) protein in brain tissue were detected by Western blot analysis. Results Compared with the control group, the PI-IBS group significantly reduced the time of first black stool, the number of fecal particles and AWR score within 6 hours significantly increased, the protein expression of CGRP in colon tissue, BDNF and CGRP mRNA significantly increased, and the protein expression of CGRP, ASIC3 and TRPV1 in brain tissue significantly increased. Compared with the control group, the PI-IBS group significantly reduced the time to first black stool, the number of fecal particles and AWR score within 6 hours significantly increased, the expression of CGRP protein in colon tissue, the expression of BDNF and CGRP mRNA significantly increased, and the protein expression of CGRP, ASIC3 and TRPV1 in brain tissue significantly increased; compared with the PI-IBS group, the first time of black stool clearance in the APETx2 group was significantly prolonged, the number of fecal particles and AWR score within 6 hours were significantly reduced, the expression of CGRP protein in colon tissue, the expression of BDNF and CGRP mRNA was significantly reduced, the protein expression of CGRP, ASIC3 and TRPV1 in brain tissue was significantly reduced, and the difference was statistically significant(P<0.05). Conclusion APETx2 can alleviate visceral sensitivity and regulate gastrointestinal motility in PI-IBS mice by downregulating the expression of BDNF, CGRP, ASIC3 and TRPV1. APETx2 may provide a new therapeutic option for the treatment of IBS.

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Objective To investigate the effect of riboflavin on inactivation of pathogens under different light time conditions. Methods Plasma containingvesicular stomatitis virus(VSV) was divided into 4 groups, and the final concentrations of riboflavin were 50, 100, 150 μmol/L and 0 μmol/L was used as the control group. The growth titers at 10, 15, 20 and 25 min were observed. The control group was treated with no light. The growth titers of VSV culture before and after inactivation were calculated by Reed-Muench method, and the effect of different concentrations of riboflavin on the inactivation effect was evaluated by log reduction factor after inactivation. Results Under the condition of no light, the addition of riboflavin had no pathogen inactivation effect. With the increase of light time from 15 min to 25 min, the inactivation effect of VSV in the non-riboflavin group increased. When the light was less than 15 min, there was no significant difference in the inactivation effect of 50 to 150 μmol/L riboflavin VSV. When the light duration was 15 to 25 min, the inactivation effect of VSV was enhanced after the addition of riboflavin, but there was no difference between different concentrations of riboflavin groups. Conclusion The inactivation effect of riboflavin pathogen within 25 min is better with the increase of light time, and has little influence on riboflavin concentration.

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Objective To investigate the changes of osteogenic genes and osteogenic capacity after overexpression of bone morphogenetic protein 2(Bmp2) in rat adipose stem cells. Methods Bmp2 was transfected into rat adipose stem cells by electrotransfection, and the blank group was used as control. The cells were divided into blank group(Control group), empty load group(Vector group), osteogenesis induction group(Inducement group), and overexpression group(Bmp2 group). Western blot, qRT-PCR and immunofluorescence were used to compare the expression of osteogenic genes, and alkaline phosphatase staining and alizarin red staining were used to observe cobalt sulfide precipitation and calcium salt deposition. Results Western blot and qRT-PCR showed that the expression of Bmp2, Bmp4, OPN, OCN, Runx2, P-Smad1/5, Dlx2, and BSP was significantly higher compared to the Control group(P<0.05), while the expression of Smad1/5 was not statistically different. Immunofluorescence showed that the fluorescence intensity of OPN and Runx2 was significantly higher compared to the Control group(P<0.05).The alizarin red staining and alkaline phosphatase staining showed that the Bmp2 group had increased pericellular calcium salt deposition, increased cobalt sulfide precipitation, and enhanced alkaline phosphatase activity compared to the Control group. Conclusion Overexpression of Bmp2 enhances osteogenic gene expression in adipose stem cells and improves their osteogenic ability.

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Objective To evaluate the expression of programmed death receptor ligand-1(SPP1), programmed cell death(PD-L1) in hepatocellular carcinoma(HCC), their correlation with M2-polarized macrophages, and their impact on prognosis. Methods Immunohistochemistry(IHC) was used to detect the expression of SPP1, PD-L1, and CD163(commonly used markers of M2-polarized macrophages) in 81 HCC patients. The correlation between the expression of SPP1, PD-L1, and CD163 and the clinical characteristics of patients and their prognosis were analyzed. The relationship between SPP1,PD-L1,and M2-polarized macrophages in HCC tissue was also analyzed. Results The expression of SSP1,PD-L1 and CD163 increased in HCC tissues compared with para-carcinoma tissues. The expression of SSP1,PD-L1 and CD163 were all related to the BCLC staging( P<0. 05),but were not related to age and sex. The expression of SSP1 was elevated in HCC patients with large tumor diameter and multiple intrahepatic tumors,PD-L1 was elevated in HCC patients with multiple intrahepatic tumors and HBV infection. SSP1,PD-L1 and CD163 were related to the prognosis,and high expression patients had lower 3-year OSR and OST compared to low expression patients. Cox analysis showed the BCLC staging,SPP1 and PD-L1 were independent prognostic factors. The expression of SPP1 and PD-L1 were positively correlated with M2 macrophages in HCC. Conclusion SPP1,PD-L1 and CD163 can be used to evaluate the biological behaviors of HCC,and the positive expression of SPP1 and PD-L1 suggests that HCC patients may have a poor prognosis.

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Objective To study the mechanism of Pirfenidone(PFD) inhibiting the invasion of biliary tract tumors through cancer associated fibroblasts(CAF). Methods Primary CAF were extracted from the tumor tissues of patients with biliary tract tumor, and the marker proteins of CAF, including vimentin(VIM), α-smooth muscle actin(α-SMA) and fibroblast activating protein(FAP) were detected by Western blot. Phalloidin experiment showed the function of fibroblast cytoskeleton. ELISA and Western blot were used to verify the difference of TGF-β expression between normal fibroblasts(NF) and CAF. The functional change of CAF was observed by adding PFD to CAF. The expression of TGF-β in CAF was verified by ELISA, quantitative real-time PCR(qRT-PCR) and Western blot. The change of TGF-β in serum was verified by subcutaneous tumor mouse model. The change of collagen contractile function in CAF was observed by collagen contractile test. The changes of MMP2 and MMP9 in CAF medium were observed by gelatin enzyme assay. The changes of SMAD signaling pathway protein in CAF were detected by Western blot. Results The related marker proteins VIM, α-SMA and FAP of CAF were highly expressed, and the filamentous actin(F-actin) of CAF was abundant. ELISA showed that the expression of TGF-β in CAF was enhanced. Western blot experiment confirmed that CAF had stronger collagen function. Western blot, PCR and related phenomenon experiments showed that PFD could inhibit collagen production and TGF-β expression in CAF. SMAD signaling pathway-related protein experiments demonstrated that PFD could affect tumor invasion by inhibiting TGF-β/SMAD signaling pathway. Conclusion The function of CAF extracted from cancer patients is dominated by collagen production, while PFD inhibits the collagen production and collagen remodeling related processes of CAF through TGF-β/SMAD signaling pathway to inhibit tumor invasion.

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Objective To reduce the biotoxicity of Au nanoparticles by encapsulating them in the metal organic framework Uio-66-NH2, and to further investigate the radiosensitizing effect of Au@Uio-66-NH2nanoparticles. Methods Au nanoparticles were assembled into the Uio-66-NH2by the method of high temperature heating. The synthesized Au@Uio-66-NH2nanoparticles were characterized, and dynamic light scattering(DLS) was performed to determine the particle size, potential, and stability; ultraviolet spectrophotometry was carried out to obtain the ultraviolet absorption spectrum; transmission electron microscopy(TEM) was conducted to visualize the spatial structure; the cell counting Kit-8(CCK-8) assay was used for biological safety analysis; the CCK-8 assay and the colony formation assay were designed for assessing the radiosensitizing effect of Au@Uio-66-NH2nanoparticles on cancer cells; a subcutaneous tumor-bearing mouse model was constructed to evaluate the radiosensitizing effect and biological safety of Au@Uio-66-NH2nanoparticlesin vivo. Results The characterization results demonstrated good stability of Au@Uio-66-NH2NPs in water solutions; the CCK-8 assay showed a more favorable biological safety profile of Au@Uio-66-NH2nanoparticles compared with Au nanoparticles. Meanwhile, the group of Au@Uio-66-NH2nanoparticles combining with radiotherapy had a favorable cell-killing effect; the animal experiment also proved supportive evidence for the radiosensitizing effect and biological safety of Au@Uio-66-NH2nanoparticlesin vivo. Conclusion The toxicity of Au@Uio-66-NH2nanoparticles is significantly lower than that of Au nanoparticles, and it shows obvious radiosensitization effect, which has good clinical application prospects.

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Objective To isolate, culture and identify adult mouse dorsal keratinocyte(KC) in vitro and to compare of the proliferation and differentiation capabilities of four adult mouse dorsal skin KC. Methods Four kinds of adult mouse dorsal skin were taken, the subcutaneous adipose tissue was peeled, and KC was obtained by 0.25% trypsin digestion; KC were identified using immunofluorescence method,cell growth status was observed under microscopy,different seed plate times and densities were designed,and cell proliferation viability was detected by CCK-8,Ed U and high connotation method. Four adult mouse KC differentiation was detected by Western blot. Results Trypsin digestion was used to extract four strains of adult mouse back skin KC,and the Kunming( KM)mice had the lowest number of KC and C57BL/6 to extract the highest number of cells for the same area of skin tissue. BALB/c,KM,and nude mouse had higher plate-laying efficiency at a density of 8 × 103/well,respectively,and C57BL/6 had higher plate-laying efficiency at a density of 1. 6 × 104/well. Among the four strains of mice,the primary KC of BALB/c mice was weaker than that of C57BL/6 mice,and there was no statistical difference in the degree of differentiation. Conclusion The proliferation and differentiation capacity of four adult mouse primary KC on the dorsal skin is successfully isolated and compared by a simple and easy method,which provided a certain experimental basis for the strain selection of mice with skin-related diseases.

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Objective To analyze the expression and immune infiltration levels of the BMP7 gene(BMP7) in esophageal squamous cell carcinoma(ESCC). Methods Initially, in 274 cases of ESCC and 242 cases of normal tissues, the level of BMP7 was verified by immunohistochemistry, and the relationship between the expression difference and the survival cycle and clinical pathological characteristics of patients with ESCC was explored, and BMP7 overexpression plasmid transfection of ESCC cells was established, and the effect of BMP7 on the biological behavior of ESCC cells was examined by CCK-8, Clone, and Transwell. Results BMP7 expression in normal esophageal tissues was higher than that of ESCC(P<0.001), the expression level of BMP7 was correlated with the degree of differentiation of patients(P=0.006) and TNM staging(P<0.001), and the survival of patients with high expression of BMP7 exceeded that of patients with low BMP7(P=0.041), and the experiments of CCK-8 and Clone showed that the proliferation effect of cells in the overexpressed BMP7 group was lower than that of the control group. Transwell experiments confirmed that the cell invasion migration capacity of the overexpressed BMP7 group was less than that of the control one. The immune infiltration results showed that BMP7 was positively correlated with macrophages(P=0.008) and negatively correlated with γ-δT cells(P=0.028). Conclusion BMP7 is low in ESCC and associated with poor prognosis and immune infiltration levels in patients.

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Objective To clarify the targeted regulatory relationship between long non-coding RNA(lncRNAs) lnc-CCDC117-1 and oncogenic transcription factor c-Myc, and to explore the effect of lnc-CCDC117-1 knockdown and overexpression on the development of tongue squamous cell carcinoma cells. Methods Lentiviral vectors carrying flag-c-Myc, plko.1-shc-Myc and their controls were transfected into HN6,SCC9 and CAL27 cells respectively, and real-time quantitative PCR(qRT-PCR) was used to detect the expression of lnc-CCDC117-1. In addition, the intracellular localization of lnc-CCDC117-1 was detected by fluorescence in situ hybridization. On this basis, the lnc-CCDC117-1 expression vector and knockdown vector were constructed and transfected with HN6, SCC9, CAL27 cells, CCK-8 method, cloning and formation to verify the proliferation of tongue squamous cell. Results LncRNAs positively regulated by c-Myc were initially screened by gene chip technology. After overexpression or c-Myc knockdown in tongue squamous cell HN6, these lncRNAs were verified to be consistent with the results of the chip, and the expression difference of lnc-CCDC117-1 was found to be the most significant. qRT-PCR test showed that c-Myc had a positive regulatory effect on lnc-CCDC117-1. When c-Myc was overexpressed, the expression of lnc-CCDC117-1 could be significantly up-regulated. lnc-CCDC117-1 expression was significantly down-regulated by c-Myc knockdown. Dual luciferase reporter genes showed that c-Myc could target the regulation of lnc-CCDC117-1, and c-Myc was involved in regulating and enhancing the transcriptional activity of lnc-CCDC117-1. lnc-CCDC117-1 mainly existed in the nucleus of the cell. qRT-PCR results showed that the expression of lnc-CCDC117-1 and c-Myc significantly decreased by sh-lnc-CCDC117-1. The expression of lnc-CCDC117-1 was significantly up-regulated by +lnc-CCDC117-1. The results of growth curve assay, CCK-8 assay, cell scratch assay and cloning formation showed that overexpression of lnc-CCDC117-1 significantly promoted the proliferation and migration of tongue squamous cell cells, while knockdown of lnc-CCDC117-1 could inhibit the growth and proliferation of tongue squamous cell carcinoma cells. Conclusion lnc-CCDC117-1 is positively regulated by c-Myc, and overexpression of lnc-CCDC117-1 can promote cell proliferation, on the contrary, inhibit cells growth.

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Objective To explore the risk factors of fever after hysterectomy in patients with uterine fibroids, and establish a nomogram prediction model to provide theoretical guidance for clinical diagnosis and treatment. Methods The data of patients who underwent hysterectomy due to uterine fibroids were collected and randomly divided into training group and validation group at 6 ∶4. Univariate and multivariate analyses were performed by χ2test and Logistic regression model analysis. The nomogram was established based on the results of multivariate analysis, the calibration curve and receiver operating characteristic, curve were used to evaluate the prediction ability and accuracy of the nomogram. The decision curve reflected the clinical application value of the model. Results Among 264 patients in the training group, 33 patients had fever, and the postoperative disease rate was 12.5%. The results of univariate and multivariate analysis showed that anemia, preoperative curettage, operation scope and operation time were independent risk factors for fever after hysterectomy in patients with uterine fibroids. The internal and external calibration curves had a good fit with the actual observation results, with an average error of 0.031 and 0.025, respectively. The area under receiver operating characteristic curve of the internal and external verification of the nomogramare 0.788 and 0.712, respectively, reflecting the good predictive ability of the model. The decision curve showed that the model had good clinical benefit in a certain threshold range. Conclusion Based on multivariate analysis, the independent risk factors of fever after hysterectomy in patients with uterine fibroids, including anemia, preoperative curettage, operation scope and operation time. In addition, we established a nomogram with good predictive ability and accuracy, which is helpful for clinicians to timely manage fever after hysterectomy in patients with uterine fibroids and guide clinical individualized treatment.

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Objective To analyze the status of glucose metabolism in obese children. Methods 266 cases of obese children were included in the study, fasting plasma glucose(FPG), fasting insulin(FINS), glycosylated hemoglobin(HbA1c) were measured, Oral glucose tolerance test(OGTT) and insulin release test(IRT) were carried out, insulin resistance index(HOMA-IR), quantified insulin sensitivity index(QUICKI) and peak insulin/fasting insulin(Ip/I0) were calculated. Two hundred non-obese healthy children were used as control group, and glucose metabolism indexes were compared between the two groups. Glucose metabolism indexes and the incidence of insulin resistance(IR) were compared in children with different degrees of obesity. Prediabetes risk factors and body mass index( BMI) correlation were analyzed in obese children. Results The proportion of prediabetes and type 2 diabetes mellitus in obese children was 18. 0%( 48/266). FINS,HOMA-IR and Hb A1c in obesity group were higher than those in the control group,but QUICKI was lower than that in the control group( P<0. 05). There was no significant difference in FPG between the two groups( P = 0. 423). FINS and HOMA-IR of severe obesity group were higher than those of mild to moderate obesity group,Ip/I0was lower than that of mild to moderate obesity group( P<0. 05),and there was no significant difference in FPG,QUICKI,Hb A1c and 2h PG of severe obesity group. There was no significant difference in IR incidence between mild to moderate obesity group and severe obesity group( P = 0. 163). Logistic regression indicated that BMI and acanthosis nigricans was great influential on prediabetes with no statistical significance( P>0. 05). Correlation analysis showed that BMI had no correlation with FPG( P = 0. 160),but was positively correlated with FINS,HOMA-IR,2h PG and Hb A1c( P<0. 05),and negatively correlated with QUICKI and Ip/I0( P<0. 05). Conclusion Abnormal glucose metabolism is available in nearly 20% of obese children. The prevalence of insulin resistance in obese children is not affected by the degree of obesity.

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Objective To analyze the metabolism of blood glucose, blood lipid and insulin in patients with recurrent pregnancy loss(RPL), and to compare the predictive efficacy of triglyceride glycemic index(TyG), triglyceride glycemic body mass index(TyG-BMI) and homeostatic model assessment of insulin resistance(HOMA-IR)for RPL. Methods A total of 573 patients with RPL were selected as the RPL group, and 652 women who received assisted reproductive technology for male infertility were selected as the control group. The general data of the two groups were retrospectively analyzed. The levels of body mass index(BMI), fasting blood glucose(FPG), fasting triglyceride(FTG), fasting insulin(FINS), TyG index, TyG-BMI index and HOMA-IR were compared between the two groups. Spearman correlation analysis was used to verify the correlation between TyG index, TyG-BMI index and HOMA-IR. Receiver operating characteristic(ROC) curve was used to evaluate the predictive efficacy of TyG index, TyG-BMI index and HOMA-IR for RPL occurrence, and the optimal predictive cut-off point was calculated. Results The BMI, FPG, FTG, TyG index, TyG-BMI index and HOMA-IR in the RPL group were significantly higher than those in the control group(P<0.05). TyG index and TyG-BMI index were positively correlated with HOMA-IR, and the correlation coefficients were 0.442 and 0.505, respectively(P<0.001). ROC curve analysis showed that the area under the curve of TyG-BMI index predicting RPL was 0.579(95%CI:0.551-0.607,P<0.001), which was greater than that of TyG index of 0.557(95%CI: 0.529-0.585,P<0.001) and HOMA-IR of 0.535(95%CI: 0.507-0.563,P<0.05), among which the difference between TyG-BMI index and HOMA-IR index area under the curve was statistically significant(P<0.05). The optimal cut-off points of TyG-BMI index, TyG index and HOMA-IR for predicting RPL were 172.3(sensitivity 75.7%, specificity 37.06%), 8.32(sensitivity 59.44%, specificity 51.61%) and 3.58(sensitivity 25.87%, specificity 81.62%), respectively. Conclusion The incidence of overweight, abnormal blood glucose, lipid and insulin metabolism in RPL patients is higher than that in normal women. TyG index and TyG-BMI index can be used as indicators of insulin metabolic status in RPL population besides HOMA-IR, and TyG-BMI index has a higher efficacy in predicting the occurrence of RPL compared to HOMA-IR.

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Objective This study aimed to assess the incidence, etiological characteristic, potential risk factors and clinical outcomes of multidrug-resistant(MDR) infection in patients with peritoneal dialysis associated peritonitis(PDAP). Methods The data of PDAP patients admitted in recent 3 years in our center were collected retrospectively to analyze the distribution of pathogenic bacteria and drug resistance. According to whether the pathogenic bacteria were MDR or not, they were divided into MDR group and non-MDR group. The data of the two groups were compared, and the risk factors and clinical outcomes of MDR infection were analyzed. Results PDAP patients were mainly infected with gram-positive bacteria. Staphylococcus was the most common pathogen isolated from the peritoneal fluid in the MDR group, accounting for 60.6%(43/71), and streptococcus was the most common pathogen isolated in the non-MDR group, accounting for 42.0%(21/50). Multivariate analysis showed that the use of wide-spectrum antibiotics for more than 5 days within 3 months before the onset of PDAP(OR=3.634,P=0.033), gram-positive bacterial infection(OR=3.741,P=0.031), higher lipoprotein(a) level(OR=1.003,P=0.003), and higher platelet-to-albumin ratio(OR=1.198,P=0.038) were independent risk factors for MDR infection in PDAP patients. The incidence of refractory peritonitis in MDR group(29.6%) was higher than that in non-MDR group(12.0%)(χ2=5.229,P=0.022). The incidence of recurrent peritonitis(16.9%) was also higher than that in the non-MDR group(4.0%)(χ2=4.773,P=0.029). Conclusion The main pathogen of MDR infection is staphylococcus in PDAP patients. Compared with the non-MDR group, the MDR group has a higher proportion of wide-spectrum antibiotics used for more than 5 days within 3 months before the onset of PDAP, a higher proportion of gram-positive bacterial infection, higher lipoprotein(a) level, higher platelet-to-albumin ratio, and an increased risk of refractory peritonitis and recurrent peritonitis.

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Objective To explore the relationship between the methylation level of RABL6 in peripheral blood and early lung cancer(LC) with a case-control study in the Chinese population.Methods The methylation levels of 7 CpG sites in RABL6 gene in peripheral blood of samples from 275 LC patients(81.5% at stage I),and age-and gender-matched 185 benign lung nodule cases and 267 matched healthy controls were measured by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Multinomial Logistic regression adjusted for covariates was used to analyze the association between the RABL6 methylation and LC. Mann-Whitney U test was applied for the comparisons of RABL6 methylation levels between clinical characteristics subgroups of LC. Results Compared to the healthy controls,the methylation of RABL6 ＿Cp G ＿17 was inversely associated with LC in females( per-10% methylation: OR = 2. 47,95% CI = 1. 19-5. 13,P = 0. 016),but positively associated with LC in males( per-10% methylation: OR = 0. 52,95% CI = 0. 29-0. 94,P = 0. 030). In addition,hypermethylation of RABL6＿Cp G＿2 and RABL6＿Cp G＿5 was significantly associated with LC in the subjects older than 55 years( for RABL6＿Cp G＿2: per-10% methylation: OR = 0. 77,95% CI = 0. 60-0. 99,P = 0. 038; for RABL6＿Cp G＿5: OR = 0. 58,95% CI= 0. 34-0. 97,P = 0. 038). Conclusion The study reveals an association between peripheral blood-based RABL6methylation levels and early LC,providing a new clue for developing peripheral blood-based DNA methylation as a potential marker for the evaluation of LC risk.

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Objective To explore the general information, clinical features, imaging findings and outcome of patients with drug-induced cholestatic. Methods The clinical data of patients diagnosed as cholestatic drug-induced liver injury in recent 5 years were analyzed retrospectively, and followed up for at least 6 months. Results There were 105 patients with drug-induced cholestatic liver injury in the hospital, the median age was 55 years old(54.3% male),most of them had no specific clinical symptoms; there were nearly 80 kinds of drugs involved, the top three were traditional Chinese medicine(34.3%),antineoplastic drugs(22.9%) and antibiotics(10.5%).Among the 49 patients with magnetic resonance cholangiopancreatography(MRCP),7(14.3%) had sclerosing cholangitis-like changes; most of the patients were combined with multiple liver-protecting and enzyme-lowering drugs; there were 53 cured(50.5%),41 improved(39.0%),7 uncured(6.7%),and 4 died(3.8%) among 105 patients, and the total effective rate was 89.5%. Conclusion Drug-induced cholestatic liver injury mainly occurs in elderly patients, involving many kinds of drugs, mainly traditional Chinese medicine.Drugs can also cause changes similar to sclerosing cholangitis, so drug-induced liver injury(DILI) should be used as one of the differential diagnosis of cholestasis and abnormal cholangiography.Liver disease and the number of drugs have no effect on the prognosis of the patients, while low Roussel Uclaf causality assessment method(RUCAM) score and hemoglobin count, short hospitalization time and high white blood cell count are poor prognostic factors.

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Objective To evaluate the application of routine ultrasound combined with contrast-enhanced ultrasound(CEUS) for the renal wound after robotic-assisted nephron-sparing surgery(RANSS)and analyze the correlation with the postoperative complications. Methods 80 patients underwent RANSS were involved. Routine ultrasound was performed to evaluate the morphological features the first day after the surgery. A classification system named the grades of ultrasound images(GUIs) was build to evaluate the severity of the renal wound. CEUS was performed on postoperative day 1 to confirm the contrast agent extravasation from the renal wound. The incidence of postoperative complications in different grades was compared, meanwhile compared in the two groups of extravasation or not. Results In GUIs, the routine ultrasound images were classified to three grades: Grade1,Grade2,Grade3. The incidence of postoperative complications increased with GUIs(P<0.05), meanwhile higher in the extravasation group(P<0.05). This trend matched up with GUIs especially in Grade 2 and 3. The incidence of the cases with extravasation increased with GUIs(P<0.05). Conclusion Routine ultrasound and CEUS are feasible to evaluate the morphologic changes of renal wound after RANSS. There is a certain correlation between the evaluation results and the postoperative complications.

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Objective To analyze the application value of metagenomic next-generation sequencing(mNGS) in the detection of pathogenic bacteria in pulmonary infection, and to provide a theoretical basis for clinical diagnosis. Methods A retrospective study was conducted on 161 patients with suspected pulmonary infection, and samples of bronchoalveolar lavage fluid, sputum and lung tissue were collected specimens, pleural effusion and blood were simultaneously subjected to routine culture, respiratory pathogen nucleic acid detection and pathogenic microorganism mNGS detection. The sensitivity, specificity, positive predictive value, and negative predictive value of the three detection methods were calculated respectively to evaluate the clinical application value of mNGS. Results Among 161 patients with suspected pulmonary infection, 113 cases were finally confirmed as pulmonary infection. Among the 161 patients enrolled, the sensitivity and negative predictive value of mNGS were improved compared with routine culture and nucleic acid detection of respiratory pathogens, and the difference was statistically significant(P<0.001). Although the specificity of mNGS was higher than that of respiratory pathogen nucleic acid detection, the difference was not statistically significant. The positive predictive value of mNGS was the highest, and was higher than that of respiratory pathogen nucleic acid detection, but had no statistical significance compared with conventional culture. Conclusion The sensitivity of mNGS in the diagnosis of pathogenic microorganisms in pulmonary infection is higher than that of conventional culture and nucleic acid detection of respiratory pathogens, which reduces the false negative rate of diagnosis and has a better comprehensive evaluation; conventional culture detection has low sensitivity, but high specificity; respiratory pathogen nucleic acid detection compared with conventional culture, the sensitivity is improved, but the specificity decreased.

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Objective To evaluate the effect of screw-retained abutment integrated crown and cement-retained in first molar tooth implant restorations. Methods A retrospective study was conducted to select 91 patients. A total of 104 implant restorations were completed, of which 52 were screw-retained abutment integrated crown(SR) and 52 were cement-retained(CR). The data of the patients on the day of implant restoration and 6th and 12th month after teeth were conducted, the bone height of the mesial and distal edges of the implants were measured, the periodontal clinical indexes around the implants and the patient satisfaction score were recorded at the time of reexamination, and the proximal contacts after implant crown restoration was evaluated according to Barnes evaluation standard. Results 6 and 12 months after restoration, the implant retention rate in SR group and CR group was 100%; in the screw retention group, the retention screws of 3 restorations were loosen at 6 months, and the torch force was applied to 35 N/cm; at 12 months after restoration, the qualified rate of proximal contacts in the screw retention group was 90.38%, which was significantly lower than that of 99.04% in the adhesive retention group(P=0.037). There was no difference in marginal bone absorption and periodontal soft tissue indexes mPLI and mSBI between the two groups. Conclusion The two retention methods can both achieve good implant effects, and cement-retained restorations has more advantages in the first molar tooth restoration of proximal contact and the prevention of food impaction.

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Objective To explore the optimal pulse width of ultra-short pulse Er: YAG laser for improving dentin bonding strength. Methods Forty isolated third molars were prepared in the shape of wedge defect in the neck, and the coronal plane was taken.The samples were randomly divided into 5 groups: group G0 was blank group, and groups G1-G4 were treated with 30 MJ,20 Hz Er: YAG laser, and the pulse width was set to 30,40,50 and 60 μs, respectively.The morphology of dentin surface was observed by scanning electron microscopy(SEM),and the ratio of calcium to phosphorus was detected by energy dispersive X-ray spectroscopy(EDX).The shear bond strength(SBS) of each group and the fracture form under the type microscope were tested by the universal mechanical testing machine.The effect of different pulse width on improving dentin bonding strength was evaluated. Results SEM observation showed that the stained layer was removed after laser treatment with different pulse widths, and the dentin tubules were opened to different degrees.The calcium-phosphorus ratio of dentin after laser treatment increased, and the difference of calcium-phosphorus ratio in G2 group was statistically significant(P<0.05).The mean value of SBS after the cold and hot cycle experiment was G2>G3>G1>G4>G0,and the difference between G2 group and G0 group, G1 group, G3 group and G4 group are statistically significant(P<0.05).The difference between group G0 and groups G1,G2,G3 and G4 was statistically significant(P<0.05).The main fracture forms of each group were type 1 and type 3. Conclusion The 30 MJ,20 Hz Er: YAG laser pulse width of 40 μs can significantly improve the bond strength of dentin.