Effect of electrochemically dealloying Ti6Al4V abutments on human gingival fibroblasts / 口腔疾病防治

Objective@#To investigate the effects of electrochemically dealloying of Ti6Al4V abutments on human gingival fibroblasts (HGFs) and to provide experimental evidence for surface modification of implant abutments.@*Methods@#The samples were divided into an NC group (negative control, no other treatment on a smooth surface), an NM-1 group (nanomesh-1, electrochemical dealloying treatment in 1 mol/L NaOH 1 h on 2 V voltage), and an NM-2 group (nanomesh-2, electrochemical dealloying treatment in 5 mol/L NaOH 1 h on 2 V voltage). The surface morphologies of the samples and the adhesion of HGFs on the sample surfaces were observed with scanning electron microscopy (SEM). The surface hydrophilicities of the samples were measured with a contact angle measuring instrument. The proliferation of HGFs on the different samples were evaluated with CCK-8, and the expression of adhesion-related genes, including collagen Ⅰ (COL1A1), collagen Ⅲ (COL3A1), fibronectin 1 (FN1), focal adhesion kinase (FAK), vinculin (VCL), integrin α2 (ITGA2), and integrin β1 (ITGB1), on the different samples was measured with qRT-PCR. The expression of vinculin on the surfaces of HGFs was observed via confocal laser scanning microscopy (CLSM) after immunofluorescent staining. Collagen fiber secretion and syntheses of HGFs from different samples were evaluated via Sirius red staining.@*Results@#SEM revealed the formation of ordered and uniform three-dimensional mesh structures on the surfaces of the NM-1 and NM-2 groups, with grid diameters of approximately 30 nm for the NM-1 group and approximately 150 nm for the NM-2 group. Compared with that of the NC group, the water contact angles of the NM-1 group and NM-2 groups were significantly lower (P<0.000 1). Cell proliferation in the NM-1 group was significantly greater than that in the NC group (P<0.01). Moreover, there was no significant difference in the water contact angles or cell proliferation between the NM-1 group and the NM-2 group. SEM revealed that HGFs were adhered well to the surfaces of all samples, while the HGFs in the NM-1 and NM-2 groups showed more extended areas, longer morphologies, and more developed pseudopodia than did those in the NC group after 24 h. qRT-PCR revealed that the expression levels of the adhesion-related genes COL1A1, COL3A1, FN1, FAK and VCL in the NM-1 group were significantly greater than those in the NC and NM-2 groups (P<0.01). The expression of vinculin protein in the NM-1 group was the highest, and the number of focal adhesions was greatest in the NM-1 group (P<0.01). The results of Sirius red staining showed that the NM-1 group had the highest secretion and syntheses of collagen fibers (P<0.000 1).@*Conclusion@#The three-dimensional nanomechanical structure of Ti6Al4V modified by electrochemical dealloying promoted the adhesion, proliferation, collagen fiber secretion and syntheses of HGFs, and electrochemical dealloying of Ti6Al4V with a grid diameter of approximately 30 nm obviously promoted HGF formation.

Molecular mechanisms of Ca2+-induced pyroptosis and adhesion changes of HK-2 cells in the formation of calcium-containing kidney stones / 天津医药

Objective To investigate the possible role and mechanism of activation of pyroptosis classical pathway and alterations in cell adhesion in calcium-containing kidney stones after the action of high concentration of Ca2+ on HK-2 cells.Methods HK-2 cells were cultured in the presence of different concentrations of CaCl2(0,0.1,0.5,1.0,2.0,4.0 and 8.0 g/L)for 24 hours,and cell counting Kit-8(CCK-8)and flow cytometry were used to determine the optimal treatment concentration.Subsequently,the ultrastructure of renal tubular epithelial cells under high Ca2+ condition was observed by transmission electron microscopy after Ca2+ treatment.DCFH-DA staining was used to detect intracellular reactive oxygen species production,and quantitative real-time polymerase chain reaction(qRT-PCR)and Western blot analysis were performed to examine the expression of pyroptosis-related proteins NLRP3,Caspase-1,gasdermin D(GSDMD),adhesive molecules osteopontin(OPN)and CD44 at mRNA and protein levels after high concentration Ca2+ treatment.The expression levels of pyroptosis-related inflammatory factors interleukin(IL)-1β,IL-18 and adhesive molecule monocyte chemotactic protein-1(MCP-1)were detected by enzyme-linked immunosorbent assay(ELISA)after high Ca2+ stimulation.Results Ca2+ showed cytotoxicity for HK-2 cell growth and can promote apoptosis.The higher the Ca2+ concentration,the more toxicity and apoptosis rate for HK-2 cell growth.High concentration of Ca2+ can promote pyroptosis-like morphological changes in HK-2 cells,including loss of cell membrane integrity,release of contents and numerous intracellular vacuoles.Compared with the control group,the expression levels of ROS were sequentially increased in the 1.0 g/L CaCl2 group and the 2.0 g/L CaCl2 group,and the expression levels of pyroptosis-related genes NLRP3,Caspase-1,GSDMD,and the pyroptosis-associated inflammatory factors IL-1β and IL-18,as well as the adhesion molecules OPN,CD44 and MCP-1 were significantly increased(P＜0.05).Conclusion High Ca2+ treatment can cause oxidative stress damage in HK-2 cells to produce ROS,which activates NLRP3 inflammasome,leads to the activation of the classical pathway of pyroptosis and increase the adhesion of cells,and ultimately leads to the formation of kidney stones.

Exploring the Ideas of Traditional Chinese Medicine in the Prevention and Treatment of Tumour Metastasis Exacerbated by Chronic Stress：from the Perspective of Abnormal Tumour Cell Adhesion / 中医杂志

Abnormal tumour cell adhesion is a key step in tumour metastasis， in which weakened homologous and enhanced heterologous adhesion of tumour cells is an important cause of tumour metastasis. Chronic stress can activate the sympathetic nervous system to link and regulate the homologous and heterologous adhesion of tumour cells and exacerbate tumour metastasis. Combining the understanding of traditional Chinese medicine （TCM） and Western medicine on tumour metastasis， it is believed that the mechanism of "qi constraint and stagnation， tumor toxin transmission and retention" in TCM theory is highly related to the abnormal adhesion of tumour cells triggered by chronic stress. Qi constraint and stagnation is closely related to chronic stress and its activation of the sympathetic nervous system， and transmission and retention of tumor toxin explained the mechanism of tumour metastasis due to abnormal adhesion of tumour cells from the perspective of TCM. By regulating the key link of sympathetic nervous system-tumour cell adhesion， application of the formulas of regulating qi and resolving toxin can improve chronic stress and inhibit tumour metastasis.

Exploring the Mechanism of Action of Qizhu Kang'ai Formula （芪术抗癌方） for the Treatment of Colorectal Cancer based on Network Pharmacology and Experimental Validation / 中医杂志

ObjectiveTo explore the potential molecular mechanism of Qizhu Kang'ai Formula （芪术抗癌方， QZKAF） for the treatment of colorectal cancer （CRC）. MethodsNetwork pharmacology was used to analyze the active ingredients and targets of QZKAF for CRC， and analyze the key targets of QZKAF for the treatment of CRC by gene function annotation （GO） and Kyoto Encyclopedia of Genomes （KEGG） pathway enrichment analysis. Molecular docking was applied to predict the binding activity of the core active ingredients to the key targets. A orthotopic transplantation tumor mice model of CRC was established to validate the key targets of QZKAF for CRC obtained from network pharmacology analysis. Forty-eight mice were randomly divided into the sham operation group， the model group， the 5-fluorouracil （5-Fu） group， and the QZKAF low-， medium-， and high-dose groups， with 8 mice in each group. Except for the sham operation group， the remaining groups underwent colon cancer orthotopic transplantation tumor modeling. The 5-Fu group was given 30 mg/kg of 5-Fu by intraperitoneal injection once every 3 days on the alternate day after modeling， while the QZKAF low-， medium-， and high-dose groups were given 2.925， 5.85， and 11.7 g/（kg·d） of QZKAF by gastric gavage， respectively， and the sham-operation group and the model group were gavaged with 0.1 ml/10 g of normal saline every day， all for 21 days. The in situ tumors mass and the number of liver metastases were compared between the groups. The pathological changes of colon tumor tissues were observed by HE staining， and the protein expression of protein tyrosine phosphatase nonreceptor type 1 （PTPN1）， vinculin， integrin subunit αν， integrin subunit β3， and E-cadherin were detected in colon tumor tissues by Western blot. ResultsNetwork pharmacology screening yielded that the top six core active ingredients of QZKAF intervening in CRC were quercetin， kaempferol， apigenin， luteolin， baicalein and ursolic acid. There were 212 targets of action， and the ranked top three were prostaglandin endoperoxide synthase 1 （PTGS1）， prostaglandin endoperoxide synthase 2 （PTGS2）， and PTPN1， which may be the key targets of QZKAF in the treatment of CRC. These key targets were significantly enriched mainly in phosphatidylinositol 3-kinase/protein kinase B （PI3K-Akt） signaling pathway， focal adhesion and adhesion junction. Molecular docking results： except for PTGS1 with better binding activity to quercetin， kaempferol， and apigenin （binding energy ≥-7.0 kcal/mol）， PTGS1 showed strong binding activity to lignans， baicalein， ursolic acid， as well as PTGS2 and PTPN1 to the six core active ingredients （binding energy ＜-7.0 kcal/mol）. Experimental validation results： the protein expression of PTPN1， vinculin， integrin subunit αν， integrin subunit β3 in the colon tumor tissues of mice in the model group significantly increased， and the expression of E-cadherin significantly decreased compared to those in the sham operation group （P＜0.05）. Compared to those in the model group， the mass of the in situ tumors was reduced， and the number of hepatic metastasis nodules decreased in the high- and medium-dose QZKAF groups （P＜0.05）； the expression levels of PTNP1， vinculin， integrin subunit αν， integrin subunit β3 and E-cadherin in all QZKAF groups and 5-Fu group showed different degrees of retracement， and the changes of the indicators in all QZKAF groups showed a certain degree of dose-dependence （P＜0.05）. HE staining showed that the nuclei of tumor cells in the model group were mostly schizophrenic， and there were different degrees of nuclear fragmentation of tumor cells in all QZKAF groups with more in the medium- and high-dose groups. ConclusionQZKAF could inhibit the growth of in situ tumors and liver metastasis of CRC. Its mechanism might be related to the regulation of tumor cell-cell and tumor cell-extracellular matrix adhesion by PTPN1.

Construction of porous structure on the surface of PLLA/β-tricalcium scaffold and its effect on cell adhesion / 西安交通大学学报(医学版)

【Objective】 To construct a 3D printed PLLA/β-tricalcium (PLLA/β-TCP) bone tissue engineering scaffold surface porous structure through simple treatment with NaOH solution, increase the roughness and hydrophilicity of the scaffold, and promote cell adhesion on the scaffold surface. 【Methods】 The PLLA/β-TCP mesh scaffold was prepared by 3D printing melt deposition molding technology, and the scaffold was roughed by NaOH etching. The effects of NaOH concentration and time on the scaffold were observed according to the microstructure, energy spectrum, contact angle, mechanics, and cell adhesion of the scaffold. 【Results】 The PLLA/β-TCP composite scaffold constructed by melt deposition technology had a pre-set porous structure, and the pores were interconnected. After NaOH etching, a porous structure with both macroscopic and microscopic pores was formed. The increase in any of the NaOH concentration and time parameters would lead to the increase of pore diameter and surface roughness. When the NaOH treatment parameter was 0.1 mol/L (9 h), it could significantly reduce the water contact angle on the surface of the scaffold, and had no significant effect on the compressive strength of the scaffold. In vitro cell testing showed that the surface porous composite scaffold etched with NaOH had more advantages in the adhesion and proliferation of BMSCs. 【Conclusion】 Using NaOH to process 3D printing of PLLA/β-TCP bone tissue engineering scaffolds can effectively improve the surface morphology of the scaffold, and optimize its hydrophilicity and cell adhesion.

Preparation of an amino hybrid mesoporous silica-based nanotopography protective coating on a titanium implant surface and evaluation of its osteogenic effect / 口腔疾病防治

Objective@#To deposit degradable amino-hybrid mesoporous silica (AHMS) in situ on the surface of titanium nanotube (TNT) and explore its protective effect on nanomorphology and osteogenesis.@*Methods@#TNT and TNT@AHMS were sequentially prepared via an anodizing method: the oil-water two-phase method (experimental group) and the acid-etched titanium method [control group (Ti)]. The parameters for synthesis were explored by changing the silicon source dosage ratio (3∶1, 1∶1, 1∶3); the surface morphology was observed by scanning electron microscope(SEM), hydrophilicity was detected by Water Contact Angle Tester, elemental composition was detected by X-ray photoelectron spectroscopy (XPS); nanoindentation test and ultrasonic oscillator were used to observe the morphological holding effect as mechanical strength of TNT@AHMS in vitro; simulated immersion experiments in vitro was used to observe the degradation behavior of the material. the MC3T3-E1 cell line was used to observe the effect of cell adhesion, proliferation and differentiation on the material; and an SD rat femoral implant model and micro-CT were used to verify the protective effect and osseointegration effect of AHMS on TNT morphology.@*Results@#The morphologies of TNT and TNT@AHMS were successfully prepared, and the silicon source ratio was 1:3. SEM showed that the titanium nanotubes were uniformly covered with AHMS coating, and the mesoporous pore size was about 4 nm. After AHMS was incorporated, the surface of the material was hydrophilic (12.78°), the presence of amino groups (NH2-) was detected, the material was completely degraded within 12 h in vitro, and the active morphology of the TNT was re-exposed with a cumulative silicon release of 10 ppm. Nanoindentation test showed that TNT@AHMS exhibited more ideal surface mechanical strength. SEM revealed that TNT maintains its own morphology under the protection of AHMS, and the TNT group suffered severe exfoliation. In addition, the early adhesion and proliferation rates, ALP activity, and bone volume fraction of cells on the TNT@AHMS surface 4 weeks after implantation were significantly higher than those in the TNT group.@*Conclusion@#By depositing AHMS on the surface of TNT, the nanotopography can be protected. It not only prevents the active base topography from exerting subsequent biological effects but also further endows the material with the ability to promote bone regeneration, laying a foundation for the future development of nanotopography-modified titanium implants.

Bushen Jianpi Jiedu Liyan Formula Regulates Homing-associated Cell Adhesion Molecules in Rat Model of IgA Nephropathy / 中国实验方剂学杂志

ObjectiveTo investigate the effect of Bushen Jianpi Jiedu Liyan formula on the expression of integrin alpha 4 beta 1 （α4 β1）， vascular cell adhesion molecule-1 （VCAM-1）， stromal-derived factor-1 （SDF-1）， and chemokine receptor-4 （CXCR4） in the small intestine and bone marrow of the rat model of immunoglobulin A（IgA） nephropathy. MethodA total of 120 male SD rats were used to establish the IgA nephropathy model by intragastric administration of bovine serum albumin （BSA）， subcutaneous injection of CCl4， and tail vein injection of lipopolysaccharide （LPS）. The successfully modeled rats were randomized into blank， model， lotensin （63 mg·kg-1）， and low-， medium-， and high-dose （10.4， 20.81， 41.62 g·kg-1， respectively） Bushen Jianpi Jiedu Liyan formula groups （n=16）. The rats were treated with corresponding drugs according to their body weight. After 7 weeks of administration， the rats were sacrificed for the collection of samples， and the protein and mRNA levels of α4 β1， VCAM-1， SDF-1， and CXCR4 in the small intestine and bone marrow were determined by immunohistochemistry and real-time fluorescence quantitative polymerase chain reaction， respectively. ResultCompared with the blank group， the model group showed increased red blood cell count in the urine at the 10th， 12th， 14th， 16th weeks （P<0.01）， and such increases were reduced in the drug intervention groups （P<0.05）， especially in the medium-dose Bushen Jianpi Jiedu Liyan formula group （P<0.05）. Compared with those in the blank group， the protein levels of α4 β1， VCAM-1， SDF-1， and CXCR4 in the intestinal lamina propria in the model group were up-regulated （P<0.05）， and such un-regulations were inhibited in the drug intervention groups （P<0.05）. Compared with the model group， medium-dose Bushen Jianpi Jiedu Liyan formula down-regulated the protein levels of SDF-1 and CXCR4 in the intestinal lamina propria （P<0.05）. Compared with the blank group， the model group showed down-regulated mRNA levels of α4 β1 and SDF-1 and up-regulated mRNA levels of VCAM-1 and CXCR4 （P<0.05）. Compared with the model group， the drug intervention groups showed down-regulated mRNA levels of SDF-1 and CXCR4 （P<0.05）. ConclusionBushen Jianpi Jiedu Liyan formula regulates the expression of α4 β1， VCAM-1， SDF-1， and CXCR4 in the intestinal lamina propria to inhibit the homing effect of plasma cells， which may be associated with the Toll-like receptor-mediated activation of immune response. Bushen Jianpi Jiedu Liyan formula can down-regulate the expression of adhesion molecules to inhibit the proliferation of plasmocytes in circulation， so as to reduce the renal injury of IgA nephropathy.

Micro/nano hierarchical structure of titanium surface promotes MC3T3-E1 adhesion proliferation and drug loading potential evaluation / 口腔疾病防治

Objective @# To investigate the effect of micro/nano hierarchical structures on the adhesion and proliferation of MC3T3-E1 cells, evaluate the drug delivery potential of titanium surfaces, and provide a reference for the modification of selected areas of titanium surfaces to enhance drug delivery and slow drug release. @*Methods @# Pure titanium samples (10 mm in diameter and 2.5 mm in thickness) were randomly divided into a polished group (T), anodized group (TO), and micro/nano hierarchical structure group (FTO) according to the surface treatment of the titanium. The T group was polished, the TO group was treated with anodic oxidation technology, and the FTO group was treated by femtosecond laser etching combined with anodic oxidation technology. The three surface morphologies were observed by scanning electron microscopy (SEM), the wettability of the surface was measured by the contact angle, and the surface chemical composition was analyzed by X-ray energy dispersive spectroscopy (EDS). The depth of the FTO structure and the surface roughness were measured by confocal laser scanning microscopy (CLSM). MC3T3-E1 cell adhesion proliferation and differentiation on the surface of each group of samples was assessed by immunofluorescence staining, CCK-8, and semiquantitative analysis of Alizarin staining. A freeze-drying method was applied to load recombinant human bone morphogenetic protein-2 (rhBMP-2), and an enzyme-linked immunosorbent assay (ELISA) was used to assess the drug-loading potential of different surface structures. @* Results@#SEM revealed that the surface of T group titanium plates showed uniform polishing marks in the same direction. The surface of the TO group was a nanoscale honeycomb-like titanium dioxide (TiO2) nanotube structure, and the FTO group formed a regular and ordered micro/nano layered structure. The contact angle of the FTO group was the smallest at 32° ± 1.7°. Its wettability was the best. The average depth of the first-level structure circular pores was 93.6 μm, and the roughness was 1.5-2 μm. The TO and FTO groups contained a high percentage of oxygen, suggesting TiO2 nanotube formation. The FTO group had the most significant surface cell proliferation (P<0.001) and the largest cell adhesion surface area (P<0.05). rhBMP-2 was slowly released for 14 d after loading in the FTO group and promoted extracellular matrix mineralization (P<0.001). @*Conclusion @#Titanium surface microprepared hierarchical structure has the effect of promoting MC3T3-E1 cell adhesion, proliferation, and osteogenic differentiation with drug loading potential, which is a new method of titanium surface treatment.

CD44-HA Mediate Adhesion of MDA-MB-231 Cells and HL60 Cells under Fluid Shear Flow / 医用生物力学

Objective To explore how hyaluronic acid ( HA) in extracellular matrix regulates the adhesion ofCD44+tumor cells. Methods MDA-MB-231 cells or HL60 cells were perfused in a parallel plate chamber. Themovement of cells over immobilized HA was observed and analyzed to obtain the characteristics of cell adhesionand rolling. Results The adhesion number of MDA-MB-231 cells on HA substrate was positively regulated by HAconcentration, but not by HA molecular weight. Compared with physically adsorbed HA, immobilized HA byavidin-biotin could significantly improve the cell adhesion ratio. With the increase of shear stress in the range of30-50 mPa, the rolling velocity of cells increased and the adhesion ratio decreased, but the tether lifetime of cellswas not affected. In the same flow field, compared with MDA-MB-231 cells, HL60 cells with low expression ofCD44 rolled more quickly on immobilized HA, with shorter tether lifetime and much lower adhesion ratio(<1. 5% ). Conclusions Fluid shear stress might mediate the rolling velocity of MDA-MB-231 cells by regulatingthe CD44-HA association rate rather than their dissociation rate. The interaction between CD44 and HA is involved in the initial adhesion of HL60 cells, but it does not play a major role. This study will provide references for the design of anti-tumor drugs.

The expression of adhesion molecules in Z310 cells induced by combined exposure to black carbon and lead / 中国职业医学

Objective: To investigate the effects of combined exposure to black carbon and lead on the expression of cell adhesion molecules and their regulating microRNAs (miRNAs) in the rat choroid plexus epithelial Z310 cells. Methods: i) Z310 cells were randomly divided into control group, black carbon exposure group, lead exposure group and combined exposure group. The lead exposure group and black carbon exposure group were treated with 10 μmol/L lead acetate and 10 mg/L black carbon, respectively, and the combined exposure group was treated with both in the above doses. After 12.0 hours, the expressions of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and mucosal vascular addressin cell adhesion molecule-1 (MAdCAM-1) in Z310 cells was detected by Western blotting. The expression of miR-326, miR-328-3p and miR-542-3p which regulated ICAM-1 was detected by real-time fluorescent quantitative polymerase chain reaction. ii) Z310 cells or Z310 cells transfected with miRNA-326 mimic were randomly divided into control group, miRNA-326 transfection control group, combined exposure group and miRNA-326 transfection combined exposure group. Cells in the two control groups were not treated. The two combined exposure groups were treated with 10 mg/L black carbon and 10 μmol/L lead acetate for 12.0 hours. The expression of ICAM-1 was detected by Western blotting. Results: i) The relative expression of ICAM-1, VCAM-1 and MAdCAM-1 in the cells of black carbon exposure group and ICAM-1 in the lead exposure group was higher than those in the control group (all P<0.05). The relative expression of ICAM-1 and MAdCAM-1 in the combined exposure group was higher than those in the other three groups (all P<0.05). The relative expression of VCAM-1 in cells of combined exposure group was higher than those in the control group and lead exposed group (all P<0.05). The relative expression of miR-326 in cells of the lead exposure group and black carbon exposure group was lower than those in the control group (all P<0.05). The relative expression of miR-326 in the combined exposure group was lower than that in the other three groups (all P<0.05). There was no significant difference between miR-328-3p and miR-542-3p in the four groups (all P>0.05). ii) The relative expression of ICAM-1 in cells of the miR-326 transfection control group cells was lower than that in the control group (P<0.05), while in the cells in the combined exposure and miRNA-326 transfection combined exposure group, it was higher than that in the control and miRNA-326 transfection control groups (all P<0.05), and lower in the miRNA-326 transfection combined exposure group than in the combined exposure group (P<0.05). Conclusion: Black carbon or lead exposure can upregulate the expression of ICAM-1, VCAM-1 and MAdCAM-1 in Z310 cells. Black carbon and lead combined exposure lead to a synergistic effect on upregulation of ICAM-1 and MAdCAM-1 expression, particularly ICAM-1. The combined exposure of black carbon and lead may upregulate the expression of ICAM-1 by downregulating the expression of miR-326.

Research progress in the biological characteristics of matrix metalloproteinase-19 in tumor / 国际生物医学工程杂志

The matrix metalloproteinases family (MMPs) are proteins related to tumor formation and metastasis that have attracted the attention of scholars in recent years. Tumor cells can secrete MMPs during malignant transformation, and the expression of MMPs in different malignant tumors is diverse, and different members of MMPs do not have exactly the same biological properties. Matrix metalloproteinase-19 (MMP-19) is a new member of MMPs whose secretion increases rapidly during the malignant transformation of cells and is released into the extracellular space to participate in biological processes such as proliferation, adhesion, invasion, migration, and angiogenesis of tumor cells. In this paper, the progress of research on the biological properties of MMP-19 in tumors was reviewed to provide a theoretical basis for exploring the development of tumors, especially for studying the invasion and metastasis of tumor cells.

Change of serum D-D, sVCAM-1 and P-selectin after limb fractures surgery and their relationship with thrombosis / 中国医师杂志

Objective:To investigate the relationship between change of serum D-dimer (D-D), soluble vascular cell adhesion molecule 1 (sVCAM-1), P-selectin and thrombosis after limb fracture surgery.Methods:289 patients with limb fractures who were treated in the emergency department of Shulan (Hangzhou) Hospital from January 2021 to January 2022 were selected as the study subjects. They were divided into deep vein thrombosis (DVT) group ( n=62) and non-DVT group ( n=227) according to whether DVT occurred after operation. The levels of prothrombin time (PT), activated partial prothrombin time (APTT), D-D, sVCAM-1 and P-selectin were measured before and after operation in all patients. The levels of PT, APTT, D-D, sVCAM-1 and P-selectin were compared between DVT group and non-DVT group. Logistic sequential stepwise regression analysis was used to analyze the risk factors of postoperative thrombosis in patients with limb fractures. Results:There was no statistically significant difference in PT and APTT between 289 patients with limb fractures after operation and before operation (all P>0.05), while the levels of serum D-D, sVCAM-1 and P-selectin after operation were higher than that before operation (all P<0.05). There was no significant difference in general data between DVT group and non-DVT group (all P>0.05); There was no statistically significant difference in PT and APTT before and after operation between DVT group and non-DVT group (all P>0.05). The levels of serum D-D, sVCAM-1 and P-selectin before and after operation in DVT group were higher than those in non-DVT group (all P<0.05). Logistic sequential stepwise regression analysis showed that high levels of D-D, sVCAM-1 and P-selectin were risk factors for thrombosis after limb fracture surgery (all P<0.05). Conclusions:High levels of D-D, sVCAM-1 and P-selectin are risk factors for thrombosis after limb fracture surgery.

The role of electroacupuncture in regulating cell adhesion molecule L1 expression and in neuroprotection in Alzheimer′s disease / 中华物理医学与康复杂志

Objective:To observe any effect of electroacupuncture on the expression of L1 cell adhesion molecule (L1CAM) in mice modeling Alzheimer′s disease (AD) and also any effect on learning and memory.Methods:Thirty male APP/PS1 mice were randomly divided into a model group, an electroacupuncture (EA) group, and a no acupuncture (NA) group, each of 10. All the animals were modeled as AD. Ten C57BL/6 mice served as a control group. The mice in the EA and NA groups were given continuous 50Hz EA at a current intensity of 1mA at and near the Baihui (GV20) and Shenshu (BL23) acupoints, respectively, once a day for 14 days, while the other two groups were not given any EA. The mice in the model and control groups continued to be routinely fed without any special treatment such as electroacupuncture. After the intervention, any behavioral changes were evaluated by using a Morris Water Maze, and the expression of L1CAM, PTEN and p53 protein in the hippocampus of each group was detected using western blotting.Results:Compared with the control group, the escape latency in positioning navigation experiments was significantly longer in the model group on the first 5 days of Morris Water Maze testing. Compared with the model group, the escape latency was significantly shorter in the EA group on days 2 to 5 of the Morris Water Maze testing, and the expression of L1CAM had increased significantly in the electroacupuncture group compared with the model group while PTEN and p53 expression had decreased significantly. The average escape latency of the NA group was significantly longer than that of the model group on days 2 to 5 of the Morris Water Maze testing. The average L1CAM expression in the NA group had decreased significantly, and the expression of PTEN and p53 protein had increased significantly more than in the EA group. The escape latency was negatively correlated with L1CAM expression but positively correlated with p53 protein and PTEN expression.Conclusion:L1CAM is involved in learning and memory processes, at least in mice. Electroacupuncture can improve the learning and memory of mice modeling Alzheimer′s, which may be due to its promoting the expression of L1CAM and inhibiting the expression of PTEN and p53.

The predictive value of combined detection of serum ANGPTL8 and VCAM-1 levels for cerebral vasospasm after intracranial aneurysm embolization / 临床外科杂志

Objective The combined detection of serum angiopoietin-like protein 8(ANGPTL8)and Vascular cell adhesion molecule-1(VCAM-1)levels was analyzed for the predictive value of cerebral vasospasm(CVS)after intracranial aneurysm embolization.Methods A total of 196 patients who underwent intracranial aneurysm embolization in our hospital from March 2019-March 2022 were selected as the study subjects,99 patients with CVS were in the CVS group,and 97 patients without CVS were in the non CVS group.Serum ANGPTL8 and VCAM-1 levels were detected by ELISA;the correlation between serum ANGPTL8 and VCAM-1 levels was analyzed by Pearson method,Logistic regression was used to analyze the influencing factors of CVS in patients undergoing intracranial aneurysm embolization;ROC curve was used to analyze the serum levels of ANGPTL8 and VCAM-1 to predict the cutoff value of CVS in patients undergoing intracranial aneurysm embolization;four grid table method was used to analyze the predictive value of ANGPTL8,VCAM-1 and their combination on the occurrence of CVS in patients undergoing intracranial aneurysm embolization.Results The differences between CVS and non-CVS groups were statistically significant in hypertension,Hunt-Hess grade,and Glasgow coma(GCS)scores(P＜0.05).The serum ANGPTL8 and VCAM-1 levels in the CVS group were significantly higher than those in the non-CVS group(P＜0.05).There was a positive correlation between serum ANGPTL8 and VCAM-1(r=0.468,P＜0.05).Multivariate analysis showed that high level of ANGPTL8(OR=3.652,95％CI:1.434-9.302),high level of VCAM-1(OR=2.619,95％CI:1.212-5.658),Hunt Hess grade Ⅲ-Ⅳ(OR=1.927,95％CI:1.104-3.362),GCS score of 3-8(OR=2.813,95％CI:1.257-6.295)were independent risk factors for CVS in patients undergoing intracranial aneurysm embolization.The AUC of serum ANGPTL8 level in predicting CVS in patients undergoing intracranial aneurysm embolization was 0.844,and the cut-off value was 189.233 U/L;the AUC of serum VCAM-1 level in predicting CVS in patients undergoing intracranial aneurysm embolization was 0.795,and the cutoff value was 17.984 mg/L.The accuracy,sensitivity and specificity of the combined prediction for CVS were 89.81％,93.94％and 85.57％,respectively,which were obviously higher than those of the single prediction.Conclusion The serum levels of ANGPTL8 and VCAM-1 in CVS group are obviously higher than those in non CVS group.The combination of the two has a high predictive value for CVS after intracranial aneurysm embolization.

A study on epithelial cell adhesion molecule targeted nucleic acid aptamer drug conjugate in gastric cancer targeted treatment / 中华消化外科杂志

Objective:To investigate the affinity and toxicity of epithelial cell adhesion molecule (EpCAM) targeted nucleic acid aptamer drug conjugate SYL3C-MMAE on human gastric epithelial cells GES-1 (hereinafter referred to as GES-1 cells) and human gastric cancer cells AGS and MKN45 (hereinafter referred to as AGS cells and MKN45 cells).Methods:The experimental study was conducted. The expression level of EpCAM in gastric cancer tissues was detected using immunohistochemistry. The mRNA expression level of EpCAM in gastric cancer tissues was detected using real-time fluorescence quantitative PCR (RT-PCR). The expression level of EpCAM protein in GES-1, AGS and MKN45 cells was detected using Western blot. The affinity of SYL3C on GES-1, AGS and MKN45 cells was detected using flow cytometry. SYL3C-MMAE was synthesized through a thiol-maleimide reaction. The toxicity of drugs on GES-1, AGS and MKN45 cells was detected using CCK-8 assay. The cell cycle condition of GES-1, AGS and MKN45 cells after drug treatment was detected using propidium iodide (PI) staining. Observation indicators: (1) expression of EpCAM in gastric cancer; (2) affinity of antibodies targeting EpCAM and SYL3C on GES-1, AGS and MKN45 cells; (3) situation of drug synthesis; (4) drug toxicity and inhibition of cell cycle. Measurement data with normal distribution were represented as Mean± SD. One-way ANOVA was used for comparison among multiple groups, and pairwise comparison was conducted using the least significant difference test. Comparison of unequal variances was conducted using the Welch' t test. Measurement data with skewed distribution were represented as M(IQR), and comparison between groups was conducted using the paired rank sum test. Count data were described as absolute numbers, comparison between groups was conducted using the paired chi-square test. Results:(1) Expression of EpCAM in gastric cancer. Results of immunohistochemistry on tissue microarrays showed that the positive rate of EpCAM was 82.9%(29/35) and 22.9%(8/35) in the 35 pairs of gastric cancer and its adjacent tissues (normal tissues), respectively, showing a significant difference between them ( P<0.05). Results of RT-PCR showed that the mRNA relative expression levels of EpCAM was 1.23 (4.13) and 4.04 (1.72) in 12 pairs of gastric cancer and its adjacent tissues respectively, showing a significant difference between them ( Z=-2.67, P<0.05). Results of Western blot showed that the relative expression levels of EpCAM protein in GES-1, AGS, and MKN45 was 0, 1.00, and 0.27, respectively, with the expression level of EpCAM protein in AGS cells as the standard. (2) Affinity of antibodies targeting EpCAM and SYL3C on GES-1, AGS and MKN45 cells. Results of flow cytometry showed that antibodies targeting EpCAM and SYL3C had good affinity on AGS and MKN45 cells but no affinity on GES-1 cells. (3) Situation of drug synthesis. Results of mass spectrometry showed that the drug solution of compound formed by connecting SYL3C with monomethylorestatin E (VcMMAE) exhibited a strong peak at the molecular weight position of 16 355, consistent with the expected molecular weight of the SYL3C-MMAE complex, indicating that SYL3C-MMAE was successfully synthesized. (4) Drug toxicity and inhibition of cell cycle. Results of CCK-8 assay showed that the half maximal inhibitory concentration (IC 50) of VcMMAE on GES-1, AGS and MKN45 cells was 123.00, 30.48 and 51.83 nmol/L, respectively. The IC 50 of SYL3C-MMAE on GES-1, AGS and MKN45 cells was 241.80, 20.66 and 27.64 nmol/L, respectively. Results of PI staining and flow cytometry showed that both VcMMAE and SYL3C-MMAE could induce G2/M phase blockage in the cell cycle of GES-1, AGS and MKN45 cells. Conclusion:The SYL3C-MMAE has a good affinity on gastric cancer cells. Compared with VcMMAE, SYL3C-MMAE exhibits efficient inhibition on gastric cancer cells, but less influence on normal cells.

Effects of emodin on proliferation, adhesion, migration and c-kit mRNA and protein expression of human epidermal melanocytes / 中华医学美学美容杂志

Objective:To investigate the effects of emodin on proliferation, adhesion, migration, c-kit mRNA and protein expression of human epidermal melanocytes.Methods:Human epidermal melanocytes cultured in vitro were treated with different concentrations of emodin, a blank control group (containing medium + human epidermal melanocytes) and an experimental group (containing medium + human epidermal melanocytes + different concentrations of emodin) were set up. Cell proliferation was measured by CCK-8 method, cell adhesion was measured by microplate assay, cell migration was measured by Transwell membrane assay, and c-kit mRNA expression was measured by reverse transcription-polymerase polymerase chain reaction; Western blot was used to detect the expression of c-kit protein.Results:Compared with the blank control group, emodin decreased the proliferation, adhesion and migration of human epidermal melanocytes, the difference was statistically significant ( F=391.48, P<0.0001; F=10.93, P=0.003; F=7.75, P=0.009). Compared with the blank control group, emodin in the experimental group had a bidirectional effect on the expression of c-kit mRNA and protein. High concentration of emodin inhibited the expression of c-kit mRNA and protein, and the expression of c-kit mRNA and protein was significantly increased by Emodin ( F=11.491, P=0.003; F=2155.11, P<0.001). Conclusions:Emodin inhibits the proliferation, adhesion and migration of human epidermal melanocytes, high concentration of emodin inhibits the expression of c-kit mRNA and protein, and low concentration of emodin promotes the expression of c-kit mRNA and protein, and the results provide a basis for the clinical application of emodin in pigmented dermatosis.

Trichostatin C attenuates TNFα-induced inflammation in endothelial cells by up-regulating Krüppel-like factor 2 / 药学学报

Krüppel-like transcription factor 2 (KLF2) plays a key regulatory role in endothelial inflammation, thrombosis, angiogenesis and macrophage inflammation and polarization, and up-regulation of KLF2 expression has the potential to prevent and treatment atherosclerosis. In this study, trichostatin C (TSC) was obtained from the secondary metabolites of rice fermentation of Streptomyces sp. CPCC 203909 as a KLF2 up-regulator by using a high throughput screening model based on a KLF2 promoter luciferase reporter assay. TSC significantly inhibited the adhesion of tumor necrosis factor-α (TNFα) induced monocytes (THP-1) to human umbilical vein endothelial cells (HUVECs). Western blot results showed that TSC decreased TNFα induced the protein expression increase of vascular cell adhesion molecule-1 (VCAM-1), and thereby inhibited endothelial inflammation. The results of histone deacetylase (HDAC) overexpression and molecular docking experiments showed that TSC upregulated the expression of KLF2 by inhibiting subtypes of HDAC 4/5/7. In conclusion, this study suggests that TSC up-regulates the expression of KLF2 through inhibiting HDAC 4/5/7 and thus inhibits TNFα induced endothelial inflammation, and it has the potential to prevent and treat atherosclerosis.

Role of HBV DNA polymerase in mediating the immune escape of tumor cells in HBV-related hepatocellular carcinoma / 临床肝胆病杂志

ObjectiveTo determine whether HBV DNA polymerase is associated with T-cell failure and thus mediates the immune escape of HBV-related hepatocellular carcinoma （HCC） tumor cells， and to investigate the specific molecular mechanisms. MethodsLiver cancer cell lines Huh7 and HepG2 stably transfected with HBV DNA polymerase expression plasmid with Flag （Flag-HBV-P） and intercellular adhesion molecule-1 （ICAM1） were co-cultured with Jurkat cells， and MTT assay， qRT-PCR， and ELISA were used to measure Jurkat cell proliferation， activation （CD69 expression）， and secretion of the cytokine IFN-γ. RNA-seq was used to screen for differentially expressed immune-associated molecules between stably transfected cell lines and control cells， and mRNA half-life and protein half-life assays were used to determine the specific levels of the immune-associated molecules that were affected by HBV DNA polymerase. Related websites were used to predict the transcription factors that may bind to the promoter region of this immune-associated molecule， Western blot was used to verify the effect of transcription factors on the immune-associated molecule， and rescue experiment was used to determine whether HBV DNA polymerase affects the expression level of the immune-associated molecule through this transcription factor. The independent-samples t test was used for comparison between two groups. ResultsThe experimental group had significant reductions in Jurkat cell proliferation， activation， and cytokine secretion compared with the control group （all P<0.01）. Compared with the control group， the experimental group （Huh7 and HepG2 cell lines） had significant reductions in the mRNA and protein expression levels of ICAM1 （all P<0.01）. Website prediction identified the ICAM1 promoter and preliminarily highlighted NFKB1， RELA， and STAT3. Compared with the control group， the experimental group （Huh7 and HepG2 cell lines） had a significant reduction in the protein expression level of p65 （all P<0.01）. After p65 overexpression， there was a significant increase in the protein expression level of ICAM1， and after the expression of p65 was reduced， there was a significant reduction in the protein expression level of ICAM1 （all P<0.01）. In the rescue experiment， there was no significant difference in the protein expression level of ICAM1 between the control group and the experimental group after p65 overexpression （all P>0.05）. After the overexpression of ICAM1， there were no significant differences in the proliferation， activation， and cytokine secretion of Jurkat cells between the control group and the experimental group （Huh7 and HepG2 cell lines） （all P>0.05）. ConclusionHBV DNA polymerase downregulates the level of ICAM1 to mediate HCC immune escape by inhibiting the expression of p65 in NF-κB.

A novel homozygous nonsense mutation in NECTIN4 gene in a Pakistani family with ectodermal dysplasia syndactyly syndrome 1

Abstract Background Ectodermal dysplasia syndactyly syndrome 1 (EDSS1) is a rare hereditary disorder characterized by defects in teeth, hair, and nails in association with a fusion of the digits. Genetically, the disease phenotypes are caused by homozygous and compound heterozygous variants in NECTIN4 gene. Objective The main objective of the study was to identify the pathogenic sequence variant(s) for family screening and identification of carriers. Methods In the present study, the authors have investigated a large consanguineous family of Pakistani origin segregating autosomal recessive EDSS1. All the coding exons of the NECTIN4 gene were directly sequenced using gene-specific primers. Results The affected individuals presented the classical EDSS1 clinical features including sparse hair, hypoplastic nails with thick flat discolored nail plates, peg-shaped, conical, and widely spaced teeth with enamel hypoplasia, proximal cutaneous syndactyly of fingers and toes. Sequence analysis of the coding region of the NECTIN4 identified a novel nonsense variant [c.163C>T; p.(Arg55*)] in exon-2 of the gene. Computational analysis of protein structure revealed that the variant induced premature termination at Arg55 located in Ig-like V-loop region leading to loss of Ig-C2 type domains and transmembrane region, and most likely Nectin-4 function will be lost. Study limitation Gene expression studies are absent that would have strengthened the findings of computational analysis. Conclusion The present study expanded the phenotypic and mutation spectrum of the NECTIN4 gene. Further, the study would assist in carrier testing and prenatal diagnosis of the affected families.

Perillyl alcohol modulates activation, permeability and integrity of human brain endothelial cells induced by Plasmodium falciparum

BACKGROUND Cerebral malaria (CM) is a severe immunovasculopathy caused for Plasmodium falciparum infection, which is characterised by the sequestration of parasitised red blood cells (pRBCs) in brain microvessels. Previous studies have shown that some terpenes, such as perillyl alcohol (POH), exhibit a marked efficacy in preventing cerebrovascular inflammation, breakdown of the brain-blood barrier (BBB) and brain leucocyte accumulation in experimental CM models. OBJECTIVE To analyse the effects of POH on the endothelium using human brain endothelial cell (HBEC) monolayers co-cultured with pRBCs. METHODOLOGY The loss of tight junction proteins (TJPs) and features of endothelial activation, such as ICAM-1 and VCAM-1 expression were evaluated by quantitative immunofluorescence. Microvesicle (MV) release by HBEC upon stimulation by P. falciparum was evaluated by flow cytometry. Finally, the capacity of POH to revert P. falciparum-induced HBEC monolayer permeability was examined by monitoring trans-endothelial electrical resistance (TEER). FINDINGS POH significantly prevented pRBCs-induced endothelial adhesion molecule (ICAM-1, VCAM-1) upregulation and MV release by HBEC, improved their trans-endothelial resistance, and restored their distribution of TJPs such as VE-cadherin, Occludin, and JAM-A. CONCLUSIONS POH is a potent monoterpene that is efficient in preventing P. falciparum-pRBCs-induced changes in HBEC, namely their activation, increased permeability and alterations of integrity, all parameters of relevance to CM pathogenesis.