Single-cell transcriptome analysis reveals development atlas of mouse molar pulp cells / 中华口腔医学杂志

Objective: Single-cell RNA sequencing (scRNA-seq) was used to analyze the developing mouse molars, in order to construct a spatiotemporal development atlas of pulp cells, and further to reveal the developmental process and regulatory mechanism of tooth development. Methods: Ten mandibular first molars from C57BL/6 mice in postnatal day (PN) 0 and 3 were respectively dissected and digested to obtain single-cell suspensions. scRNA-seq was performed on 10× Genomics platform. PN 7 mouse molar scRNA-seq data were obtained from our previous study. PN 0, 3, and 7 scRNA-seq data were integrated for following analysis. The initial quality control, mapping and single cell expression matrix construction were performed by Cell Ranger. Quality control, standardization, dimensional reduction and cluster analysis were performed by using Seurat. Monocle was used to generate the pseudotime trajectory. Scillus was used to perform gene ontology analysis. In order to detect the spatiotemporal change of different population of pulp cells, the marker genes of each cluster were demonstrated by RNAscope in situ hybridization. Results: There were twenty-six cell clusters within mouse molars, which were identified as eight different cell types, including dental pulp cells, dental follicle cells, epithelial cells, immune cells, endothelial cells, perivascular cells, glial cells and erythrocytes. We further re-clustered and analyzed dental pulp cells. Cluster 0 were mature pulp cells, which located at the upper portion of crown. The main functions of cluster 0 were osteogenesis and extracellular structure organization. Cluster 1 were apical papilla cells, which located at the apical part of roots, whose main functions were extracellular structure organization and organ development. Cluster 2 were cycling cells, which were actively proliferated, resided in the lower portion of the crown. Cluster 3 and 4 were preodontoblasts and odontoblasts, respectively. Their functions were closely related to biomineralization. The proportion of mature pulp cells increased with the development process, while the proportion of cycling cells and odontoblast lineage decreased. According to the expression pattern of marker genes of each cluster, we constructed a cell atlas of dental pulp. Pseudotime trajectory analysis found there were two development trajectories within dental pulp. They both started from SPARC related modular calcium binding 2 (Smoc2)+ dental papilla cells, then went through DNA topoisomerase Ⅱ alpha (Top2a)+ cycling cells, and finally divided into coxsackie virus and adenovirus receptor (Cxadr)+ mature pulp cells or dentin sialophosphoprotein (Dspp)+ odontoblasts two lineages. Conclusions: scRNA-seq could fully discover the intercellular heterogeneity of cells on transcriptome level, which provides a powerful tool to study the process and regulatory mechanism of organ development.

Correlation between cervical curvature and cervical disc bulging in young patients with neck pain / 中国骨伤

OBJECTIVE@#To explore the correlation between the curvature of the cervical spine and the degree of cervical disc bulging in young patients with cervical pain.@*METHODS@#The clinical data of 539 young patients with neck pain from January 2015 to December 2018 were retrospectively analyzed. There were 251 males and 288 females, aged 18 to 40 (32.2±6.3) years old. The cervical curvature and cervical disc bulging were measured by cervical X-ray and MRI. According to cervical curvature, the patients were divided into 175 cases of cervical lordosis group (cervical curvature > 7 mm), 163 cases of cervical erection group (0

Morphological changes in the midface of Beagle dogs treated by trans-sutural distraction / Cambios morfológicos en la cara media de perros Beagle tratados por distracción trans-sutural

Trans-sutural distraction is a biological process that induces the formation of new bone and changes the position of bone by pulling on growing suture under the action of external forces. Currently, therapy to midfacial hypoplasia treated by trans-sutural distraction has been applied. In this study, Beagle dogs were selected as experimental animals, and a traction device designed by ourselves was applied to Beagle dogs to simulate the treatment process of trans-sutural distraction in human face, so as to provide a basis for the subsequent research on the related mechanism of trans-sutural distraction. The objective is that the animal model can provide the basis for the follow-up study of transsutural distraction. 45 month beagle dogs were randomly divided into two groups 3 in experiment group and 3 in control group. Implant nails were implanted as the bone marker in the bilateral zygomatic temporal suture, zygomandibular maxillary suture and palatine transverse suture in experimental group. The traction of the maxilla was carried out by the external cranial traction frame with canine fossa as bearing point, 800g force each side, elastic traction for 15 days. The control group only implanted the implant nail as the bone marker on both sides of the bone suture. The distance between two implant nails was measured by vernier calipers and X-ray examination, compared with preoperative and postoperative changes. X-ray and cephalometric measurements were used to measure change in the cranial basal angle. HE staining was used to observe the width of the bone seams, the morphology and structure of the cells and the tissue of the new bone under the phase contrast microscope. Then descriptive statistical analysis and t-test between two independent samples are carried out for the measurement data. The experimental group had a good retention of the beagle traction frame. In the experimental group, the maxillaries of dogs were protrudent in the process of traction gradually and the occlusal relationship changed to type II malocclusion. When the traction is 15 days, the coverage distance is about 8~9 mm. Before and after the traction, the distance between landmark points indicated that the spacing between the transverse palatine suture was the largest (experimental group: 5.52±0.19 mm control group 1.31±0.06 mm P<0.05), and zygomaticotemporal suture was the second (experimental group: 3.12±0.15 mm, control group 0.73±0.04 mm, P<0.05), and zygomaticomaxillary suture was less (experimental group: 2.60±0.34 mm, control group 0.53±0.05 mm, P<0.05). The cranial basal angle was no change before and after operation (controlgroup: 32.3±1.3°, experimental group: 33.2±1.1° P>0.05. Histology showed that the collagenous fibers in the suture of the control group were denser and the osteoblasts were visible on the edge of the suture, showing osteogenic activity. The experimental group significantly widened suture (experimental group: 1209.388±42.714 µm, control group 248.276±22.864 µm, P<0.05), the number of fibroblasts increased significantly with loose collagen fiber. The direction of cell and fiber arrangement were parallel to the traction force. There were many small blood vessels and marrow cavities, and the bone trabecula around the bone suture was thin (experimental group: 23.684±3.774 mm, control group: 86.810±9.219 mm, P < 0.05), showing active osteogenic activity. The growing beagle dog can be used to establish a suture traction animal model for experimental study. In the experiment, Kirschner wire was used to penetrate the bottom plane of the piriform hole of the maxilla (about the position of the canine fossa at the back) and the traction direction was basically the same as the growth direction, and the maxilla was basically parallel and moved forward.

La distracción trans-sutural es un proceso biológico que induce la formación de hueso nuevo y cambia la posición del éste al tirar de la sutura en crecimiento bajo la acción de fuerzas externas. Actualmente, se ha aplicado la terapia para la hipoplasia de la cara media tratada por distracción trans-sutural. En este estudio, fueron seleccionados perros Beagle como animales experimentales, y un dispositivo de tracción fue instalado a los perros para simular el proceso de tratamiento de la distracción trans-sutural en el rostro humano. El objetivo fue proporcionar una base para la investigación posterior sobre mecanismos relacionados con la distracción trans-sutural. El modelo animal puede proporcionar la base para este tipo de estudio de seguimiento de la distracción trans-sutural. Perros Beagle de 45 meses de edad se dividieron aleatoriamente en dos grupos: 3 en el grupo experimental y 3 en el grupo control. Los clavos de implante se usaron como marcadores óseos en la sutura temporal cigomática bilateral, la sutura maxilar cigomandibular y en la sutura transversal palatina en el grupo experimental. La tracción del maxilar se realizó mediante el marco de tracción craneal externo con fosa canina como punto de apoyo, 800 g de fuerza a cada lado, tracción elástica durante 15 días. En el grupo control solo se implantó el clavo del implante como marcador óseo en ambos lados de la sutura. La distancia entre dos clavos de implante se midió mediante calibradores de vernier y examen de rayos X, en comparación con los cambios preoperatorios y postoperatorios. Se utilizaron mediciones cefalométricas y de rayos X para medir el cambio en el ángulo basal craneal. La tinción con HE se usó para observar el ancho de las suturas óseas, la morfología y la estructura de las células y el tejido del hueso nuevo bajo el microscopio de contraste de fase. Luego se realizó un análisis estadístico descriptivo y una prueba t entre dos muestras independientes para los datos de medición. El grupo experimental tuvo una buena retención del cuadro de tracción del Beagle. En el grupo experimental, los maxilares de los perros sobresalieron gradualmente en el proceso de tracción y la relación oclusal cambió a maloclusión tipo II. Cuando la tracción era de 15 días, la distancia de cobertura fue de aproximadamente 8 ~ 9 mm. Antes y después de la tracción, la distancia entre los puntos de referencia indicaba que el espacio entre la sutura palatina transversal era más grande (grupo experimental: 5,52 ± 0,19 mm, grupo de control 1,31 ± 0,06 mm, P <0,05), y la sutura cigomáticotemporal fue la segunda. (Grupo experimental: 3,12 ± 0,15 mm, grupo control 0,73 ± 0,04 mm, P <0,05), y la sutura cigomaticomaxilar fue menor (grupo experimental, 2,60 ± 0,34 mm, grupo control 0,53 ± 0,05 mm, P <0,05). El ángulo basal craneal no cambió antes ni después de la operación (grupo control 32,3 ± 1,3, grupo experimental, 33,2 ± 1,1 ° , P> 0,05). La histología mostró que las fibras colágenas en la sutura del grupo control eran más densas y los osteoblastos se observaron en el margen de la sutura, mostrando actividad osteogénica. En el grupo experimental se amplió significativamente la sutura (1209,388 ± 42,714 µm, grupo control 248,276 ± 22,864 µm, P <0,05), el número de fibroblastos aumentó significativamente con fibras colágenas dispersas. La dirección de la disposición de la celda y las fibras era paralela a la fuerza de tracción. Se observó gran cantidad de vasos sanguíneos pequeños, cavidades medulares, y trabéculas óseas alrededor de la sutura ósea (grupo experimental: 23,684 ± 3,774 mm, grupo control: 86,810 ± 9,219 mm, P <0,05), que mostró actividad osteogénica activa. El perro Beagle en crecimiento se puede utilizar para estudios experimentales y así establecer un modelo animal de tracción de sutura. En el proceso, se usó alambre de Kirschner para penetrar en el plano inferior del foramen piriforme del maxilar (aproximadamente en la posición de la fosa canina en la parte posterior) y la dirección de tracción fue básicamente la misma que en el crecimiento.

Methodology, safety and applications of colonic transendoscopic enteral tubing (with video) / 中华消化内镜杂志

Objective@#To evaluate the methodology, safety and clinical applications of colonic transendoscopic enteral tubing (TET) as a new method of fecal microbiota transplantation (FMT) and colonic administration.@*Methods@#This prospective study included patients who underwent colonic TET for FMT and(or) colonic administration in the Second Affiliated Hospital of Nanjing Medical University from October 2014 to December 2018. The TET procedure time, success rate, retention time of TET tube, factors influencing TET tube retention, adverse events and satisfaction degree were evaluated.@*Results@#A total of 257 patients underwent TET, among whom 130 patients (50.6%) for microbiota tronsplantation, 8 patients (3.1%) for colon-drip medication, 118 patients (45.9%) for FMT and colon-drip medication, and 1 patient (0.4%) without treatment after TET. The TET procedure time was 10.0±2.8 min. The number of endoscopic clips used was 3.5±1.0. The success rate of the TET procedure was 100.0% (257/257). The retention time of TET tube for 160 patients maintaining the tube for treatment was 9.3±3.8 days. Multivariate analysis indicated that endoscopic clip type (P=0.001) was an independent influencing factor for the retention time of the tube. A total of 9 patients (3.5%) reported adverse events of mild anus discomfort, 4 patients (1.6%) of mobile inconvenience, 3 (1.2%) of anal pain, 2 (0.8%) of mild abdominal pain, 2 (0.8%) of mild bloating, and 1 (0.4%) of mild anal bleeding. No severe adverse events were observed in this study. The total satisfaction degree on colonic TET was 97.3% (250/257) in all patients.@*Conclusion@#The colonic TET, a safe and easy-operating endoscopic interventional technology with a high degree of patients satisfaction, can be used for colonic delivering of FMT and medications for various diseases.

Methodology, safety and applications of colonic transendoscopic enteral tubing (with video) / 中华消化内镜杂志

Objective:To evaluate the methodology, safety and clinical applications of colonic transendoscopic enteral tubing (TET) as a new method of fecal microbiota transplantation (FMT) and colonic administration.Methods:This prospective study included patients who underwent colonic TET for FMT and(or) colonic administration in the Second Affiliated Hospital of Nanjing Medical University from October 2014 to December 2018. The TET procedure time, success rate, retention time of TET tube, factors influencing TET tube retention, adverse events and satisfaction degree were evaluated.Results:A total of 257 patients underwent TET, among whom 130 patients (50.6%) for microbiota tronsplantation, 8 patients (3.1%) for colon-drip medication, 118 patients (45.9%) for FMT and colon-drip medication, and 1 patient (0.4%) without treatment after TET. The TET procedure time was 10.0±2.8 min. The number of endoscopic clips used was 3.5±1.0. The success rate of the TET procedure was 100.0% (257/257). The retention time of TET tube for 160 patients maintaining the tube for treatment was 9.3±3.8 days. Multivariate analysis indicated that endoscopic clip type ( P=0.001) was an independent influencing factor for the retention time of the tube. A total of 9 patients (3.5%) reported adverse events of mild anus discomfort, 4 patients (1.6%) of mobile inconvenience, 3 (1.2%) of anal pain, 2 (0.8%) of mild abdominal pain, 2 (0.8%) of mild bloating, and 1 (0.4%) of mild anal bleeding. No severe adverse events were observed in this study. The total satisfaction degree on colonic TET was 97.3% (250/257) in all patients. Conclusion:The colonic TET, a safe and easy-operating endoscopic interventional technology with a high degree of patients satisfaction, can be used for colonic delivering of FMT and medications for various diseases.

Intervention Effect of Shenfu Injection on the Nuclear Translocation of HMGB 1 in LPS-induced RAW 264.7 Cells / 中国药房

OBJECTIVE：To investigate the intervention effect of Shenfu i njection（SFI）on the nuclear translocation of high mobility group box 1（HMGB1） in lipopolysaccharide （LPS）-induced RAW 264.7 cells. METHODS ： Using LPS-induced RAW264.7 cells as objects ，the histone deacetylase inhibitor RGFP 966 as positive control ，CCK-8 assay was used to screen drug dosage，and the effects of low ，medium and high doses （3，6，12 μL/mL）of SFI on HMGB 1 nuclear translocation in RAW 264.7 cells were observed by immunofluorescence method ；mRNA expression of HMGB 1 in RAW 264.7 cells were detected by real time fluorescent PCR. Western blotting assay was used to determine protein expression of HMGB 1 and Toll-like receptor 4（TLR4）；the expression of HMGB 1 were compared between nucleus and cytoplasm. The levels of HMGB 1，IL-1β and TNF-α in supernatant of cells were detected by ELISA. RESULTS ：In blank control group ，HMGB1 was mainly located in the nucleus ；after LPS induction， HMGB1 migrated from nucleus to cytoplasm. Compared with blank control group ， mRNA and protein （No.81760738） expression of HMGB 1， protein expression of TLR 4 in RAW264.7 cells as well as the levels of HMGB 1，IL-1β and TNF-α in supernatant of cells were increased significantly in LPS group （P＜0.01）. The protein expression of HMGB 1 was decreased significantly in nucleus while was in creased significantly in cytoplasm （P＜0.01）. After SFI treatment ，the nuclear translocation and secretion of HMGB 1 were inhibited in different degrees ；compared with LPS group ，mRNA and protein expression of HMGB 1 in administration groups ，protein expression of TLR 4 in RAW 264.7 cells of positive control group ，SFI medium- and high-dose groups as well as the levels of HMGB 1，IL-1β and TNF-α in supernatant of cells in administration groups were decreased significantly （P＜0.01）. In positive control group ，SFI medium- and high-dose groups ，the protein expressions of HMGB1 in nucleus were increased significantly ，while protein expressions of HMGB 1 in cytoplasm were decreased significantly （P＜0.01）. CONCLUSIONS ：SFI may inhibit the nuclear translocation and secretion of HMGB 1 in RAW 264.7 cells，thus avoiding the activation of inflammatory pathways and the production of inflammatory factors ，so as to reduce the inflammatory response induced by LPS.

Improvement and Anti-inflammation Mechanism of Shenfu Injection on Lung Tissue in Endotoxin Shock Model Rats / 中国药房

OBJECTIVE： To study the improvement and anti-inflammation mechanism of Shenfu injection on lung tissue of endotoxin shock model rats. METHODS： Totally 48 rats were randomized into control group，model group，dexamethasone group （positive control，1 mg/kg） and Shenfu injection low-dose，medium-dose and high-dose groups （5，10，15 mL/kg），with 8 rats in each group. Except for normal group， other groups were given intraperitoneal injection of lipopolysaccharide （LPS） to induce endotoxin shock model. After modeling， each group was given relevant medicine once intraperitoneally. 24 h after medication， HE staining was used to observe pathological changes of lung tissue in rats and pathological scoring was conducted. RT-PCR was used to determine mRNA levels of P65 and P50 proteins related to NF-κB signaling pathway. Western blot assay was used to determine the expression levels of P65 and P50 proteins in lung tissue， and the expression levels of P65 protein in nucleus and cytoplasm of lung tissue were also determined. The level of TNF-α in plasma in rats were determined by ELISA. RESULTS： Compared with control group， alveolar septum became thicker， obvious vascular engorgement was found， and a large number of neutrophils infiltrated the interstitium in model group. Histopathological score， mRNA and protein expression levels of P65 and P50 in lung tissues were increased significantly （P＜0.01 or P＜0.001）； the protein expression of levels P65 in nucleus and cytoplasm and level of TNF-α in plasma were increased significantly （P＜0.001）. Compared with model group， alveolar structure of rats in dexamethasone group and Shenfu injection medium-dose and high-dose groups was complete， no obvious bleeding was observed， and the degree of inflammatory cell infiltration was improved significantly. Histopathological score， mRNA and protein expression levels of P65 and P50 in lung tissue and level of TNF-α in plasma were decreased significantly （P＜0.05 or P＜0.01 or P＜0.001）. The protein expression level of P65 in nucleus and cytoplasm of lung tissue were decreased significantly in dexamethasone group and Shenfu injection low-dose and medium-dose groups were decreased significantly （P＜0.05 or P＜0.01 or P＜0.001）. CONCLUSIONS： Shenfu injection can decrease mRNA and protein expression levels of P65 and P50 in lung tissue， level of TNF-α in plasma， and protect lung tissue of endotoxin shock rats.

Morphological observation of bone marrow mesenchymal stem cells under matrigel three dimensional culture conditions / Observación morfológica de células madre mesenquimales de médula ósea en condiciones de cultivo tridimensional con matrigel

SUMMARY: Matrigel is a basement membrane matrix extracted from the EHS mouse tumor containing extracellular matrix protein, its main components are laminin, type IV collagen, nestin, heparin sulfate, growth factor and matrix metalloproteinase.At room temperature, Matrigel polymerized to form a three dimensional matrix with biological activity. It can simulate the structure, composition, physical properties and functions of the cell basement membrane in vivo, which is beneficial to the culture and differentiation of the cells in vitro, and can be used for the study of cell morphology, biochemical function, migration, infection and gene expression. In this study, Matrigel three-dimensional culture model of bone marrow mesenchymal stem cells(BMSCs) was established, and its morphology, proliferation and survival were observed. BMSCs were isolated and cultured with whole bone marrow adherence method. The Second generation BMSCs with good growth condition were selected and mixed with Matrigel to form cell gel complexes. The morphology and proliferation of mesenchymal stem cells were observed by phase contrast microscope and HE staining,Live/Dead staining was used to evaluate the cell activity.Phase contrast microscopy showed that BMSCs were reticulated in Matrigel and proliferated well, After 7 days, the matrix gel gradually became soft and collapsed, a few cell reticular crosslinking growth was seen at 14 days; HE staining showed that the cytoplasm of the cells was larger on the fourth day and the cells were elongated and cross-linked on the seventh day; Live/dead staining showed that most cells showed green fluorescence with the prolongation of culture time, on the first, 4 and 7 days, the activity of bone marrow mesenchymal stem cells in Matrigel gradually increased, and the percentages were 92.57 %, 95.54 % and 97.37 %, respectively. Matrigel three-dimensional culture system can maintain the morphology, function and proliferation ability of bone marrow mesenchymal stem cells.

RESUMEN: Matrigel es una matriz de membrana basal extraída del tumor de ratón EHS que contiene proteína de matriz extracelular. Los componentes principales son laminina, el colágeno tipo IV, nestina, sulfato de heparina, factor de crecimiento y metaloproteinasa de matriz. A temperatura ambiente, Matrigel se polimerizó para formar una matriz tridimensional. Es posible simular la estructura, la composición, las propiedades físicas y las funciones de la membrana basal celular in vivo, lo que es beneficioso para el cultivo y la diferenciación de las células in vitro, y se puede utilizar para el estudio de la morfología celular, la función bioquímica, la migración, infección y expresión génica. En este estudio, se estableció el modelo de cultivo tridimensional Matrigel de células madre mesenquimales de médula ósea (BMSC), y se observó su morfología, proliferación y supervivencia. Las BMSC fueron aisladas y cultivadas con el método de adherencia de la médula ósea completa. Se seleccionaron las BMSC de segunda generación con buenas condiciones de crecimiento y se mezclaron con Matrigel para formar complejos de gel de células. La morfología y la proliferación de las células madre mesenquimales se observaron con microscopio de contraste de fase y se tiñó con Hematoxilina-Eosina (HE); para evaluar la actividad celular se usó la tinción Live/Dead. La microscopía de contraste mostró que las BMSC se reticularon en Matrigel y proliferaron bien. Después de 7 días, se observó que el gel de matriz gradualmente se volvió blando y colapsó, y se visualizó un cruce transversal de algunas células reticulares a los 14 días. La tinción mostró que la mayoría de las células mostraron una fluorescencia verde con la prolongación del tiempo de cultivo; en los primeros 4 y 7 días, la actividad de las células madre mesenquimales de la médula ósea en Matrigel aumentó gradualmente y los porcentajes fueron de 92,57 %, 95,54 % y 97,37 %, respectivamente. El sistema de cultivo tridimensional de Matrigel puede mantener la morfología, la función y la capacidad de proliferación de las células madre mesenquimales de la médula ósea.

In vivo metabolism of three coumarins from Chimonanthi Radix based on UHPLC-QTOF-MS/MS / 中国中药杂志

Scopolin (SC-1), scopoletin (SC-2) and isofraxidin (IS-1) are the main active constituents in Chimonanthi Radix. However, the in vivo metabolism of SC-1, SC-2 and IS-1 have not been comprehensively clarified. In this study, the in vivo metabolic profiles of these three coumarins in the rat plasma, urine and feces were analyzed. Ultra-high performance liquid chromatography-quadrupole time of flight mass spectrometry (UHPLC-QTOF-MS/MS) method was applied to characterize the prototypes and metabolites of SC-1, SC-2 and IS-1 in rat feces, urine, and plasma after intravenous administration. A total of 11 metabolites of the three parent compounds were tentatively identified. The main metabolic pathways were analyzed by identification of metabolites, and it was found that these three coumarins underwent multiple in vivo metabolic reactions including glucuronidation, sulfonation, isomerism and reduction. In this study, the analysis of metabolites of three coumarins basically demonstrated their in vivo metabolic process, providing basis for the further pharmacokinetics and pharmacological evaluations of SC-1, SC-2 and IS-1.

Application of CLT guiding offline extubation in patient with tracheal intubation of mechanical ventilation / 中国医学装备

Objective: To explore the application of the cuff-leak test(CLT)guiding offline extubation in patients with tracheal intubation of mechanical ventilation.Methods: 64 patients with tracheal intubation who underwent mechanical ventilation were divided into CLT-negative group(47 cases)and CLT-positive group(17 cases)according to leakage situation of CLT.The CLT guiding offline extubation were adopted in the study and some basic situations,such as body mass index(BMI),APACHE-Ⅱ and so on,of the two groups were compared.And the relative situation of intubation,blood gas analysis index,vital signs,the occurrence rate of upper airway obstruction(UAO)post removing intubation and re-intubation rate between the two groups also were compared,and then the risk factors of influencing UAO were further analyzed.Results: The BMI of CLT-negative group was significantly smaller than that of CLT-positive group(t=2.44,P＜0.05).The occurrence rate of UAO and re-intubation rate of CLT-negative group(6.38％and 2.13％)were significantly lower than that of CLT-positive group(35.29％and 17.65％)(x2=8.63,x2=5.13,P＜0.05),respectively.The differences of BMI,APACHE-Ⅱ scores,intubation time,air sac pressure,PaO2and SpO2between patients with UAO and patients without UAO were significant(t=5.63,t=2.65,t=4.27,t=3.35,t=2.37,t=2.66,P＜0.05).The results of Logistic regression analysis showed that the BMI,APACHEⅡ score,intubation time,air sac pressure were independent risk factors for occurring UAO post extubation.Conclusion: The CLT guiding offline extubation in patient with tracheal intubation of mechanical ventilation can effectively reduce the re-intubation rate.For these patients with obesity,high APACHE-Ⅱ score,long intubation time and big air sac pressure,the risk of occurring UAO is higher.Therefore,the number of intubation pre extubation should be reduced for them,and their physiological status should be comprehensively assessed so as to decrease the occurrence rate of UAO.

PTC: The correlation between sonographic characteristics of ultrasound and metastasis of cervical lymph node / 中国医学装备

Objective: To analyze the correlation between sonographic characteristics of ultrasound and metastasis of cervical lymph node of papillary thyroid carcinoma (PTC). Methods: 498 patients with PTC who once received radical operation of thyroid cancer and cervical lymph node dissection were divided into metastasis group (n=163) and non-metastasis group (n=335) according to results post pathology. Gender, age, the number of lesion, maximum diameter, aspect ratio, margin, whether contacted with tunica, border, halo of ultrasound, whether existed hypoecho in interior, whether the echo was uniform, whether existed difference about some characteristics of calcification and blood flow between the two groups were compared and analyzed. The correlation of characteristics and cervical lymphatic metastasis was analyzed by using single factor analysis, and the malignancy phenomenon of ultrasonogram that was closely relevant with cervical lymph node metastasis of patients with PTC were screened out by using Logistic regression. Results: As the results of single factor analysis, there were a certain correlation between series of factors, such as gender, age, the number of lesion, maximum diameter, margin, whether contacted with tunica, blood flow and whether contained tiny calcification, and cervical lymph node metastasis, and these correlations were significant (x2=33.094, x2=126.566, x2=127.961, x2=145.087, x2=53.757, x2=6.807, P<0.05). The results of Logistic regression indicated that these factors, included multi- lesions, maximum diameter more than 10.0mm, irregular margin, contacted with tunica of thyroid and abundant blood flow, were closely correlative with the malignancy phenomenon of ultrasonogram of cervical lymphatic metastasis of patients with PTC, and their correlation coefficient were significant (OR=3.103, OR=3.595, OR=2.822, OR=6.317, OR=2.078, P<0.05). Conclusion: The characteristics of ultrasonogram contributes to increase the detectable rate of ultrasound for cervical lymph node metastasis of patients with PTC, and it can provide reference for clinical decision.

Establishment of a chronic rhinosinusitis model in C57BL/6J mice / 医学研究生学报

Objective Streptococcus pneumoniae is one of the main pathogenic factors of chronic rhinosinusitis (CRS). This study was to explore the method of establishing a CRS model in C57BL/6J mice.Methods Thirty C57BL/6J mice were equally randomized into a CRS model, a sham operation and a blank control group. The CRS model was made by implanting type-Ⅲ streptococcus pneumoniae in the maxillary sinuses, the mice of the sham operation group underwent opening and suturing of the maxillary sinuses without implantation of any streptococcus pneumonia, while the blank controls did not receive any operation. Two months later, all the mice were sacrificed and the nasal sinus mucosal tissue was harvested, embedded with paraffin, sectioned, and subjected to HE staining, followed by observation of the pathological changes under the microscope.Results The CRS model mice exhibited significant proliferation and disorderly arrangement of epithelial cells in the nasal mucosal tissue, with degeneration, necrosis, exfoliation, ulceration, goblet cell proliferation, gland and tissue hyperplasia, and infiltration of lymphocytes, plasmocytes and monocytes. No obvious inflammation was observed in the sham operation and blank control groups. The pathology index score of the nasal sinus mucosal tissue was significantly higher in the CRS model than in the sham operation and blank control groups (\[14.800±5.200\] vs \[2.000±2.906\] and \[1.800±2.098\], P0.05).Conclusion The CRS model was successfully established in mice, which has provided some reference for the construction of the CRS model in animals.

Stable co-expression of human kappa opioid receptor and PKAcat-EGFP in CHO cells and functional identification / 中国药理学通报

Aim To establish a cell model which stably co-ex-press human kappa opioid receptor (hKOR) and enhanced green fluorescent protein ( EGFP) labeled catalytic domain of cAMP-dependent protein kinase A(PKAcat) fusion protein (PKAcat-EGFP) in Chinese hamster ovary(CHO) cells, laying the foun-dation for the high-throughput screening of hKOR drugs and drug molecular mechanisms in vitro. Methods Hygromycin B resist-ant hKOR recombinant plasmid [ pcDNA3.1/Hygro ( + ) -hKOR] was transfected into CHO cells stably expressing PKA-cat-EGFP by a lipofectin based method. Transfected cells were selected in culture medium containing hygromycin B. The posi-tive clones were selected by PKA redistribution assay. Z’ factor was used for evaluation and validation the reliability of the cell model. PKA redistribution assay and LANCE cAMP 384 Kit were used to test the function of the receptors in selected clone. Results CHO-PKAcat-EGFP/hKOR-13 cell model exhibited stable response in PKA redistribution assay and LANCE cAMP 384 Kit. Treated with 100 nmol·L-1U-50488 for 30 min, the average value of Z’ factor was 0.596, proving the reliability of the cell model. The hKOR expression in cell model remained stable after a few generations. Conclusion The CHO-PKAcat-EGFP/hKOR-13 cell model with stable co-expression of hKOR and PKAcat-EGFP has been successfully established.

Treatment of secondary intra-and extrahepatic cholangiolithiasis with electrohydraulic shock wave lithotripsy combined with laparoscope and choledochoscope / 局解手术学杂志

Objective To explore the clinical application value of electrohydraulic shock wave lithotripsy combined with laparoscope and choledochoscope in treatment of intra-and extrahepatic cholangiolithiasis.Methods The clinical data of 42 patients with intra-and extrahepatic cholangiolithiasis treated by laparoscope and choledochoscope(control group) in hepatobiliary surgery department of affiliated hospital of Xuzhou medical university from May 2012 to December 2015,and patients with intra-and extrahepatic cholangiolithiasis treated by electrohydraulic shock wave lithotripsy combined with laparoscope and choledochoscope (combined group) from January 2016 to February 2017,were retrospectively analyzed.The rate of transferring to laparotomy (transfer to laparotomy due to inflammatory adhesion or difficulty to remove),residual rate of choledocholithiasis,incidence of bile leakage and incidence of common bile duct stricture between two groups were compared.Results All patients in both groups were recovered.In the control group,there were 6 cases(6/42) transferred to laparotomy,among which 2 cases were transferred to laparotomy due to severe abdominal inflammatory adhesion and 4 cases were due to the difficulty to remove the calculus,without bile leakage or common bile duct stricture.There were 3 cases with residual choledocholithiasis,but without bile leakage or common bile duct stricture.In the combined group,there were 2 cases(2/42) transferred to laparotomy due to severe abdominal inflammatory adhesion and 5 cases used the technique of electrohydraulic shock wave lithotripsy without post-operation residual choledocholithiasis,bile leakage or bile duct stricture.In the control group,the residual choledocholithiasis was removed with T tube fistula choledochoscope.Conclusion In the process of minimally invasive surgery to treat intra-and extrahepatic cholangiolithiasis,electrohydraulic shock wave lithotripsy under laparoscope and choledochoscope can decrease the incidence of transferring to laparotomy due to the difficulty to remove calculus and the residual rate of calculus.It can be applied safely and efficiently to patients with intra-and extrahepatic cholangiolithiasis but with laparoscopic surgery indications.

Bone marrow mesenchymal stem cells on Matrigel: growing and changing / 中国组织工程研究

BACKGROUND: Matrigel as an extracellular matrix complex can facilitate cell proliferation, differentiation and collagen secretion in a cell culture system. OBJECTIVE: To establish a three-dimensional culture model of Matrigel combined with bone marrow mesenchymal stem cells (BMSCs) and to observe the morphology, proliferation and survival of BMSCs in the Matrigel three-dimensional culture model. METHODS: BMSCs were isolated and cultured by the whole bone marrow adherent method, followed by osteogenic and adipogenic induction and identification. The growth curve of passage 3 BMSCs was determined through the CCK-8 experiment. Passage 2 BMSCs were combined with Matrigel, and the morphology and proliferation of BMSCs were observed by hematoxylin-eosin staining under a phase contrast microscope. The viability of the cells was evaluated by the Live/Dead staining. RESULTS AND CONCLUSION: (1) BMSCs cultured by the whole bone marrow adherent method had the ability to differentiate into osteoblasts and adipocytes. (2) BMSCs exhibited an "S"-shaped growth curve, which was consistent with the growth characteristics of normal cells. (3) Under the phase contrast microscope, BMSCs cultured by the Matrigel model were extended and interconnected in a three-dimensional network growth state with good proliferation. The Matrigel gradually became soft and collapsed over time (7days) and a few cells were still in a network growth after 14 days. Hematoxylin-eosin staining results showed that the cell cytoplasm was larger at 4 days and the cells became thinned and mutually cross-linked at 7 days. (4) The active percentage of BMSCs in the Matrigel model was 92.57%, 95.54% and 97.37% at 1, 4, 7 days of culture, respectively. To conclude, BMSCs cultured on the Matrigel has good proliferation and high viability.

Robot-assistedlaparoscopic ureteral reimplantation:A report of 5 cases / 医学研究生学报

Objective Reports are relatively few at home and abroad onrobot-assistedlaparoscopic ureteral reimplantation ( RAUR) .This study summarizes our experience with RAUR and assesses the feasibility and clinical effect of the strategy . Methods We retrospectively analyzed the clinical data about 5 cases of lower ureterostenosistreated by RAUR in our department from May 2015 to February 2017. Results Operations were successfully completed in all the cases , with an estimatedintraoperative blood loss of 40-100 mL but no blood transfusion , nor conversion to open surgery , nor intestinal or major vascular injury .The urethral catheter was re-moved at 5-7 days after surgery and there were no postoperative complications .The patients were followed up for over 6 months , during which all showed increased glomerular filtration rate ( 2-12 mL/min) of the involved kidney at 3 months and ultrasonography revealed no ureterostenosisorhydronephrosis . Conclusion Robot-assistedlaparoscopic ureteral reimplantation is a safe , effective and minimally invasive surgical option for distal ureteral obstruction .

Magnetic resonanceT1ρversus T2 mapping for evaluating ischemic lumbar intervertebral disc degeneration in a rabbit model / 中国组织工程研究

BACKGROUND: MRI T1ρ and T2 mapping have been applied to study lumbar intervertebral disc degeneration in human and rhesus monkey, showing that they can be used for evaluating the early degeneration, but their application in New Zealand rabbit lumbar intervertebral disc degeneration is never reported.OBJECTIVE: To compare the relaxation time values of T1ρ and T2 mapping at different time points in a New Zealand rabbit model of ischemic lumbar intervertebral disc degeneration, and to compare the sensitivity for degeneration.METHODS: Fifteen New Zealand rabbits were randomly divided into three groups, and one of intervertebral discs at L4-5,L5-6 and L6-7 was designed for ischemic lumbar intervertebral disc degenerative model and the other two discs for controls. All the rabbits underwent T1ρ and T2 mapping preoperatively, 1, 3 or 6 months postoperatively to analyze the changes in the relaxation time at different time points.RESULTS AND CONCLUSION: (1) In the modeling group, the T2 mapping relaxation time of nucleus pulposus was (118.53±20.51) ms and (85.42±11.65) ms at 3 and 6 months after modeling, respectively, which showed significant difference when compared with preoperatively (146.21±16.93) ms (P < 0.05); but the time showed no significant difference compared with 1 month after modeling (P > 0.05). (2)T1ρ relaxation time of nucleus pulposus was (64.75±14.63) ms at 6 months after modeling, which showed significant difference when compared with preoperatively (87.88±8.87) ms (P < 0.05); but the time showed no significant difference compared with 1 and 3 months after modeling (P > 0.05). (3) In the control group, there was no significant difference in the T1ρ or T2 mapping relaxation time of nucleus pulposus before operation with 1, 3 or 6 months after operation (P > 0.05). The Pfirrmann grade of lumbar intervertebral disc was changed to Pfirrmann grades II-III at 6 months after modeling. (4) These results suggest that MR T1ρ and T2 mapping both are quantitative tools for evaluating the progress of ischemic lumbar intervertebral disc degeneration in New Zealand rabbits, especially the T2 mapping MRI may be more sensitive to early degenerative changes.

Effects and mechanisms of hepatitis B virus X protein on invasion and migration of hepatocellular carcinoma cells / 中华消化外科杂志

Objective To explore the effects and mechanisms of hepatitis B virus-X protein (HBx) on invasion and migration of hepatocellular carcinoma (HCC) cells.Methods The retrospective cohort study was conducted.The clinicopathological data of 30 patients with liver tumor (20 with HCC and 10 with benign tumor of liver) who were admitted to the Affiliated Hospital of Xuzhou Medical College between July 2014 and July 2015 were collected.HCC tissues of 20 patients with HCC (with history of HBV infection) were collected by surgical resection and peritumoral normal tissues (outside of tumor capsule) of 10 patients with benign tumor of liver (without history of HBV infection) were collected.The expressions of epidermal growth factor receptor 3 (ErbB3)in HCC tissues and peritumoral normal tissues were detected by immunohistochemistry (IHC).The relative expressions of ErbB3 and HBx in HCC tissues and peritumoral normal tissues were detected by Western blot,and relative expressions of ErbB3 in HepG2 of which green fluorescent protein (GFP) and GFP-HBx were respectively transfected were detected.The relative expressions of ErbB3 mRNA in HepG2 transfected by GFP and GFP-HBx were detected by real-time polymerase chain reaction (RT-PCR).The migration and invasion of HepG2 were respectively detected by Transwell assay with and without matrix.The measurement data with normal distribution were represented as $± s.The comparisons between groups were evaluated with the independent-sample t test.Correlation analysis was done by the Pearson test.Results (1) The expressions of ErbB3 were detected by IHC:relative value of mean optical density (MOD) of ErbB3 in HCC tissues of 20 patients with HCC and peritumoral normal tissues of 10 patients with benign tumor of liver were 2.54± 1.33 and O.99±0.29,respectively,with a statistically significant difference (t =6.542,P ＜ 0.05).(2) The relative expressions of ErbB3 and HBx were detected by Western blot:relative expressions of ErbB3 and HBx were respectively 0.79±0.13,1.10±0.28 in HCC tissues of 10 patients with HCC and 1.07±0.17,0 in peritumoral normal tissues of 10 patients with benign tumor of liver,with statistically significant differences (t =3.229,19.486,P＜0.05).The results of Pearson test showed that there was a positive correlation of expression between ErbB3 and HBx in HCC tissues (r=O.637,P＜ 0.05).(3) The relative expressions and transcriptional levels of ErbB3 were detected by Western blot and RT-PCR:relative expressions of ErbB3 in HepG2 of which GFP and GFP-HBx were respectively transfected were O.75±0.11 and 1.10±0.10,respectively,with a statistically significant difference (t=4.291,P＜0.05).The relative expressions of ErbB3 mRNA in HepG2 of which GFP and GFP-HBx were respectively transfected were O.38±0.03 and O.94±0.07,respectively,with a statistically significant difference (t=11.703,P＜O.05).(4) The effects of ErbB3 on migration and invasion of HepG2:numbers of transmenbrane cell in HepG2 of which His and His-ErbB3 were respectively transfected by Transwell assay with matrix were respectively 271± 18 and 463± 31,respectively,with a statistically significant difference (t =8.202,P＜0.05).Numbers of transmenbrane cell in HepG2 of which His and His-ErbB3 were respectively transfected by Transwell assay without matrix were respectively 315±38 and 549±34,respectively,with a statistically significant difference (t =8.310,P＜0.05).Conclusion HBx protein can promote the invasion and migration of hepatocellular carcinoma cells through up-regulating expressions of ErbB3 protein.

Analysis on population health in Minhang District of Shanghai within circumstance of universal health / 复旦学报（医学版）

Objective To evaluate the population health for disease prevention and control in Shanghai Minhang District of Shanghai the data of mortality from 1993 to 2015 and communicable diseases from 2002 to 2015.Methods We adopted descriptive epidemiological method to analyze the trends of average life expectancy (ALE),specific death rate and causes of death cis-position from 1993 to 2015,and the incident rates of communicable diseases,incidence trends in Minhang District from 2002 to 2015.Results Overall,the ALE of population in Minhang District increased 11.80 years from 1993 to 2015 (from 71.78 years in 1993 to 83.58 years in 2015),including the ALE of male population increased 14.03 years (from 67.43 years in 1993 to 81.37 years in 2015) and the ALE of female population elevated 9.67 years (from 76.22 years in 1993 to 85.89 years in 2015).In 2015,Crude death rate (CDR) was 755.35/105,which was 21.45％ higher than in 1993 and 2.71％ higher than in 2014,respectively.In the same year,standardized mortality rate (SMR) was 196.07/105,which was 54.17％ lower than in 1993 and 0.51％ lower than in 2014.The top five leading causes of death were circulatory system diseases,tumors,respiratory diseases,endocrine and metabolic diseases,and injury and poisoning,which contributed 91.33％ of the population death.From 2002 to 2015,a total of 23 kinds of notifiable infectious diseases were reported in Minhang District,including 62 845 cumulative cases and 152 cases died.The total reported incidence rate of communicable diseases sharply elevated by 291.98％ during 14 years (Z=10 943.83,P＜0.001),and it increased after standardized.The top five communicable diseases were hand foot and mouth disease (HFMD),scarlet fever,syphilis,tuberculosis and hepatitis B in 2015.Conclusions Over the years,Minhang District of Shanghai comprehensive implemented "health in all policies" by integrating the resources of all levels of regional healthcare institutions.The ALE of the residents was at a high level.The control and prevention of chronic non-communicable diseases and major communicable diseases will continue to be the priority of public health.

Analysis on population health in Minhang District of Shanghai within circumstance of universal health / 复旦学报（医学版）

Objective To evaluate the population health for disease prevention and control in Shanghai Minhang District of Shanghai the data of mortality from 1993 to 2015 and communicable diseases from 2002 to 2015.Methods We adopted descriptive epidemiological method to analyze the trends of average life expectancy (ALE),specific death rate and causes of death cis-position from 1993 to 2015,and the incident rates of communicable diseases,incidence trends in Minhang District from 2002 to 2015.Results Overall,the ALE of population in Minhang District increased 11.80 years from 1993 to 2015 (from 71.78 years in 1993 to 83.58 years in 2015),including the ALE of male population increased 14.03 years (from 67.43 years in 1993 to 81.37 years in 2015) and the ALE of female population elevated 9.67 years (from 76.22 years in 1993 to 85.89 years in 2015).In 2015,Crude death rate (CDR) was 755.35/105,which was 21.45％ higher than in 1993 and 2.71％ higher than in 2014,respectively.In the same year,standardized mortality rate (SMR) was 196.07/105,which was 54.17％ lower than in 1993 and 0.51％ lower than in 2014.The top five leading causes of death were circulatory system diseases,tumors,respiratory diseases,endocrine and metabolic diseases,and injury and poisoning,which contributed 91.33％ of the population death.From 2002 to 2015,a total of 23 kinds of notifiable infectious diseases were reported in Minhang District,including 62 845 cumulative cases and 152 cases died.The total reported incidence rate of communicable diseases sharply elevated by 291.98％ during 14 years (Z=10 943.83,P＜0.001),and it increased after standardized.The top five communicable diseases were hand foot and mouth disease (HFMD),scarlet fever,syphilis,tuberculosis and hepatitis B in 2015.Conclusions Over the years,Minhang District of Shanghai comprehensive implemented "health in all policies" by integrating the resources of all levels of regional healthcare institutions.The ALE of the residents was at a high level.The control and prevention of chronic non-communicable diseases and major communicable diseases will continue to be the priority of public health.