Background::Bladder cancer, characterized by a high potential of tumor recurrence, has high lifelong monitoring and treatment costs. To date, tumor cells with intrinsic softness have been identified to function as cancer stem cells in several cancer types. Nonetheless, the existence of soft tumor cells in bladder tumors remains elusive. Thus, our study aimed to develop a microbarrier microfluidic chip to efficiently isolate deformable tumor cells from distinct types of bladder cancer cells.Methods::The stiffness of bladder cancer cells was determined by atomic force microscopy (AFM). The modified microfluidic chip was utilized to separate soft cells, and the 3D Matrigel culture system was to maintain the softness of tumor cells. Expression patterns of integrin β8 (ITGB8), protein kinase B (AKT), and mammalian target of rapamycin (mTOR) were determined by Western blotting. Double immunostaining was conducted to examine the interaction between F-actin and tripartite motif containing 59 (TRIM59). The stem-cell-like characteristics of soft cells were explored by colony formation assay and in vivo studies upon xenografted tumor models. Results::Using our newly designed microfluidic approach, we identified a small fraction of soft tumor cells in bladder cancer cells. More importantly, the existence of soft tumor cells was confirmed in clinical human bladder cancer specimens, in which the number of soft tumor cells was associated with tumor relapse. Furthermore, we demonstrated that the biomechanical stimuli arising from 3D Matrigel activated the F-actin/ITGB8/TRIM59/AKT/mTOR/glycolysis pathways to enhance the softness and tumorigenic capacity of tumor cells. Simultaneously, we detected a remarkable up-regulation in ITGB8, TRIM59, and phospho-AKT in clinical bladder recurrent tumors compared with their non-recurrent counterparts.Conclusions::The ITGB8/TRIM59/AKT/mTOR/glycolysis axis plays a crucial role in modulating tumor softness and stemness. Meanwhile, the soft tumor cells become more sensitive to chemotherapy after stiffening, that offers new insights for hampering tumor progression and recurrence.

Background and purpose:Long non-coding RNA(lncRNA)LINC01410,with a length of 647 bp,participates in a variety of tumor biological processes.However,the role and mechanism of LINC01410 involved in esophageal squamous cell carcinoma(ESCC)remain unclear.This study aimed to explore the potential mechanism of LINC01410 promoting ESCC proliferation and invasion,to provide a potential prognostic indicator and therapeutic target for individuals with ESCC.Methods:Gene Expression Profiling Interactive Analysis 2(GEPIA2)databases were used to analyze the expression of LINC01410 and overall survival in esophageal squamous cell carcinoma data set in the Cancer Genome Atlas(TCGA).Gene Set Enrichment Analysis(GSEA)was performed to identify the underlying signaling pathways involved in the biological effects of LINC01410 in ESCC.A total of 62 pairs of ESCC tissues and paracancerous tissues from ESCC patients who underwent radical surgery in the Department of Thoracic Surgery at the First Affiliated Hospital of Xinxiang Medical College and the First People's Hospital of Pingdingshan City from January 2020 to December 2021 were collected.This project has been approved by the Hospital Ethics Committee(First Affiliated Hospital of Xinxiang Medical College,No.2018036;First People's Hospital of Pingdingshan City,No.2019-018).The expression of LINC01410 in ESCC tissues was detected by real-time fluorescence quantitative polymerase chain reaction(RTFQ-PCR).We transfected EC109 cells with LV-NC or LV-over/LINC01410 and EC9706 cells with shRNA-NC or shRNA-LINC01410.Stable transfected cells(EC109/NC,EC109/OE,EC9706/NC and EC9706/KD)were selected in primary cell culture medium containing puromycin.The expression of LINC01410 was detected by RTFQ-PCR.The impact of LINC01410 on ESCC cell proliferation was determined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT)and colony formation assays.The effect of LINC01410 on ESCC cell invasion was detected by transwell migration assay.T cell factor/lymphoid enhancer factor 1(TCF/LEF1)luciferase reporter assay was performed to validate the effect of LINC01410 on the activity of canonical Wnt/β-catenin signaling pathway.The expressions of Wnt/β-catenin and epithelial-mesenchymal transition(EMT)signal pathway related proteins in ESCC cells were detected by Western blot.Results:By analyzing the LINC01410 expression from ESCC samples in TCGA by GEPIA2,we found LINC01410 was consistently increased in ESCC tumors compared with normal tissues(P＜0.05),and high LINC01410 expression was associated with poorer overall survival(OS).RTFQ-PCR assay showed that expressions of LINC01410 were higher in esophageal cancer tissues and esophageal cancer cells(EC109 and EC9706)than in precancerous tissues and HEEC cells(P＜0.05).The expression level of LINC01410 was significantly correlated with invasion range,lymph node metastasis and TNM stage in ESCC patients(P＜0.01).LINC01410 expression was also upregulated in EC109/OE,however the expression of LINC01410 in EC9706/KD was decreased(P＜0.01).MTT assay showed overexpression of LINC01410 increased the viability of EC109 cells,while knockdown of LINC01410 decreased the viability of EC9706 cells(P＜0.01).Colony formation assay indicated that overexpression of LINC01410 enhanced the clonogenic ability of ESCC cells,while knockdown of LINC01410 reduced colony formation(P＜0.01).Transwell migration assay showed that LINC01410 overexpression drastically increased the number of migratory cells,while silencing of LINC01410 suppressed the migration in EC9706 cells(P＜0.01).GSEA revealed that Wnt/β-catenin and EMT pathways were significantly enriched in ESCC samples with a high level of LINC01410.TCF/LEF1 luciferase reporter assay showed higher levels of Wnt-dependent activities were observed in EC109/OE cells,whereas silenced LINC01410 in EC9706 cells led to contrary results(P＜0.01).Western blot analysis showed that overexpression of LINC01410 in EC109 cell significantly increased the expression levels of N-cadherin,β-catenin,cyclin D1,c-Myc and decreased E-cadherin expression,while knockdown LINC01410 resulted in opposite results.Conclusion:LINC01410 promotes proliferation and metastasis of ESCC,which might be caused by activation of Wnt/β-catenin and EMT signaling pathways.

Postzygotic mutations are acquired in normal tissues throughout an individual's lifetime and hold clues for identifying mutagenic factors.Here,we investigated postzygotic mutation spectra of healthy individuals using optimized ultra-deep exome sequencing of the time-series samples from the same volunteer as well as the samples from different individuals.In blood,sperm,and muscle cells,we resolved three common types of mutational signatures.Signatures A and B represent clock-like mutational processes,and the polymorphisms of epigenetic regulation genes influence the pro-portion of signature B in mutation profiles.Notably,signature C,characterized by C＞T transitions at GpCpN sites,tends to be a feature of diverse normal tissues.Mutations of this type are likely to occur early during embryonic development,supported by their relatively high allelic frequencies,presence in multiple tissues,and decrease in occurrence with age.Almost none of the public datasets for tumors feature this signature,except for 19.6％of samples of clear cell renal cell carcinoma with increased activation of the hypoxia-inducible factor 1(HIF-1)signaling pathway.Moreover,the accumulation of signature C in the mutation profile was accelerated in a human embryonic stem cell line with drug-induced activation of HIF-1α.Thus,embryonic hypoxia may explain this novel signature across multiple normal tissues.Our study suggests that hypoxic condition in an early stage of embryonic development is a crucial factor inducing C＞T transitions at GpCpN sites;and indi-viduals'genetic background may also influence their postzygotic mutation profiles.

OBJECTIVE：To study the inhibi tory effects of genistein on the growth of human nasopharyngeal carcinoma. CNE 1 cells and predict its potential target. METHODS ：CCK-8 method was used to test the effects of 0（blank control ），12.5，25，50， 100，150 µmol/L genistein on the proliferation of CNE 1 cells after treated for 24，48，72 h. Flow cytometry was carried out to detect the effects of 0（blank control ），15，30，60 µmol/L genistein on the cell cycle and ap optosis of CNE 1 cells after treated for 24 h. Scratch test was used to investigate the effects of 0（blank control ）， 10， 20， 30 µmol/L genistein on themigration ability of CNE 1 cells after treated for 24 h. High （No.18210156） throughput sequencing was conducted to discover the differential genes in CNE 1 cells after treated with 0（blankcontrol），30 µmol/L genistein for 24 h. RT-qPCR assay was adopted to verify the mRNA expression of related differential genes in above trials. RESULTS ： Compared with blank control，12.5，25，50，100，150 µmol/L genistein sho wed significant inhibitory effect on the proliferation of CNE 1 cells（P＜ 0.01），in a concentration- time-effect manner ；15，30 µmol/L genistein could arrest CNE 1 cell cycle at G 0/G1 stage（P＜0.05 or P＜ 0.01）；30，60 µmol/L could arrest CNE 1 cell cycle at G 2/M stage and promoted cell apoptosis （P＜0.05 or P＜0.01）. 10，20，30 µmol/L genistein could significantly inhibit the migration ability of CNE 1 cells（padj＜0.01）. High throughput sequencing revealed a total of 2 271 differentialgenes（P＜0.05），1 154 of which were up-regulated while 1 117 of which were down-regulated ；8 potential target genes ，including p53，p21，STC2，FGF2，CDK6，CYCLIN D ，PI3K，AKT，were screened by cell experiment. After validated by RT-qPCR assay ，mRNA expression of p53，p21，STC2，FGF2，CDK6，CYCLIN D and AKT were significantly down-regulated（P＜0.05），which consistent with the sequencing results. CONCLUSIONS ：Genistein can effectively inhibit the growth of human nasopharyngeal carcinoma CNE 1 cells，the mechanism of which may associated with inhibiting the expression of mutant gene p53，restoring the function of wild-type P 53 protein and inhibiting the activity of PI 3K/Akt pathway.

Objective:To provide a theoretical basis for the clinical application of the posterior route through atlas occipital articular slope screw internal fixation system through the biomechanical study.Methods:Eight cadavers of healthy adults aged 35-60 years and 155-180 cm in height were selected. The specimens with complete anatomical structure and without surgical operation were established as normal models. The model of occipito-atlantoaxial complex was established by breaking the articular capsule, ligament and other connecting structures and cutting the dentate process. The device was established as an internal fixation model through the specimen of atlantooccipital joint slope screw internal fixation system. Given normal model and internal fixation of 1.5 N·m in the moment of flexion, lateral bending and axial rotation and to measure the specimen C 0-C 1 and C 0-C 2 segment of the range (range of motion, ROM), comparative analysis of pillow neck area within the normal model and fixed model changes the range of movement, after the evaluation through the slope between atlas and the occipital screw internal fixation system of mechanical properties. Results:In the normal model, the flexion, flexion and extension, lateral bend and axial rotation ranges of C 0-C 1 segments were 23.85°±2.43°, 4.74°±0.55°, 5.77°±0.75°, respectively; the corresponding activity ranges of C 0-C 2 segments were 30.66°±3.05°, 9.09°±1.37°, 70.97°±9.48°, respectively; in the internal fixation model, the flexion and extension, lateral bend and axial rotation ranges of C 0-C 1 segments were 0.71°±0.24°, 0.24°±0.06°, 0.34°±0.09°, respectively. The corresponding activity range of C 0-C 2 segment was 3.09°±0.82°, 0.74°±0.07°, 1.22°±0.10°, respectively. Compared with the normal model, the range of activity of the internal fixation model in all directions was significantly reduced (<3°), and the reduction ratio of activity was more than 90%. Conclusion:The posterior route through pillow slope screw internal fixation system can effectively reduce the range of motion of the occipital neck in flexion, extension, lateral bending and rotation, and has safe and reliable biomechanical stability.

Objective:To strengthen the consciousness of postoperative rehabilitation exercise for focal intramuscular venous malformations and to explore the necessity of postoperative rehabilitation exercise for patients with focal intramuscular venous malformations.Methods:A retrospective analysis of patients with focal intramuscular venous malformations. From June 2017 to April 2018, 19 patients with focal intramuscular venous malformation were treated in the Hemangioma & Vascular Malformation Treatment Center of Henan Provincial People’s Hospital. All patients complained of pain and discomfort. According to the location of the lesion and the anatomical relationship between the lesion and the surrounding tissues, the operation method was selected. The rehabilitation physicians and surgeons worked out the rehabilitation plan together to carry out rehabilitation exercise treatment. The VAS scores before and after operation were compared to evaluate the effect of symptom relief and postoperative functional recovery.Results:A total of 19 patients were enrolled, including 11 females and 8 males, with an average age of 17.7 years (range, 3-44 years). The patients were followed up for 6 months to 1 year. The comparison of postoperative MRI and preoperative MRI showed that the lesions were completely removed. The VAS score of all patients was lower than that at admission. The preoperative VAS was 4.84±2.32 and the postoperative VAS was 2.16±1.17, P<0.01, indicating that the preoperative and postoperative VAS score had changed, and the VAS score had decreased. After rehabilitation exercise, all patients’ body functions were improved, joint stability, muscle strength and walking ability were restored to normal life. The preoperative joint function score was 10.47±3.29, and the postoperative joint function score was 14.46±3.50, P<0.01, indicating that the preoperative and postoperative joint function score had changed, and all the joint function scores had improved. Patients’ satisfaction was achieved. Conclusions:Postoperative rehabilitation guidance can effectively prevent postoperative limb dysfunction and accelerate the recovery of body function.

Objective:To investigate more safe, effective and standard perioperative managements of infant patients with Kasabach-Merritt phenomenon (KMP).Methods:We made a retrospective analysis on the clinical data of KMP infant patients, who received surgical intervention in our department between January 2017 and September 2019. Inclusion criteria : (1) diagnosed as KMP that characterized by a large hemangioma (located in trunk or limb), profound thrombocytopenia and consumptive coagulopathy; (2) received surgical treatment in our center during January 2017 and September 2019; (3) age ≤1 year. Before surgical treatment, all the patients were given glucocorticoid and continued to the operation day in the sensitive group. The insensitive group received single large dose of platelet (PLT) transfusion 1 day before surgery, for the purpose of correcting thrombocytopenia and coagulopathy. Endotracheal intubation and intravenous anesthesia, combined with deep vein catheterization, arterial puncture catheterization and continuous invasive blood pressure monitoring were used to maintain hemodynamic stability. Radical resection of the tumor, combined with flap plasty or in situ skin grafting was carried out when necessary; after the operation, the endotracheal tube was routinely taken to ICU, and the endotracheal tube was removed as appropriate after the recovery of respiratory and circulation. The patient was kept overnight in ICU, and patient was transferred out after evaluation of stability. The dynamic changes of platelet were monitored and nutritional support was strengthened. Patients with lesions in limbs (except those with in situ skin grafting) were given passive rehabilitation training on the third day after surgery. The patients were followed up for 6-36 months. Routine blood examination, coagulation function, color Doppler ultrasonography and MRI were performed when necessary. The range of motion and muscle strength of adjacent joints were examined during the follow-up visit. Results:A total of 55 infant patients with KMP were included in this study. Peripheral blood test at 1 h before surgery showed platelets > 100×10 9/L in 54 cases and > 80×10 9/L in 1 case, and hemoglobin was corrected to more than 10 g/L. The operation time was 48-135 min, with an average of 87 min. There was no intraoperative or postoperative death. It took 4 to 36 hours for platelet to return to normal level, with an average of 8.4 hours. All surgical specimens were found to be KMP. The hospital stay was 9-30 d, with an average period of 16.7 d. Delayed incision healing in 3 cases, scar contracture in 1 case, scar hyperplasia in 3 cases. There was no death during the follow-up period, and the platelet was stable in the normal range. Conclusions:Surgical treatment of vascular tumors complicated with KMP has definite curative effect, rapid effect, short course of treatment and low cost. A series of perioperative treatments, including active preoperative preparation, effective coagulation function correction measures, perfect anesthesia and monitoring methods, stable hemodynamic support, fine surgical operation and early postoperative rehabilitation exercise are the necessary guarantee for the success of surgical treatment.

Objective:To analyze the etiological characteristics and molecular epidemiological correlation of five cases of typhoid fever during the same period in yantai city.Methods:Six S. Typhis strains were isolated from 5 typhoid patients and epidemiological samples in Yantai city in 2018. The onset time of the cases were from May 26, 2018 to July 24, 2018, distributed in Shuidao Town of Muping District, Dengzhou Street of Penglai District, Donglai Street of Longkou District, Wenhua Street of Muping District and Fulaishan Street of zhifu District. S. Typhis strains were analyzed by conventional bacterial isolation method and XbaⅠ/BlnⅠ double-enzyme digestion pulse-field gel electrophoresis (PFGE). Meanwhile, ViaB virulence gene detection and 27 common antibiotics sensitivity tests were conducted to study the etiology of S. Typhis.Results:Six strains of S. Typhi were isolated from 5 patients and the domestic egg of one patient, which were divided into 4 PFGE patterns by PFGE-XbaⅠ and PFGE-BlnⅠ and among which 3 strains had the same PFGE patterns.One multi-drug resistant strain (foreign patient), one single-drug resistant strain (patient with a history of provincial retention), and one completely sensitive strain were detected. The three strains of the same PFGE pattern exhibit the same drug-sensitive phenotype which were intermediate against aminoglycosides and quinolones and susceptibility against the other antibiotics.All of the strains carried the ViaB virulence factor except the strain from the foreign patient.Conclusion:Local S. Typhi is susceptibility or intermediate against antibiotics commonly used in clinic.Sporadic cases of typhoid fever and typhoid imported infections still need attention.

Objective:To analyze the etiological characteristics and molecular epidemiological correlation of five cases of typhoid fever during the same period in yantai city.Methods:Six S. Typhis strains were isolated from 5 typhoid patients and epidemiological samples in Yantai city in 2018. The onset time of the cases were from May 26, 2018 to July 24, 2018, distributed in Shuidao Town of Muping District, Dengzhou Street of Penglai District, Donglai Street of Longkou District, Wenhua Street of Muping District and Fulaishan Street of zhifu District. S. Typhis strains were analyzed by conventional bacterial isolation method and XbaⅠ/BlnⅠ double-enzyme digestion pulse-field gel electrophoresis (PFGE). Meanwhile, ViaB virulence gene detection and 27 common antibiotics sensitivity tests were conducted to study the etiology of S. Typhis.Results:Six strains of S. Typhi were isolated from 5 patients and the domestic egg of one patient, which were divided into 4 PFGE patterns by PFGE-XbaⅠ and PFGE-BlnⅠ and among which 3 strains had the same PFGE patterns.One multi-drug resistant strain (foreign patient), one single-drug resistant strain (patient with a history of provincial retention), and one completely sensitive strain were detected. The three strains of the same PFGE pattern exhibit the same drug-sensitive phenotype which were intermediate against aminoglycosides and quinolones and susceptibility against the other antibiotics.All of the strains carried the ViaB virulence factor except the strain from the foreign patient.Conclusion:Local S. Typhi is susceptibility or intermediate against antibiotics commonly used in clinic.Sporadic cases of typhoid fever and typhoid imported infections still need attention.

Objective:To strengthen the consciousness of postoperative rehabilitation exercise for focal intramuscular venous malformations and to explore the necessity of postoperative rehabilitation exercise for patients with focal intramuscular venous malformations.Methods:A retrospective analysis of patients with focal intramuscular venous malformations. From June 2017 to April 2018, 19 patients with focal intramuscular venous malformation were treated in the Hemangioma & Vascular Malformation Treatment Center of Henan Provincial People’s Hospital. All patients complained of pain and discomfort. According to the location of the lesion and the anatomical relationship between the lesion and the surrounding tissues, the operation method was selected. The rehabilitation physicians and surgeons worked out the rehabilitation plan together to carry out rehabilitation exercise treatment. The VAS scores before and after operation were compared to evaluate the effect of symptom relief and postoperative functional recovery.Results:A total of 19 patients were enrolled, including 11 females and 8 males, with an average age of 17.7 years (range, 3-44 years). The patients were followed up for 6 months to 1 year. The comparison of postoperative MRI and preoperative MRI showed that the lesions were completely removed. The VAS score of all patients was lower than that at admission. The preoperative VAS was 4.84±2.32 and the postoperative VAS was 2.16±1.17, P<0.01, indicating that the preoperative and postoperative VAS score had changed, and the VAS score had decreased. After rehabilitation exercise, all patients’ body functions were improved, joint stability, muscle strength and walking ability were restored to normal life. The preoperative joint function score was 10.47±3.29, and the postoperative joint function score was 14.46±3.50, P<0.01, indicating that the preoperative and postoperative joint function score had changed, and all the joint function scores had improved. Patients’ satisfaction was achieved. Conclusions:Postoperative rehabilitation guidance can effectively prevent postoperative limb dysfunction and accelerate the recovery of body function.