目的：探讨苍术素（ATR）通过调节受体相互作用蛋白激酶（RIPK）1/RIPK3/混合谱系激酶结构域样（MLKL）信号通路对非小细胞肺癌（NSCLC）A549细胞程序性死亡及裸鼠移植瘤生长的影响。方法：使用0~160 μmol/L的ATR处理A549细胞，MTT法检测细胞存活率以确定后续实验给药浓度。使用ATR和/或RIPK1抑制剂Nec-1（necrostatin-1）、caspase抑制剂Z-VAD-FMK处理A549细胞，验证ATR是否诱导A549细胞发生程序性坏死。将A549细胞分为对照组、ATR-L组、ATR-M组、ATR-H组（分别用0、10、20、40 μmol/L ATR处理）、ATR+Nec-1组（40 μmol/L ATR+50 μmol/L Nec-1处理），处理24 h后，采用PI单染及Hoechst33342/PI双染法检测细胞死亡情况、透射电镜观察细胞死亡形态、DCFH-DA荧光探针法检测细胞内ROS水平、JC-1染色法检测线粒体膜电位、WB法检测细胞中RIPK1/RIPK3/MLKL信号通路相关蛋白质的表达水平。构建A549细胞裸鼠移植瘤模型，用10 mg/kg ATR（溶于玉米油中）对裸鼠灌胃给药5周，观察ATR对移植瘤生长的影响，WB法检测移植瘤组织中RIPK1/RIPK3/MLKL信号通路相关蛋白质的表达水平。结果：10~160 μmol/L的ATR可显著抑制A549细胞增殖，选择10、20、40 μmol/L的ATR进行后续实验。ATR组A549细胞存活率显著低于对照组（P<0.01）和ATR+Nec-1组（P<0.01），而ATR+z-VAD组细胞存活率显著低于z-VAD组（P<0.01），说明ATR可诱导A549细胞发生程序性坏死而非凋亡。与对照组比较，ATR处理组A549细胞发生肿胀，线粒体内脊消失呈空泡化，细胞内容物向外泄漏，细胞核聚集，表现为坏死特征，ATR-L组、ATR-M组、ATR-H组A549细胞死亡率、ROS水平及p-RIPK1、p-RIPK3、p-MLKL表达水平均显著升高，线粒体膜电位显著降低（均P<0.01），且呈药物浓度依赖性；与ATR-H组比较，ATR+Nec-1组细胞死亡率、ROS及p-RIPK1、p-RIPK3、p-MLKL表达水平降低，线粒体膜电位显著升高（均P<0.01）。裸鼠移植瘤实验结果显示，与对照组比较，ATR组裸鼠移植瘤体积、移植瘤质量均降低（P<0.05，或P<0.01），而与瘤组织中p-RIPK1、p-RIPK3、p-MLKL蛋白表达水平均显著升高（均P<0.01）。结论：ATR可能通过激活RIPK1/RIPK3/MLKL信号通路诱导A549细胞发生程序性坏死，抑制A549细胞及其裸鼠移植瘤的生长。

Objective: To explore the pathogenic genes in a Chinese family affected by nonsyndromic tooth agenesis so as to study the pathogenesis of oligodontia. Methods : Hospital ethical approval and informed consent of the patients and family members were obtained. Clinical data of the proband and close family members were collected, peripheral venous blood was collected, and DNA was extracted. Gene sequencing was performed through whole-exome sequencing, and then the screened pathogenic genes were verified by Sanger sequencing. The three-dimensional structure of the mutant proteins was analyzed and compared with the wild-type using bioinformatics tools. Results: The two patients with congenital majority tooth loss in this family were cousins, and there were no other patients with congenital majority tooth loss in the family. Besides congenital multiple tooth loss, the two patients had no obvious hair abnormalities, finger/toe abnormalities, sweating abnormalities or other abnormal manifestations of ectodermal tissue. We found a mutant gene that in this family by carrying out gene sequencing of the patients and their close family members. A novel EDA (ectodysplasin A) missense mutation c.983C>T (p. Pro328Leu) was identified, which changed the encoded amino acid from proline (Pro) to leucine (Leu). Analysis of the mutation site showed that the site was highly conserved, and three-dimensional structure modeling also found that it changed the structure of EDA. Conclusion A novel EDA missense variant (c.983C>T, p.Pro328Leu) was first identified in a Chinese family with nonsyndromic tooth agenesis, extending the mutation spectrum of the EDA gene.

The number of investigator initiated research (IIR) is increasing. But the recognition and management of IIR in China is still in its infancy, and there is a lack of specific and operable guidance for the implementation process. Based on our practical experiences, previous literature reports, and current policy regulations, the authors took prospective IIR as an example to summarize the implementation process of IIR into 14 steps, which are as the following: study initiation, ethical review, study registration, study filing, case report form design, database establishment, standard operating procedure making, investigator training, informed consent, data collection, data entry, data verification, data locking and data archiving.

Abstract OBJECTIVE Neuroblastoma(NB) is a prevailing pediatric extracranial solid tumor that accounts for 10%-15% of all childhood cancer-related fatalities. Despite significant strides made in NB therapy through multimodal approaches, the survival rate of high-risk NB patients remains at approximately 50%. Consequently, there is an urgent need to identify novel molecular targets for NB treatment. Recent studies have shown that MYCN oncogene amplification is present in about 25% of NB cases and is a crucial determinant of poor prognosis for high-risk NB patients. Since MYC family proteins, including MYCN, are inherently disordered proteins, MYCN lacks a defined ligand binding site along with a large protein-protein interaction surface. Current treatment approaches for MYCN-amplified NB patients do not include direct targeting of MYCN itself, since the absence of a “drugable” pocket renders it challenging. Notably, no direct MYC-targeting drugs are currently available. There is an existing association between Aurora A kinase(AURKA) and MYCN, whereby they form a complex to fortify MYCN stability. However, MYCN is inherently unstable, with a half-life of only 30 min, but AURKA intervenes by facilitating its stability through a direct protein-protein interaction, hence protecting it from proteasomal degradation. This interaction potentially augments tumor cell proliferation and invasiveness. Notably, AURKA has been verified as a transcriptional target of MYCN. The present study endeavors to employ computer-aided drug design technology to probe AURKA inhibitors discerned from a natural product library of traditional Chinese medicine(TCM), thereby identifying a novel drug for treating NB. METHODS Collected from the YaTCM database, a total of 47 696 natural compounds from TCM were subjected to preprocessing including protonation, deionization, hydrogenation, stereoisomerism, conformation generation, and energy minimization. Of these, 58 048 compounds were initially screened as potential ligands for the library. Utilizing “Lipinski Ro5” and “Verber Ro3” guidelines, 22 227 hit compounds were selected from the library that met the screening criteria. Initially, crystal structures of AURKA and its inhibitor AA35 were downloaded from the RCSB PDB database. The spatial coordinates of AA35 were set as the center of the binding pocket for AURKA, and a 10 Å * 10 Å * 10 Å space around the pocket was designated as the active space. A comprehensive drug screening platform integrating lead-likeness filtering, pharmacokinetic prediction, molecular docking, flexible docking, and molecular dynamics(MD) simulations were established to excavate potential aurora kinase A inhibitors from the TCM compound library, which were further validated by MD simulations. RESULTS A grand total of 6 220 Chinese herbal remedies had been meticulously curated within the YaTCM database. Out of these, an impressive 47 696 Chinese herbal monomers had undergone a rigorous series of flexible docking tests, resulting in the selection of the top ten molecules with the most favorable docking scores. The aforementioned molecules underwent AMDE parameter and toxicity predictions. It was discovered that with the exception of a few compounds such as Tryptophane, 3&#39;-Methoxydaidzein, and Burttinol D(which might elicit liver toxicity), 3&#39;-Methoxydaidzein and Pratensein(which might elicit kidney toxicity), and 3-Deoxysappanone B(which had moderate oral toxicity), as well as Tryptophane(with an oral bioavailability of less than 50%), five compounds including Compound X, (+)-Sesamin dicatechol, Tuberosin, Abrine, and Maackiain, displayed favorable pharmacokinetic parameters and low toxicity predictions. Moreover, all of these compounds exhibited a high binding affinity with the inhibitor active pocket of AURKA. In this study, Compound X, despite its cumbersome name, was referred to as “Compound X”. Upon focusing on Compound X as the subject of investigation, it was discovered that its phenolic framework could readily interact with the hydrophobic cavity constituted of hydrophobic amino acids, namely TYR199, VAL182, LEU178, LEU208, and VAL206. Notably, Compound X could partake in Pi-Pi interactions with TYR199 and create hydrogen bonds with HIS201, GLU175, and LYS166. Computational studies via MD simulations confirmed that Compound X could form a stable receptor-ligand complex with the receptor. Impressively, the inclusion of Compound X significantly reduced the stability of the AURKA-MYCN complex. CONCLUSION This study concludes that Compound X can be used as an AURKA inhibitor for the treatment of NB, which is a novel finding based on the combination of various virtual screening techniques from the natural product database of TCM.

OBJECTIVE To establish characteristic HPLC chromatograms for Jubei mixture, and to simultaneous determine its six components by HPLC-ELSD. METHODS An HPLC specific chromatogram of the Jubei mixture was established. The chromatographic column was Agilent ZORBAX-C18(250 mm×4.6 mm, 5 μm), gradient elution was conducted with acetonitrile(A)-0.1% phosphoric acid aqueous solution(B) as the mobile phase, the flow rate was 1.0 mL·min-1, and the column temperature was 30℃, the detection wavelength was 250 nm. An HPLC-ELSD method was established and the chromatography was accomplished on a Waters T3 C18(250 mm×4.6 mm, 5 μm) column, the mobile phase consisted of acetonitrile(A)-1% acetic acid aqueous solution(B) in a gradient mode, the flow rate was 1.0 mL·min-1, the column temperature was set at 30℃. RESULTS The characteristic chromatograms of the HPLC with baicalin as the reference peak was established, and eight common peaks were defined. The linear ranges of amygdalin, liquiritin, peimine, peiminine, platycodin D and ophiopogonin D were good within their own ranges(r ≥ 0.999). The average recovery were 99.1%, 98.6%, 100.9%, 97.7%, 98.9% and 98.0%, RSD were 0.4%, 1.0%, 1.1%, 1.2%, 0.7% and 1.3%, respectively. CONCLUSION The established methods can be used to qualitative identification and quantitative determination of Jubei mixture, can effectively used for evaluation and quality control of Jubei mixture.

Objective To analyze the feasibility of six-minute walk test (6MWT) before pulmonary lobectomy and prediction for postoperative outcome. Methods A total of 580 patients who were hospitalized in the department of lung surgery from May, 2017 to May, 2019 were reviewed, and 274 eligible patients were selected, who underwent first surgery and the surgical method was pulmonary lobectomy. They were divided into two groups based on the results of 6MWT before operation. The cut-off value of six-minute walk distance (6MWD) was obtained by receiver operating characteristic curve (ROC) area under curve (AUC). The postoperative outcome and the occurrence of cardiopulmonary complications in the two groups were analyzed. Results Compared to patients with 6MWD > 449 meters, the age was significantly older (P < 0.001), the forced expiratory volume in the first second (FEV1) was poor in patients with 6MWD ≤ 449 meters (P < 0.05), and other factors such as surgical resection site, pathological stage, gender, etc., were not significantly different (P > 0.05). The incidence of postoperative cardiopulmonary complications was significantly higher (OR = 2.672, 95%CI 1.488 to 4.798, P = 0.002), and the postoperative extubation time and hospital stay was longer in patients with 6MWD ≤ 449 meters than in patients with 6MWD > 449 meters (P < 0.05). 6MWD ≤ 449 meters was an independent risk factor for postoperative cardiopulmonary complications (OR = 2.395, 95%CI 1.299 to 4.415, P = 0.005). Conclusion As a simple function test, 6MWT can be routinely used to assess the physiological function of patients undergoing pulmonary lobectomy. Patients with 6MWD ≤ 449 meters may be in higher risks of postoperative cardiopulmonary complications.

Objective    To summarize the experience of surgical treatment of asphyxiating thoracic dysplasia (Jeune syndrome). Methods    A total of 15 patients with asphyxiating thoracic dysplasia from August 2018 to April 2020 in our hospital were retrospectively included. There were 7 males and 8 females, aged 1-25 (8.87±6.71) years. Special steel bars were used to correct the growth direction of the rib and costal cartilage. Meanwhile, the concave and convex deformities of the chest wall on both sides were corrected to increase the chest volume and correct the thoracic deformity. Results    The contour appearance of the chest wall of all patients changed after the operation. The shape was close to normal, and the symptoms of hypoxia were improved. The operation time was 147.73±59.78 min, intraoperative bleeding volume was 105.67±91.90 mL, ICU stay time was 14.20±13.54 d and hospital stay time was 26.00±17.87 d. Eleven patients were directly extubated after the operation, 4 patients underwent tracheotomy and received assisted respiration, and the assisted respiration time was 19, 13, 22 and 12 days, respectively. The postoperative chest circumference was significantly increased, and the blood oxygen saturation was significantly improved. There were 5 patients with cardiac insufficiency, and 3 of them were improved by cardiotonic therapy, 2 of them died of heart failure on the 2nd and 31st day after the operation, respectively. Abdominal distention occurred in 10 patients after operation, and 5 of them were obstinate and eliminated by comprehensive treatment. All patients were followed up. The appearance of thorax was improved obviously and there was no sign of compression in lungs. One 13-year-old patient developed respiratory discomfort 3 months after the operation, and the symptoms were relieved after self-administration of oxygen. A 25-year-old patient developed cardiac insufficiency half a month after the discharge, and the symptoms disappeared after cardiotonic treatment. Four patients took out the steel bars in 13, 13, 15 and 17 months after the operation, respectively. The appearance of thorax remained well after the operation. The imaging examination showed that the position of bone structure was normal, the lung field was clear, and there was no sign of chronic inflammation. Conclusion    This technique is a safe and simple operation method. It can not only eliminate the deformity of chest wall, but also increase the volume of chest obviously. However, the long-term effect needs to be further evaluated.

How to effectively repair esophageal fistulas, caused by esophageal perforation, rupture and anastomotic leakage after esophagectomy has always been a key problem for the digestive surgeon. Although there are many clinical treatment methods, the therapeutic effect is still completely unsatisfactory, especially when severe mediastinal purulent cavity infection is associated with the esophageal fistula. In recent years, foreign centers have promoted a new minimally invasive endoscopic treatment technology to repair the esophageal fistula, endoscopic vacuum-assisted closure therapy, with significantly curative effect. In this article, we will review the specific operation, advantages and disadvantages, as well as the clinical efficacy of endoscopic vacuum-assisted closure therapy in treating the esophageal fistulas, to provide a new therapeutic technique for esophageal fistulas and expand the new field of minimally invasive endoscopic therapy.

Cognitive impairment is the most common non-motor symptom of Parkinson&#39;s disease (PD), which may associate with the disorders of transmitters, cortical atrophy, proteinosis, etc., and seriously influence the motor function, ability of daily life of the patients. Transcranial direct current stimulation (tDCS), which may adjust the cerebral excitability, regional blood flow, plasticity of synapses, etc., has been used for PD cognitive impairment.