Effect of azacitidine on HOXA9 gene expression and biological behaviors of A375 melanoma cells / 中华皮肤科杂志

Objective:To investigate the effect of the methyltransferase inhibitor azacitidine (5-azaC) on the expression of homeobox A9 (HOXA9) gene in, as well as proliferation, invasion and migration of A375 cells.Methods:In vitro cultured A375 cells were treated with 5-azaC at various concentrations of 1, 5, 10 and 20 μmol/L, while routinely cultured A375 cells receiving no drug intervention served as control group. Methylation-specific PCR was performed to analyze methylation status of the HOXA9 gene promoter region after the treatment with different concentrations of 5-azaC, in order to screen the optimal concentration of 5-azaC for following experiments. Cell counting kit-8 (CCK8) assay was conducted to evaluate the proliferation of A375 cells, Transwell and wound healing assays were performed to estimate the invasion and migration of A375 cells, and real-time fluorescence-based quantitative PCR (qRT-PCR) and Western blot analysis were conducted to determine the mRNA and protein expression of HOXA9 in A375 cells after 5-azaC treatment. Two-independent-sample t test was used for comparisons between two groups. Results:Methylation was observed in the HOXA9 gene promoter region in A375 cells in the control group. After 5-azaC treatment, methylated and unmethylated states coexisted in the HOXA9 gene promoter region in A375 cells, and the higher the concentration of 5-azaC, the higher the degree of demethylation of the HOXA9 gene. Therefore, 20 μmol/L 5-azaC was selected to treat A375 cells for 72 hours, which served as 5-azaC treatment group in subsequent experiments. Compared with the control group, the 5-azaC treatment group showed significantly decreased cellular proliferative ability (72.46% ± 2.19% vs. 100%, t = 28.09, P < 0.001) , significantly decreased number of invasive cells (242.70 ± 29.19 vs. 466.00 ± 22.65, t = 10.47, P < 0.001) , significantly decreased migratory ability (27.56% ± 2.74% vs. 35.69% ± 2.50%, t = 3.79, P = 0.019) , significantly increased HOXA9 mRNA expression (1.73 ± 0.28 vs. 1.01 ± 0.15, t = 3.93, P = 0.017) , and significantly increased HOXA9 protein expression (0.62 ± 0.03 vs. 0.50 ± 0.01, t = 3.82, P = 0.019) . Conclusion:5-azaC can inhibit the proliferative, invasive and migratory ability of A375 melanoma cells, and one of the possible mechanisms underlying this process may be that 5-azaC reverses the methylation in the HOXA9 gene promoter region, activates HOXA9 gene expression, and participates in the regulation of biological behaviors of melanoma cells.

Role of BDNF/TrkB signaling pathway in pre-injection of young rat plasma-induced reduction of sevoflurane-caused cognitive dysfunction in aged rats / 中华麻醉学杂志

Objective:To evaluate the role of brain-derived neurotrophic factor (BDNF)/tropomyosin-related kinase B (TrkB) signaling pathway in pre-injection of young rat plasma-induced reduction of sevoflurane-caused cognitive dysfunction in aged rats.Methods:Eighty SPF healthy male Sprague-Dawley rats, aged 18 months, weighing 550-650 g, were divided into 4 groups ( n=20 each) using a random number table method: control group (group C), sevoflurane anesthesia group (group S), young rat plasma group (group Y) and BDNF/TrkB signaling pathway inhibitor K252a group (group K). The plasma 100 μl obtained from 3-month-old young rats was injected via the tail vein in group Y and group K, while the equal volume of normal saline was given via the tail vein in group C and group S, twice a week, for 4 weeks.In S, Y and K groups, 3% sevoflurane was inhaled for 3 h starting from the end of treatment, and BDNF/TrkB signaling pathway inhibitor K252a was injected via the tail vein before anesthesia in group K. The open field test and Morris water maze test were performed at 3 days after anesthesia to assess the spontaneous motor ability and cognitive function.Then the rats were sacrificed, and the hippocampal tissues were isolated for determination of the expression of BDNF, phosphorylated TrkB (p-TrkB), postsynaptic dense protein-95 (PSD-95) and synaptic vesicle protein (SYN) (by Western blot), dendritic length and dendritic ridge density of neurons in hippocampal CA1 area (by Golgi staining), and the number of synapses and length of synaptic active area (with a transmission electron microscope). Results:Compared with group C, the escape latency was significantly prolonged, the number of crossing the original platform was reduced, the expression of p-TrkB, BDNF, PSD-95 and SYN was down-regulated, and the dendritic length, dendritic ridge density, the number of synapses and length of synaptic active area were decreased in group S ( P<0.05). Compared with group S, the escape latency was significantly shortened, the number of crossing the original platform was increased, the expression of p-TrkB, BDNF, PSD-95 and SYN was up-regulated, and the dendritic length, dendritic ridge density, the number of synapses and length of synaptic active area were increased in group Y ( P<0.05). Compared with group Y, the escape latency was significantly prolonged, the number of crossing the original platform was reduced, the expression of p-TrkB, BDNF, PSD-95 and SYN was down-regulated, and the dendritic length, dendritic ridge density, the number of synapses and length of synaptic active area were decreased in group K ( P<0.05). Conclusions:The mechanism by which pre-injection of young rat plasma reduces sevoflurane-induced cognitive dysfunction is related to activation of BDNF/TrkB signaling pathway and improvement in synaptic plasticity in the hippocampus of aged rats.

Transoral endoscopic resection of benign tumors in parapharyngeal space via medial pterygomandibular raphe approach / 中华耳鼻咽喉头颈外科杂志

Objective: To explore the surgical methods and effects of transoral endoscopic resection of benign tumors in parapharyngeal space via medial pterygomandibular raphe approach. Methods: The clinical data of 23 patients who underwent resection of benign tumors in parapharyngeal space by endoscopic medial pterygomandibular raphe approach from January 2016 to July 2020 in the Department of Otorhinolaryngology Head and Neck Surgery, the Affiliated Hospital of Qingdao University were analyzed retrospectively. There were 14 males and 9 females with a median age of 43 years. The tumors were located in the anterior space of the styloid process in 13 cases and in the posterior space in 10 cases. The smallest tumor volume was 7.3 ml and the largest was 80.2 ml. The preoperative imaging features, the characteristics and risks of this approach in the operation were analyzed, and the feasible mode of operation was explored. Results: All patients completed the operation successfully. The intraoperative blood loss was 20 to 50 ml, with an average of 28.3 ml. The operation time was 40 to 110 min, with an average of 75.4 min. The incision length was 2 to 4 cm, with an average of 3.0 cm. The postoperative pain score was 2 to 4, with an average of 3.2. The postoperative hospital stay was 4 to 9 d, with an average of 6.7 d. Postoperative pathological diagnosis included pleomorphic adenoma (n=12), neurilemmoma (n=10) and basal cell adenoma (n=1). The patients were followed up for 6 to 60 months. There was no postoperative complication such as infection or serious bleeding, and there was no tumor recurrence after operation. Conclusion: Endoscopic resection of benign tumor in parapharyngeal space via medial pterygomandibular raphe approach is a safe, effective, and minimally invasive surgical method for the treatment of tumors in parapharyngeal space.

Study of clinical characteristics in patients with gp210 antibody-positive primary biliary cholangitis / 中华肝脏病杂志

Objective: To analyze the clinical characteristics and prognostic value of liver function in a large samples of patients with anti-glycoprotein 210 (gp210 antibody) positive primary biliary cholangitis (PBC). Methods: A retrospective study was performed on 931 PBC cases in Beijing You'an Hospital affiliated to Capital Medical University from 2010 to 2019. According to the detection of gp210 antibody, 318 cases were divided into gp210 antibody positive group (positive group) and 613 cases were divided into gp210 antibody negative group (negative group). The differences in demographic, medical history, clinical indicators, B-ultrasound and pathological indicators as well as the histopathological basis were compared between the two groups. SPSS 16.0 software was used for statistical analysis. Measurement data were analyzed by t-test or rank sum test, and enumeration data by χ2 test. Multivariate analysis was used for logistic test, and and survival analysis was used for prognosis. Results: The positive and the negative groups were compared. The ratio of male to female was significantly higher in positive than negative group (1:5.35 vs. 1:9.73, P＜0.05), and the difference was statistically significant. The proportion of hormone use in history of past diagnosed and treated was higher in positive than negative group (12.9% vs. 3.47%, P＜0.05), and the difference was statistically significant. The detection of biochemical indexes such as alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBIL), alkaline phosphatase (ALP), glutamyl transpeptidase (GGT) were higher in positive than the negative group (51.1 U/L vs. 41.1 U/L, 62.6 U/L vs. 49.6 U/L, 24.1 μmol/L vs. 17.9 μmol/L, 228.3 U/L vs. 169.6 U/L, 203.9 U/L vs. 147.6 U/L), (P＜0.05), and the differences were statistically significant. Antinuclear antibody (ANA)-positive rate, high titer ratio and immunoglobulin G (IgG) levels were higher in positive than negative group (95.2% vs. 81.6%, 69.7% vs. 48.8%, 17.2 g/L vs. 16.2 g/L), (P＜0.05), and the differences were statistically significant. The incidence of liver failure was higher in positive than negative group (P＜0.05). CK7 and inflammation score were higher in positive group than negative group in liver histopathological observations (0.83±0.53 vs. 0.28±0.47; 1.06±0.39 vs. 0.54±0.65), (P＜0.05), and the differences were statistically significant. Conclusion: The illness condition of patients with gp210 antibody positive PBC is more severe than patients with gp210 antibody negative PBC, and the incidence of liver failure is significantly increased. Cholangiocytes may be the histopathological basis of the clinical characteristics of gp210 antibody positive PBC patients.

Association between clinicopathological characteristics and gene mutations in 94 cases of cutaneous melanoma / 中华皮肤科杂志

Objective:To investigate associations between clinicopathological characteristics and mutations in susceptibility genes in cutaneous melanoma (CMM) .Methods:A total of 94 patients with confirmed CMM were collected from People′s Hospital of Xinjiang Uygur Autonomous Region from January to December in 2019, and their clinical and histopathological characteristics were retrospectively analyzed. In 48 paraffin-embedded melanoma tissue specimens, Sanger sequencing was performed to detect mutations in the BRAF, NRAS, c-KIT genes and the promoter region of human telomerase reverse transcriptase (hTERT) gene, and the association between gene mutations and clinicopathological characteristics was analyzed. Measurement data were compared using t test, and enumeration data were compared using chi-square test or Fisher′s exact test. Results:Among the 94 patients with CMM, there were 46 (48.9%) males and 48 (51.1%) females, with the age being 58.5 ± 16.0 years; 41 (43.6%) patients were of Han nationality, and 53 (56.4%) were of ethnic minorities. Skin lesions were located at the acral sites in 50 (53.2%) patients, including 27 (28.7%) of Han nationality; non-acral skin lesions occurred in 44 (46.8%) , including 14 (31.8%) of Han nationality; there was a significant difference in the nationality distribution between the acral CMM group and non-acral CMM group ( χ2 = 5.25, P = 0.022) . Histopathological examination showed CMM of Clark grades Ⅳ or Ⅴ in 41 (43.6%) cases, ulcers in 52 (55.3%) cases, and lymph node metastasis in 32 (34.04%) cases at the first clinic visit. Gene sequencing revealed BRAF gene mutations in 11 (22.9%) of 48 cases, including c.1799 T>A (p.V600E) , c.1790 T>A (p.L597Q) and c.1394 C>T (p.S465F) ; NRAS gene mutation c.182 A>G (p.Q61R) was identified in 5 (10.4%) cases; c-KIT gene mutations were identified in 6 (12.5%) cases, including c.1727 T>C (p.L576P) and c.1669 T>C (p.W557R) ; mutations in the promoter region of hTERT gene were identified in 7 (14.6%) cases, including 4 cases with a mutation at 124 bp upstream of the ATG start codon (C228T) and 3 cases with a mutation at 146 bp upstream of the ATG start codon (C250T) . Among 26 patients aged < 60 years, BRAF gene mutations were found in 9, and the incidence of BRAF gene mutations was significantly higher in the patients aged < 60 years than in those aged ≥ 60 years (2/22, P < 0.05) , but significantly lower in the patients with acral CMM (3/27) than in those with non-acral CMM (8/21, P < 0.05) ; the incidences of the NRAS, c-KIT and hTERT gene mutations were all significantly higher in the patients with lymph node metastases (3/10, 4/10, 4/10, respectively) than in those without (2/38, 2/38, 3/38, respectively, all P < 0.05) . Conclusion:CMM lesion locations significantly differed among different ethnic groups; the BRAF gene mutation was associated with the age of patients and lesion locations of CMM; NRAS, c-KIT gene mutations and hTERT promoter mutations were closely related to lymph node metastasis.

Chemical genetics technology regulates the activity of GLP-1 neurons and its effect on appetite / 中华内分泌代谢杂志

Objective:To conduct a glucagon like peptide-1(GLP-1)controllability model rat by chemical genetics, and observe the impact of GLP-1 neuron excitability on appetite.Methods:Fifteen rats were evenly divided into Green fluorescent protein(GFP)group, HM3D group, and HM4D group. Various combinations of adeno-associated virus(rAAV)were injected into the nucleus tractus solitarius(NTS). rAAV-GLP-1-cre and rAAV-GFP-dio were administered in rats of GFP group. The rats of HM3D group were injected with rAAV-GLP-1-cre and rAAV-HM3D-mCherry-dio while rAAV-GLP-1-cre and rAAV-HM4D-mCherry-dio were injected in rats of HM4D group . The optimal dose of clozapine N-oxide(CNO)was selected based on feeding behavior and body weight changes of rats after intraperitoneal injection of different doses of CNO. The controllability of GLP-1 neurons was confirmed by comparing with intraperitoneal injection of saline. The number of activated GLP-1 neurons in the NTS area and the expression of POMC neurons in the hypothalamus were detected 30 minutes after CNO injection.Results:GLP-1 neurons in the NTS area of rats were successfully labeled. The rat of HM3D group revealed a decrease in food intake( P=0.021)while the rat of HM4D group showed an increase( P=0.002), when given 1 mg/kg of CNO, no changes at the dose of 0.5 mg/kg and 3.0 mg/kg. Immunofluorescence showed that the activity of GLP-1 neurons in NTS of GFP group was lower than that of HM3D group( P=0.022), and higher compared with that of the HM4D group( P=0.049). The expression of GLP-1 neurons in NTS and POMC neurons in the hypothalamus of the HM3D group after intraperitoneal injection of CNO was also higher than that in the HM4D group( P=0.003). Conclusion:Using chemical genetics technology, GLP-1 controllability model rat could be successfully established via injecting varying combinations of rAAV into the NTS area of rat. Injection of 1 mg/kg CNO can effectively activate or inhibit the neuron to regulate appetite.

Role of RhoA/ROCK2 signaling pathway in multiple exposures to sevoflurane-induced long-term cognitive impairment in neonatal rats / 中华麻醉学杂志

Objective:To evaluate the role of RhoA/ROCK2 signaling pathway in multiple exposures to sevoflurane-induced long-term cognitive impairment in neonatal rats.Methods:Sixty SPF healthy neonatal Sprague-Dawley rats of either sex, aged 6 days, weighing 12-20 g, were divided into 3 groups ( n=20 each) using a random number table method: control group (group C), multiple exposures to sevoflurane group (group S) and RhoA/ROCK2 signaling pathway inhibitor Y-27632 group (group Y). Group S and group Y inhaled 3% sevoflurane for 2 h at days 6, 7 and 8 after birth.In group Y, Y-27632 5 mg/kg was intraperitoneally injected before sevoflurane anesthesia.The spontaneous activity was evaluated by open field test on day 35 after birth.The cognitive function was detected by Morris water maze test at day 36 after birth.The rats were sacrificed after Morris water maze test, and the hippocampal tissues were isolated for determination of the apoptosis rate of hippocampal neurons and cytoplasmic calcium concentration ([Ca 2+ ] i) (by flow cytometry) and expression of phosphorylated RhoA (p-RhoA), ROCK2 and cleaved-caspase-3 (by Western blot) and for microscopic examination of the ultrastructure of hippocampal neurons (with a transmission electron microscope). Results:There was no significant difference in movement speed, distance and time of stay in the open field center in the open field test among the three groups ( P>0.05). Compared with group C, the escape latency was significantly prolonged, the number of crossing the original platform was reduced, the apoptosis rate of hippocampal neurons and [Ca 2+ ] i were increased, the expression of p-RhoA, ROCK2 and cleaved-caspase-3 was up-regulated ( P<0.05), and the pathological injury to hippocampal neurons was found in group S. Compared with group S, the escape latency was significantly shortened, the number of crossing the original platform was increased, the apoptosis rate of hippocampal neurons and [Ca 2+ ] i were decreased, the expression of p-RhoA, ROCK2 and cleaved-caspase-3 was down-regulated ( P<0.05), and the pathological injury to hippocampal neurons was attenuated in group Y. Conclusions:The mechanism by which multiple exposures to sevoflurane induces long-term cognitive impairment is related to activation of RhoA/Rock2 signaling pathway and induction of apoptosis rate of hippocampal neurons in neonatal rats.

Application effect of severe ultrasound combined with extravascular lung water and intrathoracic blood volume in volume management of mechanical ventilation / 中国医师进修杂志

Objective:To explore the application effect of critical ultrasound combined with extravascular lung water (EVLW) and intrathoracic blood volume (ITBV) on volume management of mechanically ventilated patients.Methods:From May 2017 to January 2020, 98 patients treated with mechanically ventilated admitted to Hebei Petro China Central Hospital were selected and divided into two groups by random number table method, with 49 cases in each group. Both groups were treated with fluid resuscitation, the control group was guided by central venous pressure (CVP) and the observation group was guided by critical ultrasound combined with EVLW and ITBV. The acute physiology and chronic health score Ⅱ(APACHE Ⅱ) and sequential organ failure assessment (SOFA) scores, hemodynamic indexes, respiratory system indexes , urine output and fluid intake at 6 h and 24 h after resuscitation and mechanical ventilation time, ICU stay, complications and prognosis were compared between the two groups.Results:The scores of APACHE Ⅱ and SOFA in the observation group at 6 h and 24 h after resuscitation were lower than those in the control group: 6 h after resuscitation: (22.02 ± 4.29) scores vs. (23.94 ± 3.56) scores, (10.02 ± 3.11) scores vs. (11.64 ± 2.30) scores; 24 h after resuscitation: (19.66 ± 2.85) scores vs. (21.78 ± 3.60) scores, (7.64 ± 2.15) scores vs. (9.83 ± 2.07) scores, the differences were statistically significant ( P<0.05). The mean arterial pressure (MAP) and CVP in the observation group at 6 h and 24 h after resuscitation were higher than those in the control group: 6 h after resuscitation: (69.44 ± 5.25) mmHg(1 mmHg=0.133 kPa) vs. (65.98 ± 4.33) mmHg, (13.64 ± 2.30) mmHg vs. (11.89 ± 3.07) mmHg; 24 h after resuscitation: (72.89 ± 4.69) mmHg vs. (69.26 ± 5.53) mmHg, (13.07 ± 2.15) mmHg vs. (11.89 ± 3.07) mmHg; the heart rate was lower than those in the control group: 6 h after resuscitation: (98.58 ± 9.32) bpm vs. (105.03 ± 8.76) bpm; 24 h after resuscitation: (94.97 ± 8.46) bpm vs.(101.44 ± 7.34) bpm, the differences were statistically significant ( P<0.05). The central venous oxygen saturation (ScvO 2) and oxygenation index (OI) in the observation group at 6 h and 24 h after resuscitation were higher than those in the control group: 6 h after resuscitation: 0.749 ± 0.043 vs. 0.711 ± 0.047, (258.18 ± 20.75) mmHg vs. (234.66 ± 25.42) mmHg; 24 h after resuscitation: (77.68 ± 4.09)% vs. (73.54 ± 4.23)%, (376.29 ± 22.39) mmHg vs. (234.66 ± 25.42) mmHg; the blood lactic acid was lower than that in the control group: 6 h after resuscitation: (3.04 ± 0.52) mmol/L vs. (4.22 ± 0.39) mmol/L; 24 h after resuscitation: (1.01 ± 0.34) mmol/L vs. (1.87 ± 0.41) mmol/L, the differences were statistically significant( P<0.05). The urine output at 6 h and 24 h in the observation group was higher than that in the control group: 6 h after resuscitation: (0.49 ± 0.08) ml/(kg·h) vs. (0.35 ± 0.06) ml/(kg·h); 24 h after resuscitation:(0.54 ± 0.05) ml/(kg·h) vs. (0.42 ± 0.07) ml/(kg·h); the fluid intake was lower than that in the control group: 6 h after resuscitation: (1 230.2 ± 562.3) ml vs. (1 782.4 ± 534.7) ml; 24 h after resuscitation: (3 065.5 ± 521.2) ml vs. (3 642.0 ± 507.8) ml; the mechanical ventilation time, and ICU stay in the observation group were lower than those in the control group: (3.3 ± 0.9) d vs. (5.0 ± 0.7) d, (9.7 ± 2.1) d vs. (10.9 ± 1.8) d, the differences were statistically significant ( P<0.05). There was no significant differences in complication rate and 28-day survival curve between the two groups ( P>0.05). Conclusions:Critical ultrasound combined with EVLW and ITBV has a good application effect on volume management of patients with mechanical ventilation, which can help maintain hemodynamic stability, improve oxygenation status.

Risks and intervention strategies of pulmonary embolism in children / 中国小儿急救医学

There are more and more cases of pulmonary embolism in children, most of whom have etiology or risk factors, and the clinical features are not specific.At present, there is a lack of large samples of clinical researches in children.The methods of evaluation, diagnosis and treatment of pulmonary embolism in children mostly refer to adult experience and standards.No unified standards are applied to children.This review analyzed the risks factors, diagnosis, evaluation and treatments of pulmonary embolism in children, in order to help clinicians judge and treat their patients.

Early changes within the lymphocyte population are associated with the long term prognosis in severely injured patients / 北京大学学报(医学版)

OBJECTIVE@#To investigate the relationship between early lymphocyte responses and the prognosis in severely injured patients.@*METHODS@#Consecutive patients with severe trauma who were treated in Peking University People's Hospital Trauma Medical Center between June 2017 and June 2020 were enrolled in this restropective chart-review study. According to the responses of lymphocyte after severe injury, the patients were divided into three groups, group 1: lymphopenia-returned to normal; group 2: persistent lymphopenia; group 3: never lymphopenic, and the outcome of 28 d were recorded. Clinical data such as gender, age, base excess, mechanism of injury, Glasgow coma scale (GCS), injury severity score (ISS) and massive blood transfusion were collected. Perform statistical analysis on the collected clinical data to understand the trend of lymphocyte changes in early trauma and the relationship with prognosis. In order to eliminate the interference of age, stratification was carried out according to whether the age was ≥ 65 years old, in different age groups, they were grouped according to whether the length of stay was ≥ 28 d, and the relationship between lymphocyte trend and length of stay was discussed.@*RESULTS@#A total of 83 patients were included, 66 males and 17 females. The main injury mechanisms were traffic accident injuries and high-altitude fall injuries. The average ISS was (30±11) points. 65 patients had lymphopenia on the day of injury, 32 of them returned to normal on the 5th day, and the rest did not recover; the other 18 patients had normal lymphocyte levels after injury. Patients which are failure to normalize lymphopenia within the first 5 days following admission was related with the long hospitalization time and higher 28 d mortality rate. After further stratification by age, failure to normalize lymphopenia within the first 5 days following admission in the elderly group (age ≥65 years) was a risk factor for prolonged hospital stay (≥28 d), P=0.04. While in younger group, a high level of neutrophils within the first 5 d following admission was a risk factor for bad outcome.@*CONCLUSION@#A failure to normalize lymphopenia in severely injured patients is associated with significantly higher mortality and longer hospital stay. This study reveals lymphocytes can be used as a reliable indicator for the prognostic evaluation.

Application research of individualized teaching based on PBL in gynecology practice teaching / 中华医学教育探索杂志

Objective:To explore the teaching effect of individualized teaching based on problem-based learning (PBL) that applied in gynecology practice teaching under the background of conflicts between postgraduate examination preparation and clinical internship for medical undergraduates.Methods:A total of 157 medical students of the five-year program who started gynecology internship and meanwhile prepared for the postgraduate entrance examination in 2018 were enrolled in the study and divided into two groups randomly. One group received traditional teaching (control group), and the other group received individualized teaching mode based on PBL (observation group). After the internship, the two groups of students were assessed for their theoretical and clinical skills, and the students' evaluation of the teaching effect was acquired through a questionnaire survey. SPSS 22.0 was used to perform chi-square test.Results:The students in observation group had statistically significant higher scores in theory and skill tests than those in the control group ( P<0.001). The evaluation of boosting their enthusiasm for internship, advancing self-learning ability, conducing to improving learning methods in the future, and enhancing clinical skills and thinking ability in the observation group was significantly better than that in the control group ( P<0.05). Conclusion:The individualized teaching model based on PBL could efficiently alleviate the conflicts between internship and postgraduate entrance examination preparation and improve the effect of gynecology practice teaching.

Screening of DNA methylation sites associated with melanoma and construction of an aberrant methylation profile: a preliminary study / 中华皮肤科杂志

Objective:To screen aberrant DNA methylation sites associated with melanoma using gene chip technology, and to preliminarily construct a melanoma-specific methylation profile.Methods:The Illumina Human Methylation 450K whole-genome methylation chip was used to detect the whole-genome DNA in 6 melanoma tissues and their paralesional skin tissues, and DNA differentially methylated sites were obtained. Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) -based pathway analysis were carried out to investigate gene functions.Results:Gene chip testing showed that there were 27 779 differentially methylated sites between melanoma tissues and paralesional tissues, of which 16 673 were hypermethylated sites and 11 106 were hypomethylated sites in melanoma tissues. According to more stringent screening criteria " P < 0.01 and |Δβ| > 0.2", a total of 4 883 differentially methylated sites were screened out after filtering out all single nucleotide polymorphism-related probes, probes located on the XY chromosomes and cross-reactive probes, 1 459 (30%) of which were located in the promoter region including TSS1500, TSS200, 5′UTR and 1st Exon. GO enrichment analysis showed that differentially methylated genes were involved in many biological processes, including cell growth, differentiation, adhesion, movement and migration, signal transduction, transcriptional regulation, etc. KEGG-based pathway analysis showed that differentially methylated genes were mainly involved in signaling pathways, such as focal adhesion pathway, cancer pathways, transforming growth factor-β signaling pathway, phosphatidylinositol signaling pathway, melanogenesis pathway, chemokine signaling pathway, adhesion junction pathway, calcium signaling pathway, cell adhesion molecule pathway, mitogen-activated protein kinase signaling pathway, Wnt signaling pathway, Janus kinase-signal transducer and activator of transcription signaling pathway. Based on the criteira "the top 16 most differentially methylated genes related to hypermethylated sites in the promoter region, the genes with the highest methylation frequency (CpG sites ≥ 7) , the genes with certain functions or involved in a certain signaling pathway", 8 genes (KAAG1, DGKE, SOCS2, TFAP2A, GNMT, GALNT3, ANK2 and HOXA9) were selected as candidate biomarkers for melanoma. Conclusion:There are many hypermethylated genes in melanoma tissues, and 8 differentially methylated genes may serve as biomarkers for melanoma.

Role of reactive oxygen species-mediated mitochondrial pathway of apoptosis in long-term cognitive impairment induced by multiple exposures to sevoflurane in neonatal rats / 中华麻醉学杂志

Objective:To evaluate the role of reactive oxygen species (ROS)-mediated mitochondrial pathway of apoptosis in long-term cognitive impairment induced by multiple exposures to sevoflurane in the neonatal rats.Methods:Sixty SPF healthy neonatal Sprague-Dawley rats, weighing 12-20 g, were divided into 3 groups ( n=20 each) using a random number table method: control group (group C), multiple exposures to sevoflurane for anesthesia group (group S) and ROS inhibitor group (group A). Group S and group A inhaled 3% sevoflurane for 2 h starting from 6, 7 and 8 days after birth, while group C inhaled air.In group A, ROS inhibitor N-acetylcysteine (NAC) 150 mg/kg was intraperitoneally injected before each anesthesia with sevoflurane.The spontaneous activity was evaluated by open field test on day 35 after birth.The cognitive function was determined by Morris water maze test on day 36 after birth.The rats were sacrificed after the end of Morris water maze test, and the hippocampal tissues were obtained for determination of the apoptosis rate of hippocampal neurons, reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) (by flow cytometry) and levels of Cyt c and cleaved caspase-9 and caspase-3 (by Western blot). The expression of Bcl-2 and Bax mRNA was detected by real-time polymerase chain reaction.The ultrastructure of mitochondria in hippocampal neurons was observed with a transmission electron microscope. Results:Compared with group C, the escape latency was significantly prolonged, the number of crossing the original platform was reduced, the apoptosis rate of hippocampal neurons and levels of ROS and MMP were increased, the expression of Cyt c, cleaved caspase-9, cleaved caspase-3 and Bax mRNA was up-regulated, the expression of Bcl-2 mRNA was down-regulated, the ratio of Bax/Bcl-2 was increased ( P<0.05), mitochondria were swollen, and mitochondrial cristae structure was broken in group S. Compared with group S, the escape latency was significantly shortened, the number of crossing the original platform was increased, the apoptosis rate of hippocampal neurons and levels of ROS and MMP were decreased, the expression of Cyt c, cleaved caspase-9, cleaved caspase-3 and Bax mRNA was down-regulated, the expression of Bcl-2 mRNA was up-regulated, the ratio of Bax/Bcl-2 was decreased ( P<0.05), and the mitochondrial swelling and rupture of cristae structure were improved in group A. Conclusion:The mechanism by which multiple exposures to sevoflurane induce long-term cognitive impairment may be related to activating the ROS-mediated mitochondrial pathway of apoptosis in neonatal rats.

The clinical features and survival analysis of 109 patients with neuropsychiatric systemic lupus erythematosus / 中华风湿病学杂志

Objective:To investigate the clinical characteristics, prognosis, and risk factors for poor prognosis of neuropsychiatric systemic lupus erythematosus (NPSLE) .Methods:Patients who were diagnosed as NPSLE between January 2009 to January 2019 in Peking University First Hospital were included. Patients with neuro-psychiatric symptoms caused by other reasons such as infection and metabolic disorders were excluded. Patients were retrospectively followed up by telephone or medical records. Continuous variables were compared by student t test or Wilcoxon rank sum test. Quantitative variables were compared by chi-square test. Survival was analyzed by Kaplan-Meier curve. Predictive factors of prognosis was estimated by using Cox regression analysis. Results:One hundred and nine NPSLE patients were included. Thirteen (11.9%) were male and 96 (88.1%) were female with a median age of 33 years old. Central nervous system involvement was predominant (89/109, 81.7%) . The most common types were headache, cerebrovascular disease and epilepsy. Cranial neuropathy was the most common type at the initial onset of systemic lupus erythematosus (SLE) , while cerebrovascular disease was more common when SLE relapsed. Patients who demonstrated NPSLE at the initiation of SLE had shorter survival time than those who got NPSLE when SLE relapsed [ (32±26) months vs (197±79) months, t=2.834, P=0.037]. Among the 105 patients with complete followed up data, the follow up time was 118.0 (1.4, 525.7) months and 53.1 (0.4, 363.0) months from the onset of SLE and NPSLE, respectively. The mortality rate was 14.3% (15/105) . The survival rates of 1-5 years were 96.2%, 94.3%, 91.0%, 89.9% and 88.3%, respectively. The survival time was (180±138) months and (33±32) months, t=3.861 , P<0.01) from the onset of SLE and NPSLE, respectively. The major causes of death were infection, NSPLE and cardiovascular disease. Cerebrovascular disease was the independent risk factor for death [ RR=3.413, 95% CI (1.049, 11.102) , P=0.041]. Conclusion:Cranial neuropathy is the most common type at the initial onset of SLE, while cerebrovascular disease is more common when SLE relapsed. Patients who had NPSLE at the initiation of SLE have shorter survival time than those who got NPSLE when SLE relapsed. Cerebrovascular disease is the independent risk factor of death of NPSLE patients.

Research methods of miRNA in plants and research progress of miRNA in medicinal plants / 药学学报

microRNA (miRNA) is a class of endogenous ~21nt non-coding single-strand small RNAs which play important roles in plant growth and development, signal transduction, stress response, and secondary metabolism. In recent years, a large number of miRNAs have been identified in various medicinal plants, and the regulatory effects of these miRNAs have been preliminarily studied. In medicinal plants, most of the active components are secondary metabolites, so it is of great significance to study the regulatory effects of miRNA on the formation of secondary metabolites. In this paper, the general research methods of plant miRNA and the research progress of medicinal plant miRNA and their regulatory effects on the formation of bioactive metabolites were reviewed, and the future direction of medicinal plant miRNA was prospected, so as to provide reference for the future research of medicinal plants.

Transcriptome analysis and validation of key genes involved in biosynthesis of iridoids in Gentiana lhassica / 中国中药杂志

As the main chemical constituents, iridoids are widely distributed within Gentiana, Gentianaceae, with promising bioactivities. Based on the previous work, the transcriptome of G. lhassica, an original plant of Tibetan herb "Jieji Nabao", was sequenced and analyzed in this study, and the transcriptome databases of roots, stems, leaves, and flowers were constructed so as to explore unigenes that may encode the key enzymes in the biosynthetic pathway of iridoids. Then, qRT-PCR was used to validate the relative expression levels of 11 genes named AACT, DXS, MCS, HDS, IDI, GPPS, GES, G10H, 7-DLNGT, 7-DLGT, and SLS in roots, stems, leaves, and flowers. Also, the total contents of gentiopicroside and loganic acid were determined by HPLC, respectively. The results are as follows:(1)a total of 76 486 unigenes with an average length of 852 bp were obtained;(2)335 unigenes were involved in 19 stan-dard secondary metabolism pathways in KEGG database, with phenylpropanoid biosynthesis having the maximum number(75 unigenes), and no isoflavone biosynthetic pathway was annotated;(3)171 unigenes participatedin 27 key enzymes encoding in the biosynthetic pathway of iridoids, and 1-deoxy-D-xylulose-5-phosphate reductoisomerase(DXR) gene was highly expressed;(4)qRT-PCR results were approximately consistent with RNA-Seq data and the relative expression levels of the 11 genes were higher in the aboveground parts(stem, leaf, and flower) than in the underground part(root);(5)the total contents of gentiopicroside and loganic acid were higher in the aboveground parts(stem, leaf, and flower) than in the underground part(root), and the difference was significant. This study provides basic scientific data for accurate species identification, evaluation of germplasm resources, research on secondary pro-duct accumulation of medicinal plants within Gentianaceae, and protection of endangered alpine species.

Alterations in thymocyte populations under conditions of endotoxin tolerance / 中华医学杂志(英文版)

BACKGROUND@#Endotoxin tolerance (ET) is a protective phenomenon in which pre-treatment with a tolerance dose of lipopolysaccharide (LPS) leads to dramatically elevated survival. Accumulating evidence has shown that peripheral T cells contribute to the induction of ET. However, what happens to T cell development in the thymus under ET conditions remains unclear. The purpose of this study was to analyze the alterations in thymocyte populations (double-positive [DP] and single-positive [SP] cells) under ET conditions.@*METHODS@#Mice were intraperitoneally injected with LPS at a concentration of 5 mg/kg to establish an LPS tolerance model and were divided into two groups: a group examined 72 h after LPS injection (72-h group) and a group examined 8 days after LPS injection (8-day group). Injection of phosphate-buffered saline was used as a control (control group). Changes in thymus weight, cell counts, and morphology were detected in the three groups. Moreover, surface molecules such as CD4, CD8, CD44, CD69, and CD62L were analyzed using flow cytometry. Furthermore, proliferation, apoptosis, cytokine production, and extracellular signal-regulated kinase (ERK) pathway signaling were analyzed in thymocyte populations. The polymorphism and length of the T-cell receptor (TCR) β chain complementarity-determining region 3 (CDR3) were analyzed using capillary electrophoresis DNA laser scanning analysis (ABI 3730).@*RESULTS@#Thymus weight and cell counts were decreased in the early stage but recovered by the late stage in a murine model of LPS-induced ET. Moreover, the proportions of DP cells (control: 72.130 ± 4.074, 72-h: 10.600 ± 3.517, 8-day: 84.770 ± 2.228), CD4+ SP cells (control: 15.770 ± 4.419, 72-h: 44.670 ± 3.089, 8-day: 6.367 ± 0.513), and CD8+ SP cells (control: 7.000 ± 1.916, 72-h: 34.030 ± 3.850, 8-day: 5.133 ± 0.647) were obviously different at different stages of ET. The polymorphism and length of TCR β chain CDR3 also changed obviously, indicating the occurrence of TCR rearrangement and thymocyte diversification. Further analysis showed that the expression of surface molecules, including CD44, CD69, and CD62L, on thymocyte populations (DP and SP cells) were changed to different degrees. Finally, the proliferation, apoptosis, cytokine production, and ERK pathway signaling of thymocyte populations were changed significantly.@*CONCLUSION@#These data reveal that alterations in thymocyte populations might contribute to the establishment of ET.

Transcriptome analysis and exploration of genes involved in the biosynthesis of iridoids in Gentiana crassicaulis (Gentianaceae) / 药学学报

italic>Gentiana crassicaulis Duthie ex Burk. in Gentiana (Sect. Cruciata), Gentianaceae, is one of the original plants of both Gentianae Macrophyllae Radix and Tibetan herb Jie-Ji Na-Bao, which contain such bioactive iridoids as gentiopicroside, loganic acid and others. In this study, based on previous work, the transcriptome of G. crassicaulis was sequenced and analyzed to construct transcriptome databases of roots, stems, leaves and flowers. qRT-PCR verification was conducted for parts of unigenes that may be key enzymes in the pathway of iridoid biosynthesis. The results are as follows: ① a total of 159 534 unigenes were obtained, with an average length of 679 bp. According to the functional classification of GO, unigenes can be divided into 3 categories with 67 branches. The unigenes were aligned in the KOG database and were classified into 25 categories according to function. ② In the KEGG database, 215 unigenes were implicated in 20 standard secondary metabolism pathways. The analysis shows that 305 unigenes encoded 28 key enzymes in the pathway of iridoid biosynthesis, and their expression in different organs is different; and ③ qRT-PCR was approximately consistent with RNA-Seq results. The 7 annotated unigenes identified in this study, HMGS, DXS, MCS, GPPS, G10H, 7-DLNGT and STR, all had higher relative expression levels in the above-ground parts (stem, leaf and flower) than in the underground part (root). Iridoids are common active and index components of such traditional Chinese medicines as Qinjiao, Longdan, Dangyao, and Qingyedan, among others. Therefore, this work provides basic scientific data for further development including obtaining active components or intermediates through biotechnology, exploring the accumulation of effective components, evaluating the quality of different ecotype varieties, and identifying authentic biosynthesis pathways of medicinal materials.

The effects of spinal manipulation on chronic, non-specific neck pain / 中华物理医学与康复杂志

Objective:To observe the clinical effect of spinal manipulation on chronic, non-specific neck pain.Methods:Thirty patients with chronic, nonspecific neck pain were divided randomly into an observation group ( n=15) and a control group ( n=15). Patients in the observation group were given 20 minutes of a novel 4R spinal manipulation (resetting joint malalignment, resetting abnormal muscle, resetting joint stabiliazation, resetting sensorimotor control) twice a week for 2 weeks while the control group were given 20 minutes of medium frequency and high frequency conventional physiotherapy 4 times a week, also for 2 weeks. Before the treatment, right after, and one and three months later, both groups were evaluated using a visual analogue scale (VAS) and a neck disability index (NDI). Right before and after the treatment, cervical flexion and extension range of motion (ROM) were measured. The surface electromyography was employed to record the root mean square (RMS) of the EMG amplitude and the median frequency (MF) from the erector spinae and upper trapezius. Results:Before the treatment no significant differences were found in any of the measurements between the two groups. Afterward and one and three months later the average VAS, NDI and cervical ROM results of both groups had improved significantly, with the improvements in the observation group significantly greater than those in the control group on average. After 2 weeks of treatment, the average RMS and MF values had improved in both groups, again with the observation group′s average values significantly better than those of the control group.Conclusion:Spinal manipulation can effectively improve the strength and stamina of cervical muscle groups in patients with chronic, non-specific neck pain.

Risk factor analysis of non-acute symptomatic portal vein thrombosis after endoscopic gastric variceal injection / 中国医师杂志

Objective:To analyze the incidence and risk factors of non-acute symptomatic portal vein thrombosis (PVT) after endoscopic gastric variceal injection (GVI) in the treatment of liver cirrhosis with gastric variceal bleeding (GVB).Methods:66 patients with GVB who were treated with GVI for the first time from July 2017 to October 2019 in Minhang Hospital Affiliated to Fudan University were retrospectively analyzed. The data of gender, age, preoperative Child-Pugh grade, preoperative platelet count, preoperative plasma D-dimer concentration, preoperative splenic length, preoperative portal vein velocity, preoperative splenic vein velocity, preoperative portal vein diameter, preoperative splenic vein diameter, treatment times, total number of injection points, total dose of sclerosing agent and tissue adhesive agent were collected. The patients were divided into PVT group and non-PVT group according to the occurrence of non-acute symptomatic PVT within one year after GVI. Univariate analysis was performed first, and then the factors with P<0.2 were included in the binary logistic regression model to screen the risk factors of PVT after GVI. Results:There were 25 cases (37.88%) in PVT group and 41 cases (62.12%) in non-PVT group. There were significant differences in D-dimer concentration, spleen length, Child-Pugh grade and total dose of sclerosing agent between the two groups ( P<0.05). The D-dimer concentration ( OR=2.319, 95% CI:1.359-3.956), spleen length ( OR=1.044, 95% CI:1.007-1.081) and total dose of sclerosing agent ( OR=1.075, 95% CI:1.004-1.152) were independent risk factors for PVT ( P<0.05). Conclusions:Preoperative D-dimer concentration, spleen length and total dose of sclerosing agent can predict the risk of PVT after GVI. In order to reduce the risk of PVT after GVI, the dose of sclerosing agent should be reduced as much as possible.