Objective To approach the clinical features of refractory Mycoplasma pneumoniae pneumonia (RMPP) in children and provide evidence for clinical treatment.Methods A retrospective analysis was conducted.Two hundreds and twenty children with Mycoplasma pneumoniae pneumonia (MPP) admitted to the Integrated Chinese and Western Medicine Hospital of Zhejiang Province were enrolled from January 2013 to June 2016.According to the criteria of diagnosis of MPP and RMPP,the patients were divided into a general group (172 cases) and a refractory group (48 cases).Their general information,incidence of pulmonary positive sign,laboratory data,radiological findings,complications,therapeutic situations and prognoses,etc.between the two groups were observated.Results There were no significant difference in gender distribution between the two groups,but the age of the refractory group was much older than that of the general group (years:7.15 ± 1.89 vs.4.86±2.16),and the fever duration in the refractory group was obviously longer than that in the general group (days:11.17±2.82 vs.6.27±2.29,P ＜ 0.01).the levels of C-reactive protein [CRP (mg/L):46.8 ± 18.6 vs.13.5 ± 8.4],erythrocyte sedimentation rate [ESR (mm/1 h):56.9 ± 14.8 vs.28.7 ± 10.2],incidence of positive pulmonary sign [100.0％ (48/48) vs.33.7％ (58/172)],the incidence of pulmonary segmental consolidation [81.3％ (39/48) vs.27.9％ (48/172)] and incidence of extra-pulmonary complications [83.3％ (40/48) vs.16.3％ (28/172)] were all higher in refractory group than those in general group (all P ＜ 0.01).All the patients were treated with macrolide antibiotics and traditional Chinese medicine,the patients in the refractory group was additionally treated with glucocorticoids,and a few of them were treated with gamma immunoglobulin and bronchoalveolar lavage;all of them recovered well,high temperature had returned to normal and the imageology findings recovered well,then they all discharged without any sequela such as bronchiectasis and pulmonary atelectasis.Conclusions Physicians should pay more attention to the MPP patients with longer fever duration (especially longer than 10 days),serious signs and symptoms,more complications,higher CRP and ESR and severe radiological findings.Besides early application of macrolide antibiotics,rational use of glucocorticoids,gamma immunoglobulin and bronchoalveolar lavage can be considered.

BACKGROUND: There are differences in physical and biological activity between the antibody from mammals and egg yolk antibody (IgY) from chicken. IgY is acid- and heat-resistant, and can prevent and cure the infectious diseases in animals and human being, which is also benefit to develop routine diagnostic immunoassays. Conventional ELISA assay for IgY takes much more time than dot-immunobinding assay.OBJECTIVE: To detect the IgY stability byusing dot-immunobinding assay.DESIGN: Open trail.SETTING: Department of Transfusion, Kunming General Hospital of Chinese PLA.MATERTALS: The experiment was completed in the Kunming General Hospital of Chengdu Military Area Command of Chinese PLA from January to June 2006. Two White Leghorn hens (30 weeks old) were selected. HLA-A*0201 α chain served as the antigen. The total protein concentration of the purified antigen was 0.04 g/L with the molecular mass of 32 000(self-prepared); nitrocellulose filter (NC, import and divided); nonfat dry milk (Anyi Corp. No. 20051220); DAB (Boshide Corp.);caprylic acid (made by Shanghai Xinghuo Chemical Factory); ammonium sulfate (Shantou Guanghua Chemical product).METHODS: ①HLA-A*0201 α chain with the total protein concentration of 0.04 g/L was purified with egg yolk antibody,and identified by SDS-PAGE. ②1 μL antigen was spotted into the center of NC membrane and dried in the incubator at 37 ℃. Then the NC membrane was blocked in 1 mL PBST and put in the incubator at 37℃ with shaking in 90 r per minute for 15 minutes. Then the liquid was exchanged with 1 mL PBST and added the primary antibody at a final concentration of 10 mg/L. After 30 minutes shaking in the incubator at 37 ℃, the NC membrane was washed in PBST for three times. The second antibody, mouse anti-chicken IgY conjugated to horseradish peroxidase (HRP) was added and after 30 minutes incubation, the NC membrane was washed three times in PBST. Binding was revealed by incubation with a DAB reagent. A positive reaction was represented by adeep brown spot,Irdlcating that IgY had better activity; if the spot became lighter IgY lost part activity, and when the spot disappeared, the IgY lost a the activty.According to intensity (gray degree)of the dot compared tothe standard, the remained percent of activity of the IgY was calculated. ③IgY was adjusted to three different protein concentrations with PBS: 1, 0.1, 0.01 g/L and stayed at room temperature for four months. 10 μg lgY was taken out from each concentration sample every month to detect the activity by dot-immunobinding assay. ④IgY was put into seven EP tubes with 100 μL per tube and numbered 1-7. Number 1 to 3 was adjusted pH to 5, 3 and 2, respectively with 1 mol/L HCI; Number 4 to 6 was to 9, 11 and 12, respectively with 1 mol/L NaOH. The pH of number 7 was neutral without adding acid or base. The samples were stayed in incubator at 37 ℃ for 3 hours. 10 μg IgY from each tube was taken every hour to detect the stability at different pH by dot-immunobinding assay. ⑤IgY was added to six EP tubes (10 μL per tube) and numbered 1-6. Number 1-6 was put into waterbath at 30, 40, 50, 60, 70 and 80 ℃ for 15 minutes. After cooled in refrigerator at 4 ℃, 10 mg samples from each tube and standard sample (untreated sample) taken to check the thermal stability by dot-immunobinding assay.MAIN OUTCOME MEASURES: ①SDS-PAGE of IgY. ②IgY stability at room temperature. ③IgY stability at different pH. ④ Detection of IgY thermal stability.RESULTS: ①Purified IgY after SDS-PAGE had two major binds, the molecular mass of the heavy chain was 66.000,and the light chain was 25 000. ②1, 0.1, 0.01 g/L IgY still had partial activity after staying at room temperature for four months. ③When pH ranged from 5 to 9, IgY still had partial activity after staying in 37 ℃ for 3 hours. If pH was lower than 5 or higher than 9, it lost the whole activity in above condition. ④Purified IgY was added to six EP tubes, the number 1-4 still had partial activity, but number 5 and 6 showed some white precipitate, which was caused by protein denaturation at higher temperature.CONCLUSION: IgY stability is higher than others. The dot-immunobinding assay described a rapid and simple method for the demonstration and characterization of functional activity of egg yolk antibody. With only small volume antigen and antibody, and specific dot, the dot-immunobinding assay method could process many samples at the same time.

Medical education should combine clinical professional skills with humanities skills,integrate humanities knowledge into vocational education.Taking 2008 grade seven-year program clinical medicine students in the 2nd affiliated hospital of Harbin Medical University as fostering object,we made researches into students' understanding of doctor-patient relationship before practice and their mastering of skills.Through conducting questionnaire,we got to know the effect of humanities practice skill training for seven-year program clinical medicine students.Meanwhile,we compared students' self evaluation results before and after training,discussed on how to improve medical students' communication skills,cultural skills and the reform direction in an aim to guide students to transit from students to clinical doctor.

Objective To establish the screening platform of circulating tumor cells (CTCs) using acridine orange fluo?rescent (AO-F) dyeing method, and to apply it in the screening of peripheral blood CTCs in patients with kidney cancer. Methods Twenty-seven patients with metastatic renal cell carcinoma was included in this study. Primitive tumor cells and kidney cancer cell line 769-P were cultured with different concentrations of fetal bovine serum. Smears were prepared and observed under fluorescence microscopy. The percentage of AO-F positive staining of 769-P cells under 5 random sights was calculated. The sensitivity of AO-F staining to cells was evaluated. The 5 mL morning fasting venous blood was obtained from 10 subjects with healthy check-up. The 1×106 cell suspension was prepared. The logarithmic phase of renal tumor cells was used to prepare tube containing 500, 200, 100, 50 and 10 tumor cell suspension, which were mixed with 1×106 nucleated cells to establish CTCs model of renal cancer. AO-F staining method was used to detect the expression of AO-F positive cells. The correlation between expression of AO-F positive cells and clinical parameters was analyzed. Results The prima?ry cells and cell line 769-P showed similar bright color and morphological characteristics. The percentage of AO-F positive staining in 769-P cells was 93%±3%under 5 random sights. The recovery rates (%) of four groups (500, 200, 100 and 50 tu?mor cell suspension) were 10.2±3.8, 9.2±2.3, 10.8±2.6 and 10.5±1.9, respectively. There were no significant differences in recovery rates between four groups (P>0.05). The group of 10 tumor cell suspension could find AO-F positive staining cells occasionally. Zero case was positive in controls. Nine of 27 patients were positive and the rate was 33.33%. There were no significant statistical differences in AO-F positive rates between gender, age, tumor size, pathological pattern, Furhman stage, metastasis of lung and presence of tumor (P>0.05). Conclusion It is confirmed that the method of CTCs staining with AO-F, which has high specificity and reproducibility, is feasible to detect CTCs and worthy of being studied. There is a certain reference value to predict tumor recurrence and metastasis.

Objective:To investigate the prognosis of severe hyperbilirubinemia in full-term infants who met the exchange transfusion criteria and were treated by blood exchange transfusion and phototherapy.Methods:A total of 168 full-term infants with severe hyperbilirubinemia who met the criteria for exchange transfusion and were hospitalized in the Neonatology Department of seven tertiary hospitals in Hebei Province from June 2017 to December 2018 were retrospectively included. According to the treatment protocol, they were divided into two groups: exchange transfusion group (38 cases) and phototherapy group (130 cases). Two independent sample t-test and Chi-square test were used to compare the clinical manifestations and follow-up results between the two groups. Multivariate logistic regression was used to analyze the risk factors for poor prognosis. Results:Neonatal severe hyperbilirubinemia in the exchange transfusion and phototherapy group were both mainly caused by hemolytic disease [42.1%(16/38) and 29.2%(38/130)], sepsis [28.9%(11/38) and 11.5%(15/130)] and early-onset breastfeeding jaundice [15.8%(6/38) and 11.5%(15/130)]. Total serum bilirubin level on admission in the exchange transfusion group was significantly higher than that in the phototherapy group [(531.7±141.3) vs (440.0±67.4) μmol/L, t=3.870, P<0.001]. Moreover, the percentage of patients with mild, moderate and severe acute bilirubin encephalopathy in the exchange transfusion group were higher than those in the phototherapy group [15.8%(6/38) vs 3.8%(5/130), 7.9%(3/38) vs 0.8%(1/130), 13.2%(5/38) vs 0.0%(0/130); χ2=29.119, P<0.001]. Among the 168 patients, 135 were followed up to 18-36 months of age and 12 showed poor prognosis (developmental retardation or hearing impairment) with four in the exchange transfusion group (12.9%, 4/31) and eight in the phototherapy group (7.7%, 8/104). Multivariate logistic regression analysis showed that for full-term infants with severe hyperbilirubinemia who met the exchange transfusion criteria, phototherapy alone without blood exchange transfusion as well as severe ABE were risk factors for poor prognosis ( OR=14.407, 95% CI: 1.101-88.528, P=0.042; OR=16.561, 95% CI: 4.042-67.850, P<0.001). Conclusions:Full-term infants who have severe hyperbilirubinemia and meet the exchange transfusion criteria should be actively treated with blood exchange transfusion, especially for those with severe ABE, so as to improve the prognosis.

Objective:To investigate the clinical, neuropsychological, and neuroimage characteristics in patients with corticobasal syndrome (CBS), and to elucidate the exact diagnosis of CBS patients.Methods:Twelve CBS cases admitted to the Department of Neurology, Huashan Hosiptal,Fudan University from April 2019 to July 2021 were retrospectively enrolled in this study. Those data, including clinical features (demographic data and clinical characteristics of cortical dysfunction and movement disorder), neuropsychological assessment [Mini-Mental State Examination (MMSE) and Montreal Cognitive Assessment (MoCA) scales score], brain magnetic resonance imaging (MRI) and multi-mode positron emission tomography (PET)/CT, were collected and carefully reviewed. Exact diagnosis of these patients was given according to the disease diagnosis criteria.Results:Cortical dysfunction and asymmetrical movement disorders were found in all cases, with poor response to levodopa. Patients suffered from cognitive impairment (MMSE score 16.16±9.82, MoCA score 13.44±7.35). The cranial MRI demonstrated significant asymmetric atrophy of frontal and parietal lobes, especially in the pre- and post-central gyrus. Fluorodeoxyglucose PET of 12 patients showed asymmetric frontal lobe and basal ganglia (especially caudate and putamen) hypometabolism (obviously on the contralateral side of the affected limb). Tau PET was implemented in 11 patients and displayed that abnormal tau protein deposition was positive in the cortex and/or subcortex in all patients. Of the 4 cases, who completed amyloid PET, amyloid protein deposition was positive in the cortex of 2 patients. As a result, 6 patients were diagnosed as progressive supranuclear palsy, 1 patient was diagnosed as corticobasal degeneration, and 5 patients were diagnosed as Alzheimer′s disease.Conclusions:The etiology of CBS is heterogeneous. The combination of clinical manifestation, cranial MRI and multi-mode PET/CT helps the differential diagnosis of CBS.

Objective:To analyze the effect of intestinal fluid reinfusion after anterior resection of rectum and preventive ileostomy in patients with low rectal cancer.Methods:A prospective research method was used, and 60 patients with low rectal cancer in Zhejiang Jinhua Guangfu Tumor Hospital from January 2021 to June 2022 were enrolled. The patients were divided into control group (treated with anterior resection of rectum and preventive ileostomy) and observation group (treated with anterior resection of rectum and preventive ileostomy combined with intestinal fluid infusion) by random number table method with 30 cases each. On the next day and 3 months after surgery, the low anterior resection syndrome (LARS) scale was used to evaluate anal function, the patient-generated subjective global assessment (PG-SGA) method was used to evaluate nutritional status, the stoma-quality of life (stoma-QOL) was used to evaluate the quality of life. The complications were recorded.Results:Both groups of patients were successfully completed the scheduled surgery. There were no statistical differences in LARS score and incidence of LARS on the next day after surgery between the two groups ( P>0.05); the LARS score and incidence of LARS 3 months after surgery in observation group were significantly lower than those in control group: (18.63 ± 3.15) scores vs. (24.90 ± 6.11) scores and 23.33% (7/30) vs. 46.67% (14/30), and there were statistical differences ( P<0.01 or <0.05). There were no statistical differences in PG-SGA score and incidence of malnutrition on the next day after surgery between the two groups ( P>0.05); the PG-SGA score and incidence of malnutrition 3 months after surgery in observation group were significantly lower than those in control group: (3.07 ± 0.82) scores vs. (5.13 ± 1.01) scores and 26.67% (8/30) vs. 46.67% (14/30), and there were statistical differences ( P<0.01 or <0.05). There were no statistical differences in the scores of psychological burden, social interaction, stoma management and daily living of stoma-QOL on the next day after surgery between the two groups ( P>0.05); the scores 3 months after surgery in observation group were significantly higher than those in control group: (27.70 ± 4.28) scores vs. (21.47 ± 5.16) scores, (14.33 ± 2.03) scores vs. (11.90 ± 1.64) scores, (14.87 ± 1.92) scores vs. (11.57 ± 2.38) scores and (15.30 ± 1.03) scores vs. (12.37 ± 2.11) scores, and there were statistical differences ( P<0.01). The incidence of complications in observation group was significantly lower than that in control group: 30.00% (9/30) vs. 60.00% (18/30), and there was statistical difference ( χ2 = 5.45, P<0.05). Conclusions:For the patients with low rectal cancer, the anterior resection of rectum and preventive ileostomy combined with intestinal fluid reinfusion can reduce LARS, improve nutritional status, improve quality of life, and also reduce complications.

Objective:To study the current status of longitudinal extrauterine growth restriction (EUGR) in extremely preterm infants (EPIs) and to develop a prediction model based on clinical data from multiple NICUs.Methods:From January 2017 to December 2018, EPIs admitted to 32 NICUs in North China were retrospectively studied. Their general conditions, nutritional support, complications during hospitalization and weight changes were reviewed. Weight loss between birth and discharge > 1SD was defined as longitudinal EUGR. The EPIs were assigned into longitudinal EUGR group and non-EUGR group and their nutritional support and weight changes were compared. The EPIs were randomly assigned into the training dataset and the validation dataset with a ratio of 7∶3. Univariate Cox regression analysis and multiple regression analysis were used in the training dataset to select the independent predictive factors. The best-fitting Nomogram model predicting longitudinal EUGR was established based on Akaike Information Criterion. The model was evaluated for discrimination efficacy, calibration and clinical decision curve analysis.Results:A total of 436 EPIs were included in this study, with a mean gestational age of (26.9±0.9) weeks and a birth weight of (989±171) g. The incidence of longitudinal EUGR was 82.3%(359/436). Seven variables (birth weight Z-score, weight loss, weight growth velocity, the proportion of breast milk ≥75% within 3 d before discharge, invasive mechanical ventilation ≥7 d, maternal antenatal corticosteroids use and bronchopulmonary dysplasia) were selected to establish the prediction model. The area under the receiver operating characteristic curve of the training dataset and the validation dataset were 0.870 (95% CI 0.820-0.920) and 0.879 (95% CI 0.815-0.942), suggesting good discrimination efficacy. The calibration curve indicated a good fit of the model ( P>0.05). The decision curve analysis showed positive net benefits at all thresholds. Conclusions:Currently, EPIs have a high incidence of longitudinal EUGR. The prediction model is helpful for early identification and intervention for EPIs with higher risks of longitudinal EUGR. It is necessary to expand the sample size and conduct prospective studies to optimize and validate the prediction model in the future.