Expression of water-soluble nucleocapsid protein of SARS-CoV-2 and analysis of its immunogenicity.

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to be a major public health concern. Nucleocapsid (N) protein is the most abundant structural protein on SARS-CoV-2 virions and induces the production of antibodies at the early stage of infection. Large-scale preparation of N protein is essential for the development of immunoassays to detect antibodies to SARS-CoV-2 and the control of virus transmission. In this study, expression of water-soluble N protein was achieved through inducing protein expression at 25°C with 0.5 mM IPTG for 12 h. Western blot and ELISA showed that recombinant N protein could be recognized by sera collected from subjects immunized with Sinovac inactivated SARS-CoV-2 vaccine. Four monoclonal antibodies namely 2B1B1, 4D3A3, 5G1F8, and 7C6F5 were produced using hybridoma technology. Titers of all four monoclonal antibodies in ELISA reached more than 1.28×10 6.0. Moreover, all monoclonal antibodies could react specifically with N protein expressed by transfection of pcDNA3.1-N into BHK-21 cells in IPMA and IFA. These results indicated that water-soluble N protein retained high immunogenicity and possessed the same epitopes as that of native N protein on virions. In addition, the preparation of water-soluble N protein and its monoclonal antibodies laid the basis for the development of immunoassays for COVID-19 detection.

[Parameningeal or non-parameningeal head and neck rhabdomyosarcoma: a study based on propensity score matching and survival analysis].

Objective: To compare the prognoses between parameningeal and non-parameningeal head and neck rhabdomyosarcoma based on propensity score matching and to explore the prognostic factors of overall survival in patients with head and neck rhabdomyosarcoma. Methods: The medical records of 64 patients with pathologically diagnosed as head and neck rhabdomyosarcoma from January 2016 to May 2020 in Peking Union Medical College Hospital were retrospectively retrieved, including 31 males and 33 females, with an average age of (8.0±8.9) years. Kaplan-Meier method was used to draw and compare survival curves in subgroup analysis according to different histopathological characteristics. Patients were divided into non-parameningeal (27 cases) and parameningeal (37 cases) group based on the location of primary lesion. Patients were further selected using 1∶1 propensity score matching method. The basic clinical data and overall survival were compared before and after matching. Prognostic factors were anlysed using Cox's proportional hazards regression model. Results: In 64 patients with head and neck rhabdomyosarcoma, lower risk stratification, and lower TNM stage indicated higher overall survival (all P<0.05). Before matching, patients in parameningeal group presented with higher T stage and IRS (Intergroup Rhabdomyosarcoma Study) staging (all P<0.05). There were no significant differences in basic clinical data and 1-, 2-, and 3-year overall survival rates between two groups after matching(P>0.05). Tumor size smaller than 5 cm, embryonal histology, negative FOXO1 fusion gene, lower risk stratification, and lower TNM stage were associated with higher overall survival (all P<0.05). Among these, tumor size and histology were independent prognostic factors (HR=2.36, 95%CI:1.07-5.20, P=0.033; HR=5.54, 95%CI: 1.18-25.95, P=0.030). Conclusions: There is no significant difference in overall survival between patients with parameningeal and non-parameningeal rhabdomyosarcomas. Tumor size smaller than 5 cm and embryonal histology are two independent prognostic factors.

[The effects of sequence of radiotherapy and chemotherapy on prognosis of patients with early-stage extranodal NK/T cell lymphoma (nasal type) and its individualized application].

Objective: To analyze the effects of the sequence of radiotherapy and chemotherapy on the efficacy of early-stage extranodal NK/T-cell lymphoma (nasal type, ENKTCL) patients, and to provide a quantitative evaluation method for individualized radiotherapy and chemotherapy. Methods: The Chinese Lymphoma Collaborative Group (CLCG) collected the clinical data of 2 008 patients with early-stage Ⅰ/Ⅱ ENKTCL who received radiotherapy and chemotherapy from January 2000 to early September 2019 from 21 hospitals across the country, including 1 417 males and 591 females, aged 2 to 83 (42±14) years. According to the sequence of radiotherapy and chemotherapy, patients were divided into radiotherapy-first group (388 cases) and chemotherapy-first group (1 620 cases). Survival rate was estimated using Kaplan-Meier method, and multivariate Cox proportional risk model was used to screen and identify independent prognostic factors. The prognostic prediction models of the two therapies were constructed separately, and the models were used to predict the individualized mortality risk of all patients to determine the appropriate radiotherapy and chemotherapy regimen for each patient. Results: The 5-year overall survival rate was 74.2% (95%CI: 69.6%-79.2%) in the radiotherapy-first group and 69.7% (95%CI: 67.1%-72.4%) in the chemotherapy-first group. Although the 5-year overall survival rate of patients in the radiotherapy-first group was numerically higher than that of the chemotherapy-first group, the difference was not statistically significant (χ2= 2.26, HR=0.84 (95%CI: 0.68-1.05), P=0.133). Six variables including age, gender, ECOG score, LDH, Ann Arbor staging, and PTI (primary tumor invasion) were screened out as independent prognostic factors (the chemotherapy-first group: HR were 1.01, 1.25, 2.07, 0.77, 1.34, 1.49, respectively, all P<0.05; radiotherapy-first group: HR were 1.02, 1.31, 1.66, 0.78, 1.37, 1.29, all P>0.05). The mean 5-year predicted mortality risk for all patients receiving radiotherapy-first regimen was lower than those receiving chemotherapy-first regimen (26.8% vs 30.2%, P<0.001). There were individualized differences in the predicted mortality risk of patients with different clinical characteristics who received radiotherapy-first regimen or chemotherapy-first regimen. Conclusion: Patients with stage Ⅰ/Ⅱ ENKTCL treated with radiotherapy-first regimen had a better expected prognosis than patients treated with chemotherapy-first regimen. The quantitative assessment of the differential effects of the sequence of radiotherapy and chemotherapy on the mortality risk of individual patients based on their clinical characteristics was helpful for the clinical development of the optimal radiotherapy and chemotherapy plan for each patient.

[A survey on human resources of dentists who deal with periodontal disease in Beijing].

Objective: To investigate the status quo of human resources of dentists who deal with periodontal disease in Beijing area through an online survey, which may hopefully provide a preliminary basis for the decision-making of administrative departments and the formulation of periodontal professional development plan. Methods: The dentists who deal with periodontal disease at least half a day per week in Beijing area were investigated. A questionnaire was designed by the chairman of the Periodontology Committee of Beijing Stomatological Association. The questionnaire was sent to and finished by the dentists via "WenJuanXing" online survey software. The contents of the survey included general condition, the property of practice unit, title and position of the dentist, membership of professional society, time and content of periodontal treatment, adoption of new technology and new method of periodontal therapy during the past one year, status of periodontal treatment in the local population and reasons, understanding and influencing factors of periodontal professional development. Results: A total of 1 255 dentists completed the survey, who came from all 16 districts in Beijing, mainly Haidian, Chaoyang, Dongcheng and Xicheng Districts [The total percentage of these four main districts was 70.3% (882/1 255)]. The mean age of the dentists was (36.1±8.3) years. Among the dentists, 71.1% (892/1 255) were females, 88.1% (1 106/1 255) got a Bachelor's degree or above. It was estimated that 35.4% (444/1 255) of the dentists had received standardized periodontal training ever. The percentage of dentists carrying out new technology in the past one year was as high as 68.1% (855/1 255). There were only 163 periodontal specialists (13.0%) out of the dentists in the survey. Only 15.9% (200/1 255) of the dentists routinely performed periodontal surgery. The majority of the dentists [82.8% (1 039/1 255)] were from the state-owned hospitals. Fifty-four point seven percemt (686/1 255) of the dentists thought that lack of knowledge was the main reason why the general public failed to receive periodontal treatment. As for the biggest bottleneck affecting periodontal professional development, fifty-one point zero percent (640/1 255) of the dentists attributed it to the public neglect. Conclusions: The periodontal practitioners in Beijing are young, highly educated, unevenly distributed in 16 districts and mostly females. State-owned oral health institutions are an important force in periodontal diagnosis and treatment services in Beijing. The number of periodontal specialists need to be improved. Promotion of standardized periodontal surgery and the popularization of healthcare knowledge on periodontal disease should also be the focus in the future.

[Outcome of radiotherapy for low-risk early-stage patients with extranodal NK/T-cell lymphoma, nasal-type].

Objective: To evaluate the prognosis and determine the failure patterns after radiotherapy for low-risk early-stage patients with extranodal NK/T-cell lymphoma, nasal-type (ENKTCL). Methods: A total of 557 patients from 2000-2015 with low-risk early-stage ENKTCL who received radiotherapy (RT) with or without chemotherapy (CT) from China Lymphoma Collaborative Group were retrospectively reviewed. Among them, 427 patients received combined modality therapy, whereas 130 patients received RT alone. Survivals were calculated by Kaplan-Meier method and compared with Log-rank test. Overall survival (OS) was compared with age and sex-matched general Chinese population using expected survival and standardized mortality ratio (SMR). Cox stepwise regression model was used for multivariate analysis. Results: The 5-year OS and progression-free survival (PFS) were 87.2% and 77.2%. The SMR was 3.59 (P<0.001) at 1 year after treatment, whereas it was 1.50 at 4 years after treatment, without significant difference between ENKTCL group and country-matched general population (P=0.146). Compared with RT alone, CMT did not result in significantly superior 5-year OS (87.0% vs 87.4%, P=0.961) or PFS (76.1% vs 80.7%, P=0.129). Local failure (11.5%, 64/557) and distant failure (10.8%, 60/557) were the main failure modes, while regional failure was rare (2.9%, 16/557). The 5-year locoregional control rate (LRC) was 87.2% for the whole group, with 89.5% for ≥50 Gy versus 73.7% for <50 Gy (P<0.001). Radiotherapy dose was an independent factor affecting LRC(P<0.05). Conclusions: Radiotherapy achieves a favorable prognosis in patients with low-risk early-stage ENKTCL. The incidence of either locoregional or distant failure is low. Radiation dose still is an important prognostic factor for LRC.

[Clinical features and treatment outcome of extranodal nasal-type NK/T-cell lymphoma of the extra-upper aerodigestive tract].

Objective: To investigate the clinical features and prognosis of extranodal nasal-type NK/T-cell lymphoma of the extra-upper aerodigestive tract (extra-UADT NKTCL). Methods: The clinical data of 159 patients with extra-UADT NKTCL from the China Lymphoma Collaborative Group (CLCG) database between November 2001 and December 2015 were retrospectively analyzed. Kaplan-Meier survival analysis and Log-rank test were used to evaluate the prognosis. The Cox regression model is used for multi-factor analysis. Results: Extra-UADT NKTCL commonly occurs in skin and soft tissues (106/159, 66.7%) and gastrointestinal tract (31/159, 19.5%). The incidences of elevated lactate dehydrogenase (LDH) and Ann Arbor Ⅲ~Ⅳ stage were 47.8% (76/159) and 64.2% (102/159), respectively. The 3-year overall survival (OS) and progression-free survival (PFS) rates were 43.6% and 27.9%, respectively. The corresponding OS rates of primary skin/soft tissue site and gastrointestinal tract site were 41.0% and 59.4% (P=0.281), while the PFS rates were 24.8% and 48.3%, respectively (P=0.109). Combined modality treatment improved the 3-year OS of all the patients (58.4% vs 33.9%, P=0.001) and 3-year PFS (40.7% vs 20.7%, P=0.008) when compared with chemotherapy alone. LDH elevation, Ann Arbor synthesising and ≥2 junction external bits were intrusive as independent risk factors for total survival (P<0.05), LDH elevation and ≥2 junction outer bits were intrusive as independent risk factors for progressionless survival(P<0.05). The distant extranodal dissemination was the primary failure patterns. Conclusions: Extra-UADT NKTCL appears to have distinct clinical characteristics and poor outcome. Compared with chemotherapy alone, combined modality treatment may improve the prognosis of patients with extra-UADT NKTCL.

Energy value of rice, broken rice, and rice bran for broiler chickens by the regression method.

The objective of this study was to determine the ileal digestible energy (IDE), ME, and MEn of rice, broken rice, and rice bran. The birds were fed a standard starter diet from day 0 to 14 and experimental diets from day 15 to 21 after hatching. A total of 336 birds were grouped by BW and assigned to 7 diets, each diet comprised 8 replicates with 6 birds per replicate. The diets comprised a reference diet (RD) and 6 test diets (TD). The TD contained 2 levels of rice, broken rice or rice bran that partly replaced the energy sources in the RD at 120 or 240 g/kg (rice and broken rice) or 150 or 300 g/kg (rice bran). Addition of rice or broken rice to RD linearly increased (P < 0.01) ileal digestibility of DM, energy, as well as total tract metabolizability of DM, energy, and N-corrected energy in the TD. The inclusion of rice bran in the TD linearly decreased (P < 0.01) energy digestibility and utilization in the test diet. Regressions of rice-associated, broken rice-associated, or rice bran-associated IDE, ME, or MEn intake in kcal against rice, broken rice, or rice bran intake were as follows: IDE = Y = 2 (6) + 3,185 (73) × Rice + 3,199 (72) × Broken Rice + 2,562 (61) × Rice Bran, r2 = 0.98; ME = Y = 8 (6) + 3,103 (72) × Rice + 3,190 (71) × Broken Rice + 2,709 (60) × Rice Bran, r2 = 0.98; MEn = Y = 4 (5) + 3,014 (68) × Rice + 3,092 (101) × Broken Rice + 2,624 (57) × Rice Bran, r2 = 0.98; Based on the regression equations, the IDE, ME, MEn values (kcal/kg of DM) of rice were 3,185, 3,103 and 3,014, respectively, while for broken rice, the values were 3,199, 3,190, and 3,092 and for rice bran, the values were 2,562, 2,709, and 2,624, respectively.

[The relationship between HPV integration and prognosis of cervical cancer].

Objective: To investigate the relationship between human papillomavirus (HPV) integration and prognosis of cervical cancer patients. Methods: The data of 82 patients with cervical cancer treated in the Radiotherapy Department of Peking Union Medical College Hospital from October 2004 to June 2012 were retrospectively analyzed.The patients were divided into poor prognosis group (recurrence or metastasis after surgery and adjuvant radiotherapy) and good prognosis group based on a propensity score matching strategy.The HPV integration of the two groups were detected by whole exome sequencing to determine whether the integration sites were located in the common fragile sites (CFSs). HPV integration and integration into CFSs were compared between the two groups. Results: Among the enrolled 82 patients, 37 were divided in poor survival group and 45 in good survival group. A total of 90 integration breakpoints were identified, 30 of them occurred in poor prognosis group and 60 occurred in good prognosis group. In the poor prognosis group, HPV integration occurred in 20 patients, 13 of them were inserted in CFSs of 11 patients, and the numbers in good prognosis group were 26, 17, 11, respectively. There were no significantly statistical differences in the number of HPV integration events (P=0.289), HPV integration patients (P=0.735), CFSs integration events (P=0.427), and CFSs integration patients (P=0.591) between the two groups. In poor prognosis group, more CFSs integration events occurred in patients with metastasis than those in patients with only local recurrence (9 vs 2, P=0.003). Conclusions: No significant differences are observed in HPV integration and HPV integration into CFSs between cervical cancer patients with different prognoses. HPV integration into CFSs may be associated with distant metastasis.

[Clinical characteristics and prognostic analysis of 458 children with high-risk neuroblastoma in a single center].

Objective: To summarize the clinical characteristics of high-risk neuroblastoma (HR-NB) in a single center, analyze the prognostic factors of HR-NB. Methods: The clinical data of children with HR-NB who were treated and followed up at the hematology-oncology center of Beijing Children's Hospital from February 1, 2007 to June 30, 2018 were analyzed retrospectively. The clinical features were summarized. Kaplan-Meier method was used for survival analysis and Cox regression was used to analyze the prognostic factors. The last follow-up time was June 30, 2019. Results: A total of 458 children with HR-NB were enrolled in this study, including 265 males (57.9%) and 193 females (42.1%), the age at diagnosis was 40.0 months (4.5-148.0 months), the follow-up time was 22.0 months (0.2-138.0 months) and the time of tumor progression or recurrence was 15 months (1-72 months). The 5-year event-free survival (EFS) rate was (31.2±2.6)% and the 5-year overall survival (OS) rate was (43.9±3.2)%. The 5-year EFS rate and 5-year OS rate in 142 hematopoietic stem cell transplantation (HSCT) patients with bone marrow metastases were better than that in 196 non-transplantation cases with bone marrow metastases ((26.5±4.5)% vs. (25.1±3.6)%, χ²=13.773, P=0.001; (38.1±5.5)% vs. (35.7±4.7)%, χ²=9.235, P=0.002); 128 transplantation patients with bone metastases had higher 5-year EFS rate and 5-year OS rate than 188 non-transplantation cases with bone metastases ((28.5±5.0)% vs. (26.7±3.8)%, χ²=10.222, P=0.001; (37.1±6.0)% vs. (36.2±4.8)%, χ²=7.843, P=0.005). The 5-year EFS rate was higher in 37 HSCT patients with MYCN amplification than in 49 non-transplantation cases with MYCN amplification ((26.8±8.0) % vs. (20.5±6.4) %, χ²=5.732, P=0.017). No significant difference was found in 5-years OS rate between transplantation group with MYCN amplification and non-transplantation group with MYCN amplification ((31.4±8.6) % vs. (26.2±7.4) %, χ²=3.230, P=0.072). Univariate survival analysis showed that lactate dehydrogenase (LDH)≥1 500 U/L was associated with poor prognosis of patients with MYCN amplification (χ²=6.960, P=0.008). Multivariate Cox analysis showed bone marrow metastasis and LDH≥1 500 U/L were independent risk factors for poor prognosis of patients with non-MYCN amplification (HR=2.427, 1.618;95%CI：1.427-4.126, 1.275-2.054, P<0.05) for both comparisons. Conclusions: LDH≥1 500 U/L was the poor prognostic factor for patients with MYCN amplification. The bone marrow metastasis and LDH≥1 500 U/L were the poor prognostic factors for HR-NB patients with non-MYCN amplification. HSCT can improve the prognosis of patients with bone or bone marrow metastasis. It can also retard the time of progression or recurrence for patients with MYCN amplification.

[Progress of researches on global prevalence of Blastocystis hominis human infections and its subtypes].

Blastocystis hominis is a parasite that parasitizes in the intestines of humans and animals, and is closely related to a variety of gastrointestinal diseases such as abdominal pain and diarrhea. B. hominis is distributed worldwide, and the prevalence of B. hominis infections and dominant subgenotypes vary in countries and in regions from the same country. This paper reviews the global prevalence of B. hominis human infections, its subtypes and geographical distribution, so as to provide insights into the understanding of the global epidemiology of B. hominis and the management of B. hominis infections.

[Progress of researches on animal models of Blastocystis hominis infections].

Blastocystis hominis is a common parasitic protozoa in human and animal intestines; however, its pathogenicity remains controversial. Construction of animal models is of great significance to investigate the pathogenicity, pathogenic mechanisms and drug screening of B. hominis. Experimental animals, mode of infections, parasite strains and host immune status are important factors affecting the successful modeling of B. hominis infections in animals. Hereby, we review the progress of researches on animal models of B. hominis infections, and summarize the influencing factors and application of animal models of B. hominis infections, in order to provides insights into the selection of animals models of B. hominis infections.

Antidepressant Drugs Correct the Imbalance Between proBDNF/p75NTR/Sortilin and Mature BDNF/TrkB in the Brain of Mice with Chronic Stress.

Depression is a worldwide problem with a great social and economic burden in many countries. In our previous research, we found that the expression of proBDNF/p75NTR/sortilin is upregulated in patients with major depressive disorder. In addition, the treatment of proBDNF antibodies reversed both the depressive behaviors and the reduced BDNF mRNA detected in our rodent chronic stress models. Antidepressant drugs are usually only effective in a subpopulation of patients with major depression with a delayed time window of 2-4 weeks to exert their efficacy. The mechanism underlying such delayed response is not known. In this study, we hypothesize that antidepressant drugs exert their therapeutic effect by modulating proBDNF/p75NTR and mature BDNF/TrkB signaling pathways. To test the hypothesis, C57 mice were randomly divided into normal control, chronic unpredictable mild stress (CUMS), vehicle (VEH), fluoxetine (FLU), and clozapine (CLO) groups. Behavioral tests (sucrose preference, open field, and tail suspension tests) were performed before and after 4 weeks of CUMS. The gene and protein expression of proBDNF, the neurotrophin receptor (p75NTR), sortilin, and TrkB in the cortex and hippocampus were examined. At the protein level, CUMS induced a significant increase in proBDNF, p75NTR, and sortilin production while the TrkB protein level was found to be lower in the cortex and hippocampus compared with the control group. Consistently, at the mRNA level, p75NTR expression increased with reduced BDNF/TrkB mRNA in both cortex and hippocampus, while sortilin increased only in the hippocampus after CUMS. FLU and CLO treatments of CUMS mice reversed all protein and mRNA expression of the biomarkers in both cortex and hippocampus, except for sortilin mRNA in the cortex and proBDNF in the hippocampus, respectively. This study further confirms that the imbalance between proBDNF/p75NTR/sortilin and mBDNF/TrkB production is important in the pathogenesis of depression. It is likely that antidepressant FLU and antipsychotic CLO exert their antidepressant-like effect correcting the imbalance between proBDNF/p75NTR/sortilin and mBDNF/TrkB.

[Characteristics and outcomes of primary angiosarcoma].

Objective: The study was designed to analyze the clinicopanthologic characteristics, treatments and outcomes of a series of patients with primary angiosarcoma. Methods: The clinical, surgical and pathological data and treatment of 68 patients with pathologically confirmed angiosarcoma admitted to Peking Union Medical College Hospital from January 1990 to June 2017 was retrospectively analyzed. Kaplan-Meier method and Log rank test were used for univariate survival analysis and Cox regression model was used for multivariate survival analysis. Results: A total of 68 patients were enrolled, 38 were male, 30 were female. The median age at diagnosis was 50.5 years. The time from symptom onset to diagnosis was (7.5±7.5) months. The primary sites included face and scalp, breast, chest wall, lung, heart, liver, spleen, extremities, bones and so on. At diagnosis, the mean size of tumors were (7.4±7.3) cm, 28 patients (41.2%) had localized disease (stage Ⅰ+ Ⅱ) and 40 patients had metastatic disease (stage Ⅲ+ Ⅳ). There were 37 patients treated with surgery alone, three receiving radiotherapy alone, five receiving chemotherapy alone and sixteen receiving comprehensive treatment with 5 underwent surgery plus radiotherapy, three treated by surgery plus chemotherapy, four had surgery plus interventional therapy, two had chemoradiotherapy, one had radiotherapy and interventional therapy and 1 had surgery plus chemoradiotherapy and targeted therapy. Five patients received only palliative treatment, and 2 patients lost follow-up after diagnosed. Fifty patients were followed up with a median overall survival time of 8.5 months. The median survival time of patients with metastatic angiosarcoma was 6.6 months, significantly shorter than that of patients with localized disease (15.0 months, P=0.020). The median survival time of patients with cardiac angiosarcoma was 3.0 months, significantly shorter than that of patients with angiosarcoma at other sites (11.5 months, P=0.010). The median survival time of patients receiving comprehensive treatment was 31.0 months, significantly longer than that of patients without comprehensive treatment (5.6 months, P=0.007). Multivariate analysis showed that staging, heart occurrence and comprehensive treatment were independent factors for the prognosis of primary angiosarcoma (all P<0.05). Conclusions: Angiosarcoma is a rare malignancy, and patients with metastatic disease or cardiac occurence have poor prognosis. Comprehensive treatment can improve the prognosis of patients with angiosarcoma.

MiR-27a alleviates osteoarthritis in rabbits via inhibiting inflammation.

OBJECTIVE: To investigate the regulatory effect of micro ribonucleic acid-27a (miR-27a) on the nuclear factor-kappa B (NF-κB) pathway and to explore its effect on rabbits with osteoarthritis (OA). MATERIALS AND METHODS: Anterior cruciate ligament (ACL) cross-section method was adopted to establish OA rabbit models. Cartilage specimens were collected to detect expression levels of miR-27a in OA cartilage and normal cartilage tissues. Meanwhile, chondrocytes were isolated and cultured, and transfected with miR-27a mimics and miR-27a inhibitor. Blank control group was set up. Next, the changes in chondrocyte proliferation were detected using 5-ethynyl-2'-deoxyuridine (EdU) staining and cell counting kit-8 (CCK-8). Quantitative Real Time Polymerase Chain Reaction (PCR) was applied to detect the messenger RNA (mRNA) expression of inflammatory factors interleukin 6 (IL-6) and tumor necrosis factor-α (TNF-α) in chondrocytes. Also, Western blot was adopted to detect the differential expression of NF-κB pathway-related proteins NF-κB and matrix metalloproteinase 13 (MMP-13). RESULTS: Compared with that in normal cartilage tissues, miR-27a in OA cartilage tissues was decreased evidently (p<0.05). The expression level of miR-27a was higher in miR-27a mimics group than in control group, while it significantly declined in miR-27a inhibitor group (p<0.05). EdU staining and CCK-8 method results showed that miR-27a mimics could promote the proliferation of chondrocytes, while miR-27a inhibitor inhibited the proliferation of chondrocytes. Compared with those in control group, the expression levels of inflammatory factors TNF-α and IL-6 in chondrocytes in miR-27a inhibitor group were increased significantly (p<0.05). MiR-27a mimics could evidently reduce the expression of inflammatory factor IL-6 (p<0.05), but did not significantly reduce the expression of TNF-α. Besides, the results of Western blot suggested that the expression levels of MMP-13 and NF-κB proteins were decreased significantly in miR-27a mimics group (p<0.05) and increased significantly in miR-27a inhibitor group (p<0.05). CONCLUSIONS: MiR-27a in OA cartilage tissues is evidently lower than in normal cartilage tissues. Transfection of miR-27a mimics can promote proliferation of chondrocytes, lower the expression of inflammatory factors, and reduce the expression of MMP-13 and NF-κB proteins. Therefore, the up-regulation of miR-27a can benefit the treatment of bone joints through the NF-κB pathway.

[Clinical analysis of right atrial appendage aneurysms complicated by atrial tachyarrhythmias in three children].

Objective: To investigate the clinical features and results of treatment for right atrial appendage aneurysms complicated by atrial tachyarrhythmias in children. Methods: This retrospective study included three children with right atrial appendage aneurysm complicated by atrial tachyarrhythmias (aged 1.0 to 5.3 years, weight 10 to 17.1 kg) who were hospitalized at the Pediatric Cardiology Department (Heart Center) of the First Hospital of Tsinghua University (Beijing Huaxin Hospital) during the period from January 2016 through April 2018. The patients' clinical features, the results of electrocardiogram (ECG) and echocardiography, the effects of therapeutic interventions (antiarrhythmics, radiofrequency ablation, and surgical resection of right atrial appendage aneurysm) and the results of pathological assessment were analyzed. Results: Three cases of right atrial appendage aneurysm were diagnosed at 36 weeks of gestational age, 1 month and 4 months after birth respectively. In two cases, ECG showed alternating episodes of atrial tachyarrhythmias including atrial tachycardia, atrial flutter, and atrial fibrillation, and echocardiography showed aneurysmal dilatation of right atrial appendage. These two cases underwent right atrial appendage aneurysm resection. In the remaining one case of atrial tachycardia, echocardiography did not visualize important lesions in the right atrium, thus the intracardiac electrophysiologic study and radiofrequency ablation were performed; and focal atrial tachycardia originating from the apex of right atrial appendage was mapped but failed to be ablated; consequently, the patient received the right atrial appendage resection, in which the right atrial appendage aneurysm was found. Preoperative multiple antiarrhythmics showed only modest or no efficacy for all the three cases. The atrial tachyarrhythmias disappeared in all the three cases after right atrial appendage aneurysm resection. Postoperative atrial tachycardias associated with new foci of impulse formation developed in two cases. These two patients reverted to normal sinus rhythm and remained in this rhythm by using antiarrhythmics. Pathological assessment showed cystic dilation of parts of atrial cavity, fibrosis of cyst wall, generalized fibrosis of atrial myocardium combined with myocardial atrophy and cystic dilation, as well as uneven myocardial thickness with generalized myocardial interstitial fibrosis. Conclusions: For patients with congenital right atrial appendage aneurysm, atrial tachyarrhythmias might develop during fetal stage or early postpartum period. Reliance on echocardiography might often lead to the missed diagnosis. These patients with atrial tachyarrhythmias responded poorly to antiarrhythmics. Radiofrequency ablation might be associated with a high risk and limited efficacy. Surgical resection of right atrial appendage aneurysm showed satisfactory results and should be highly recommended.

[Survival and prognostic factors analysis of primitive neuroectodermal tumor].

Objective: To investigate the clinical features and prognosis of primitive neuroectodermal tumor (PNET). Methods: The clinical data of 99 patients with PNET from February 1, 1998 to February 1, 2017 were retrospectively analyzed. Univariate analysis was performed using Kaplan-Meier and Log rank test. Multiviate Cox regression was applied to analyzed independent prognostic factor for patient survival. Results: Among the 99 patients, 81 were peripheral PNET(pPNET) and 18 were central PNET (cPNET) . Biopsy was performed exclusively in 16 cases, with R0 resection in 61 cases, with R1 resection in 4 cases, and with R2 resection in 18 cases. Twelve patients underwent surgery only, nine had chemotherapy only, and one received radiotherapy only. There were 72 patients who had combined treatment including chemotherapy, and 48 patients had combined therapy including radiotherapy. The one-year, three-year and five-year overall survival(OS) rates of the 99 PNET patients were 79.2%, 63.9% and 56.1% respectively, and median OS time was 14.0 months. The one-year, three-year and five-year progression free survival (PFS) rates of these patients were 42.7%, 25.7% and 19.8% respectively, and median PFS time was 8.0 months. The univariate analysis revealed that lymph node metastasis, surgical resection, numbers of cycles of chemotherapy and radiotherapy dose were the main factors affecting the OS (all P<0.05). Gender, age, lymph node metastasis, staging, and chemotherapy cycles were related to PFS in PNET patients (P<0.05). Multivariate analysis showed that the degree of surgical resection, chemotherapy cycle, and radiotherapy dosage were independent influencing factors of OS in PNET patients (risk ratio=1.856, 0.398, and 0.408, respectively, all P<0.05), and gender was an independent factor influencing PFS in PNET patients (risk ratio=0.494, P<0.05). Conclusions: Comprehensive therapy is the main therapy for PNET patients. The surgical resection, cycles of chemotherapy and radiotherapy dosage are independent prognostic factors for patient's OS.

Effects of continuous low dose infusion of lipopolysaccharide on inflammatory responses, milk production and milk quality in dairy cows.

The objective of this study was to evaluate the effects of continuous low dose infusion of lipopolysaccharide (LPS) on inflammatory responses and milk production and quality in lactating dairy cows. Eight Holstein cows were assigned to two treatments in a cross-over experimental design. Cows were infused intravenously either with saline solution or with saline solution containing LPS from Escherichia coli O111:B4 at a dose of 0.01 µg LPS/kg body weight for approximately 6 hr each day during a seven-day trial. The clinical symptoms and milk production performance were observed. Milk samples were analysed for conventional components, fatty acids and amino acids. And jugular vein and mammary vein plasma samples were analysed for concentrations of cytokines and acute phase proteins. LPS infusion decreased feed intake and milk yield. An increase in body temperature was observed after LPS infusion. LPS infusion also increased plasma concentrations of interleukin-1ß, serum amyloid A, LPS-binding protein, C-reactive protein and haptoglobin. LPS infusion decreased the contents of some fatty acids, such as C17:1, C18:0, C18:1n9 (trans) and C18:2n6 (trans), and most amino acids except for methionine, threonine, histidine, cysteine, tyrosine and proline in the milk. The results indicated that a continued low dose infusion of LPS can induce an inflammatory response, decrease milk production and reduce milk quality.

[Application of negative molds technology based on three-dimensional printing in digital maxillofacial prostheses].

Objective: To explore a digital negative molds technique based on three-dimensional (3D) printing to assist in the manufacture of maxillofacial prostheses, and to improve the deficiency of the current clinical treatment. Methods: Seventeen patients with maxillofacial defects (including nasal defects, orbital defects, cheek defects, auricle defect) were scanned by means of facial optical scanning and computer tomography (CT). The 3D models were then reconstructed and global registration was made to merge the reconstructed models into a new digital model for 3D design. The 3D design of the prostheses was implemented in software. The mechanical connection structure was designed by forward engineering technology for 3 patients with intra-oral defects in maxilla who needed to make removable partial dentures, so that the silicone prostheses and removable partial denture could be combined. The removable partial dentures were made by conventional method and connected with the prostheses. According to the 3D data of the prostheses, the digital negative molds were designed, and the 3D printing technology was used to finish the processing of the resin molds. Silicone for prostheses were filled and cured in the resin molds to fabricate the clinical restorations for the patients. The margin adaptation and retention of the prostheses was detected. Results: Twenty patients with varying degrees of maxillofacial defects were rehabilitated using the courses developed in the study. All patients reported no pain or discomfort during the treatment; and they were satisfied with the final prostheses of the shape, color, retention, stability, etc. Eighteen of the prostheses showed good marginal adaptation, and sixteen of the prostheses showed good retention effect. Conclusions: The digital negative molds technique used in this study could greatly reduce the intensity of manual operation and provided a good therapeutic effect for patients with maxillofacial defects.

[Quantity of Treg cells and Th17 cells in spleen of primary immune thrombocytopenic purpura patients].

Objective: To observe the quantity of Treg cells and Th17 cells in spleen of adult primary immune thrombocytopenic purpura (ITP) patients. Methods: 43 ITP cases with splenectomy treatment were enrolled from December 2008 to June 2016 at Union Hospital of Fujian Medical University, including 20 males and 23 females with a median age of 36 (18-76) years. The controls were thirty patients who underwent splenectomy because of pancreatic diseases or splenic impairment, including 21 males and 9 females with a median age of 47 (21-69) years. The quantity and ratio of Treg cells and Th17 cells were examined by immunohistochemistry between ITP patients and controls. Results: ①The quantity of Treg cells in ITP were less than controls[ (11.3±4.7) /mm(2) vs (59.0±15.0) /mm(2), t=-22.894, P<0.001], but Th17 cells were more than controls[ (235.2±69.4) /mm(2) vs (181.1±23.7) /mm(2), t=13.768, P<0.001]. So the ratio of Treg/Th17 in ITP was lower than controls (0.048±0.027 vs 0.328±0.086, t=19.522, P<0.001) . ② The quantity of Treg cells in cases without response after splenectomy were less than cases with response[ (9.5±5.0) /mm(2) vs (11.6±4.7) /mm(2), t=2.723, P=0.010], and there is no statistical differences between the two groups about the quantity of Th17 cells and the ratio of Treg/Th17 cells[ (232.3±80.8) /mm(2) vs (239.6±66.9) /mm(2), t=1.108, P=0.277; 0.040±0.024 vs 0.053±0.027, t=0.540, P=0.592]. Conclusions: There is a significant difference about the quantity of Treg cells and Th17 cells in spleen between ITP patients and healthy controls, and they are relevant to the response after splenectomy.

Milk protein responses to balanced amino acid and removal of Leucine and Arginine supplied from jugular-infused amino acid mixture in lactating dairy cows.

This study was undertaken to evaluate the milk protein response when cows were supplied a balanced AA profile and to determine whether a deficiency of Leucine (Leu) or Arginine (Arg) had a negative effect on milk protein. Eight mid-lactation Holstein cows were randomly assigned to 5-day continuous jugular infusions of saline (CTL), EAA mixture prepared on the profile of casein and supplied (in % of lysine (Lys)) 100% of Lys, 33.3% of methionine (Met), 110.2% of Leu, 43.6% of Arg, 50.8% of threonine (Thr), 81.6% of valine (Val), 69.7% of isoleucine (Ile), 61.4% of phenylalanine (Phe) and 34.2% of histidine (His) (Casein, 160 g/d), EAA mixture excluding Leu (-Leu, 163 g/d) or EAA mixture excluding Arg (-Arg, 158 g/d) in a duplicated 4 × 4 Latin square design with four infusion periods separated by 7-day interval period. The basal diet supplied 1.6 Mcal NEL and 94.4 g MP per 1 kg DM to meet requirements for lactation. The Casein treatment provided a balanced supply (in % of MP) of 10.3% Leu and 5.3% Arg, whereas in the two subsequent -Leu and -Arg treatments, the concentration of Leu and Arg was reduced to 8.4 and 4.6% respectively. Dry matter intake (15.4 kg/day) was not affected by treatments. The Casein treatment increased milk yield (14.9%, p < 0.001), milk protein yield (120 g, p < 0.001) and milk protein efficiency (0.03, p = 0.099) than CTL treatment. However, the -Leu treatment decreased the responses of above-measured parameters by 6.25%, 70 g, 0.05 (p < 0.06) (compared with Casein). These effects of Leu were related to decreased Leu concentration and improved concentration of Ile and Val in plasma. The -Arg treatment decreased the plasma Arg concentration than the Casein treatment, whereby resulted in the decrease of milk yield (5.7%, p = 0.073), milk protein yield (60 g, p = 0.011) and milk protein efficiency (0.04, p = 0.037). In conclusion, supply of EAA profile of casein can increase the lactation production in dairy cows, and 8.6% of Leu in MP partly limits the milk protein response when the requirements of Lys, Met and His were met. The level of Arg at 4.6% MP is not deemed to an ideal profile, as evidenced by decreased milk protein efficiency.