Objective: To study the influence of inserting glycines(Gly) on biological properties of HIV Tat-(Gly)n-thymidine kinase (-TK) fusion proteins. Methods: Different fragments containing 0, 2, 4 or 6 Gly were inserted between the HIV Tat gene and TK using gene splicing by overlap extension (SOEing) PCR, and the products were cloned into PBK vector. The vectors were then transferred into E. coli after sequencing. After IPTG induction, bacilli were collected and destructed by ultrasound; the fusion protein was collected and identified by monoclonal antibody of HIV protein. HepG2 cells were incubated with DMEM supplemented with 1 μg/ml fusion protein containing 0, 2, 4 or 6 Gly for 24 h. HepG2 cells of different groups were detected by immunofluoreseence assay with HIV Tat monoclonal antibody; the apoptosis rate of HepG2 cells was determined by cell flow cytometry after they were incubated with gencilovir (10 μg/ml) for 3 d and the survival rate of cells was recorded by trypan blue in different groups. Results: The recombined genes containing 0, 2, 4 or 6 Gly were successfully constructed, inserted into PBK vectors, and expressed into E. coli. Their proteins were obtained and purified. The level of fluorescence in different groups was similiar, but the cell survival rate and apoptosis rate were different. The highest apoptosis rate was 14.77%, which was found in the group containing 4 Gly, followed by 12.69% in 2 Gly group, 8.31% in HIV Tat-TK group, 4.36% in 6 Gly group, and 1.0% in group containing no Gly. Significant differences were found between each 2 groups (P<0.05). Trypan blue showed similar results in the cell death rate of different groups: the highest cell death rate was 80.2%, which was found in the group containing 4 Gly, followed by 65.4% in 2 Gly group, 58.4% in HIV Tat-TK group, 56.7% in 6 Gly group, and 9.1% in the group containing no Gly. Conclusion: The number of Gly inserted into HIV Tat-TK protein does not alter the transcellular function of upstream Tat protein, but does substantially influence the TK protein-mediated cytoxic effects of gencilovir, and the influence is the smallest when 4 Gly are inserted.

Objective To investigate of the radiological manifestations in congenital cystic adenomatoid malformation (CCAM) of lung in adult and to improve the diagnostic accuracy of CCAM of lung in adult.Methods Five cases with pathologically proved CCAM of lung in adult were retrospectively analyzed.Chest X-ray was available in 5 cases and chest CT was performed in 2 cases.Results On plain chest radiography,thin wall air cystic lesions with air-fluid level were detected in 3 cases.Honeycomb like small cystic lesion was detected in 1 case.Multiple round cystic lesions were found in 1 case.CT scan of the chest demonstrated a round thin wall air cystic lesion in the lateral segment of right lung's middle lobe,and a thin wall air cystic lesion with the wall merged into the shadow of chest wall in the left apicoposterior segment in one case.Small cystic lesions just like honeycomb were found in bilateral basal segments of the inferior lobes,with a 0.8—1.0 cm sized round mass revealed in the right lung′s inferior lobe basal segment, and the mass was spiculated in another case.Conclusion The imaging signs of CCAM of lung in adult is cyst or cyst-solid and at the risk of developing carcinoma.

One leading research target of modern tumor radiotherapy is to increase radiosensitization of tumor and improve curative effect of radiotherapy.Histone deacetylase inhibitors are epigenetic drugs that can play a part in radiosensitization through means of induction of apoptosis,inhibition of repair of DNA doublestrand breaks,cell cycle arrest,improvement of tumor cell hypoxia and increase of reactive oxygen species.There is an urgent need to develop biomarkers based on these pathways,which can promote clinically individualized treatment.

AIM: To study the effect of bioactive compounds from Entomogenous fungi (BCEF0083), on AVP content of hypothalamus, pituitary and expression of glucocorticoid receptor (GR) mRNA of hippocampus in chronic unpredictable stress model of depression in rats. METHODS: The depression animal model was induced by chronic unpredictable stress. The effect of BCEF0083 on AVP content in hypothalamus and pituitary was tested by radioimmunoassay. RT-PCR was used to test the expression of GR mRNA in hippocampus. RESULTS: BCEF0083 decreased the AVP content of hypothalamus and pituitary in chronic unpredictable stressed rats, and increased the expression of GR mRNA of hippocampus in chronic stressed rats. CONCLUSION: BCEF0083 exhibits antidepressant effect by attenuating the stimulated function of hypothalamus-pituitary-adrenal axis in chronic unpredictable stress model of depression in rats.

Objective:To analyze the clinical features and genetic factors of neonatal 17β-hydroxysteroid dehydrogenase type10 (HSD10) deficiency.Methods:The clinical characteristics and genetic test results of a child with HSD10 deficiency coming from Children′s Hospital of Nanjing Medical University in April 2019 were retrospectively analyzed.The keywords" 17β-hydroxysteroid dehydrogenase type 10 deficiency" or " 2-Methyl3-Hydroxybutyryl-CoA dehydrogenase deficiency" or " HSD10" , etc.were searched in various databases, including CNKI, Wanfang, Weipu, Embase and PubMed to review the cases collected from all published data until May 31, 2020.Results:The patient was a newborn male who developed symptoms on the first day after birth.The main signs were metabolic acidosis, increased blood ammonia and lactate, and hypotonia.Trio whole exom sequencing in the patient and his parents identified hemizygous NM001037811: c.650G>A, p.R217Q in the HSD17B10 gene that is inherited from the mother.Since the child died on the third day after birth, no further central nervous system examination was performed.The mother of the child has intellectual disability, the sibling sister is normal and the HSD17B10 locus is wild type.By lite-rature reviewing, 5 newborn cases with clear medical records and genetic test results were listed.All patients were male, and had onset of HSD10 deficiency within 1 week after birth.The main phenotypes include metabolic acidosis (increased blood ammonia and lactate), hypoglycemia, hypotonia, and convulsions.All 6 children died in early infancy.The corresponsive HSD17B10 variants were c. 740A>G/p.N247S, c.677G>A/p.R226Q, c.257A>G/p.D86G and c. 650G>A/p.R217Q, which did not indicate the hot spots of mutation. Conclusions:HSD10 deficiency in the neonatal period is relatively rare.The clinical diagnosis is difficult due to the serious condition and short course of the disease.Severe metabolic acidosis, hypotonia, and convulsions in neonatal patients are the main reasons for the poor prognosis, which can be attributed to the hemizygous variation and heterogeneity of the mutation site in male patients.c.650G>A may be closely associated with severe neonatal HSD10 deficiency, but the molecular biological mechanism needs to be further clarified.HSD10 deficiency has a poor prognosis and lacks effective treatment.

OBJECTIVETo observe the inhibitory effect of a eukaryotic expression vector expressing human IFN-gamma (pcDNA3.1- IFN-gamma) on HBV replication in hepG2.2.15 cells.

METHODSThe eukaryotic expression vector expressing human IFN-gamma was constructed using PCR and gene recombination technique. hepG2.2.15 cells were transfected with pcDNA3.1-IFN-gamma and the culture supernatant was collected to determine the expression of IFN-gamma protein by ELISA. The HBV DNA copies and the concentration of HBeAg and HBsAg were measured by fluorescence real-time PCR and ELISA kit, respectively.

RESULTSCompared with that of negative control and blank 2.2.15 cells, the concentration of HBeAg in the supernatant of 2.2.15 cells transfected with pcDNA3.1- IFN-gamma were decreased by 49%, and HBsAg concentration was lowered by 35% and 33%, respectively. A significant decrease of HBV DNA copies was observed in pcDNA3.1- IFN-gamma-transfected cells in comparison with the two control cells. No significant differences were noted in all the results between the two control groups.

CONCLUSIONWe have successfully constructed the eukaryotic expression vector expressing human IFN-gamma, which provides a basis for anti-HBV gene therapy using human IFN-gamma.

Genetic Vectors ; Hep G2 Cells ; Hepatitis B virus ; drug effects ; Humans ; Interferon-gamma ; genetics ; pharmacology ; Transfection ; Virus Replication ; drug effects

Aim To explore the differences in hyper-trophic marker genes such as atrial natriuretic peptide ( ANP) , brain natriuretic peptide ( BNP) and β-myo-sin heavy chain (β-MHC) genes in different models of cardiac hypertrophy. Methods Respectively using re-nal abdominal aortic coarctation ( AAC) , arteriovenous fistula ( AVF) and isoproterenol ( ISO) methods to es-tablish C57BL/6 mouse model of cardiac hypertrophy. After modeling, each mouse ’ s body weight ( BW ) , heart weight ( HW) and left ventricular weight ( LVW) were weighed, and the heart weight ( HW/BW) and left ventricular index ( LVW/BW ) were calculated;myocardium by HE staining, pathological morphologi-cal changes were observed; myocardium by immuno-histochemistry, ANP, BNP and β-MHC protein ex-pression was observed;myocardium by Real-time PCR detection, ANP, BNP and β-MHC mRNA expression was observed. Results Compared with control group, HW/BW and LVW/BW were increased in three mod-els. Through the light microscope, each mouse model showed varying degrees of cardiac hypertrophy. ANP, BNP and β-MHC were increased in the protein and mRNA expression. Compared with AAC group, AVF and ISO groups’ myocardial tissue ANP, BNP and β-MHC expression were decreased in the protein and mRNA expression. Conclusions Three cardiac hy-pertrophy models are successful. Cardiac tissue ANP, BNP and β-MHC expression in AAC model exceeds AVF and ISO model.

Objective To explore the clinical effect of anteromedial thigh perforator flap for repairing soft tissue defect in lower extremity.Methods From January 2009 to December 2012,12 patients with soft tissue defect of lower extremity were treated with free anteromedial thigh perforator flap.Among them,there were 5 cases of wound in front of tibia,4 cases of dorsal foot wound,3 cases of heel wound;the wound size was 3.0 cm ×3.5 cm-7.0 cm × 9.5 cm;3 cases were repaired with free perforator flap of descending genicular artery,2 cases with free anteromedial thigh perforator flap,and 7 cases with medial vastus muscle perforator flap.Results All the transplants survived.Among them,1 case of anteromedial thigh perforator flap and 1 case of descending genicular artery perforator flap appeared vascular crisis postoperative.After anticoagulation and antispasmodic treatment aggressively,the flaps survived.One case of descending genicular artery perforator flap necrosis in edge,and then were cured by changing dressing.Twelve cases were followed up 6-12 months after operation (average 8.6 months),and the texture of flaps were good,the appearance appropriate ;7 sensation of 5 free flaps after nerve anastomosis came back to S; the donor sites in all cases were sutured directly and healed,leaving only the linear scar,and no effect on function.Conclusion The anatomy of anteromedial thigh perforator flap is constant,and the operation is convenient.The clinical result of repairing soft tissue defect of lower extremity is satisfying.

Background and purpose:It has a signiifcant effect for erlotinib on treatment of patients with epidermal growth factor receptor mutation in advanced non-small cell lung cancer (NSCLC). But almost all patients will eventually progress for the resistance of drug. This study was to evaluate the efifcacy and safety of retreatment of erlotinib in patients with advanced NSCLC. Methods:It was a retrospective analysis of the 46 advanced NSCLC patients who previously treated with erlotinib and had clinical beneift. The patients were given erlotinib 150 mg orally once daily after failure to other medications until disease progression or the occurrence of intolerable toxicity. The clinical features, therapeutic effect and survival were analyzed. Results: The objective response rate of retreatment with erlotinib was 28.3%. The disease control rate was 60.9%. The rate of symptom relief was 45.7%. The median progression-free survival was 3.6 months. The median overall survival was 7.3 months. One-year survival rate was 8.7%. The median progression-free survival was signiifcant longer in the patients who stopped taking erlotinib more than 6 months than those less than 6 months (P=0.002). The median overall survival was signiifcant longer in the patients whose ECOG ≤2 than those ECOG >2 (P=0.038). The most common drug-related adverse events were rash and diarrhea. Conclusion:The retreatment of erlotinib could possibly prolong the survival time of patients who previously treated with erlotinib and had clinical beneift.

This is to report the screening, extracting and validating antitumor components and compounds from Stellera chamaejasme L. under the case of discrete distribution of active data. In this work, different components from Stellera chamaejasme L. were collected by HPD macroporous resin and polyamide resin column, and their antitumor activity on A549 were tested by MTT assay. Activity results indicate that activity of components at 30-39 min is more potent than that of Stellera chamaejasme L. extract, and the activity of components at 33.97 min is equivalent to positive drug, cis-platinum at 100 microg x mL(-1), but with totally different mode of action. Under the case of discrete activity, the weight analysis is capable of screening active components and compounds from natural products.