Chinese hamster ovary (CHO) cells are the preferred host cells for the production of complex recombinant therapeutic proteins. Adenine phosphoribosyltransferase (APRT) is a key enzyme in the purine biosynthesis step that catalyzes the condensation of adenine with phosphoribosylate to form adenosine phosphate AMP. In this study, the gene editing technique was used to knock out the aprt gene in CHO cells. Subsequently, the biological properties of APRT-KO CHO cell lines were investigated. A control vector expressed an enhanced green fluorescent protein (EGFP) and an attenuation vector (containing an aprt-attenuated expression cassette and EGFP) were constructed and transfected into APRT-deficient and wild-type CHO cells, respectively. The stable transfected cell pools were subcultured for 60 generations and the mean fluorescence intensity of EGFP in the recombinant CHO cells was detected by flow cytometry to analyze the EGFP expression stability. PCR amplification and sequencing showed that the aprt gene in CHO cell was successfully knocked out. The obtained APRT-deficient CHO cell line had no significant difference from the wild-type CHO cells in terms of cell morphology, growth, proliferation, and doubling time. The transient expression results indicated that compared with the wild-type CHO cells, the expression of EGFP in the APRT-deficient CHO cells transfected with the control vector and the attenuation vector increased by 42%±6% and 56%±9%, respectively. Especially, the EGFP expression levels in APRT-deficient cells transfected with the attenuation vector were significantly higher than those in wild-type CHO cells (P < 0.05). The findings suggest that the APRT-deficient CHO cell line can significantly improve the long-term expression stability of recombinant proteins. This may provide an effective cell engineering strategy for establishing an efficient and stable CHO cell expression system.
Adenine/metabolism* ; Adenine Nucleotides ; Adenine Phosphoribosyltransferase/genetics* ; Adenosine Monophosphate ; Animals ; CHO Cells ; Cricetinae ; Cricetulus ; Recombinant Proteins/genetics*

【Objective】 To investigate the clinical application of self-developed magnetic anchoring device for assisting thoracoscopic pulmonary resection. 【Methods】 Eleven patients underwent thoracoscopic pulmonary assisted with resection magnetic anchoring technique at the Department of Thoracic Surgery, The First Affiliated Hospital of Xi’an Jiaotong University, from March to May 2019. Their clinical data were retrospectively analyzed. The operation time, blood loss, blood transfusion volume, postoperative hospital stay, and postoperative complications were recorded. 【Results】 There were seven male and four female patients, with the average age of (51.6±13.9) years (range from 22 to 69 years). Three single-port and eight single-utility-port thoracoscopic surgeries were performed. Magnetic instruments provided good surgical field exposure in all operations. Among 11 surgeries, one was converted to thoracotomy and one to three-hole surgery due to enlargement and adhesion of hilar lymph nodes. The operation time was (107.8±63.1) minutes (range of 27-182 minutes). The blood loss was 50 (10-50)mL (range of 5-1 000 mL). No blood transfusion was needed during the operation. The postoperative hospital stay was (5.0±1.8) days (range of 3-9 days). No postoperative complications occurred in all the patients. 【Conclusion】 Magnetic anchor technique can effectively alleviate the "chopstick effect" in thoracoscopic surgery. Magnetic anchor technique is safe and feasible in assisting thoracoscopic pulmonary resection.

Objective To evaluate the clinical efficacy of arthroscopic bursal layer-only double-row suture-bridge repair for delaminated rotator cuff tear which is difficult to reposit in comparison with separate double-layer repair and whole-layer repair.Methods From May 2013 through June 2016,82 patients with delaminated rotator cuff tear difficult to reposit were treated at Department of Joint Surgery,The Affiliate Hospital to Chengde Medical University.They were 47 males and 35 females with a mean age of 53.0 ± 7.9 years.They were divided into 3 groups according to their surgical procedures.In group A,28 cases were treated by arthroscopic whole-layer double-row suture-bridge procedure;in group B,29 cases were treated by arthroscopic separate double-layer double-row suture-bridge procedure;in group C,25 cases were treated by arthroscopic bursal layer-only double-row suture-bridge procedure.The 3 groups were compared in terms of University of California Los Angeles (UCLA) score,American Shoulder and Elbow Surgeons (ASES) score,visual analogue scale (VAS),Constant shoulder score,range of motion of shoulder joint and rotator cuff retear preoperatively and postoperatively.Results The patients in the 3 groups were comparable because their preoperative general data showed no significant significances (P ＞ 0.05).The operation time for groups A,B and C was respectively 105.5 ±5.6 min,117.4 ±6.9 min and 88.0 ±4.2 min,showing significant differences between the 3 groups (P ＜ 0.05).The 82 patients were followed up for 21 to 24 months (average,23.3 months).At 24 months postoperatively,UCLA,ASES,VAS,Constant score,shoulder anteflexion and lateral extorsion were respectively 32.4 ± 2.5,12.8 ± 0.9,1.0 ± 1.1,93.4 ± 5.6,158.3° ± 9.3°and 58.9°±5.0°in group A,32.2±2.5,12.9±1.0,0.9±1.0,92.8±6.0,156.4°±9.5°and 59.3°± 5.6° in groups B,and32.4±2.4,12.9±0.9,0.7±0.9,94.3±5.2,156.0°±9.5°and57.6°°5.4°in group C,showing no significant differences between the 3 groups (P ＞ 0.05).The occurrence of rotator cuff retear in groups A,B and C were respectively 17.9％ (5/28),13.8％ (4/29) and 12.0％ (3/25),showing no significant differences between the 3 groups (P ＞ 0.05).Conclusions In repair of delaminated rotator cuff tear difficult to reposit,although the arthroscopic bursal layer-only double-row suture-bridge repair is similar to conventional arthroscopic whole-layer double-row suture-bridge repair and arthroscopic separate double-layer double-row suture-bridge repair in functional recovery and range of motion of the shoulder and incidence of rotator cuff retear,it can reduce obviously operation time and make the operation easier.

Objective To analyze the clinical effects of arthroscopic en masse repair with footprint ending shift using double-row suture-bridge technique for delaminated rotator cuff tears under tension.Methods A total of 58 patients with delaminated rotator cuff tears under tension from August 2013 to August 2016 who underwent arthroscopic en masse repair using doublerow suture-bridge technique were retrospectively analyzed.There were 33 males and 25 females with a mean age of 53.0±7.8 years (range 39-74) with 24 patients left side involved and 34 right side.They were divided into 2 groups to receive en masse repair either footprint ending shift or on the footprint.There were 28 patients with footprint ending shift and 30 patients on the footprint.Clinical effects were evaluated by University of California Los Angeles (UCLA) score,American Shoulder and Elbow Surgeons (ASES) score,visual analogue scale (VAS),Constant-Murley score and shoulder range of motion at preoperatively and postoperatively.Results The average follow-up duration was 23.2±0.8 months (range 21-24).The two groups were compatible with no significant difference in age,gender,tear size,follow-up duration,preoperative function and range of motion of the shoulder joint (P＞0.05).At the last follow up,the UCLA,ASES,VAS,Constant-Murley scores and shoulder range of motion in the group footprint ending shift were respectively 32.4±2.5,12.8±0.9,1.0±1.1,93.4±5.6,158.3°±9.3°,58.9°±5.0° with significantly differences compared with preoperative scores (P ＜ 0.05).The postoperative value in the group on footprint were respectively 31.6±2.9,12.8±0.9,0.7 ± 1.2,91.3±7.1,156.1°± 10.7°,59.6°±4.6° with significantly differences compared with the preoperative scores (P ＜ 0.05).There were no significant difference between the two groups (P ＞ 0.05).The operation duration in the group footprint ending shift was 100.9±6.0 min,while that in the group on footprint was 106.6±6.1 min.There was significantly difference in the operation duration between two groups (t=-3.600,P=0.001).Conclusion Arthroscopic en masse repair using double-row suture-bridge technique can successfully treat delaminated rotator cuff tears under tension.Compared with arthroscopic en masse repair on footprint using double-row suture-bridge technique,the footprint ending shift is easier and time saving without significant difference in function of the shoulder joint and the range of motion in repair of delaminated rotator cuff tear under tension.

Objective To explore the clinical results of the arthroscopic acromioplasty in the rotator cuff repair.Methods Sixty-five patients (42 males and 23 females) to receive suture bridge repair under the arthroscopy between May 2012 and May 2014 were selected and randomly divided into an experimental group and a control group.The experimental group was given suture bridge repair with acromioplasty,while the control group underwent the suture bridge repair without acromioplasty.The time of operation was recorded.The range of motion (ROM) of the shoulder,the UCLA shoulder scoring system (ULCA) score,American orthopedic surgeon scoring system (ASES) score,the visual analogue scale (VAS) score and Constant score were observed 3 and 12 months after the operation.Results There were no significant differences in age,sex and the affected side between the two groups.Significant differences were found between the two groups in the duration of the operation (t=-18.5,P＜0.05).There were no significant differences in the ROM and the ULCA,ASES,VAS and constant scores of the shoulder 3 and 12 months after the operation.Conclusion No significant differences were found in the ROM and the shoulder function scoring between giving acromioplasty under the arthroscopy or not.However,the operation time of undergoing acromioplasty under the arthroscopy was longer.

Objective To explore the characters of lower urinary tract symptoms (LUTS) in patients with Parkinson's Disease (PD).Methods From Oct 2013 to Jun 2016,after evaluating of movement disorder by modified Hoehn-Yahr(H-Y) scale and LUTS by international prostate symptom score (IPSS),urodynamic study was performed in PD patients with LUTS.The incidence of every symptom of LUTS,the relationships between the IPSS categories and urodynamic study were analyzed.Results 64 patients (containing 26 male and 38 female) with 40-80 (62.7 ± 10.2) years old were included.2,4,30,19,12 and 6 patients were belonged to modified H-Y scale 1-4,respectively.Frequency (50 patients,78.1％) was the most common LUTS,while frequency all day (20 patients,31.3％) was the most common symptom for the most severe LUTS.IPSS was 1 7.5 ± 7.8 (4-35) and quality of life was 5.1 ± 0.6 (4-6) for the patients.There was no significant correlation between modified H-Y scale and IPSS (P ＞ 0.05).According to the criteria of IPSS,28 patients (43.8％) only had irritative symptoms,3 patients (4.7％) only had obstructive symptoms,while 26 patients (40.6％) had mixed symptoms and 7 patients (10.9％) belonged to no one.Urodynamic study showed 11 patients (39.3％) with only irritative symptoms had detrusor overactivity(DO),6 patients (23.1％) with mixed symptoms had DO + bladder outlet obstruction (BOO) or DO + detrusor underactivity,however,there was no one with BOO in the three patients with only obstructive symptoms.Conclusion Frequency was the most common LUTS,while frequency all day was the most common symptom for the most severe LUTS in PD patients.Irritative and mixed symptoms were common in PD patients with LUTS,but the urodynamic results were not consistent with the category of LUTS in most of the patients.LUTS severity was not correlated with movement disorders in PD patients.

Objective This study is designed to determine whether an association exists between single nucleotide polymorphism (SNP) variant rs2252673 of insulin receptor(INSR) gene and polycystic ovarian syndrome (PCOS) in Han Chinese in order to identify INSR as a genetic susceptibility factor for PCOS.Methods A total of 224 women with PCOS,192 controls and 672 participants consisting of 224 trios (mother,father and offspring with PCOS) were recruited from the Hospital for Reproductive Medicine Affiliated to Shandong University,from July 2007 to April 2013.Genomic DNA was extracted according to the manufacturer' s protocol.SNP rs2252673 of INSR gene was amplified by PCR and then sequenced on an automated sequencer.Moreover,clinical and metabolic features of the patients with PCOS were compared according to the genotypes.The subjects were divided into twot groups according to body mass index (BMI),and then the results were compared between two groups.And the transmitted disequilibrium test (TDT) was applied for data analysis.Results (1) There were three kinds of genotype of CC,CG and GG.Genotype frequencies of rs2252673 were 8.0％,38.8％,53.1％ and 14.6％,42.2％,43.2％ in the PCOS group and the control group,respectively.The allele frequencies of C and G were 27.5％,72.5％ and 35.7％,64.3％ in the PCOS group and the control group,respectively.There were statistical differences in genotype frequencies and allele frequencies between two groups (all P＜0.05).(2)No significant differences were observed in the different genotype according to clinical and metabolic characteristics of women with PCOS (P＞0.05).But when merging the genotype CG and GG,carriers of the CG and GG genotypes in women with PCOS were slightly associated with total cholesterol (TC) levels (t=2.072,P=0.048) and lower high-density lipoprotein (HDL) levels (t=2.274,P=0.026).Although statistical significance was not achieved,there was an increased tendency in fasting blood glucose (FBG) and fasting insulin (FINS) levels in CG and GG genotypes in PCOS cases.(3)Between the obesity and the non-obesity with PCOS,there was no statistical significance in the genotype and allele frequencies (x2=0.054,P=0.974; x2=0.022,P=0.883).(4)The results of families based analysis shown that genotype distribution of the SNP rs2252673 was in Hardy-Weinberg equilibrium (P＞0.05).After the TDT,the G allele in SNP rs2252673 was over transmitted in families (transmitted∶ non-transmitted=120∶ 88; x2=4.923,P=0.027).There was a transmitted disequilibrium in rs2252673,which implies the association of INSR and PCOS were independent of population stratification.Conclusions There were a association between the SNP variant rs2252673 of INSR gene and the susceptibility to PCOS in Han Chinese women,which was independently of body mass index.The carrier of G allele frequency of rs2252673 may have higher risk of PCOS.

Objective To transform the vocabulary based Chinese personality (VBCP) to a sentence based Chinese personality inventory.Methods Based on the dimension theory and mapping sentence,347 sentences were developed and quantified with equal weight.772 subjects participated in a pilot study and an exploratory factor analysis was conducted to the collected data.A trial version of the Chinese Personality Inventory with 7 factors was developed.With the trial version as the survey instrument,3190 subjects from 22 provinces and centrally administered municipalities in China participated in the survey,the exploratory factor analysis,confirmatory factor analysis,annova,t-test,correlation and cronbach test were utilized in this research.The Chinese seven factors personality inventory (CSPI) was developed,which included seven factors with 130 items.Results The 7 factors of Chinese personality inventory were emotion,innovation,persistency,virtue,extraversion,agreeableness and tradition.42.27％ of the total variance of the seven factors was explained.The internal reliability coefficient was Cronbach' s α =0.902(P＜0.001),while the internal reliability coefficients among the factors were between 0.794-0.947.The retest reliability coefficients of the factors were between 0.696-0.921 (P＜ 0.001).Significant positive correlations (r ranged from 0.151 to 0.548,P＜0.01)were found between 5 of the 7 factors of CSPI,but innovation and tradition.Significant positive correlations (r ranged from 0.193 to 0.715,P＜0.05,P＜0.01)were found between the 7 factors and CSPI.Significant correlations between CSPI and EPQ-RSC were found (P＜0.05).Confirmatory factor analysis showed that thc inventory was valid,and NFI and CFI were greater than 0.90.Conclusion CSPI met the measurement requirement in validity,reliability and explanation ability.

Objective To examine the relationship of marital quality,personalities,coping style in spouses.Methods Total 923 spouses were assessed with the marriage perception scale (MPS),Eysenck personality questionaire(EPQ) and simplified coping style questionnaire(SCSQ).Results The results revealed that psychoticism and neuroticism were both negatively correlated with marital quality(r=-0.24,-0.31,P＜ 0.01),but intraversion/extraversion and lie were both positively correlated with marital quality (r =0.24,0.14,P ＜ 0.01).And coping style was positively correlated with marital quality(r=0.33,P＜ 0.01).Significant partial mediation of coping style on the association between personality and marital quality were revealed,and the mediating effect was accounted for 8.8％ of the total effect.Conclusion Coping style is the mediator between personalities and marital quality.

Objective To explore the effect of three-dimensional conformal radiation dose fractionation treatment of advanced non-small cell lung cancer(NSCLC).Methods 75 NSCLC patients were treated with hypofractionated 3DCRT( observation group),while 73 cases were treated with conventional fractionated radiotherapy (control group).The efficacy and adverse reactions were observed;Survival after treatment were followed up in 1,2,3 years.Lung function was detected before and after radiotherapy treatment,including FVC,FEV1 and CLCO.Results The total effective cases of control group were 47 cases patients ( 64.4％ ),observation group's was 60 cases ( 80.0％ ),total effective rate had statistically significant difference ( x2 =4.50,P ＜ 0.05 ).Survival of control group after treatment in 1,2,3year were 50.7％,24.7％,8.2％,the median survival was 13 months,observation group's were 73.3％,45.3％,20.0％,and 19 months.The survival of these two groups was statistically different (x2 =8.07,6,94,4.22,all P ＜ 0.05 ).The patients blood system side effects of observation group were significantly lower than the control group ( x2 =4.73,P ＜0.05 ) ;acute radiation pneumonia,esophagitis of these two groups had no significant difference in the incidence( P ＞ 0.05 ).Conclusion Hypofractionated 3DCRT treatment of advanced NSCLC had good effect,and its adverse reactions was low,and it was worthy of clinical application.