Abstract:Objective: Based on the comparison of transcriptional profiles between porcine primed embryonic stem cells(pESC), porcine inner cell mass(ICM) and porcine embryonic fibroblasts(PEF) in our previous study, five transcriptional factors(OCT4, TBX3, REX1, LIN28 and DPPA5) were selected due to their expressions were significantly higher in ICM than pESC and PEF. To establish porcine induced pluripotent stem(iPS) cell line, three expression vectors with five porcine transcriptional factors connected via 2A peptide gene sequence were constructed. The induced expression vectors and pEF1a-Tet3G were cotransfected into PEF followed by supplement with doxycycline hyclate to obtain more efficient pluripotent stem cell induction strategy. Methods: Firstly, the cDNA sequences of transcription factors REX1, LIN28, DPPA5 were cloned from PEF through PCR and linked together with E2A and T2A sequence(RLD). The cDNA sequences of transcription factors OCT4 and TBX3 were synthesized. Secondly, OCT4, TBX3 and RLD were transfected respectively into the TET-ON induced expression plasmid(pTRE3G-Zs), followed by plasmid extraction and restriction endonuclease digestion to verify the correct construction of three recombinant vectors. Finally, the transgenic cell lines were obtained via nucleofection of these three vectors and pEF1a-Tet3G, and were screened with G418. Results: 975bp REX1 sequence, 727bp LIN28 sequence and 408bp DPPA5 sequence were obtained by PCR. The correct construction of three recombinant vectors(pTRE3G-Zs-OCT4, pTRE3G-Zs-TBX3, and pTRE3G-Zs-RLD) was verified by restriction endonuclease. A total of 70 cell lines including 29 cell lines with multicopy of foreign genes were established by drug screening and PCR. Conclusion: We have established transgenic cell lines with five porcine transcription factors (OCT4, TBX3, REX1, LIN28, and DPPA5) and pEF1a-Tet3G plasmid successfully.

Abstract:Objective：To study the effect of apigenin on A549/DDP cells，and to explore the effect and mechanism of apigenin reverse drugs resistance on A549/DDP cells. Methods：The cells were cultured with different concentrations apigenin. A549/DDP cells proliferation and drug sensitivity was detected by MTT asssy. The drug extracellular transport was detected by Rhodamine-123 retention experiment, the P-gp and the LRP expression in the tumor cells was detected by Western blot, and the transcription of MDR1 mRNA and LRP mRNA in the tumor cells was detected by RT-PCR. Results: The apigenin can inhibit the A549/DDP cells proliferation. There were significant differences between the apigenin groups(20,40,80 μmol/L) and the DDP group（P＜0.05）. The DDP’s IC50 with the apigenin was (5.76±0.36) μg/mL in A549/DDP cells. The DDP’s IC50 without the apigenin was (14.33±0.41)μg/mL in A549/DDP cells, and the reversal index was 2.48. The apigenin can make high Rhodamine-123 accumulate in A549/DDP cells, down-regulate the P-gp expression in A549/DDP cells, and decrease the transcription of MDR1 mRNA in A549/DDP cells. Conclusion: The apigenin may inhibit the A549/DDP cells proliferation and to reverse drug resistance in A549/DDP cells, and the mechanism was ralated to decrease the MDR1 mRNA transcription and down-regulate the expression of P-gp that mediated drug extracellular transport.

Abstract:Objective: To explore the expression level of tumor suppressor gene FoxO3 in the recurrent glioblastoma tissues and its potential function in temozolomide(TMZ) resistance. Methods: The expression level of FoxO3 mRNA and protein were detected and compared in primary and recurrent glioblastoma tissues via microarray, qPCT and Western blot. And then to explore the potential mechanism of FoxO3 mediated in TMZ resistance by screening its target using a microarray. Results: The expression level of FoxO3 mRNA and protein is decreased both in recurrent glioblastoma tissues and TMZ- resistance glioblastoma cell lines. Overexpression of FoxO3 could sensitize glioblastoma cells to TMZ via modulating the expression of REV3L. Conclusion: The expression level of FoxO3 is decreased in recurrent glioblastoma tissues, which might play a pivotal role in chemotherapy-resistance. Overexpression of FoxO3 could sensitize glioblastoma cells to TMZ via suppressing the expression of REV3L.

Abstract:Objective: To detect the expression level of angiopoietin-like protein2(ANGPTL2) in esophageal squamous cell cancer (ESCC) tissues and investigate the clinical value for diagnosis. Methods：Immunohistochemistry(IHC) was used to determine the relative ANGPTL2 expression levels in ESCC tissues, and enzyme-linked immunosorbent assays were used to measure the levels of ANGPTL2. Results： Compared with controls， ANGPTL2 expression levels were significantly up-regulated in ESCC tissues and serum (P<0.001)． Higher expression level of ANGPTL2 was found associated with larger tumor size, more lymph node metastasis and TNM stage． ELISA assay showed that serum ANGPTL2 was significantly elevated in ESCC patients and positively associated with high T-stage, high TNM stage and lymph node metastasis. Conclusion： ANGPTL2 plays a crucial role in proliferation and invasion of ESCC. Serum ANGPTL2 has potential value for early detection of ESCC.

Abstract:Objective: This study investigated the impacts of oligomeric proanthocyanidins (OPC) on patient with asthma to develop new therapeutic options for asthma treatment. Methods: A total of 60 non-acute attack patients with asthma who conventionally received budesonide/formoterol inhalation were randomly divided into the placebo, high-, intermediate-and low-dosage OPC groups. The duration of OPC treatment ranged 8 weeks. Asthma control test(ACT) score, eosinophils(EOS) count and pulmonary function were analyzed. Results: A significant decrease of EOS in peripheral blood was observed in the high-dosage group who received OPC treatment after 4 weeks(P<0.05). When the treatment expended up to 8 weeks, an elevated ACT score (P<0.05) and forced expiratory volume in one second to forced vital capacity ratio(FEV1%) as well as a declined EOS count (P<0.05) were found in the high-dosage group, while intermediate-dosage OPC only presented improvements in EOS count and FEV1% (P<0.05). Dosage and intervention duration were positively correlated with ACT score and FEV1% (P<0.05), and negatively correlated with the number of peripheral blood EOS(P<0.05). Conclusion: Our results exhibited a novel profile of OPC as a potent option for airway inflammation relief and ventilation rescue, highlighting a promising role of OPC in asthma management.

Abstract:Objective: To assess the value of QRS-VHIS interval and QRS-VRVA interval on differentiating the origin of outflow tract premature ventricular contractions. Methods： Sixteen consecutive patients underwent radiofrequency ablation were enrolled from June 2016 to December 2016. Two quadrupolar catheters were positioned at the region of His bundle and right ventricular apex (RVA). With the guidance of the electroanatomic mapping system and fluoroscopy, pace mapping was conducted at different sites in the right ventricular outflow tract(RVOT) and aortic sinus cusp(ASC). Standard 12-lead electrocardiograph(ECG) and intracardiac electrogram of HIS and RVA were recorded simultaneously. Meanwhile, the QRS-VHIS and QRS-VRVA intervals were measured respectively. Results： A total of 139 pacing sites were obtained from 16 patients, and were divided into the RVOT group(n=94) and the ASC group(n=45). The QRS-VHIS interval in the ROVT group was significantly longer than that in the ASC group［(42.98±14.83) ms vs.(19.66 ± 16.14) ms, P<0.001)］. However, there was no significant difference in QRS-VRVA interval between the two groups. Receiver operating characteristic(ROC) analysis showed that the QRS-VHIS interval exhibited a greater area under the curve (AUC) (0.860) than the QRS-VRVA interval(0.626). The QRS-VHIS interval ≤36.4 ms predicted an ASC origin with a sensitivity of 67.1% and specificity of 85.0%. The QRS-VRVA interval ≥57.4 ms had a sensitivity of 54.3% and specificity of 72.0%. Conclusion： The QRS-VHIS interval could be used as a new approach to differentiate the origin of outflow tract premature ventricular contractions.

Abstract:Objective：To investigate the value of interleukin 18 (IL-18) and urinary kidney injury molecule 1 (KIM-1) in blood and urine in early diagnosis of acute kidney injury (AKI) after coronary intervention. Methods: A total of 243 eligible patients who underwent coronary intervention were divided into the AKI group (48 cases) and the non-AKI group (195 cases) according to the diagnostic criteria of AKI. Blood and urine samples were collected at different time points (0, 2, 4, 6, 8, 10, 12, 24, 48, and 72 h) after coronary intervention, and then the concentrations of serum creatinine(SCr), IL-18 and KIM-1 in serum and urine were measured. Results: The levels of SCr from 12 h to 72 h were higher than that of 0 h in the AKI group(all P<0.05). The levels of IL-18 and KIM in serum and urine were significantly higher than those of 0 h. IL-18 level reached peak at 2 h(blood) or 4 h(urine), and KIM-1 level reached peak at 6 h (blood) or 8 h (urine) (all P<0.05). Pearson correlation analysis showed the levels of 2 h-serum IL-18, 4 h-urine IL-18, 6 h-serum KIM-1 and 8 h-urine KIM-1 were positively correlated with that of SCr(24 h) (all P<0.05). The ROC analysis showed the levels of IL-18 and KIM-1 in serum and urine had a good diagnostic efficiency. The joint detection had a higher AUC than single marker detection. Conclusion： The joint detection of 2 h-serum IL-18, 4 h-urine IL-18, 6 h-serum KIM-1 and 8 h-urine KIM-1 after intervention had a better diagnostic performance than single marker detection, which may be an important reference for early diagnosis of AKI.

Abstract:Objective: To compare the efficacy of total thoracoscopic minimally invasive cardiac surgery and conventional open heart surgery in the treatment of congenital heart disease. Methods: A total of 67 cases of congenital heart disease received from April 2013 to December 2016 were randomly divided into the observation group (34 cases) which with three holes minimally invasive surgery and the control group (33 cases) which with traditional thoracotomy surgery. One patient in the observation group was converted to conventional open chest surgery for complex congenital heart disease. The two groups were compared in patients with cardiopulmonary bypass time, aortic clamping time, operation time, postoperative ventilation time, 24 hours postoperative of thoracic drainage, postoperative ICU time, postoperative hospitalization time and complication rate. The life quality one week after the operation of the two groups was also compared with the Chinese version of the European Organization for Research on Treatment of Cancer (EORTC) life quality scale QLQ-C30(V3.0). Results: The patients in the observation group were better in postoperative ventilation time, 24 hours postoperative of thoracic drainage, postoperative ICU time, postoperative hospitalization time and the life quality one week after the operation（P<0.05）, and worse in the cardiopulmonary bypass time and aortic clamping time than those in the control group（P<0.05）. There was no significant difference in the total operation time and the incidence of complications between the two groups. All the patients recovered smoothly without serious complications or death, and had no long-term death or atrioventricular recanalization after 3-24 months of follow-up. Conclusion: Total thoracoscopic minimally invasive cardiac surgery is safe and effective for simple congenital heart disease, and has the advantages of less destruction, less trauma, less bleeding, faster recovery, high early postoperative life quality and less impact on the appearance than traditional thoracotomy surgery.

Abstract:Objective: To evaluate the performance of European system for cardiac operative risk evaluation(EuroSCORE)Ⅱ and the Society of Thoracic Surgeons(STS) score in patients with infective endocarditis(IE) undergoing cardiac surgery. Methods: Data were retrospectively collected from adult patients undergoing cardiac surgery for IE between July 2012 and March 2016 in our hospital. EuroSCOREⅡ and STS score were calculated. The discrimination and calibration of these two scoring system were assessed by receiver-operating characteristic(ROC) curve analysis and Hosmer-Lemeshow goodness-of-fit test. According to the EuroSCOREⅡ, patients were divided into the low risk group(<3%), the medium risk group(3%~6%) and the high risk group(≥6%). Differences in prognosis among these three groups were compared. Results: Among 80 IE patients undergoing cardiac surgery, 3 patients(3.75%) died in-hospital and total 6 deaths(7.5%) were observed during follow-up. The area under the ROC curve(AUC) for the EuroSCOREⅡ, STS, and combination of them was 0.836, 0.833, and 0.846 to predict late mortality respectively; 0.946, 0.980, and 0.980 to predict in-hospital mortality respectively, which indicated good discriminative power. Hosmer-Lemeshow goodness-of-fit test showed significant P-values(P>0.05) indicating good calibration and accuracy. The predictive in-hospital mortality was similar to actual mortality in the high risk group(11.8% vs. 10.3%, P=0.358), and significantly higher than that in the low risk and medium groups(P<0.01). The predictive late mortality was similar to actual mortality in the medium risk group(4.1% vs. 4.2%, P=0.718), whereas was underestimated in the high risk group(11.8% vs. 17.2%，P<0.05) and overrated in the low risk group(1.9% vs. 0%，P<0.01). Conclusion: Both EuroSCOREⅡ and STS scoring system can satisfactorily predict in-hospital mortality and late mortality in patients with IE undergoing cardiac surgery.

Abstract:Objective: To investigate the predictive value of B type natriuretic peptide in evaluating fluid volume in the patients receiving fluid resuscitation with early phase of septic shock, and to find the relative markers of fluid volume which can accurately reflect the blood volume of septic shock. Methods: A prospective study was performed on 60 patients with septic shock enrolled in the intensive care unit of our hospital. All the patients receiving early goal-directed therapy(EGDT), the hemodynamic parameters such as intrathoracic blood volume index(ITBVI), global end diastolic volume index(GEDVI), extravascular lung mater index(EVLWI) were record by using PICCO system before and after EGDT, and the parameters of BNP were record at the same time. The correlations between changes in BNP and changes in ITBVI,GEDVI and EVLWI were tested by Pearson correlation analysis． Results: ITBVI,GEDVI and BNP in all patients were significantly elevated after EGDT. BNP level was not correlated with ITBVI,GEDVI and EVLWI at the time point 0 h and 6 h in the 60 patients receiving fluid resuscitation. The changes in BNP (ΔBNP) showed significant correlation with changes in ΔITBVI and ΔGEDVI. ΔEVLWI was not correlated with ΔBNP. Conclusion: The predictive value of B type natriuretic peptide in evaluating fluid volume in the patients receiving fluid resuscitation with early phase of septic shock is uncertain.

Abstract:Objective: To evaluate the efficacy of treatment with monosialotetrahexosyl ganglioside(GM1) combined with mouse nerve growth factor(NGF) on early stage of treatment and short-term prognosis of acute Bell’s palsy. Methods: A retrospective study was conducted to analyze clinical data of 219 patients with Bell’s palsy from January 2009 to December 2015. The patients with acute Bell’s palsy were divided into four groups, including the GM1 combined with NGF group(58 cases), the GM1 group(55 cases), the NGF group(51 cases), and the basic group(without any special neurotrophic drugs, 55 cases). All cases had the same basic treatment. We used the House-Brackmann facial nerve grading system(HB) and the Sunnybrook grading system to evaluate the efficacy of treatment before starting treatment, after 1 week and 2 weeks of treatment and 1 month after onset. Results: There was greater improvement in the mean HB and Sunnybrook scores of the GM1 combined with NGF group in comparison to the other three groups at 2 weeks and 4 weeks(P<0.01). After 4 weeks, 91.4% patients had good recovery in the GM1 combined with NGF group compared with the other three groups(P<0.01), the recovery rates of the GM1 group, NGF group and basic group at the same time point were 74.5%, 78.4% and 50.9%, respectively. Conclusion: The combination of GM1 and NGF treatment increases the possibility of recovery in acute Bell’s palsy.

Abstract:Objective: To explore diagnosis value of the abnormal red blood cell morphology and red blood cell parameters test combined with bone marrow iron staining and serum ferritin in children with iron deficiency anemia (IDA). Methods: A total of 388 children with IDA were administrated by abnormal red blood cell morphology and ferritin(SF) content detection, and the bone marrow smear were detected by iron staining. We selected 388 patients with IDA and 255 patients with chronic diseases anemia(ACD) on detection of red blood cell parameters ［mean corpuscular volume(MCV), mean corpuscular hemoglobin(MCH), mean corpuscular hemoglobin concentration(MCHC) and red blood cell distribution width(RDW)］ and SF, and compared with 203 cases of healthy controls. Results: Abnormal morphology of red blood cells in 388 children accounted for about 56.25% to 92.34%. Iron staining of bone marrow cells increased with the increasing of serum ferritin detection index. Compared with the ACD group, MCV, MCH, SF in the IDA group were significantly decreased, RDW was increased, and the differences were statistically significant(all P<0.05). Compared with the control group, MCV and SF in the IDA group were significantly decreased; while in the ACD group, no obvious change of the MCV was found, SF was increased obviously, the difference was statistically significant(P<0.05). Combinding MCV, RDW and SF in diagnosis of IDA had the highest sensitivity, up to 99.9%. Combinding MCV, RDW and SF play a great role in diagnosis and differential diagnosis in IDA. Conclusion: Abnormal red blood cell morphology and red blood cell parameters test combined with bone marrow iron staining and serum ferritin in IDA in children has significant clinical value in the differential diagnosis.

Abstract:Objective: The aim of this study was to evaluate the clinical effects of fixation of bone block graft with single titanium screw adjacent to alveolar ridge crest for bone augmentation in maxillary anterior implant treatment. Methods: From 2013.01 to 2013.06, five patients with serious alveolar ridge defect in maxillary anterior zone were collected in this survey. The ages of the patients ranged from 19 to 24 with a mean age of 21.6 years old. Among them, 3 cases lost single tooth and 2 cases lost multiple teeth. Autogenous chin bone blocks were taken and fixed with single titanium screw adjacent to the alveolar ridge crests. Postoperative follow-ups were made periodically. After 6-9 months, cone beam computed tomography (CBCT) was used to evaluate their alveolar ridge augmentations and measure the width of alveolar ridges and the distance from titanium screw to alveolar ridge crest. Based on the CBCT measurements, implants were then inserted. After the osseointegration period, final prosthetic restorations were finished. Results: The successful rate of the bone augmentation was 100% and no relative clinical complications occurred in this study. Adequate bone augmentations were obtained in maxillary anterior areas and the fixed titanium screws were adjacent to the alveolar ridge crest. Conclusion: The augmentation of chin bone block graft fixed with single titanium screw adjacent to alveolar ridge crest is a predictable way to improve bone defects in maxillary anterior areas for satisfactory implant treatments.

Abstract:Objective: To establish an experimental animal model of peri-implantitis after immediate implantation of mandibular anterior teeth in rabbits. Methods: A total of 12 healthy male rabbits were anesthetized in the study. A tapered titanium implant was inserted immediately after removal of the left mandibular anterior tooth in each rabbit. Three rabbits were randomly selected and executed after 8 weeks of implantation. Micro-CT was used to evaluate the osseointegration. Then, the rest of animals were randomly divided into three groups. The blank group was not given stimulation, while the control group and the experimental group were injected phosphate buffer solution and E.coli-LPS solution around the implant respectively. After 2 weeks of stimulation, all rabbits were sacrificed to collect the mandible samples with implants and gingival tissues. Micro-CT was used to evaluate bone mass around implants, and hematoxylin and eosin staining was used to observe the histological changes of gingival tissues. Results: After 8 weeks of implantation, satisfactory osseointegration of implants were formed with healthy gums. After 2 weeks of stimulation, the blank and control groups showed pink tough gums with scattered inflammatory cells and good osseointegration of implants without bone resorptions. The experimental group showed red swollen gums with massive inflammatory cells and apparent bone resorptions on the top of implants with reduced bone mineral density and percent bone volume. Conclusion: An experimental animal model of peri-implantitis in rabbits could be established rapidly by local injections of E.coli-LPS solution around the implant.