Objective To identify,construct and express scFv CD133,verify its biological function.Methods VL and VH were isolated from hybridoma of mAb CD133 by using antibody engineering technology.Its DNA sequencing and CDR were determined.scFv CD133 was then cloned into pET32a,transformed into Origami,induced by IPTG,purified by Ni2+-NTA His resin.Its affinity and specificity were tested by NH4SCN elution and ELISA.Results The size of VL and VH of scFv CD133 was 339 bp and 342 bp,which coded 113 and 114 amino acid separately.Its VL belonged to mouse Igκ chain and VH belonged to mouse IgG heavy chain subtype I.The molecular weight of scFv CD133 was about 27 × 103 which was testified by SDSPAGE and Western blot.Its affinity and specificity were also verified.Conclusion scFv CD133 has been successfully constructed and expressed in Origami,which could supply basis for target therapy of CD+133 cancer stem cell.

Objective To observe the effect of prostaglandin E1 on soluble interleukin-6 receptor(SIL-6R) and soluble interleukin-6 receptor β strands (sgp 130) in serum in patients with chronic severe hepatitis.Methods Sixteen patients with chronic severe hepatitis accepted therapy with prostaglandin E1.Serum levels of SIL-6R and sgp130 were detected by enzyme linked immunosorbent assay (ELISA) method before and after therapy.Results The content of SIL-6R and sgp 130 in serum were increased significantly (P<0.001) in patients with chronic severe hepatitis compared with normal control group and decreased obviously (P<0.01) after therapy with prostaglandin E1 compared with before therapy,and the rate of delth were decreased obviously (P<0.01) in PGE1 therapy group.Conclusion The levels of SIL-6R and sgp130 in serum can reflect the degree of liver damage in patients with chronic severe hepatitis and guide the assessment of prognosis;PGE1 can correct the immune disorder,attenuating hepatocyte inflammation and necrosis.

Objective To evaluate the effects of morphine preconditioning-postconditioning on ischemia-reperfusion (I/R) injury in isolated rat hearts. Methods Male SD rats weighing 180-200 g were killed after intraperitoneal injection of heparin 500 U/kg. The hearts were immediately removed and perfused in a Langendorff apparatus with K-H solution gassed with 95%O2-5%CO2 .HR and left ventricular systolic pressure (LVSP) were measured from a fluid-filled latex balloon in the left ventricle. Global myocardial ischemia was induced by interrupting perfusion for 45 min followed by 60 min reperfusion. Forty isolated rat hearts were randomly divided into 5 groups (n = 8 each): group 1 (I/R); group II morphine preconditioning (M1 ); group Ⅲ morphine postconditioning (M2); group IV M1 + M2; group V 5-hydroxydecanoate (5-HD) + M2. Group M1 was perfused with K-H solution containing morphine 3.0 μmol/L for 20 min 30 min before ischemia followed by 10 min normal K-H solution perfusion. Group M2 was perfused with K-H solution containing morphine 3.0 μmol/L for 10 min at the beginning of reperfusion followed by 50 min normal K-H solution perfusion. Group 5-HD + M2 was perfused with K-H solution containing morphine 3.0 μmol/L+ 5-HD 10-4 mmol/L for 10 min at the beginning of reperfusion followed by 50 min normal K-H solution perfusion. Myocardial CK-MB activity was measured and myocardial infarct size (IS/AAR) detennined (by 2,3,5-triphenyl tetrazolium staining) at the end of 60 min reperfusion. Results The preconditioning, postconditioning and combination of preconditioning and postconditioning with morphine 3.0 μmol/L perfusion for 10 min all provided cardio-protective effects in terms of IS/AAR and myocardial activation of CK-MB. Conclusion Although the combination of morphine preconditioning and postconditioning can protect the heart against I/R injury, the effects are similar to those of either of them alone, and the reason may be that either of them alone protects the heart against I/R injury via activating mitoKATP .

Objective To determine whether morphine postconditioning (MP) could protect the heart against ischemia reperfusion (I/R) injury and which specific type(s) of the opioid receptor is involved in the cardioprotective effect produced by hiP. Methods Male SD rots weighing 180-200 g were killed after intraperitoneal heparin 500 U/kg. The hearts were immediately removed and passively perfused in a Langendorff apparatus with K-H solution gassed with 95% O2-5% CO2. HR and left ventricular systolic pressure (LVSP) were measured from a fluid-filled latex balloon in the left ventricle. Global myocardial ischemia was induced by interrupting perfusion for 45 min followed by 60 min reperfusion. The experiment was performed in 3 parts. In Part Ⅰ 32 isolated rat hearts were randomly divided into 4 groups (n = 8 each): group Ⅰ control received no treatment; group Ⅱ ,Ⅲ,Ⅳ were first perfused with K-H solution containing morphine 0.3, 3.0 and 30 μmol/L respectively for 10 min immediately after the end of ischemia followed by 50 min normal K-H solution perfusion. In part Ⅱ,the concentration of morphine in K-H solution which provided the best cardio-protective effects was chosen according to the result of Part Ⅰ , 32 isolated rat hearts were randomly divided into 4 groups ( n = 8 each) : group Ⅰ received no treatment; gvoup Ⅱ,ⅢⅣ were first perfused with K-H solution containing morphine for 5, 10, 20 min respectively immediately after ischemia followed by 50 min peffusion with normal K-H solution. In part Ⅲ,the MP method which provided the best cardio-protective effects was chosen according to the result of Part Ⅱ , 37 isolated rat hearts were randomly divided into 5 groups: group Ⅰ control (n=8);group Ⅱ-Ⅴ were first perfused for 10 min with K-H solution containing morphine (Ⅱ,n = 8)/morphine + naloxone 10 μmol/L(Ⅲ, n = 7)/morphine + nor-binaltorphimine 5 μmol/L (specific κ receptor antagonist, n = 7)/morphine + nalu'indole 5 μmol/L (specific δ receptor antagonist, n = 7) followed by 50 min reperfusion with normal K-H solution. Myocardial CK-MB activity was measured and myocardial infarct size (IS/AAR) determined (by 2,3,5-triphenyl tetrazolium staining) at the end of 60 min reperfusion.Results The postconditioning with morphine 3.0 μmol/L perfusion for 10 min provided the best cardio-protective effects in terms of IS/AAR and myocardial release of CK-MB. Nuloxone completely abolished the cardio-protective effects of MP. Nor-binaltorphimine partly reversed the protective effect of MP, while naltrindole had no effects on MP. Conclusion MP protects the heart against I/R injury via activating κ receptor.

Objective To describe the characteristic endoscopic ultrasonography (EUS) features of gastric ectopic pancreas.Methods Totally 23 patients were diagnosed pathologically as having gastric ectopic pancreas.The EUS images of the lesions were retrospectively reviewed regarding layer of origin,size,growth pattern,margin,and internal echo pattern.Results The coincidence of EUS and pathologic diagnosis was 91%(21/23).EUS revealed that the lesions originated from the second,third,and/or fourth layers of the gastric wall.Most lesions were heterogenous,mainly isoechoic.The borders of the lesions were indistinct in 70%(16/23) patients. Anechoic cystic or tubular structures within the lesions appeared in 10 of the 23 lesions(43%).The mean longest/shortest diameter ratio of ectopic pancreas was 1.7.Conclusions Characteristic EUS features of gastric ectopic pancreases include:mural lateral growth pattern,third layer (submucosa) origin,indistinct margin,intermediate echogenecity,anechoic areas,and fourth-layer thickening.Careful assessment of the EUS findings may be a useful aid in the diagnosis of gastric ectopic pancreas.

Objective To compare the clinical effect and safety of peroral endoscopic myotomy (POEM) with surgical therapy in treatment of esophageal achalasia. Methods 78 patients diagnosed as esophageal achalasia from January 2012 to October 2014 were enrolled in the study and divided into POME group and Heller group. There were 42 patients in POEM group and 36 patients in Heller group. The clinical symptom remission rate, LES resting pressure, Eckardt scores, complication rate, length of hospital stay and the hospitalization expenses were analyzed between the two groups. Results The patients in POEM group and Heller group both got clinical remissions after the treatment. There was no statistical difference in the rate of complication occurrence, Eckardt scores and LES resting pressure between the two groups. Patients in POEM group had shorter operation time, hospital stay and less expenses compared with the Heller group. Conclusions Compared with Heller group, the POEM group has the similar curative effect in treatment of esophageal achalasia. The POEM as a minimally invasive surgery has the advantages of less pain and trauma, shorter hospital stay, well tolerated and low cost. Therefore, the POEM is worth to be popularized and applied in treating esophageal achalasia.

Five patients with mediastinal bronchogenic cyst (MBC) were treated with endoscopic submucosal tunnel dissection (ESTD) between January 2018 and October 2019 at the Department of Gastroenterology in Henan Provincial People′s Hospital. Lesions were located in the middle or lower esophageal level, with diameters≤3.5 cm. The tunnel establishment time was 13-18 min, and the tumor stripping time was 30-51 min. The number of titanium clips used for tunnel closure was 5-8. The tumors were completely resected. No major bleeding or hypoxemia occurred during the operation and no serious adverse reactions occurred after the operation. Postoperative pain scores were all ≤3. The hospital stay was 4-7 days. There was no tumor residue and recurrence during 4-23 months of follow-up.It is suggested that ESTD for MBC which derived from post mediastinum and located at the lower or middle level esophagus is safe and effective.

Objective To investigate cartilage oligomeric matrix protein( COMP) gene mutation in a three-generation pedigree with two cases of pseudoachondroplasia, and to definitize genotype-phenotype correlation. Methods The clinical data and peripheral blood were collected from the patients with pseudoachondroplasia and their family members. All the 19 exons and their flanking sequences of COMP gene in two patients and three unaffected family numbers and 50 unrelated individuals were analyzed by PCR amplification and direct sequencing. Results The proband, a 6-year-old girl presented with typical clinical features of pseudoachondroplasia, including disproportionate short limb dwarfism, staggering gait, double genu varus deformity, and wider clinical and imaging long bone metaphysis. The 33-year-old father showed a similar manifestation including disproportionate short limb dwarfism and double genu varus deformity, and was performed correcting operation on lower limbs for double genu varus at the age of 10 years. DNA sequencing analysis of the COMP gene revealed a del mutation ( c. 1417 1419delGAC)in exon 13 in two patients with pseudoachondroplasia, but not in the other unaffected members from the pedrgree and 50 control subjects. Conclusion A del mutation c. 1417 1419delGAC of COMP gene may contribute to the disease in the pedigree.

OBJECTIVETo investigate the effects of D-galactose (D-gal) on aging of rat marrow mesenchymal stem cells (MSCs) and its mechanism.

METHODSMSCs isolated from young (7 d) SD rats were randomly divided into four groups:control group, 1g/L, 10g/L and 50g/L D-gal treatment groups. In control group MSCs were cultured in DMEM containing 10% FBS for 48 h. In the D-gal treatment groups, MSCs were cultured in DMEM containing 10% FBS with 1g/L, 10g/L or 50g/L D-gal for 48 h. The senescence-associated changes were examined with SA-β-galactosidase (SA-β-gal) staining, the expressions of p53, p21 and p16 were detected by Western blot. The living and apoptotic cells were determined by AO/EB staining. Cell proliferation was detected by MTT assay. SOD activity was measured by xanthine oxidase method, and the MDA content was estimated with thiobarbituric acid (TBA) method.

RESULTSCompared to control group, the number of SA-β-gal positive cells and the expression of p53, p21 and p16 were significantly increased in the 10g/L and 50g/L D-gal treatment groups. The apoptosis rate in 50g/L D-gal group was significantly higher than that in control group (P<0.01). The proliferation of MSCs was decreased in the 10g/L and 50g/L D-gal groups compared to control group (P<0.05). After 10g/L and 50g/L D-gal treatment, SOD activity was significantly decreased (P<0.01), and MDA level was increased (P<0.01).

CONCLUSIONThe aging of MSCs can be induced by 10g/L and 50g/L D-gal, which may be associated with the elevated levels of oxidative stress.

Animals ; Apoptosis ; drug effects ; Cells, Cultured ; Cellular Senescence ; drug effects ; Galactose ; pharmacology ; Male ; Mesenchymal Stromal Cells ; drug effects ; physiology ; Oxidative Stress ; Rats ; Rats, Sprague-Dawley

Objective To analyze the effects of structured fat emulsion and medium/long chain fat emulsion on blood lipids,immune cells and acute inflammation after hepatectomy for hepatocellular carcinoma.Methods Total of 60 patients with hepatocellular carcinoma who underwent hepatectomy in Henan People's Hospital (Zhengzhou University People's Hospital) from January 2013 to March 2017 were divided into experimental group (using structured fat emulsion) and control group (using medium/long chain fat emulsion),30 cases in each group.Triglyceride (TG),high density lipoprotein cholesterol (HDL-C),low density lipoprotein cholesterol (LDL-C),total cholesterol (TC),T lymphocyte level,fibrinogen (FIB),C-reactive protein (CRP),prostaglandin E2 (PGE2) were detected before and 2,4,6 and 8 days after infusion.Results There were no significant differences in LDL-C,HDL-C,TG and TC between the two groups before infusion (P＞0.05).On the 2nd day of parenteral nutrition infusion,the level of blood lipids in both groups was higher than experimental group before infusion;on the 4th,6th and 8th day of infusion,LDL-C,HDL-C,TG and TC in the control group were higher than those in the experimental group (P＜0.05).After parenteral nutrition infusion,the levels of CD3+,CD4+,CD8+,CD4+/CD8+ in both groups were higher than experimental group before infusion,and the experimental group was higher than the control group,the differences were statistically significant (P＜0.05).Compared with before infusion,level of FIB,CRP and PGE2 began to increase on the 2nd day of infusion,and the differences were statistically significant (P＜0.05).On the 2nd,4th,6th and 8th day,CRP in the control group was higher than experimental group.And resepeatively (19.12±5.84) mg/ml vs.(13.76±2.36) mg/ml,(31.67±8.68) mg/ml vs.(17.21±2.66) mg/ml,(22.15±8.33) mg/ml vs.(12.48±0.63) mg/ml,(9.65±4.66) mg/ml vs.(7.52±0.99) mg/ml,and PGE2 were also higher than that in the experimental group (P＜0.05).Conclusion Structured fat emulsion is superior to medium/long chain fat emulsion in improving blood lipid,immune cells and inflammatory reaction in patients after hepatectomy.