Alkaloids ; Antineoplastic Agents, Phytogenic ; Chemical Phenomena ; Chemistry ; Harringtonines

Objective To search for a good method for generation of hematopoietic stem cells (HSC) by embryonic stem cells (ESC),and to investigate the potential of HSC derived from ESC to reconstruct hematopoiesis in vivo.Methods Using a three-step method to induce a mice ESC line, E14.1-into HSC.And identifying HSC by flow cytometry analysis cell markers with CD34 +/Sca-1+, then HSC (1×109L-1)from ESC differentiation were injected into severe combined immunodefieency (SCID) mice for observing teratoma formation.To validate function of HSC by colonngenic cells assay and to reconstitute the hematopoiesis in lethally irradiated mice.Results Combining to use more hematopoietic stimulating factor to availably promote the El4.1 cell into embryoid body (EBs) with abundant hematopoietic progenitors.EBs were induced after 14 day to fast differentiate for HSC with peak percentages of CD34+/Sca-1+ cells reached to (13.72±2.07)%.To harvested ceils from EBs by day 14 for second -step HSC differentiation, percent of CD34+/Sca-1+ cells rise to(24.62±2.50) % after day 16 induction.Cloning forming units (CFU) analysis showed that more Erythro -myeloid cloning generation was observed at this period.Cells obtained in the second step are subsequently plated on monolayer of mice bone marrow stromal cells, in the presence of TPO, FLt3 ligand and superoatant of mice fetal liver stromal cells, cultured for additional 15 days, followed fast expansion of CD34+/Sca-1+ to maximally (58.64±4.20 ) % with more CFU-E, CFU-GM and CFU-GEMM population.Wright-Giemsa stain showed that its had the character of hematopoietic progenitors.Cells from the third-step were injected into SCID mice, but no teratomas were recovered in 2 mices after 6 weeks.Positive selection of CD34+/Sca-1+ cells by magnetic sorting from the third - step differentiation were transplanted into 7 lethally irradiated female mice while predominant hematopoietic reconstitution were observed in 10 days after transplantation, with 71.4% successful engraftment rate.And 3 recipients showed that the cell population of the peripheral blood leukocytes,red cells, hemoglobin approached normal index at 40 days after transplant, hut followed relative'slow renew in platelet count.Survival rate of transplant group is 43.0%, compared to 100% mortality in control mice.Karyotyping assays confirmed female mice with XY.Conclusions The results showed that the three-step differentiation and the culture conditions described here good support differentiation of ESC into HSC.HSC derived from ESC were safe without teratomas formation in body, and its can reconstruct hematopoiesis.

Animals ; Cell Culture Techniques ; Cell Differentiation ; Cells, Cultured ; Embryonic Stem Cells ; cytology ; Female ; Hematopoiesis ; Hematopoietic Stem Cells ; cytology ; Male ; Mice

Hepatectomy ; Humans ; Liver Neoplasms ; surgery ; Preoperative Care ; Safety

Objective To determine the effect of knocking down zebrafish faf1 gene by CRISPR/Cas9 editing technique.Methods gRNA was designed and prepared for the faf1 gene of zebrafish,and gRNA was mixed with Cas9 mRNA by microinjection into zebrafish single cell embryos.The mutant F0 generation zebrafish was screened out by enzyme digestion and gene sequencing.The mutant F0 was genetically outcrossed with the wild-type zebrafish to get the F1 heterozygous zebrafish,and the genotype of zebrafish was detected by microscopic observation.Results The faf1 gRNA and Cas9 mRNA were successfully prepared.The gRNA (gRNA6) located in the exon 6 of faf1 could shift the faf1 gene into frameshift mutations.The mutation type MU1 was screened out and the somatic cytochrome deposition delay was observed in this heterozygous zebrafish.At 4 d post fertilization (dpf),there were sarcomeric dysplasia and head shrinkage,increased hyoid angle and other craniofacial cartilage deformities.And the zebrafish died at 8 ～9 dpf.Conclusion CRISPR/Cas9 knocking out thefaf1 gene produces a new phenotype for zebrafish,with delayed pigment deposition and nodule-like change in tail muscle section.

Objective To explore the effects of extract of Gingko biloba leaves (EGb) on learning, memory and hippoeampal heine oxygenase-1 (HO-1) expression in diabetic rats.Methods The behaviors of streptozotoein-induced diabetic rats were observed by Morris water maze for learning and memory after 6 months of diabetes.The HO-1 mRNA expression and protein expression in hippocampus of diabetic rats were detected by RT- PCR and immunohistochemistry respectively.Results The escape latency time in Morris water maze of diabetic rats was prolonged markedly.HO-1 mRNA and HO-1 protein expressions in hippocampus of diabetic rats were increased (1.635?0.326 vs 0.978?0.214,7.2?1.7 vs 1.9?0.5,respectively,both P＜0.01).The escape latency times in EGb (100,50 mg/kg) treated groups were shortened.HO-1 mRNA and HO-1 protein expressions in hippocampus were decreased at the same time as compared with diabetic group (100 mg/kg:0.954?0.144,2.0?0.8;50 mg/kg:0.988?0.154,2.5?0.6,all P＜0.01).Conclusion EGb can significantly inhibit HO-1 expression in hippocampus and improve learning and memory dysfunction in diabetic rats.

Objective To evaluate the conventional ultrasound (US) and contrast-enhanced ultrasound (CEUS) in diagnosis of splenic trauma including its grading diagnosis. Methods US and CEUS in 42 patients with splenic trauma confirmed by CT and/or operation were performed during 9.2004-10.2007. All the data were compared and analyzed retrospectively. Results Of 42 patients with splenic trauma, 28 cases were detected and 14 cases were missed on US examination, whereas, 40 cases were detected and only 2 mild cases were missed on CEUS examination. The detection rate of lesions with CEUS was significantly higher than that of US (P<0.001) . Ten cases in grading the injury were underestimated by US, however, none of them were underestimated by CEUS. CEUS had good concordance with CT and/or operation in grading diagnosis of 42 cases except two mild cases. Conclusions CEUS has very good concordance with CT and/or operation in detecting injury and grading the splenic trauma compared with conventional US. CEUS as a new imaging technology has made great advance of ultrasoongraphy in evaluation of splenic trauma including injury grading,and it is very useful in clinical application.

Objective To study the effects of vitamin B_6(VitB_6) injection on small intestinal peristalsis in mice and its mechanisms.Methods The mice were divided into 12 groups:calcium chloride injection group(1 mg/10 g),neostigmine methylsulfate injection group(0.001 5 mg/10 g),atropine sulfate injection group(0.005 mg/10 g),their combination with VitB_6 injection and high/low dose treated groups,high dose VitB_6 injection group(5 mg/10 g),low dose VitB_6 injection group(0.5 mg/10 g) and physiologic saline group(0.1 mL/ 10 g ).After administration 30 minutes,mice were intragastric administration Indian ink(0.1 mL/g),and they were luxated and put to death 20 minutes later.The mice belly were cut open,the length of intestine and distance of Indian ink that had moved were measured,and then the ink progradation rate were calculated.Results Compared with control group,the high dose VitB_6 injection could inhibit normal intestinal peristalsis of mice markedly(P0.05).Conclusions VitB_6 injection can inhibit hyperanakinesia of small intestine in mice,especially high dose.And this will be provided as theory foundation on enterospasm treatment.

Twelve cases of Bartter's syndrome were reported and reviewed retrospectively.Usually vomiting was the first sympton in children,while fatigue was common in adults.Bartter's syndrome was characteristic of hypokalemia,metabolic alkalosis,elevations of plasma renin activity,serum angiotersinⅡand aldosterone and juxtaglomerular apperatus hyperplasia.Supplementation of potassium choloride was the main manner of therapy.

AlM: To investigate the association between the degree of anterioruveitis and related factors including inflammatory markers as well as sacroiliac joint imaging in patients with ankylosing spondylitis ( AS) .
METHODS: Anterior changes evaluated by slit lamp, erythrocyte sedimentation rate ( ESR ) , C - reactive protein ( CRP ) and magnetic resonance imaging of 55 cases with AS associateduveitis were retrospectively analyzed. A modified endotoxin-induced uveitis ( ElU ) clinical standard was used for uveitis grading. SPARCC sacroiliac scoring was used to evaluate bone edema of sacroiliac joint. The correlation between the degree of uveitis and sacroiliitis was assessed.
RESULTS: ln the 55 patients with AS, ElU grading scored 2-10, and SPARCC index scored 0-22. Further analysis showed that the severity of uveitis was significantly correlated with ESR (r=0. 869, P<0. 001) and CRP (r=0. 485, P<0. 001). The degree of anterior uveitis in AS patients was not correlated with inflammation of sacroiliac joint (r=0. 237, P=0. 081).
CONCLUSlON: Local autoimmunity of uveitis and sacroiliac joint inflammation with subsequent bone formation in AS might be mutually independent processes.