Objective To investigate the effects of Thalidomide(THD)on transdifferentiation of human fetal lung fibroblast(HFL-F) to myofibroblast(MF) induced by Transforming Growth Factor-?1(TGF-?1) and the effects on trans differentiated MF.Methods HFL-F to MF trans-differentiation was induced with 5 ?g/L TGF-?1.The effect of 50 ?g/L THD on HFL-F to MF transdifferentiation was evaluated by measuring hydroxyproline(HYP) content with alkaline hydrolysis colorimetry,?-smooth muscle actin(?-SMA) protein with Western Blot,?-SMA and collagen Ⅲ(COL Ⅲ) mRNA with semiquantitative RT-PCR.Results THD inhibited TGF-?1 induced up-regulation of HYP and COLⅢ mRNA expressions(all P0.05).For HFL-F treated with 5 ?g/L TGF-?1 for 96 h,THD inhibited COLⅢ mRNA expression(P

Objective: To analyze the CYP2C19 gene polymorphism in patients with upper digestive system diseases in Anhui Province, so as to provide evidence for individual treatment. Methods: The 307 patients with upper digestive system diseases in the Department of Gastroenterology, The 901st Hospital of Combined Service Force of People＇s Liberation Army were selected. The CYP2C19 genotypes were detected by DNA microarray microarray. The CYP2C19 genotypes and metabolic types in different genders, ages and diseases were analyzed. Results: There were 197 males ( 64.17% ) and 110 females ( 35.83% ) , with the age of ( 58.00±16.13 ) years old. The gene frequency of CYP2C19*1, CYP2C19*2 and CYP2C19*3 was 62.70%, 32.25% and 5.05%, respectively. There were 119 cases (38.76%) of *1/*1 ( 636GG, 681GG ), 129 cases ( 42.02% ) of *1/*2 ( 636GG, 681GA ) , 18 cases (5.86%) of *1/*3 ( 636GA, 681GG ) , 29 cases ( 9.45% ) of *2/*2 ( 636GG, 681AA ) , 11 cases ( 3.58% ) of *2/*3 ( 636GA, 681GA ) , and 1 cases ( 0.33% ) of *3/*3 ( 636AA, 681GG ). In terms of metabolisms, there were 119 cases ( 38.76% ) of fast metabolism type, 147 cases (47.88%) of intermediate metabolism type and 41 cases (13.35%) of slow metabolism type. There were no significant differences in CYP2C19 genotypes and metabolic types among the patients with different gender, age and digestive system diseases ( P＞0.05 ). Conclusion The CYP2C19 genotypes of patients with upper digestive system diseases were polymorphic, mainly the fast metabolism type and the intermediate metabolism type, which could provide reference for the clinical medication of individualized treatment of proton pump inhibitors.

Objective To observe the curative effect of autologous bone marrow stem cells implantation to bone inducing active material combined with core decompression in the treatment of early femoral head osteonecrosis (FHON).Methods From April,2010 to March,2012,in Department of Orthopaedics,the First Affiliated Hospital of Zhengzhou University,a total of 79 adult patients with 108 hips suffered from the early stage FHON were treated with autologous bone marrow stem cells implantation to bone inducing active material combined with core decompression through the core of the femoral canal,male of 65 cases,female of 14 cases,the mean age was 29.5 (20-50) years old.According to the etiology classification:the alcohol-induced FHON was in 54 patients with 66 hips,steroid-induced FHON in 14 patients with 20 hips,steroid and alcohol-induced ONFH was in 11patients with 22 hips.According to association research circulation osseous (ARCO)classifying,Ⅰ-A,Ⅰ-B,Ⅱ-A,Ⅱ-B phases were 6,16,8,and 78 hips,respectively.There were 43 hips in left side and 65 hips in the right side.Results All patients were followed up from 4 to 6 (4.8 ± 0.6) years.Compared with before operation,the scores of all patients were significantly increased (P ＜ 0.05).All patients with hip pain symptoms were relieved or disappeared.The healing tine of the patients in all age groups was statistically significant (P ＜ 0.05),and with the increase of age,the healing time was prolonged.The excellent and good rates of Ⅰ-A,Ⅰ-B,Ⅱ-Aand Ⅱ-B were 100％ (6 / 6),100％ (16/16),100％ (8/8),and 98.7％ (77/78).The X-ray showed that coarse channel osteogenic phenomenon is obvious,there is 1 case collapse of femoral head of stage Ⅱ-B,the rest were not collapse.Conclusion The treatment of early osteonecrosis of the femoral head with autologous bone marrow stem cells implantation to bone inducing active material combined with core decompressionis definitely effective,especially in patients with ARCO:Ⅰ-A,Ⅰ-B and Ⅱ-A phase,and the effect of ARCO:Ⅰ-A and Ⅱ-A is the best.

Objective To measure Ihe effect on rats spinal neuron flow according nerve roots repair time.Methods We adopted the experimental rats on the root avulsion and extravertebral foramen nerve root divison of C_(5～7).We divided them into four groupsin each which there were 16 ratsaccording the type of nerves root injury and repair timeGroup AC:the avulsed roots were reimplanted into the spinal cord and the transeeted roots were sutured to the proximal stump immediately.Group B,Dthe avulsed roots and the transected roots were reimplanted into the spinal cord or were sutured to the proximal stump in delayed 3 weeks each with 16 rats.At the different time point(3 weeks3 months6 months)through pathological examina- tion and immunohistological lechniques and nerve tracing techniqueswe examined the spinal cord and distal nerve trunk in order to observe the pathologic changes and axonal regeneration.Results Group A、C were much better than group B、D in the numberthe conformation and the degree of abatement of spinal motoneu- rons and nissl body.It is the same on the number and the development level of regenerating nerve fiber. Conclusion It had the advantage of neuronal protection and nerve regeneration that reparing the injured nerve roots earlv after nerve roots injury.

Objective: To investigate the effect of RNA interference (RNAi) targeting AKT1 and PI3K P85 on the proliferation and invasion of breast carcinoma MCF-7 cells. Methods: The recombinant adenovirus expression vector, which contained short hairpin RNA (shRNA) targeting open reading frames of AKT1 and PI3K P85 (rAd5-siAKT1-siPI3K), was transfected into human breast carcinoma MCF-7 cells. AKT1 and PI3K P85 mRNA and protein expressions were detected by real-time PCR and Western blotting analysis. The expressions of PCNA, cyclinD1, and P53 were also detected by Western blotting analysis. The proliferation and apoptosis of MCF-7 cells were measured by MTT, flow cytometry and 2-dementinal and 3-dementional matrigel assay. Results: Recombinant adenovirus vector rAd5-siAKT1-siPI3K dramatically down-regulated AKT1 and PI3K P85 mRNA and protein expressions in MCF-7 cells; the downstream factors PCNA and cyclin D1 were also down-regulated, while P53 was up-regulated. Growth of MCF-7 cells was inhibited by over 50% in rAd5-siAKT1-siPI3K group as measured by MTT assay, and cell cycle was arrested in G_1/G_0 phase compared with untransfected and rAd5-siCtrl transfected groups. Cell growth on matrigel matrix showed normal cell shapes, while the cells in rAd5-siAKT1-siPI3K transfected group were detached from the matrix or grew in scattered clustering patterns, forming only small aggregates. Conclusion: shRNA targeting AKT1 and PI3K P85 can significantly down-regulate the expression of AKT1 and PI3K P85 in breast carcinoma MCF-7 cells, and inhibit the growth of MCF-7 cells in vitro.

OBJECTIVETo investigate the relationship between the changes of the copy numbers of mtDNA in peripheral blood mono-nucle- ar cell(PBMC) and the disordered of antioxidant capacity of hepatocellular carcinoma (HCC) patients.

METHODSThe Ficoll Hypaque method was used to isolate the PBMC from blood specimens. The ND1 gene of the mitochondrial was amplified by real-time PCR; meantime β-actin was served as a quantitative standard marker; the difference of mtDNA copy number in PBMC was compared between HCC and healthy control group. The level of reactive oxygen species (ROS) in PBMC was determined by flow cytometry. The change of total antioxidant capacity (T- AOC) of plasma was detected by the biochemistry examination.

RESULTSThe copy numbers of ND1 gene in PBMC of HCC was 73% that of the healthy control group,which suggested a decrease of the copy numbers of mtDNA in HCC. The levels of ROS of PBMC in HCC was (417. 82 ± 110.62) and (301.82 ± 75.54) in control group, which showed that the levels of ROS of PBMC in HCC were significant higher than that in control group (P < 0.01).Plasma T-AOC in HCC was (1.30 ± 0.85), and (3.20 ± 1.62) in control. The T-AOC of plasma of HCC was significantly lower than in control group (P < 0.01).

CONCLUSIONThere was a certain relationship between the decrease of the copy numbers of mtDNA and the disordered antioxidant capacity in hepatocellular carcinoma, which may be associated with the development of hepatocellular carcinoma.

Actins ; Antioxidants ; metabolism ; Carcinoma, Hepatocellular ; blood ; genetics ; Case-Control Studies ; DNA Copy Number Variations ; DNA, Mitochondrial ; genetics ; Humans ; Leukocytes, Mononuclear ; metabolism ; Liver Neoplasms ; blood ; genetics ; Reactive Oxygen Species ; metabolism

In order to study the relationship between lengths of homologous fragments and chromsomic recombination rate in Saccharopolyspora erythraea, three homologous sequences, with mutant loci and different flanking sequences, (26bp + 27bp), (500bp + 576bp) and (1908bp + 1749bp), were synthesized by chemical reaction or PCR amplification, and cloned into pWHM3 to construct homologous recombination plasmids, pWHM1113, pWHM1116 and pWHM1119. When the plasmids were transformed into protoplast of Saccharopolyspora erythraea A226 under PEG mediated, on an average 30, 69 and 170 transformants grew on each plate for the three plasmids respectively, but chromosomic integration frequency were 0, 2% and 19% among corresponding transformants. Both pWHM1116 and pWHM1119 could take double crossover recombination, and exchange the mutant loci in the chromosome. It was concluded that when the flanking sequences were equal or more than (500bp + 576bp), they could take effective single and double recombination with Saccharopolyspora erythraea chromosome.
Chromosomes, Bacterial ; genetics ; Polymerase Chain Reaction ; Recombination, Genetic ; genetics ; Saccharopolyspora ; genetics

OBJECTIVETo observe the effect of a new biomaterial in promoting the bone regeneration for repairing critical-size cranial defects in SD rats.

METHODSCritical-size cranial defects were induced in 3-month-old male Sprague-Dawley rats and repaired with the implants of calcium phosphate from growth factor enhanced matrix 21 (CaPfromGEM21, control), CaPfromGEM21 preloaded with 10 ng bone morphogenetic protein-2 (BMP-2), CaPfromGEM21 preloaded with 100 ng BMP-2, CaPfromGEM21 preloaded with 0.3 µg platelet-derived growth factor-BB (PDGF-BB), or CaPfromGEM21 preloaded with 3 µg PDGF-BB. The defects were examined 6 weeks after the surgery with X-ray, micro-CT, HE staining and quantitative assessments.

RESULTSX-ray showed defect repair in all the groups. The fracture line became obscure, and the defects were almost fully repaired by the regenerated bone tissues in PDGF-BB group. Micro-CT demonstarted new bone formation in the defects. The new bone volume was significantly greater in PDGF-BB groups than in BMP-2 groups (P<0.05). HE staining revealed the presence of new bones in the defects and new vessels in and around the new bones without inflammatory cells. The new bone area fraction was significantly greater in 10 ng BMP-2 group and 0.3 µg PDGF-BB group than in the control group (P<0.05), and the new vessel density was similar in the all the 4 cytokine-preloaded groups and all significantly greater than that in the blank and CaPfromGEM21 control group (P<0.05).

CONCLUSIONCaPfromGem21 combined with BMP-2 or PDGF-BB has good biocompatibility and can better promote bone regeneration for repairing bone defects.

Animals ; Biocompatible Materials ; Bone Morphogenetic Protein 2 ; pharmacology ; Bone Regeneration ; drug effects ; Calcium Phosphates ; pharmacology ; Male ; Prostheses and Implants ; Proto-Oncogene Proteins c-sis ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Skull ; pathology ; Wound Healing

The purpose of this study was to establish real-time based methods for detection of NPM1 gene mutation in acute myeloid leukemia (AML). Primers/probes were designed according to the clustered region of NPM1 mutations on exon 12. Two real-time PCR assays, including high resolution melting curve (HRM) and allele-specific PCR (AS-PCR), were developed and clinically evaluated with 89 AML samples, which were parallelly detected by capillary electrophoresis (CE) and sequencing. The results showed that a total of 17 mutation-positive samples were detected, including type A (15 cases), type B (1 case) and type Nm (1 case). HRM assay could detect all mutant types, and the analytical sensitivity was around 5%. In contrast, AS-PCR assay detected only 95% mutant types, but its sensitivity was as high as 0.01%. It is concluded that considering the characteristics of each method as well as the clinical evaluation results, HRM may be used for screening of NPM1 mutations at diagnosis, while the AS-PCR can be used for the MRD quantification during follow-up.
Alleles ; DNA Mutational Analysis ; Electrophoresis, Capillary ; methods ; Genome ; Humans ; Leukemia, Myeloid, Acute ; diagnosis ; genetics ; Mutation ; Neoplasm, Residual ; diagnosis ; Nuclear Proteins ; genetics ; Plasmids ; Polymerase Chain Reaction ; methods ; Sensitivity and Specificity

OBJECTIVETo study the effects of Kangai injection on the enzyme activities of macrophages and morphology of spleen and thymus from rats.

METHODSTwenty four male SD rats were randomly divided into two groups (n = 12), normal control group and experimental group. The rats in experimental group were injected with Kangai injection at the dosage of 5 ml/kg x d for 30 days peritoneally and those in control group were injected with nomal saline at the same volume. The content of supermicro protein was assayed by BCA method, the activities of lactate dehydrogenase (LDH), glutathione peroxidase(GSH-Px), and inducible nitric oxide synthase (iNOS) from alveolar macrophages(AM) and peritoneal macrophages (PM) were detected biochemically. The activities of acid phosphatase (ACP), superoxide dismutase(SOD) and succinate dehydrogenase (SDH) from AM and PM were detected by ELISA. The morphology of spleen and thymus were observed by light microscopy.

RESULTSThe activities of LDH, GSH-Px and iNOS within AM and PM from experimental group were increased significantly compared with those of control group (P < 0.05). The activities of ACP, SOD and SDH in AM and PM from experimental group were also higher than those from control group (P < 0.05). Microscopically, there was thickening of peripheral arterial lymphatic sheath, enlargement of splenic lymphoid nodules with expended germinal center in the spleen of experimental group. There was no significant difference in the mophology of thymus between the two groups.

CONCLUSIONKangai injection may improve immune function by activating macrophages.

Animals ; Drugs, Chinese Herbal ; pharmacology ; Glutathione Peroxidase ; metabolism ; L-Lactate Dehydrogenase ; metabolism ; Macrophages ; drug effects ; enzymology ; Male ; Nitric Oxide Synthase Type II ; metabolism ; Rats ; Rats, Sprague-Dawley ; Succinate Dehydrogenase ; metabolism ; Superoxide Dismutase ; metabolism