AIMTo explore the effects of interleukin-1beta (IL-1beta) on inducible nitric oxide synthase (iNOS)-nitric oxide (NO) system activity in arginine vasopressin (AVP)-induced rat cardiac fibroblasts (CFs).

METHODSCFs were isolated by trypsin digestion method. Nitric acid reductase method, spectrophotometry and reverse transcription-polymerase chain reaction (RT-PCR) were used to detect NO contents, NOS activity and iNOS mRNA expression.

RESULTSAVP significantly increased iNOS mRNA expressions, NOS activity and NO contents (P < 0.05) in CFs. IL-1beta enhanced the effects of AVP on iNOS-NO system activity in a concentration-dependent manner, moreover the iNOS mRNA expressions, NOS activity and NO contents of AVP + 3 ng/ml, AVP + 5 ng/ml IL-1beta group were both significantly higher than those of AVP group (P < 0.05). But when IL-1beta concentration increased to 5 ng/ml, the iNOS mRNA expressions, NOS activity and NO contents did not increase accordingly, slightly decreased instead.

CONCLUSIONWithin certain range of concentrations IL-1beta cooperates with AVP to increase iNOS-NO system activity in CFs.

Animals ; Arginine Vasopressin ; pharmacology ; Fibroblasts ; drug effects ; metabolism ; Interleukin-1beta ; pharmacology ; Male ; Myocytes, Cardiac ; drug effects ; metabolism ; Nitric Oxide ; metabolism ; Nitric Oxide Synthase Type II ; metabolism ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo investigate the influence of down-regulating Smoothened (SMO) gene expression through short hairpin RNA (shRNA) on the proliferation of breast cancer stem cells.

METHODSHuman SMO shRNA was designed, synthesized chemically, and transfected into MCF-7 cells to down-regulate SMO gene. By using G418, stable cells with down-regulated SMO were selected. In vitro proliferation of these cells was measured by CCK8 assay. The proportion of CD44(+)/CD24(-) cells was detected by flow cytometry and the mammospheres formation was determined by suspension sphere culture. The expression of SMO, GLI1 and Oct4 was detected by Western blot. In vivo, the volume of tumor was measured every 3 days and the expression of SMO, GLI1 and Oct4 detected by Western blot.

RESULTSIn vitro, the cells were transfected with SMO-shRNA and selected by G418 after 21 days. SMO-shRNA effectively down-regulated the expression of SMO gene and protein, and inhibited the proliferation of MCF-7 and markedly reduced the proportion of CD44(+)/CD24(-) cells and mammospheres. In vivo, SMO-shRNA treatment of MCF-7 significantly inhibited the volume of tumor. The positive rate of SMO in negative control and SMO-shRNA group was 5/5 and 2/5, respectively. The expression of SMO, GLI1 and Oct4 in different groups were 0.72 ± 0.17 and 0.21 ± 0.09, 1.21 ± 0.21 and 0.47 ± 0.12, 0.83 ± 0.13 and 0.25 ± 0.07. SMO, GLI1 and Oct4 down-regulation significantly suppressed at protein levels (P < 0.05).

CONCLUSIONThe shRNA by chemical synthesis can effectively down-regulate SMO gene expression and inhibit the proliferation of breast cancer stem cells.

Animals ; Cell Proliferation ; Down-Regulation ; Female ; Gene Expression Regulation, Neoplastic ; Humans ; Hyaluronan Receptors ; metabolism ; MCF-7 Cells ; Mice ; Mice, Nude ; Neoplasm Transplantation ; Neoplastic Stem Cells ; pathology ; Octamer Transcription Factor-3 ; metabolism ; RNA, Small Interfering ; genetics ; Receptors, G-Protein-Coupled ; genetics ; metabolism ; Smoothened Receptor ; Transcription Factors ; metabolism ; Transfection ; Tumor Burden ; Zinc Finger Protein GLI1

Adolescent ; Adult ; Child ; Child, Preschool ; Female ; Humans ; Laser Coagulation ; Male ; Porokeratosis ; pathology ; surgery

Objective To investigate the effects of chronic hyperglycemia on the proliferation,secretion function and expression of endoplasmic reticulum stress(ERS) related molecules in pancreatic ? cell line MIN6. Methods MIN6 cells were treated with different concentrations of glucose(5.6,25 and 33.3mmol/L) and were harvested at indicated time(24h and 96h) for examinations.Cell proliferation was tested using CCK-8 solution,insulin and proinsulin secretion function was determined by ELISA,and mRNA expression of ERS related molecules(Total XBP-1,Spliced XBP-1) and protein expression of phosphorylated IRE1? were detected by Real-time PCR and Western blotting,respectively. Results After treatment with hyperglycemia for 96h, cell proliferation was significantly lower than that treated for 24h(P

OBJECTIVE: To research the value of polymorphism of salivary esterase(Set) in paternity and personal identification. METHODS: Phenotype and genotype of human salivary esterase were detected in 114 liquid saliva samples from the Chinese population by disc electrophoresis and fast blue RR staining assay. RESULTS: The frequency of Set type was F 22.81%, FS 50.88%, S2 6.31%. The estimated gene frequency of SetF was 0.4825 and SetS was 0.5175. The PE was 0.1875 and the DP was 0.6199. CONCLUSION Polymorphism of salivary esterase (Set) was practical in paternity and personal identification.
Esterases/genetics* ; Forensic Anthropology/methods* ; Gene Frequency ; Humans ; Paternity ; Polymorphism, Genetic ; Saliva/enzymology*

Objective To investigate the inhibition effect of Minocycline on hippocampal microglia in epileptic rats. Methods Forty male SD rats were equally randomized into normal saline control group (NS), penicillin inducement group, Minocycline post-treatment group and Minocycline pre-treatment group (n=10). Rat epilepsy models in the later 3 groups were induced by intraperitoneal injection of penicillin G at a dosage of 740 million to 7.6 million units/kg. The level of hippocampal microglia in rats of the 4 groups on the 1st and 3rd d of inducement was detected by immunofluorescence and the tumor growth factor-α (TNF-α) protein level was detected by Western blotting on the 1st and 3rd d of inducement. Results Seizure could activate microglia. As compared with those in rats of the penicillin inducement group, the activation and hyperplasia of microglia in the hippoeampus in rats of the minoeyeline post- and pre-treatment groups were obviously inhibited on the 1st and 3rd d of inducement (P≤0.05), and the effects were much obvious in the pretreatment group. The level of TNF-α protein in the penicillin inducement group, minoeycline post- and pre-treatment groups was significantly higher than that in the NS group on the 1st and 3rd d of inducement (P≤0.05); as compared with that in the penicillin inducement group, the level of TNF-α protein in the minocycline post- and pre-treatment groups decreased significantly on the 1st and 3rd of inducement (P≤0.05), especially that in the pretreatment group. Conclusion Minocycline can effectively inhibit the activation and hyperplasia of hippocampal microglia and the releasing of inflammatory factor TNF-αt in epileptic rats.

BACKGROUNDGene therapy and epigenetic therapy have gained more attention in cancer treatment. However, the effect of a combined treatment of gene therapy and epigenetic therapy on head and neck squamous cell carcinoma have not been studied yet. To study the mechanism and clinical application, human laryngeal carcinoma cell (Hep-2) tumor-bearing mice were used.

METHODSA xenograft tumor model was established by the subcutaneous inoculation of Hep-2 cells in the right armpit of BALB/c nu/nu mice. The mice with well-formed tumor were randomly divided into six groups. Multisite injections of rAd-p53 and/or 5-aza-dC were used to treat tumor. Tumor growth was monitored by measuring tumor volume and growth rate. p53 and E-cadherin protein levels in tumor tissues were detected by immunohistochemical staining. The mRNA levels were monitored with FQ-PCR.

RESULTSGene therapy was much more effective than single epigenetic therapy and combined therapy. The gene therapy group has the lowest tumor growth rate and the highest expression levels of p53 and E-cadherin.

CONCLUSIONSThe combined treatment of gene and epigenetic therapy is not suggested for treating head and neck carcinoma, because gene therapy shows an antagonistic effect to epigenetic therapy. However, the mechanisms of action are still unclear.

Animals ; Azacitidine ; analogs & derivatives ; therapeutic use ; Cadherins ; analysis ; DNA Modification Methylases ; antagonists & inhibitors ; Epigenesis, Genetic ; Genes, p53 ; Genetic Therapy ; Humans ; Laryngeal Neoplasms ; genetics ; pathology ; therapy ; Male ; Mice ; Mice, Inbred BALB C ; Tumor Suppressor Protein p53 ; analysis ; Xenograft Model Antitumor Assays

OBJECTIVETo report the results of clinical characteristics, enzyme activity determination and mutation analysis of GLB1 gene in a Chinese patient with mucopolysaccharidosis (MPS) type IVB (Morquio B disease).

METHODA 14-year-old Chinese boy with MPS type IVB was firstly diagnosed by blood leucocytes galactosamine-6-sulfate sulfatase (GALNS) and β-galactosidase (GLB1) determination, who was characterized by short stature, multiplex skeletal abnormalities, difficulty in walking. PCR-sequencing analysis was applied to detect the mutations in GLB1 of the patient.

RESULTThe patient was characterized by dwarfism, pectus carinatum, kyphosis, normal intelligence, and no neurologic damage of spasms, linguistic capacity and so on. The patient had normal GALNS enzyme activity and very low GLB1 enzyme activity [5.03 nmol/(h·mg) vs. normal value 118 - 413 nmol/(h·mg) ] in leukocytes. A compound heterozygous missense mutations c.442C > T(p.R148C)/c.1454A > G(p.Y485C) in GLB1 gene were detected in this patient. The mutation p.Y485C is a novel variant. With the method of gene analysis of new variant, the mutation p.Y485C was considered to be a pathogenic mutation.

CONCLUSIONThe MPS IVB patient showed severe multiple skeletal deformities, normal intelligence, no neurologic damage and very low GLB1 enzyme activity, who carries compound heterozygous mutations p.R148C/p.Y485C. The mutation p.Y485C in GLB1 gene may be a novel pathologic mutation of MPS type IVB.

Adolescent ; Amino Acid Sequence ; Asian Continental Ancestry Group ; genetics ; Chondroitinsulfatases ; genetics ; metabolism ; DNA Mutational Analysis ; Humans ; Joints ; pathology ; Male ; Molecular Sequence Data ; Mucopolysaccharidosis IV ; enzymology ; genetics ; pathology ; Mutation, Missense ; Pedigree ; Polymerase Chain Reaction ; Radiography ; Spine ; diagnostic imaging ; pathology ; beta-Galactosidase ; genetics ; metabolism

OBJECTIVEAsthma is considered as a typical psychosomatic disease. This study aimed to investigate the temperament of asthmatic children and risk factors for asthma.

METHODSTemperamental type and dimensionality were investigated by Carry Temperament Scale in 106 children with asthma. Logistic regression analysis was used to study the risk factors for the development of asthma. One hundred and six age and sex-matched normal children served as controls.

RESULTSThere were significant differences in the adaptability, mood value and attention persistence of temperament between asthmatic patients and normal controls. Higher proportion of inter-high difficult temperamental type (17.0% vs 5.7%) and lower proportion of easy temperamental type (16.0% vs 29.2%) were found in children with asthma when compared with controls (P < 0.01). Logistic regression analysis showed that the frequency of cold between 3 and 7 years old, allergic history, idiosyncratic physique, parental history of asthma, house decoration and mood value and attention persistence of temperament were risk factors for the development of asthma.

CONCLUSIONSThere were differences in the temperamental type and dimensionality between asthmatic children and normal controls. Children with inter-high difficult temperament and suffered from the above risk factors showed a higher risk for developing asthma.

Adaptation, Psychological ; Affect ; Asthma ; etiology ; psychology ; Attention ; Child ; Child, Preschool ; Humans ; Logistic Models ; Risk Factors ; Temperament

OBJECTIVETo study the relativity between La protein and the stability of HBV mRNA and the expression of HBV protein.

METHODSFour specific siRNAs were obtained by transcription in vitro. After transfection with the siRNAs into HepG2.2.15 cells for 3 days, the inhibitive effects of La protein were analyzed by Western blot; the content changes of HBsAg, HBeAg and HBV-DNA were detected by ECL and RT-PCR.

RESULTSIn comparison to normal cells, La protein was less in the cells. There was less La protein in the cells trans-infected with siRNAs. HBsAg, the HBeAg and HBV-DNA secreted by the cells transfected with siRNA were also less than that in the normal cells.

CONCLUSIONThere is a correlation between La protein and HBV mRNA and the expression of HBV protein.

Autoantigens ; metabolism ; Cell Line, Tumor ; DNA, Viral ; Hepatitis B Surface Antigens ; Hepatitis B virus ; genetics ; metabolism ; Humans ; RNA, Messenger ; genetics ; metabolism ; RNA, Small Interfering ; RNA, Viral ; Ribonucleoproteins ; metabolism