Objective To evaluate the possibility and efficiency of nanoparticles as a new vector in vascular endothelial growth factor (VEGF) gene transfection. Methods Nanoparticle-VEGF (Np/VEGF)complex was prepared with poly (D, L-lactide-co-glycolide) (PLGA) loading VEGF165 gene using the multiple emulsion (w/o/w) technique. The envelopment efficiency and size of the complex were determined. Rat myocardial cells were cultured in vitro, and the Np/VEGF was transfected into the cultured myocardial cells. Then RT-PCR and ELISA were used to evaluate whether the Np/VEGF increased the level of gene expression. Four New Zealand rabbits were used, the suspension of Np/VEGF was injected into myocardial tissue of rabbits after thoracotomy. 96h after the operation, the tissue sections of the implant sites were observed with transmission electron microscope (TEM) to determine the process of nanoparticles as vectors for gene transfer to cardiac myocytes. Results The envelopment efficiency and size of the Np/VEGF complex thus prepared were 1.87% and 25-300nm respectively. RT-PCR and ELISA showed that VEGF gene could be successfully transfected into myocardial cells by nanoparticle, and NP/VEGF significantly enhanced gene transfection efficiency, and it was more effective than plasmid. 96h after the operation, a great number of nanoparticles were observed in myocardial cytoplasm and nucleus with TEM, and many nanoparticles began to dissolve and degrade, suggesting that the DNA was released slowly from the nanoparticles localized in the cytoplasmic compartment, and was then transferred into the nucleus. Conclusions NP/VEGF can act as a vector to transfect VEGF gene in vitro and in vivo, it significantly enhanced gene transfection efficiency, and it was more effective than plasmid.

Antineoplastic Agents ; administration & dosage ; adverse effects ; Asparaginase ; administration & dosage ; adverse effects ; Child ; Child, Preschool ; Combined Modality Therapy ; Dose-Response Relationship, Drug ; Drug Hypersensitivity ; prevention & control ; Female ; Humans ; Hyperglycemia ; chemically induced ; prevention & control ; Male ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; drug therapy ; Severity of Illness Index ; Time Factors ; Treatment Outcome

Objective To investigate the effect of microRNA-21 (miR-21) antisense oligonucleoti-de on collagen synthesis in the rat hepatic satellite cells ( HSCs) .Methods Rat hepatic stellate cells were isolated and cultured; the miR-21 antisense oligonucleotide was transfected into HSCs with lipofectamine 2000;after incubation 48 h, the HSCs were collected.The expression of miR-21 was detected with reverse transcription polymerase chain reaction ( RT-PCR) , andα-smooth muscle actin (α-SMA) protein in HSCs with Western blot.The cell proliferation was assayed with methyl thiazolyl tetrazolium ( MTT) method.Re-sults Compared to scrambled control group, the expression of miR-21 was reduced by 76%( P <0.01), the proliferation activity of HSCs was reduced by(26 ±3)%( P <0.01), the expressions of type I and III collagen proteins were reduced by(61 ±7)%and (48 ±6)%( P <0.01).Conclusions The miR-21 an-tisense oligonucleotide could reduce significantly the expression of miR-21, and inhibit HSC proliferation and extracellular matrix production.

Objective To investigate the effect of miRNA-200b-specific inhibitor on hepatic stellate cells(HSCs) activation,proliferation,and extracellular matrix production.Methods The miRNA-200b-specific inhibitors were designed,synthesized,and transfected into HSCs with lipofectamine 2000.The supernatant and HSCs were collected after incubation for 48 h.The expression of miR-200b was detected by quantitative reverse transcription polymerase chain reaction (qRT-PCR).The expression ofα-smooth muscle actin (oα-SMA) protein in HSCs was detected by Western blotting.The cell proliferation was assayed by methyl thiazolyl tetrazolium (MTT) method.Contents of type Ⅲ procollagen and hyaluronic acid in supernatant were determined by radioimmunoassay.Results Compared to the control group,miRNA-200b expression was decreased in the miRNA-200b inhibitor group by 82％ (P ＜ 0.01),α-SMA protein expression was reduced in the miRNA-200b inhibitor group by (19 ± 3) ％ (P ＜ 0.05),and the activity of HSCs proliferation was reduced by(33 ± 5)％ (P ＜0.01),and the contents of type Ⅲ procollagen and hyaluronic acid in supernatant were reduced in miRNA-200b inhibitor group by (35 ± 4)％ and (31 ± 2)％,respectively(P ＜0.01).Conclusions The miRNA-200b-specific inhibitor could significantly reduce the expression of miRNA-200b,and inhibit HSC proliferation,activation,and extracellular matrix production.

OBJECTIVEExtracellular signal-regulated kinases (ERKs) can be activated by calcium signals. In this study, we investigated whether calcium-dependent kinases were involved in ERKs cascade activation after global cerebral ischemia.

METHODSCerebral ischemia was induced by four-vessel occlusion, and the calcium-dependent proteins were detected by immunoblot.

RESULTSLethal-simulated ischemia significantly resulted in ERKs activation in N-methyl-D-aspartate (NMDA) receptor-dependent manner, accompanying with differential upregulation of Src kinase and Ca2+/calmodulin-dependent protein kinase II (CaMKII) activities. With the inhibition of Src family tyrosine kinases or CaMKII by administration of PP2 or KN62, the phosphorylation of ERKs was impaired dramatically during post-ischemia recovery. However, ischemic challenge also repressed ERKs activity when Src kinase was excessively activated.

CONCLUSIONSrc family tyrosine kinases and CaMKII might be involved in the activation of ERKs mediated by NMDA receptor in response to acute ischemic stimuli in vivo, but the intense activation of Src kinase resulted from ischemia may play a reverse role in the ERKs cascade.

Analysis of Variance ; Animals ; Brain Ischemia ; enzymology ; pathology ; Calcium-Calmodulin-Dependent Protein Kinase Type 2 ; Calcium-Calmodulin-Dependent Protein Kinases ; metabolism ; Disease Models, Animal ; Extracellular Signal-Regulated MAP Kinases ; metabolism ; Gene Expression Regulation ; physiology ; Hippocampus ; cytology ; enzymology ; Male ; Neurons ; enzymology ; pathology ; Phosphorylation ; Rats ; Rats, Sprague-Dawley ; Receptors, N-Methyl-D-Aspartate ; metabolism ; Signal Transduction ; physiology ; Statistics, Nonparametric ; src-Family Kinases ; metabolism

OBJECTIVETo observe the clinical efficacy of Wangbi Tablet (WT) on the treatment of knee osteoarthritis (KOA), and to study its treatment program by integrative medicine.

METHODSThe randomized, parallel control, and multi-center clinical observation method was used. 160 KOA patients of Gan-Shen deficiency complicated with stasis-blood blockade syndrome were assigned to three groups. Group A (72 cases) was treated by WT, Group B (42 cases) was treated by Votalin Tablet, and Group C (46 cases) was treated by WT + Votalin Tablet. All patients were treated for two months. Before and after treatment the clinical symptoms, the knee function activity, adverse reactions were observed and recorded. Erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) of each patient were respectively detected, statistically analyzed and compared.

RESULTSAfter eight weeks of treatment significant difference was shown in the time of morning stiffness in joints, joint tendemess, swelling, pain, joint functional activities, ESR, and CRP between Group A and Group B (P<0.01). Besides, better effects were obtained in Group C (P<0.01). Improvement of ESR and CRP: They were obviously improved in the three groups after treatment (P<0.01). As for the markedly effective rate, better effects were obtained in Group C (P<0.05).

CONCLUSIONSIn the treatment of KOA, WT showed favorable effects with no obvious adverse reaction. The combination of WT and Voltaren Tablet was significantly superior to use of WT or Votalin Tablet alone. It could reduce the dosage of Voltaren Tablet and avoid adverse reactions of the gastrointestinal tract.

Adult ; Aged ; Drugs, Chinese Herbal ; therapeutic use ; Female ; Humans ; Integrative Medicine ; Male ; Middle Aged ; Osteoarthritis, Knee ; drug therapy ; Phytotherapy ; Tablets

To observe therapeutic effects of acupuncture on female's climacteric depression and to study on the mechanism. Methods Sixty cases enrolled were randomly divided into an acupuncture group and a control group, 30 cases in each group. In the acupuncture group, acupuncture were given at Ganshu (BL 18), Shenshu (BL 23), Xinshu (BL 15), etc. with uniform reinforcing-reducing method, once each day, and the control group were treated with oral administration of fluexertine hydrochloride, 20 mg, once daily. HAMD scale was used for assessment before treatment and 2,4,6 weeks after treatment. Blood dopamine (DA), norepinephrine (NE) and 5-hydroxy-indoleacetic acid (5-HIAA) contents were detected before treatment and after one therapeutic course. Results The total effective rate was 86.7% in the acupuncture group and 92.9% in the control group with no significant difference between the two groups (P > 0.05). After treatment, DA content increased significantly in the acupuncture group with a significant difference as compared with that of the control group (P < 0.05); and after treatment NE and 5-HIAA contents in the two groups significantly increased as compared with that before treatment (P < 0.05, P < 0.01), with no significant difference between the two groups (P > 0.05). Conclusion Acupuncture can benignly and comprehensively regulate general functions, and increase contents of monoamines in the body, so as to play the role of anti-depression.
Acupuncture Therapy ; Adult ; Climacteric ; psychology ; Depression ; blood ; therapy ; Dopamine ; blood ; Female ; Humans ; Hydroxyindoleacetic Acid ; blood ; Medicine, Chinese Traditional ; Middle Aged ; Norepinephrine ; blood

0bjective To establish a standard curve method with more accuracy employed in fluorescence real-time PCR(RT-PCR)as a alternation of the general method.Methods β-actin and KLK11 plasmid DNA for quantitative standard curve were constructed in our study,and Plasmids of β-actin was employed as a internal control.After serial dilution these plasmid were used as DNA standard to obtained slope.Expression of these two genes in malignant prostate cancer cell line LNCAP were tested by real-time PCR,and we analyzed the RT-PCR results with two different methods and compared their accuracy.Results Thestandards curves made from these linear DNA standards showed good linearity (R2=0.991 and 0.992 for β-actin and KLK11 standards graphs),but also displayed a discrepancy in their PCR efficiency(β-actin 123% and KLK11 99%).There were different results after two different stand curve analytical method:the expression of KLK11 mRNA in LNCAP was downregulated in general standard curve method.In the new analytical method,howerer,KLK11 upregnlated for 4.46-fold.And there was a significant difference between aplification efficiency of targt gene and internal control gene(t=4.829,P<0.05).Conclusions Compared with general standard curve method,the new advanced standard curve method described here avoids an error which considers there is identical amplification efficiency between target gene and internal reference gene.It is considered to be a more correct analytical method in fluorescence real-time PCR.

ObjectiveToevaluatethefeasibilityandresultsof transorallaparoscopic thyroidectomy. MethodThyroidectomy was attempted in 5 cases,including 4 females and 1 male with the average age of 42 years( range 35 -60 years).All patients was diagnosed as single nodule of the thyroid gland confirmed by B-mode ultrasound examination before the operation.The average diameter of nodule was 2.5 cm (range 2 - 3.4 cm). ResultTransoral laparoscopic thyroidectomy was perfoormed successfully in all 5 patients.The operation time was 120 - 210 min,averaging 170 min,blood loss during the operation was 15-60 ml,the postoperative hospitalization was 4 days. There was no conversion to open surgery,no recurrent laryngeal nerve injury,nor parathyroid glands dysfunction. ConclusionsTransoral laparoscopic thyroidectomy is feasible and safe,giving excellent cosmetic results.