The β-Glucuronidase gene (sbGUS) cDNA firstly from Scutellari abaicalensis leaf was cloned by RT-PCR, with GenBank accession number KR364726. The full length cDNA of sbGUS was 1 584 bp with an open reading frame (ORF), encoding an unstable protein with 527 amino acids. The bioinformatic analysis showed that the sbGUS encoding protein had isoelectric point (pI) of 5.55 and a calculated molecular weight about 58.724 8 kDa, with a transmembrane regions and signal peptide, had conserved domains of glycoside hydrolase super family and unintegrated trans-glycosidase catalytic structure. In the secondary structure, the percentage of alpha helix, extended strand, β-extended and random coil were 25.62%, 28.84%, 13.28% and 32.26%, respectively. The homologous analysis indicated the nucleotide sequence 98.93% similarity and the amino acid sequence 98.29% similarity with S. baicalensis (BAA97804.1), in the nine positions were different. The expression level of sGUS was the highest in root based on a real-time PCR analysis, followed by flower and stem, and the lowest was in stem. The results provide a foundation for exploring the molecular function of sbGUS involved in baicalcin biosynthesis based on synthetic biology approach in S. baicalensis plants.
Amino Acid Sequence ; Base Sequence ; Cloning, Molecular ; Computational Biology ; Glucuronidase ; chemistry ; genetics ; metabolism ; Molecular Sequence Data ; Open Reading Frames ; Phylogeny ; Plant Proteins ; chemistry ; genetics ; metabolism ; Protein Structure, Secondary ; Scutellaria baicalensis ; chemistry ; enzymology ; genetics ; Sequence Alignment

OBJECTIVETo observe the three-dimensional distribution of vessels, and to establish a new method for measurement of blood flow velocity in mice cerebral cortex using two-photon laser scanning microscopy and fluorescence probe labeling technique.

METHODSThe mouse was made cranial window surgery and injected Texas-Red through tail vein after anesthetized. The three-dimensional imaging of vessel was obtained through z-stack scanning, and blood flow velocity was quantified through line scanning.

RESULTSWe could detect vascular distribution for more than 500 µm depth using two-photon microscopy. The velocity of blood flow was (0.59 ± 0.12) mm/s in capillary.

CONCLUSIONThe method for observing the brain blood flow by two-photon microscopy was established, which could achieve quantification of single vascular blood flow velocity and provide experimental evidence for basic research and medical applications.

Animals ; Blood Flow Velocity ; Brain ; blood supply ; Capillaries ; Cerebrovascular Circulation ; Fluorescent Dyes ; Hemodynamics ; Mice ; Microscopy, Fluorescence

Phytantriol (PT), ethanol (ET) and water were used to prepare in situ cubic liquid crystal (ISV2). The pseudo-ternary phase diagram of PT-ET-water was constructed and isotropic solution formulations were chosen for further optimization. The physicochemical properties of isotropic solution formulations were evaluated to optimize the composition of ISV2. In situ hexagonal liquid crystals (ISH2) were prepared based on the composition of ISV2 with the addition of vitamin E acetate (VitEA) and the amount of VitEA was optimized by in vitro release behavior. The phase structures of liquid crystalline gels formed by ISV2 and ISH2 in excess water were confirmed by crossed polarized light microscopy and small angle X-ray scattering, respectively. Rheological properties of ISV2 and ISH2 were studied by a DHR-2 rheometer. In vitro drug release studies were conducted by using a dialysis membrane diffusion method. Pharmacokinetics was investigated by determination of sinomenine hydrochloride (SMH) concentration in synovial membrane after intra-articular injection of SMH-loaded ISH2 in adjuvant-induced arthritis rats. The optimal ISV2 (PT/ET/water, 64 : 16 : 20, w/w/w) loaded with 6 mg x g(-1) of SMH showed a suitable pH, injectable and formed a cubic liquid crystalline gel in situ with minimum water absorption in the shortest time. The optimal ISV2 was able to sustain the drug release for 144 h. The optimal ISH2 system was prepared by addition of 5% VitEA into PT in the optimal ISV2 system. This ISH2 (PT/VitEA/ET/water, 60.8 : 3.2 : 16 : 20, w/w/w/w) was an injectable isotropic solution with suitable pH. The new ISH2 was able to sustain the drug release for more than 240 h. Local pharmacokinetics study indicated that the retention time and AUC(0-∞) of ISH2 group were increased significantly compared with that of SMH solution group and the AUC(0-∞) of ISH2 group was 6.01 times higher than that of SMH solution group. The developed ISH2 was suitable for intra-articular injection that may apply to patients in the treatment of rheumatoid arthritis.
Animals ; Chemistry, Pharmaceutical ; Diffusion ; Ethanol ; Fatty Alcohols ; Gels ; Injections, Intra-Articular ; Liquid Crystals ; Morphinans ; administration & dosage ; chemistry ; Rats ; Rheology ; Water ; alpha-Tocopherol

Objective To investigate dynamic changes of heme oxygenase-1 and carbon monoxide in acute liver injury induced by carbon tetrach loride(CCl_4)in rats.Method Male SD rats were randomly allocated to induce acute liver injury by CCl_4 injection.Hepatic HO activity was examined at different time point following CCl_4 treatment.Expression and location of HO-1 protein was determined by western blot and immunohistochemical methods.Serum ALT,AST levels and hepatic SOD,MDA concentrations were also analyzed.Results Administration of CCl_4 to rats caused a marked hepatic damage,characterized by significant elevation of serum ALT,AST levels and liver MDA content combined with a remarkable reduction in liver SOD activity.HO activity was elevated significanfly in a time-dependant manner after CCl_4 injection,while the expression of HO-1 protein increased remarkably from 6 to 36 hours.CO concentration in the liver homogenate of control rats remained very low but was elevated significantly after CCl_4 treatment,which was in accordance with changes of HO-1. Conclusions HO-1 activity and protein expression as well as CO production are higher in rats with acute liver injury induced by CCl_4 than in control group.HO-1/CO system plays an important role in the pathogenesis of acute hepatic damage and may have potent protective effect against liver injury.

Objective To observe the clinical efficacy of acupoint thread embedding in preventing urinary retention after cervical cancer surgery.Method Fifty eligible patients were randomized into a treatment group and a control group, 25 cases each. The control group was given catheterization after gynecological surgery; the treatment group was intervened by thread embedding at Zusanli (ST36), Sanyinjiao (SP6), Yinlingquan (SP9), Guanyuan (CV4), and Shenshu (BL23) on the 7th day after the surgery in addition to the basic intervention given to the control group. The indwelling catheter was removed on the 14th day after the surgery, and the bladder function, urination, and residual urine volume were compared between the two groups.Result After the treatment, the bladder function in the treatment group was significantly better than that in the control group (P<0.05); the markedly effective rate of spontaneous micturition was 92.0% in the treatment group versus 72.0% in the control group, and the difference was statistically significant (P<0.05).Conclusion Early intervention with acupoint thread embedding after cervical cancer surgery can effectively decrease the incidence of post-surgery urinary retention, and it's easy-to-operation and worth application in clinic.

Objective To investigate the effect of glutathione peroxidase-1 (GPX1) on cispatin resistance in endometrial carcinoma cells and its related mechanism.Methods The HEC-1-A cisplatin resistance (HEC-1-A CR)cell was constructed by intermittent concentration gradient method.The control HEC-1-A cell was constructed with equal amount of normal saline.Lentivirus transfection was used to up-regulate the expression of GPX1 in the HEC-1-A CR cell as HEC-1-A CR GPX1 cell,and blank vector transfected was regarded as HEC-1-A CR control cell.The half maximal inhibitory concentration (IC50) value of cisplatin in cells was detected by thiazolyl blue (MTS) assay.The expression of mRNA and protein of GPX1 and HIF-1 α were detected by fluorescent quantitative chain reaction (qPCR) and Western blotting.Results The IC50 of HEC-1-A cells and HEC-1-A CR cells were (2.13 ±0.24),(7.89 ± 1.17)μg · mL-1.The IC50of HEC-1-A CR control cells and HEC-1-A CR GPX1 cells were (7.91 ±0.98),(3.87 ±0.52)μg · mL-1.The expression level of GPX1 mRNA in HEC-1-A cells and HEC-1-A CR cells were 0.14 ±0.01,0.07 ±0.01.The expression level of HIF-1α mRNA in HEC-1-A cells and HEC-1-A CR cells were 0.32 ±0.03,0.72 ±0.08.The expression level of GPX1 mRNA in HEC-1-A CR control cells and HEC-1-A CR cells were 0.09 ± 0.02,0.24 ± 0.03.The expression level of HIF-1 α mRNA in HEC-1-A CR control cells and HEC-1-A CR cells were 0.76 ±0.08,0.54 ±0.05.The above indexes were compared between groups,the difference was statistically significant (all P ＜ 0.05).The results of the changes of GPX1 and HIF-1 α protein levels were in accordance with the changes of mRNA levels.Conclusion GPX1 can enhance the sensitivity of cisplatin in endometrial cancer cells via down-regulating the expression of HIF-1 α.

A two-dimensional liquid chromatography method was developed for the analysis of rice leaves proteomics based on the coupling of hydrophilic interaction liquid chromatography-reversed-phase liquid chromatography with online tandem mass spectrometry. The influence of pH value of chromatographic mobile phase on the orthogonality of the hydrophilic interaction-reversed-phase two-dimensional liquid chromatography was evaluated by the changes of standard peptide retention. The results indicated that the better orthogonality (R2=0.34113) was achieved from the system with hydrophilic interaction columns(pH 9.3) in the first and C18columns(pH 3.3) in the second LC dimension. Coupled with multiple fraction concatenation strategy,the orthogonality of two-dimensional liquid chromatography was further evaluated in the analysis of complex rice leaf proteins. The results showed that more than 50% of the total peptides were identified less than two times, and the peptides obtained from first-dimension were well distributed across the elution window,indicating that the method showed significant orthogonality in the identification of complex rice leaf proteins. Based on the proteome discoverer software,207345 peptides belonged to 2930 protein clusters were identified.

OBJECTIVETo assess the diagnostic value of core needle biopsy (CNB) before neoadjuvant chemotherapy for breast cancer.

METHODSOne hundred and nineteen breast cancer cases underwent neoadjuvant chemotherapy in our hospital during the period from June, 2005 to January, 2007 were analyzed. CNB was carried out before starting chemotherapy. The hematoxylin and eosin-stained slides of CNB taken before and after neoadjuvant chemotherapy were reviewed independently by two pathologists, and the rate of consistency was verified.

RESULTSAmongst the 119 cases studied, 110 cases were confirmed to be carcinoma, including 105 cases of invasive carcinoma and 5 cases of ductal carcinoma-in-situ. The rate of consistency was 97.22% (105/108).

CONCLUSIONCNB has important value in distinguishing benign from malignant lesions, as well as in confirming the diagnosis of invasive carcinoma before starting neoadjuvant chemotherapy.

Biopsy, Needle ; methods ; utilization ; Breast ; pathology ; Breast Neoplasms ; diagnosis ; drug therapy ; Carcinoma, Ductal, Breast ; diagnosis ; pathology ; Female ; Humans ; Neoadjuvant Therapy ; methods

OBJECTIVETo investigate the histological criteria of breast cancer response to neoadjuvant therapy.

METHODSOne hundred and fifty-four cases of breast cancer receiving neoadjuvant therapy were collected from June, 2005 to June, 2007 and the clinical data were analyzed. All patients were operated on within 4 weeks after neoadjuvant therapy. All specimens were assessed by the standard method of Miller and Payne (MP) grading system. The response to neoadjuvant therapy were assessed by two pathologists independently, using MP grading system and common grading system separately.

RESULTSThe response rate using the MP grading system were grade 1 in 12 cases (7.8%), grade 2 in 33 cases (21.4%), grade 3 in 64 cases (41.6%), grade 4 in 31 cases (20.1%) and grade 5 in 14 cases (9.1%). Using the common grading system, the response were mild in 51 cases (33.1%), moderate in 71 cases (46.1%) and severe in 32 cases (20.8%). MP grading system may be related to common grading system (chi2 = 186.660, P < 0.01). Follow up information were available in 147 cases, with 14 cases showing recurrence, metastasis or death from the disease. The MP grading system may be related to the outcome (chi2 = 11.612, P = 0.020), but not the common grading system (chi2 = 0.881, P = 0.644).

CONCLUSIONMP grading system may be one of the prognostic factors in the neoadjuvant therapy of breast cancer.

Biopsy, Needle ; Breast ; pathology ; Breast Neoplasms ; drug therapy ; pathology ; surgery ; Carcinoma, Ductal, Breast ; drug therapy ; pathology ; surgery ; Female ; Follow-Up Studies ; Humans ; Mastectomy, Modified Radical ; Mastectomy, Radical ; Neoadjuvant Therapy ; Neoplasm Recurrence, Local ; Preoperative Period

OBJECTIVETo investigate the infection rate and subtypes of human papilloma virus(HPV) in patients with oropharyngeal squamous cell carcinoma (OSCC) and analyze the clinicopathologic features of patients with or without HPV infection.

METHODSA total of 66 biopsy or surgical specimens of OSCC archived in the Pathology Department of Chinese Academy of Medical Sciences were analyzed by polymerase chain reaction (PCR), and the generic amplification products were detected by DNA enzyme immunoassay (DEIA) and typed by reverse hybridization line probe assay.

RESULTSHPV-DNA was detected in 11 (16.7%) of all specimens. Among them, 7 were infected with HPV-16,and the remaining 4 patients were infected with HPV-16/11, HPV-35, HPV-58/52, and HPV-33/52/54, respectively. HPV-16 was detected in 72.7% of all positive specimens. There were more females in HPV-positive group than HPV-negative group (36.4% vs. 1.8%,P=0.002). Patients with HPV-positive tumors were more likely to be non-smokers (36.4% vs. 0,p=0.001) and non-drinkers (45.5% vs. 1.8%,p=0.001) than those with HPV-negative tumors. The proportion of moderately or poorly differentiated tumors was higher in HPV-positive patients than HPV-negative patients (81.8% vs. 63.7%), although without statistical significance (p=0.409). No difference was observed in T classification, N classification, and overall tumor stage.

CONCLUSIONSHPV infection rate was 16.7% in this cohort. HPV-positive OSCC has its unique etiologic and clinicopathological characteristics.

Adult ; Aged ; Aged, 80 and over ; Carcinoma, Squamous Cell ; virology ; DNA, Viral ; isolation & purification ; Female ; Humans ; Male ; Middle Aged ; Oropharyngeal Neoplasms ; virology ; Papillomaviridae ; classification ; isolation & purification ; Papillomavirus Infections ; virology