Proteolytic degradation has been a severe problem when Pichia pastoris is employed to express recombinant proteins.One alternative method to circumvent this problem is to construct protease gene disruptant.However,the main study of gene disruption is focused on nonrecombinant Pichia pastoris rather than recombinant strain.In our study,we established two different methods to directly disrupt PRC1 and KEX1 gene in recombinant Pichia pastoris.On the basis of this,we further discussed and compared the application and advantages of both methods.

Adrenalectomy ; Adult ; Angiomyolipoma ; pathology ; surgery ; Follow-Up Studies ; Humans ; Kidney ; pathology ; Kidney Neoplasms ; pathology ; surgery ; Lymph Node Excision ; Lymph Nodes ; pathology ; Lymphatic Metastasis ; Male ; Nephrectomy ; Ureter ; surgery

OBJECTIVETo study the effect of solid dispersion technology and inclusion technology on dissolution performance of Pulsatillae total saponins, and preliminarily investigate its mechanism.

METHODThe solid dispersion of Pulsatillae total saponins-PEG 4000 was prepared by the melting method. The inclusion compound of Pulsatillae total saponins-hydroxypropyl-beta-cyclodextrin ( HP-beta-CD) was prepared by the freeze-drying method. The properties of solid dispersion and inclusion compound were identified by using IR, DSC and NMR. And the dissolution of solid dispersion and inclusion compound were also determined by the small glass method.

RESULTIR, DSC and NMR results showed the formation of solid dispersion and inclusion compound. In terms of the dissolution, the inclusion compound ranked first, which was followed by solid dispersion and bulk pharmaceutical chemicals.

CONCLUSIONThe inclusion technology could significantly increase the dissolution of Pulsatillae total saponins, whereas the solid dispersion showed no notable solubilization effect.

Drug Compounding ; methods ; Drugs, Chinese Herbal ; chemistry ; Saponins ; chemistry ; Solubility ; Spectrophotometry, Infrared

Objective To investigate the clinical diagnostic value of magnetic resonance tomographic angiography (MRTA) on cranial neurovascular compression syndrome,and evaluate the ability of 3D-FIESTA and 3D-TOF-SPGR sequences in demonstrating the relation of three-dimensional space between cranial nerves and blood vessels.Methods The data of 41 patients with cranial neurovascular compression syndrome,admitted to our hospital from May 2007 to May 2009,were analyzed.These patients were planed to perform micro vasular decompression (MVD).Before the operation,MRTA,3D-FIESTA and 3D-TOF-SPGR sequence scanning were performed to observe the relation of three-dimensional space between cranial nerves and blood vessels;these results were compared with the intraoperative results to evaluate the advantages and disadvantages of 3D-FIESTA and 3D-TOF-SPGR sequence scanning.Results MRTA could demonstrate such cranial nerves as trigeminal nerve,facial nerve and glossopharyngeal nerve,and responsible blood vessels clearly and simultaneously.The 3D-FIESTA imaging showed high signal in the cerebrospinal fluid and moderate signal in the nerves and blood vessels.The 3D-TOF-SPGR imaging showed low signal in the cerebrospinai fluid,moderate signal in the nerves and brain parenchyma,and high signal in the blood vessels.Closed relation between the nerves and the blood vessels in the lesion side were found in 34 patients (82.9%) by 3D-FIESTA sequence scanning,and that was found in 35 patients by 3D-TOF-SPGR sequence scanning; no significant difference between 3D-FIESTA and 3D-TOF-SPGR sequence scanning was found in displaying the relation of nerves and blood vessels (P>0.05).Conclusion MRTA technology may clearly show the relation of cranial nerves and responsible blood vessels;combined application of 3D-FIESTA and 3D-TOF-SPGR sequence scanning can help making the preoperative diagnosis and determining the surgical indications in patients with cranial neurovaseular compression syndrome.

OBJECTIVETo explore the influence of non-gonococcal Neisseria on the diagnosis and treatment of male genitourinary infection.

METHODSThe samples of urethral exudates, prostatic secretions or/and semen were collected from 8 cases of male patients with acute urethritis or chronic prostatitis, then inoculated into gonococcal agar medium, blood agar medium, Sabouraud agar medium and Mycoplasma agar medium, respectively. Neisseria gonorrhoeae, Mycoplasmae, fungi and other bacteria were isolated, Chlamydiae examined by Gemenez staining, and the gram-negative diplococci from the samples identified by oxidase test, biochemical examination and drug sensitivity test. The PCR products of the cryptic plasmid pJD1 gene of the isolated strains were amplified for the identification of Neisseria gonorrhoeae. Based on the results of drug sensitivity tests, intravenous or oral antibiotics were selected for treatment.

RESULTSEight strains of gram-negative diplococci were isolated in this study, 3 identified as N. mucosa, 4 as N. cinerea and the other 1 as N. lactamica. The PCR identification test of the cryptic plasmid pJD1 gene showed the same positive results in all the strains as in N. gonorrhoeae. The non-gonococcal Neisseria isolated from the male genital tract secretions exhibited a multidrug resistance, especially to quinolones and fosfomycin. All the symptoms disappeared and no pathogens were detected in the patients after a 7-day treatment with Cephalosporins or/and Minocycline.

CONCLUSIONSome Neisseria species normally parasitizing the upper respiratory tract can also cause male genitourinary infections, such as gonorrhea-like urethritis and chronic prostatitis. Neisseria gonorrhea could be clinically and etiologically misdiagnosed through such conventional methods as morphological examination, oxidase test and PCR identification test of cryptic plasmid and other nonspecific genes. Intravenous and/or oral antibiotic medication based on the results of drug sensitivity tests can cure acute urethritis and chronic prostatitis induced by non-gonococcal Neisseria in males. Drug resistance of non-gonococcal Neisseria directly affects the success of treatment.

Adult ; Anti-Bacterial Agents ; therapeutic use ; Humans ; Male ; Middle Aged ; Neisseria ; isolation & purification ; Neisseriaceae Infections ; diagnosis ; drug therapy ; microbiology ; Prostatitis ; diagnosis ; drug therapy ; microbiology ; Urethritis ; diagnosis ; drug therapy ; microbiology

Actins ; analysis ; Antigens, CD34 ; analysis ; Carcinoma, Papillary ; classification ; metabolism ; pathology ; Female ; Humans ; Immunohistochemistry ; Keratin-19 ; analysis ; Keratin-20 ; analysis ; Muscle, Smooth ; chemistry ; Pancreatic Neoplasms ; classification ; metabolism ; pathology ; Proto-Oncogene Proteins c-kit ; analysis ; Receptors, Estrogen ; analysis ; Receptors, Progesterone ; analysis

Objective To explore the necessity and teaching methods of the selective course:interpersonal relationships and communication skills.Methods Two hundred and two medical students were surveyed with a community questionnaire before and after this course.The questionnaire contained the purpose of taking the selective course, advice of teaching methods and their learning harvest.Results More than 90% students thought it was necessary to set up the course, and the purpose that they selected this course was to improve their abilities of interpersonal communications.92.9% students suggested the teaching methods should give prominence to the practice.93.9% students wished to adopt the method of composing the paper through the situation simulation or in combination with their true-life.97.9% students gained more harvest.Conclusions It is very necessary for medical students to set up the selective course: interpersonal relationships and communication skills.The teaching methods should take the medical students' difficulties of interpersonal communications into account and pay attention to cultivate their skills of practice.

OBJECTIVETo develop a headspace gas chromatography method for the determination of residual organic solvents in Panax notoginseng extracts.

METHODSThe samples were injected into HP-INNOWAX capillary column by headspace sampler and analyzed with FID detector using standard addition method.

RESULTSThere was a good linearity in the experimental concentration (r=0.9932-0.9999). The rate of recovery was in the rang of 81.74%-111.2%. The numbers of theoretical plates were more than 15000 and the resolutions between the adjacent peaks were more than 2. The RSD of precision and accuracy were both less than 10 %.

CONCLUSIONThe method is simple,accurate and sensitive with good reproducibility, which can be used for the determination of the residual organic solvents in Panax notoginseng extracts.

Chromatography, Gas ; methods ; Drug Contamination ; prevention & control ; Panax notoginseng ; chemistry ; Plant Extracts ; analysis ; Reproducibility of Results ; Solvents ; analysis

Objective To explore the protective effect of insulin-like growth factor 1 (IGF-1) on apoptosis of dopaminergic neurons induced by L-dopa via JAK/STAT signaling pathway. Methods PC12 cells were induced to differentiate into dopaminergic neurons with 100 μg/L β-NGF; MTT assay was employed to identify the changes in the viability of PC12 cells following L-dopa treatment at 0, 10,20, 50, 100, 150 and 200 μmol/L, and the different concentrations of IGF-1 at 0, 10, 25, 50 and 100 nmol/L with the same concentration of L-dopa (150 μmol/L); Western blotting was used to detect the levels of P-JAK2/P-STAT3 in PC12 cells treated with PBS (controls), L-dopa, L-dopa+IGF-1 and L-dopa+IGF-1+AG490 for 24 h, and then the apoptosis rate was assessed by flow cytometry and Hchest33258 staining. Results Western blotting showed that the expressions of P-JAK2 and P-STAT3 were detected in the L-dopa+IGF-1 and L-dopa+IGF-1+AG490 treatment groups but not in the control group or L-dopa treatment group; the expression of P-STAT3 in the L-dopa+IGF-1+AG490 treatment group was obviously lower than that in the L-dopa+IGF-1 treatment group (P<0.05). Hchest33258 staining indicated that L-dopa treatment group had the most obvious karyopyknosis and karyorrhexis,much more apoptotic bodies than the L-dopa+IGF-1 and L-dopa+IGF-1+AG490 treatment groups. Flow cytometry showed that the apoptosis rate was significantly different among the 4 groups (F=180.991,P=0.000): as compared with the control group, the other 3 groups had a higher apoptosis rate (P<0.05);L-dopa treatment group (38.13 ±2.54 %) enjoyed the highest level, followed by L-dopa+IGF-1 +AG490treatment group (25.60±1.30 %) and L-dopa+IGF-1 treatment group (20.17±1.54 %). Conclusion L-dopa has toxic effect on PC12 cells; IGF-1 could protect the PC12 cells from the neurotoxic effect of L-dopa and JAK2/STAT3 signaling pathway is activated in this procedure.

Objective To study the role of SIRT 1 in apoptosis of PC 12 neuronal cells induced by lipopolysaccharide (LPS). Methods PC12 cells were cultured with different concentrations of LS (50 μg/mL,500 μg/mL,750 μg/mL,1000 μg/mL and 1250 μg/mL),and some other PC12 cells were routinely cultured as controls. MTT assay was employed to identify the cell survival 24 h after the inducement,and accordingly,the suitable LPS concentration for subsequent experiments was determined based on MTT results. And then, cell apoptosis in the experimental groups under the suitable LPS concentration at different times (1/2,2,18,24,and 48 h) and control group was noted by flow cytometry and Hoechst 33258 staining; Western blotting was used to detect the SIRT1 level in PC12 cells. Results Hoechst 33258 staining indicated that a few apoptotic bodies were noted 1/2 h after inducement,expressing as karyopyknosis and karyorrhexis; apoptotic bodies began to increase 18 h after inducement,reaching their peak level 24 h after inducement; and a decreased trend was observed 48 h after inducement. Flow cytometry indicated that significantly higher apoptosis rate at each time point was noted as compared with that in the control group (P＜0.05); and Hoechst 33258 staining showed the same result. Western blotting revealed that the SIRT1 expression was (1.84±0.04) in the control group,decreasing to (1.17±0.09) 1/2 h after the inducement,and reaching the lowest level (0.62±0.03) 24 h after the inducement; and then, the expression was increased to (0.77±0.02) 48 h after the inducement;significant difference on the expression at each time point was noted as compared with that in the control group (P＜0.05). Conclusion LPS can induce PC12 cell apoptosis and SIRT1 protein expression is inhibited,indicating that SIRT1 may take part in the apoptosis and play a protective role to PC12 cells.