Aim To amplify P30 gene and express P30 fusion with GST Methods P30 gene was smplified from T. gondii chromosomal DNA and ligated to pGEM-T and pGEX-4T-1. Screening-positive recombinants were induced for expression, which was subsequently detected by WB Results P30 gene was amplified and GST-fusion was confirmed by rabbit antiT. gondii serum. Conclusions The construction of pGEM-T-P30 and pGEX-4T-1-P30, together with the recombinant protein would lay a base for further investigation of P30 at a molecule-level and application to diagnosis and vaccination

AIMTo establish an assay method for the determination of three kinds of biologically active components, five compounds (emodin, chrysophanol, baicalin, wogonin and berberine hydrochloride) simultaneously in Sanhuang tablets.

METHODSHPLC was carried out, using a C18 column (150 mm x4. 6 mm ID, 5 microm) set at 30 degrees C, acetonitrile-0.02 mol x L(-1) acetic ammonium (adjusted pH to 3. 50 with acetic acid glacial) as mobile phase (using gradient) with flowing rate 1. 00 mL x min(-1) and detected at 270 nm.

RESULTSThe calibration curve of emodin was linear from 0.020 7 microg to 0.207 microg with r = 0.999 9, the average recovery was 99.65% with RSD 1.25%. The calibration curve of chrysophanol was linear from 0.052 microg to 0.52 microg with r = 0.999 9, and the average recovery was 100.36% with RSD 0.96%. The calibration curve of baicalin was linear from 0.250 5 microg to 2.505 microg with r =0.999 8, and the average recovery was 100.22% with RSD 1.29%. The calibration curve of wogonin was linear from 0.047 6 microg to 0.476 microg with r = 0.999 9, and the average recovery was 98.97% with RSD 1.20%. The calibration curve of berberine hydrochloride was linear from 0.053 12 microg to 0.531 2 microg with r = 0.999 5, and the average recovery was 96.02% with RSD 2.02%. The established method had also been used in the determination of the 5 compounds in 10 different batches of Sanhuang tablets.

CONCLUSIONThis method was proved to be accurate and quick, and can be used for the quality control of the preparation all-around.

Anthraquinones ; analysis ; Berberine ; analogs & derivatives ; analysis ; Chromatography, High Pressure Liquid ; methods ; Coptis ; chemistry ; Drug Combinations ; Drugs, Chinese Herbal ; administration & dosage ; chemistry ; isolation & purification ; Emodin ; analysis ; Flavanones ; analysis ; Flavonoids ; analysis ; Plants, Medicinal ; chemistry ; Quality Control ; Reproducibility of Results ; Rheum ; chemistry ; Scutellaria baicalensis ; chemistry ; Tablets

The mainly antigenic sites for the adenovirus neutraliation are present on Loop1 and Loop2 of hexon.Majority research were focus in the human adenovirus.Little was known on infectious canine hepatitis virus (ICHV), which was also called canine adenovirus typeⅠ.Here,ICHV (the isolated strain) DNA was isolated and purified from the cultured MDCK cells.The Loop1 and Loop2 fragments were amplified by polymerase chain reaction(PCR) method,and then was connected by ligase T4.The target fragment was then connected with vector pET28a.The nucleotide sequence ecoding Loop1 and Loop2 was determined.The nucleotide sequence identity of Loop1 region between the isolated strain and CLL, RI261 and Toronto A26/61 strains is 100%, 100% and 83.8%, and the nucleotide sequence identity of Loop2 region between the isolated strain and CLL, RI261 and Toronto A26/61 strains is 88.1% , 88.1% and 99.3%, and amino acid identity is 93.6%, 93.6% and 98.6%.The recombinant Loop protein was expressed in E.coli and was approximately 36kDa in size,and then was purified. Then BALB/c mice were injected subcutaneously in the back and armpit with the recombinant Loop protein.The anti-ICHV antibody titers of immunized serum was tested by indirect ELISA and the titers were up to 1:320.Western blot demonstrated that immunized sera could specifically combine with ICHV. The research laid a foundation for creating new genetic engineering products of infectious canine hepatitis virus.

Testis specific serine/threonine protein kinase 4 (TSSK4) belongs to the TSSK family, and its members play an important role in spermatogenesis and/or spermiogenesis. Mouse TSSK4 has been reported to be expressed exclusively in the testis and can maintain its kinase activity through autophosphorylation at Thr-197. However, its biological function remains poorly understood. Here we found that GFP-TSSK4-overexpressed HeLa cells showed apoptotic bodies, indicating TSSK4 can lead to apoptosis in vitro. Furthermore, TSSK4 induced apoptosis in different cell lines including HeLa, Cos-7 and H1299 tested by flow cytometry but not its kinase-dead mutant TSSK4-K54M. TSSK4 knockout mice showed increased testes weight and decreased apoptotic spermatogonia and spermatocytes at 21st day after birth tested by TUNEL technology. So TSSK4 was able to induce cell apoptosis in vitro depending on its kinase activity, which leads to abnormal testes weight and apoptosis, shedding light on its function in the process of spermatogenesis and/or spermiogenesis.
Animals ; Apoptosis ; physiology ; Base Sequence ; Cell Line ; DNA Primers ; Flow Cytometry ; Humans ; In Situ Nick-End Labeling ; Male ; Mice ; Polymerase Chain Reaction ; Protein-Serine-Threonine Kinases ; physiology

The fine structure of enoxaparin sodium samples with different degree of 1,6-anhydro derivatives were analyzed with polyacrylamide gel electrophoresis, high performance liquid chromatography, ultraviolet spectroscopy, infrared spectroscopy and nuclear magnetic resonance spectroscopy. A further study of anticoagulation activity of enoxaparins was performed, including those on their inhibition activities of coagulation factor Xa (FXa) and thrombin (FIIa). The results showed that the anti-FXa and -FIIa activities of enoxaparins with different degree of 1,6-anhydro derivatives (20.0%-39.7%) with similar structure characteristics, had decreasing tendency when the degree of 1,6-anhydro derivatives increased. Especially, the anti-FXa activity was sensitive to the change of the degree of 1,6-anhydro derivatives.
Anticoagulants ; chemistry ; Enoxaparin ; chemistry ; Factor Xa Inhibitors ; chemistry ; Thrombin ; antagonists & inhibitors

The aim of this research is to refine the protocol of purification of SA and identify the character of SA. By utilizing the cold-denaturing method, most of other kinds of protein were screened out and SA was purified from the fermentation broth of L-183 by using the refined affinity chromatography method. The rate of recollection was checked to be 75%～85%. By identification, it is indicated that the molecular weight of self-made SA was 74.5kD, the biotin-combining number 3.2, the activity 11.2u/mg, the pI around 7.4. So, the essential characters of SA are same as described by documents.

OBJECTIVETo study the cost-effectiveness, benefit and utility of infant hepatitis B vaccination in Shanghai from 1992 to 2001.

METHODSTo calculate the cost of hepatitis B vaccination by cost analysis method. Both the numbers of persons with HBsAg positive and patient with hepatitis B, cirrhosis and liver cancer decreased as the index of direct effect. To study the sick-time and the cost of treating hepatitis B, cirrhosis and liver cancer patients, a face to face questionnaire was used and quasi method was adopted to understand the effect of cure and the course of hepatitis B. The cost benefit analysis method was also used to calculate the cost benefit of HBV vaccine. The disability adjusted life years (DALY) was regarded as an index of utility to measure the disease burden.

RESULTSInput of 501,129.49 Yuan might have the result of reducing one liver cancer patient, ten cirrhosis patients, one hundred chronic hepatitis B patients and one thousand HBsAg positive people. The cost of hepatitis B vaccination was 0.24 hundred million Yuan during the past ten years in Shanghai, which had obtained the total benefit value of 41.22 hundred million Yuan, with a cost benefit ratio of 1:172 Yuan. It was estimated that the total disease burden of hepatitis B, cirrhosis and liver cancer patients was 59,762.55 DALY in order to reduce one DALY loss cost of 402.50 Yuan.

CONCLUSIONHBV vaccine inoculation in infants seemed to be a low-cost input and high-effect output strategy.

Adult ; Cost-Benefit Analysis ; Female ; Hepatitis B ; complications ; economics ; prevention & control ; Hepatitis B Vaccines ; economics ; therapeutic use ; Humans ; Immunization Programs ; economics ; Infant, Newborn ; Liver Cirrhosis ; economics ; etiology ; Liver Neoplasms ; economics ; etiology ; Male ; Markov Chains ; National Health Programs ; Quality of Life ; Surveys and Questionnaires ; Vaccination

OBJECTIVETo observe amplitude changes of low frequency fluctuation in brain spontaneous nervous activities induced by needling at Hand Taiyin Lung Channel, and to preliminarily explore the possible brain function network of Hand Taiyin Lung Channel.

METHODSBy using functional magnetic resonance imaging (fMRI), 16 healthy volunteers underwent resting-state scanning (R1) and scanning with retained acupuncture at Hand Taiyin Lung Channel (acupuncture, AP). Data of fMRI collected were statistically calculated using amplitude of low frequency fluctuations (ALFF).

RESULTSUnder R1 significantly enhanced ALFF occurred in right precuneus, left inferior parietal lobule, bilateral superior temporal gyrus, bilateral middle frontal gyrus, left superior frontal gyrus, left inferior frontal gyrus, left medial frontal gyrus. Under AP significantly enhanced ALFF occurred in right precuneus, bilateral superior frontal gyrus, cerebellum, bilateral middle frontal gyrus, right medial frontal gyrus, and so on. Compared with R1, needing at Hand Taiyin Lung Channel could significantly enhance ALFF in right gyrus subcallosum and right inferior frontal gyrus. Significant decreased ALFF appeared in right postcentral gyrus, left precuneus, left superior temporal gyrus, left middle temporal gyrus, and so on.

CONCLUSIONNeeding at Hand Taiyin Lung Channel could significantly change fixed activities of cerebral cortex, especially in right subcallosal gyrus, right inferior frontal gyrus, and so on.

Acupuncture Therapy ; Brain ; physiology ; Brain Mapping ; Humans ; Magnetic Resonance Imaging

OBJECTIVETo observe the effect of acupuncture at the whole points of Hand Jueyin pericardium meridian on the amplitude of low frequency fluctuations (ALFF) of healthy people in resting state (R1) functional magnetic resonance imaging (fMRI).

METHODSTotally 16 healthy subjects received structure scan of T1 and T2. Then two fMRI scans were conducted for each participant. fMRI included the resting-state scan (R1; the scanning time was 8 min 6 s), the stimulating-acupoint scan (AP; the scanning time was 8 min 6 s). fMRI data acquisition from structure scanning and function scanning were processed with format conversion and statistical analysis.

RESULTSUnder R1 state, brain regions with activated ALFF signals included bilateral superior frontal gyrus, medial frontal gyrus, middle occipital gyrus, precuneus, superior temporal gyrus, and cingulate gyrus. Under the AP state, brain regions with activated ALFF signals were bilateral superior frontal gyrus, medial frontal gyrus, middle temporal gyrus, left fusiform gyrus, precuneus, posterior cingulate, and declivis. Compared with R1 state, obvious difference of ALFF signal areas of the brain caused by acupuncture at pericardium were: bilateral cuneus, precuneus, left posterior cingulate gyrus, right middle occipital gyrus, and right occipital lingual gyrus.

CONCLUSIONAcupuncture at the whole points of Hand Jueyin pericardium meridian could significantly change inherent activity states of the cerebral cortex, especially in bilateral superior frontal gyrus, medial frontal gyrus, and precuneus.

Acupuncture ; Acupuncture Points ; Brain ; physiology ; Brain Mapping ; Frontal Lobe ; Humans ; Magnetic Resonance Imaging ; methods ; Pericardium

Objective To investigate the serum alkaline phosphatase (ALP) and γ-glutamyl transpeptidase (GGT) levels in patients with neuromyelitis optica (NMO) and multiple sclerosis (MS).Methods Serum ALP and GGT levels in patients with NMO and MS from the database of demyelinating diseases in our hospital were analyzed.Eighty-five healthy controls were chosen.The differences of serum ALP and GGT levels in patient groups and controls were compared, and the correlations between clinical features (age of the subjects, course of disease, times of relapse and scores of EDSS) and both ALP and GGT levels were analyzed. Results The serum ALP and GGT levels in patients with NMO were significantly higher than those in patients with MS and the controls (P<0.05).Patients with NMO still had significantly higher serum ALP level in acute phase than patients with MS,and the serum ALP levels in male patients with NMO and the serum GGT levels in female patients with NMO were, respectively,statistically higher than that of male patients with MS and female patients with MS (P<0.05). In patients with NMO, significantly positive correlations between serum GGT level and both age and times of relapse were noted (P<0.05). Conclusion Serum ALP and GGT levels differ in patients with NMO and MS, indicating their differential diagnostic value in NMO and MS to certain extent.