Screening of Chinese Herbal Medicines Resistant to Chicken Escherichia coli and Infectious Laryngotracheitis Virus

[Objective] This study aimed to screen Chinese herbal medicines resistant to Chicken Escherichia coli and infectious laryngotracheitis virus. [Methed] Conven- tional punch method, test tube method and plate dilution method were adopted for in vitro susceptibility test of chicken E, coil strains O5 and O8 using 13 kinds of Chi- nese herbal medicines including Sanguisorba officinalis, Coptis chinensis, Anemar- rhena asphodeloides, Strobilanthes cusia, Agastache rugosa, etc.; chicken embryo inoculation experiment was adopted to screen Chinese herbal medicines resistant to chicken infectious laryngotracheitis virus. [Result] Sanguisorba officinalis, Fructus mume, Rheum officinale, Coptis chinensis, Herba Taraxaci, Anemarrhena asphode- Ioides, Scutellaria baicalensis and Rhizoma Fagopyri Cymosi had ideal antibacterial effect against chicken E. coil strain O5; Sanguisorba officinalis, Fructus mume, Rheum officinale, Coptis chinensis, Herba taraxaci and Rhizoma Fagopyri Cymosi had ideal antibacterial effect against chicken E. coil strain 08; other Chinese herbal medicines showed relatively poor or no antibacterial effect. Results of chicken embryo inoculation experiment showed that nine kinds of Chinese herbal medicines showed relatively strong anti-lLTV effect, including Forsythia suspensa, Radix Isatidis, Fofium isatidis, Flos Ionicerae, Radix codonopsis, Radix astragali, Atractylodes, Radix gly- cyrrhizae, and Pericarpium granati. [Conclusion] The study laid the foundation for fur- ther development of Chinese herbal compound preparations to treat chicken cofibacil- Iosis, infectious laryngotracheitis and other bacterial, viral diseases.

A review on in vitro corrosion performance test of biodegradable metallic materials

Extensive in vitro corrosion test systems have been carried out to simulate the in vivo corrosion behavior of biodegradable metallic materials. Various methods have their own unique benefits and limitations. The corrosion mechanism of biodegradable alloys and in vitro corrosion test systems on biodegradable metallic materials are reviewed, to build a reasonable simulated in vitro test system for mimicking the in vivo animal test from the aspects of electrolyte solution selection, surface roughness influence, test methods and evaluation methodology of corrosion rate. Buffered simulated body fluid containing similar components to human blood plasma should be applied as electrolyte solution, such as simulated body fluid （SBF） and culture medium with serum. Surface roughness of samples and ratio of solution volume to sample surface area should be adopted based on the real implant situation, and the dynamic corrosion is preferred. As to the evaluation methodology of corrosion rate, different methods may complement one another.

Modulatory Effect of Pollen Extract on Apoptosis of Lymphocytes in the Elderly

Aim In this study, we investigated the changes of lymphocyte subpopulationand apoptosis process of lymphocytes in the elderly, and the modulatory effect of pollen extract(PE) on apoptosis. Methods Lymphocyte phenotypes were detected using indirect immunofluorescencetechnique. The proliferation responses were determined by MTT method. Flow cytometry and automaticimage analysis were performed to evaluate the apoptosis of lymphocytes. Results The proliferationresponses of lymphocytes in the elderly were lower than that in young adults. Decreased CD_(45) RA^+cells and increased CD_(45) RO^+ cells were found in lymphocyte population of aged people, comparedwith that of young adults. The CD_(45) RO^+ cells were prone to apoptosis. There is an inhibitoryeffect of PE on apoptosis of lymphocytes in the elderly. Conclusion It is implied that thesusceptibility of lymphocyte in the elderly to apoptosis depends on activation, so called,activation-induced cell death. Present results suggest that apoptosis of lymphocytes in aged peopleplay an important role in the pathogenesis of immunosenescence. Thus, a possibility is open fordevelopment of apoptosis-modulating drugs from pollen.

在发达国家通过检测宫颈涂片中的Brn-3a发现宫颈异常

We have previously shown that Brn- 3a is elevated in biopsies from women in the United Kingdom with high grade cervical neoplasia, and that it specifically trans-activates the HPV URR in vitro and in vivo. The aim of this study is to establish the relationship of Brn- 3a, HPV E6 and pathological diagnosis in cervical smear from women in a developing country with a high prevalence of cervical disease. This is a follow-up of our previous work in which for the first time Brn- 3a and E6 levels in cervical smears from women in United Kingdom were shown to correlate with the histological diagnosis of cervical neoplasia and were even better in predicting underlying pre-malignant disease than conventional procedures. Method. Cervical smears from 295 women with cervical abnormalities attending gynecological clinics in Brazil were used to make RNA. The mRNA levels of Brn- 3a and HPV E6 were measured and the data obtained were used to establish the relationship between Brn- 3a and the histological diagnosis. Results. The cellular transcription factor Brn- 3a was readily measured in cervical smears from the Brazilian population. Its presence was shown to be frequently associated with the expression of HPV E6. The measured level of Brn- 3a parallels the severity of the cervical ailment and predicts the pathological categories. Conclusions. The ability of Brn- 3a to predict for cervical ailments is independent to the geographical characteristics of the population, and hence it could be used routinely as an adjunct to colposcopy and pathological diagnosis in developing and developed countries.

The culture and establishment of embryonic germ(EG)cell lines from Chinese mini swine

As a part of a basic research project on Xeno-transplantion. we have been engaged in the derivation of embryonic stein cell lines from Chinese mini swine. Here, we reported for the first time the establishment of two porcine EG cell lines (BPEG1 and BPEG2) from primordial germ cells of genital ridges of a 28 and a 27 d embryos respectively. Their pluripotent nature has been identified by colony morphology, marker characterization as well as by in vitro and in vivo differentiation. These porcine EG cells are potentially useful for further basic studies.

Inhibition of hepatitis B virus replication by pokeweed antiviral protein in vitro

AIM: To explore the inhibitory effects of pokeweed antiviral protein seed (PAP-S) and PAP encoded by a eukaryotic expression plasmid on hepatitis B virus (HBV) replication in vitro. METHODS: HepG2 2.2.15 cells in cultured medium were treated with different concentrations of PAP-S. HBsAg, HBeAg and HBV DNA in supernatants were determined by ELISA and fluorescent quantitative PCR respectively. MTT method was used to assay for cytotoxicity. HepG2 were cotransfected with various amounts of PAP encoded by a eukaryotic expression plasmid and replication competent wild-type HBV 1.3 fold over- length plasmid. On d 3 after transfection, HBsAg and HBeAg were determined by using ELISA. Levels of HBV core-associated DNA and RNA were detected by using Southern and Northern blot, respectively. RESULTS: The inhibitory effects of PAP-S on HBsAg, HBeAg and HBV DNA were gradually enhanced with the increase of PAP concentration. When the concentration of PAP-S was 10 μg/mL, the inhibition rates of HBsAg, HBeAg and HBV DNA were 20.9%, 30.2% and 50%, respectively. After transfection of 1.0 μg and 2.0 μg plasmid pXF3H-PAP, the levels of HBV nucleocapside- associated DNA were reduced by 38.0% and 74.0% respectively, the levels of HBsAg in the media by 76.8% and 99.7% respectively, and the levels of HBeAg by 72.7% and 99.3% respectively as compared with controls. Transfection with 2 μg plasmid pXF3H-PAP reduced the levels of HBV nucleocapside-associated RNA by 69.0%.CONCLUSION: Both PAP-S and PAP encoded by a eukaryotic expression plasmid could effectively inhibit HBV replication and antigen expression in vitro, and the inhibitory effects were dose-dependent.

Optimization for Production of Intracellular Polysaccharide from Cordyceps ophioglossoides L2 in Submerged Culture and Its Antioxidant Activities in vitro

Cordyceps ophioglossoides is a valuable traditional medicinal material.We have found that intracellular polysaccharide(IPS) is the major biologically active ingredient in Cordyceps ophioglossoides.This study is the first time to optimize the yield of IPS from Cordyceps ophioglossoides.The optimal medium for IPS production consists of glucose 54.50 g·L·1,yeast powder 25.50 g·L·1,NaH2PO4 0.4 g·L·1 and K2HPO4 0.4 g·L·1.The suggested culture conditions are 24 ℃,initial pH 4.5 with a rotary speed of 120 r·min·1 for 168 h.The yield of IPS is 737.93 mg·L·1,which is 50% higher than the yield under the conditions prior to optimization.The anti-oxidative activities of IPS in Cordyceps ophioglossoides L2 are also characterized using various in vitro assay.The anti-oxidative activity may explain the reason why IPS from Cordyceps ophioglossoides can be used to fight against neurodegenerative dis-eases and menopausal symptoms.

Synthesis and anticancer effects of 6-nitro-4-anilinoquinazolines and 6-amino-4-ani-linoquinazolines

Objective: To synthesize inhibitors of the epidermal growth factor receptor tyrosine kinase such as 6-nitro-4-anilinoquinazolines and 6-amino-4-anilinoquinazolines,and to compare their anticancer effects in vitro. Methods: The 4-anilinoquinazolines compounds were prepared by hydrolyzed, ringed, halagenated, substituded in turn from 2-amino-5-nitrobenzylcarbonitril. The synthesized 4- anilinoquinazoline compounds has been rudimentarily screened by using A431 tumor cell line which overexpresses epidermal growth factor receptor as model adopted MTT method. Results: Five 6-nitro-4-halo-sbstituted anilinoquinazolines and five 6-amino-4-halo-substituted anilinoquinazolines have been obtained,and all of them had anticancer activity. The anticancer activity of 6-amino substituted inhibitors was higher than that of 6-nitro substituted inhibitors. However, the difference of anticancer activity between two series of quinazoline was much less than that of their inhibiting EGFR tyrosine kinase activity. Conclusion: The probable reason for 6-nitro-4-anilinoquinazolines having anticancer activity in vitro was that they had been partially transformed to 6-amino-4-anilinoquinazolines through endocellular cytochrome oxidation-reduction system.

Design and activity evaluation of deoxyribozymes specifically targeting hepatitis C virus RNA

Objective: To explore the cleaving and inhibitory activity of hepatitis C virus ( HCV) -specific deoxyri-bozymes (DRz) at both molecular and transgeneic cellular levels. Methods: According to the secondary structure of HCV 5'-noncoding region (5'-NCR) and the sites characterized with 5'…Y ↓ R…3'(Y = A/G,R = U/C) , HCV-spe-cific naive deoxyribozymes were designed and named DRz-232, DRz-127, DRz-84, DRz1, and the phosphorothioate deoxyribozymes (PSDRz) and mutated phosphorothioate deoxyribozymes (MPSDRz) were also designed. HCV RNA 5'-NCR was transcribed in vitro from linearized plasmid pHCV-neo and radiolabelled at its 5'-end. DRz, PSDRz or MPSDRz was respectively mixed with the substrate RNA and incubated under appropriate conditions, the cleaved products were displayed by 8% denaturated polyacrylamide gel electrophoresis (PAGE) and autoradiography, and the optical density of each band was measured to calculate cleavage rates. After that, every kind of DRz was added respectively to the cultured transgeneic HepG2 cells containing luciferase gene controlled by HCV 5'-NCR. The cells were lysed at intended time points and the activity of luciferase was measured with chemiluminescence method for calculating inhibition rates. Results: After incubated for 90 min in vitro, the cleavage rates of DRz-127, PSDRz-127, DRz1 and PS-DRz1 reached 32.6% , 30. 8% , 24. 3% and 21. 5% , respectively. No cleavage product was observed in any MPSDRz. DRz-127, PSDRz-127, DRz1 and PSDRzl had an inhibitory rate of 53. 2% , 50. 6% , 44. 7% and 43. 3% respectively in transgeneic HepG2 cells in the first 24 h when the final dose of the DRz was 0. 5μmol/L, higher than that of the corresponding MPSDRz. There was no significant difference between the inhibitory effect of each DRz and its PSDRz in HepG2 cells, but the inhibitory rate of DRz decreased more rapidly than that of the latter with the elapse of time. The results from transfection groups were significantly better than those of non-transfection groups. Conclusion: Rationally-designed HCV-specific deoxyribozymes are able to cleave target RNA at molecular level in vitro, and efficiently inhibit the expression of luciferase gene controlled by HCV 5'-NCR in transgeneic cells. Appropriate PSDRz may be more stable, and thus more suitable than the naive DRz in the application to cells. Introduction of the deoxyribozymes with transfection is more efficient than with direct delivering ways.

In-vitro and In-vivo Anti-trypanosomal Activity of Terminalia chebula Retz Dried Fruits

African trypanosomosis had caused lots of havocs to both humans and animals over a century with successes and failure in curtailing it. This study was aimed at screening medicinal plant, Terminalia chebula dried fruits against Trypanosoma evansi for trypanocidal activity. Twenty grams of powdered Terminalia chebula dried fruits was cold extracted with methanol. Obtained MPE （methanolic plant extract） was in vitro tested against Trypanosoma brucei （1 × 10^6 trypanosomes/mL of the medium in each ELISA plate wells） at concentrations （250～1,000 μg/mL） on Vero cells grown in DMEM （Debecco＇s Modified Eagle Medium） in appropriate conditions for trypanocidal activity. In-vitro cytotoxicity test of MPE of Terminalia chebula was conducted on Vero cells grown in DMEM. In-vivo assay for trypanocidal activity, each mouse was inoculated with 1 × 10^4/mL of trypanosomes and treated （48 h post inoculation） with MPE of Terminalia chebula at concentrations （12.5, 25, 50, 100 and 200 mg/kg body weight） were administered at dose rate of 100 BL per mouse via intraperitoneal route to different groups of mice, 6 mice per concentration. In-vitro cytotoxicity test was done on Veto cells at concentrations （1.58～100 μg/mL） of MPE of Terminalia chebula. Results of in-vitro trypanocidal activity varied from immobilization, reduction and to the killing of the trypanosomes. At 250 μg/mL ofMPE ofTerminalia chebula dried fruits, there was significant trypanocidal activity at 4 h of incubation and trypanosomes were not detected in corresponding ELISA plate wells at 5 h of incubation, which was statistically equivalent to reference drug, diminazine aceturate （50 μL/mL） at 4 h of incubation. Results of in-vivo trypanocidal activity revealed that at concentrations （l 2.5～25 mg/kg body weight） of MPE of Terrninalia chebula, mice in these groups survived for 6 days. While at 50 and 100 to 200 mg/kg body weight, mice in these groups survived up to 7 and 8 days, respectively. In-vitro cytotoxicity test showed that all concentrations of MPE of Terminalia chenula and diminazine aceturate were cytotoxic to cells except at 1.56 μL/mL and 6.25 μL/mL. In conclusion, MPE of Terminalia chebula dried fruits possessed trypanocidal compounds. Further study （bioassay-guided purification） is required to know the full potential of Terrninalia chebula as future trypanocide candidate.

Impact of MTT based tumor chemosensitivity assay in vitro

Objective： The aim of the study was to evaluate the impact of the chemosensitivity assay in vitro and establish a standard process of measuring the anti-cancer drug sensitivity with MTT assay. Methods： Some influencing factors of MTT assay in studying the sensitivity of human peripheral blood mononuclear cells （PBMC） to anti-cancer drugs were observed, including red blood cells, platelets, three different kinds of DMSO and different concentrations of MTT. Meanwhile the stability of tumor drug-coated plate was monitored. Results： The red blood cells and platelets may affect the results at a certain range of concentration. Analytical pure DMSO, both imported and domestic reagents showed the same color with MI3-, and the A values of the reaction were dependent on MTT dose. The stability of the freeze-drying drug-coated plates was superior to non freeze-drying ones. Conclusion： To make clear and definite all kinds of influencing factors might contribute to a kind of standard MTT assay for drug sensitivity test in vitro.

Comparison of the unlabeled and labeled pre-mRNA splicing assays in vitro

Pre-mRNA splicing is a fundamental process required for the expression of most metazoan genes. It is carried out by the spliceosome that catalyzes the removal of non-coding intron sequences to ligate exons into mature mRNA prior to transport and translation. The purpose of our study is to explore whether the in vitro unlabeled pre-mRNA splicing assay could be performed as an alternative method of splicing reaction other than the radiolabeled one. Two different splicing methods in vitro, 32p labeled and unlabeled pre-mRNA as the substrates in the reaction, were investigated. The radiolabeled products were visualized by autoradiography while the unlabeled products were observed by Ethidium Bromide （EB） staining. As a result, although there are more unspecific bands in the EB staining assay than 32p labeled one, the RNA products of in vitro splicing could be observed clearly. This suggests that the unlabeled pre-mRNA splicing assay can be an optional substitution for the isotope-labeled assay.

Effect of Plumeria alba＇s Bioactive Extract against Colletotrichum gleosporoides in Vitro and in Vivo

Plant species in the genus Plumeria of Apocynaceae family are well known for their uses in traditional medicine antimicrobial activity. Plant materials from P. alba were extracted using Soxhlet apparatus and screened for the control of anthracnose disease caused by Colletotrichum gleosporoides on fruit during post harvest storage. For in vitro, the crude extract of P. alba （stem bark and leaves） were completely inhibited conidial germination （sporulation test method） of Colletotrichum gleosporoides at all concentration tested except the lowest concentration （4.4 uL）. In vivo studies show that percentage of infection on fruit after treated with both stem bark （11.11 ± 3.8） and leaves （15.55 ± 3.8） part, at 32,000 mg/L have no significant differences during seven days of storage at ambient temperatures. These results proved that P. alba extracted （stem bark and leaves） were found to be promising as an antifungal agent and to control anthracnose disease in fruits tested.

Study on the Current Physical Quality of Modem College Students and the Countermeasures

In recent years, the unexpected accidents of college students have occurred frequently in the playground. A student in Shanghai Sanda University suddenly fell at tile basketball class at the end of last year. In the same month, a male student in Shanghai East China University fell on the ground when taking part in the physical test of lkm running, and then passed away after the rescue was not valid. In this paper, the physical quality of the current college students is discussed on the basis of studying college students＇ quality education. This study is expected to play a role in reference.

Pro-protein convertase-2/carboxypeptidase-E mediated neuropeptide processing of RGC-5 cell after in vitro ischemia

Objective To observe the change of the neuropeptide pro-protein processing system in the ischemic retina ganglion cell-5(RGC-5) cells,pro-protein convertase-2(PC2),carboxypeptidase-E(CPE) and preproneuropeptide Y(preproNPY) protein levels in the ischemic RGC-5 cells and conditioned medium were analyzed. Methods The RGC-5 cell was differentiated in 0.1 μmol/L staurosporine for 24 h and then stressed by different doses of oxygen and glucose deprivation(OGD). The acute or chronic OGD-induced cell death rates w...

Comparison of dissolution profile characteristics of 11 berberine hydrochloride tablet brands in different dissolution media

Berberine hydrochloride is commonly used to treat bacterial dysentery,gastroenteritis and other diseases.Many manufacturers are available on the market today,while the production process and formulation are quite different,which may directly affect the therapeutic effect of the drug.To this end,11 different production producers of berberine hydrochloride tablets were collected according to the pharmacopeia berberine hydrochloride dissolution method(basket method).In addition the dissolution process was carried out in four elution media with different pH,and the difference was similar(f2).Factors were calculated to evaluate in vitro dissolution requirements,and in vitro dissolution of different manufacturers of berberine hydrochloride tablets was determined by high performance liquid chromatography(HPLC).The method was verified by linearity,precision,stability and robustness.Based on the f2 value,there was a significant difference in the dissolution behavior of the formulations of most berberine hydrochloride tablet brands.This research provided the basis for further in-depth research in the later period.Although the drug specifications(0.1 g)were the same,the dissolution curve was different.This phenomenon may be attributed to the fact that the excipients and crystal form of the tablets affected the release and dissolution of the tablets in vitro.

Effects of Chinese herbs combined with in vitro fertilization and embryo transplantation on infertility:a clinical randomized controlled trial

OBJECTIVE： To assess the effects of using Chinese herbs in assisted reproductive technology. METHODS： Four hundred and thirty-three subjects aged less than 42 years with infertility due to Fallo- pian tube or male-related factors who were willing to undertake in vitro fertilization and embryo trans- plantation were randomly allocated to a Chinese herb intervention group （n=216） or a conventional treatment control group （n=217）. All subjects re- ceived one of four routine ultra-ovulation-promot- ing therapies at the Reproductive Center in the Third Hospital Affiliated to Beijing University ac- cording to their physician＇s assessments. The sub- jects in the intervention group received various Chi- nese herbs depending on their conventional treat- ment. Endometrial thickness, number of acquired eggs, and rates of normal fertility, high-quality em- bryos, biochemical and clinical pregnancy of sub- jects were assessed in both groups.RESULTS： The high-quality embryo rate of 51.9%, biochemical pregnancy rate of 51.0%, clinical preg- nancy rate of 44.2% and endometrial thickness of （10.84± 1.75） mm in the intervention group were all significantly higher than those in the control group [48.7%, 38.9%, 34.8%, and （10.52±1.50） mm, respec- tively; P〈O.05]. The normal fertility rate of 58.5% in the Chinese herb group was also significantly supe- rior to the 54.7% achieved in the control group （P〈 0.01）. There were no statistically significant differ- ences （P〉0.05） in the average number of acquired eggs within a single cycle, incidence of excessive stimulation of ovary, rates of embryo transplanta- tion or early abortion and birth of living babies be- tween the two groups. CONCLUSION： Our findings indicate that Chinese herbs increase endometrial thickness, improve the quality of fertility and embryo, and promote embry- onic nidation, thus enhancing the success rate of in vitro fertilization/intracytoplasmic sperm injec- tion-embryo transplantation cycle. Using Chinese herbs improves the outcomes and safety of assist- ed reproductive technologies.