Progress and perspective of silkworm as a model of human diseases for drug screening / 药学学报

As a typical representative of Lepidopteran insects, the silkworm, Bombyx mori, has numerous advantages, such as simple husbandry, highly prolific nature, short generation time, easily handled to be operated with moderate body size, clear genetic background and abundant mutation resources. Silkworm has not only been studied by the geneticists, but also been used as a new laboratory animal model of human disease and drug screening. There is a plenty of genetic resources in silkworm, some of which could be used as models of human genetic diseases, such as Phenylketonuria, Parkinson 's disease, Hermansky-Pudlak syndrome and so on. Silkworm has also played a significant role in the study of pathogenesis of human pathogenic microorganisms. Moreover, silkworm could be used to evaluate the pharmacokinetic/pharmacodynamics properties and safety of a new drug comprehensively and systematically. At the same time, it can be used in the high throughput drug screening assays to shorten the period of the new drug research and development. This review summarizes that the silkworm is an excellent model in the drug screening assays, and has a potential in application to the large-scale drug screening.

Validation of a modified fluorimetric assay for the screening of trichomonacidal drugs

A fluorimetric microassay that uses a redox dye to determine the viability of the flagellate Trichomonas vaginalis has been optimised to provide a more sensitive method to evaluate potential trichomonacidal compounds. Resazurin has been used in recent years to test drugs against different parasites, including trichomonadid protozoa; however, the reproducibility of these resazurin-based methods in our laboratory has been limited because the flagellate culture medium spontaneously reduces the resazurin. The objective of this work was to refine the fluorimetric microassay method previously developed by other research groups to reduce the fluorescence background generated by the media and increase the sensitivity of the screening assay. The experimental conditions, time of incubation, resazurin concentration and media used in the microtitre plates were adjusted. Different drug sensitivity studies against T. vaginalis were developed using the 5-nitroimidazole reference drugs, new 5-nitroindazolinones and 5-nitroindazole synthetic derivatives. Haemocytometer count results were compared with the resazurin assay using a 10% solution of 3 mM resazurin dissolved in phosphate buffered saline with glucose (1 mg/mL). The fluorimetric assay and the haemocytometer counts resulted in similar percentages of trichomonacidal activity in all the experiments, demonstrating that the fluorimetric microtitre assay has the necessary accuracy for high-throughput screening of new drugs against T. vaginalis.

The effect of brucine on hepatocellular carcinoma cell lines in vitro / 中华普通外科杂志

Objective To study the effect of brucine on the growth of a hepatocellular carcinoma cell line in vitro. Methods Brucine was added into a liver cancer cell line of SMMC-7721 in vitro, at drug concentration of brucine from 2. 5 μg/ml to 400 μg/ml. The inhibition rate of cell growth was measured by MTT technique after the cells were cultured for 72 hours. The protein and mRNA expression of PCNA,cyclin D1 and FAS were respectively assayed with Western blotting and fluorescent quantitation RT-PCR techniques at 24, 48, 72 h. Results The inhibition rate of liver cancer cell was near 100% when the brucine concentration was at 320 μg/ml. The protein and mRNA expression of FAS were of no significant difference at 24 h vs 48 h ( seperately F = 2. 547,1. 582, all P ＞ 0. 05 ), and significant difference existed at 24 h vs 72 h( seperately F = 1. 036, 1. 137, all P ＜ 0. 05 ). The protein and mRNA expression of PCNA,Cyclin D1 were of no significant difference between various time period( seperately PCNA F = 3.612,2. 174,3.029;Cyclin D1 F=2.361,2.915,1.976,all P＞0.05). Conclusions Brucine inhibits the growth of liver cancer cells, by inducing increased apoptosis of the cells probably through FAS overexpression.

Heterogeneous Chemosensitivity of Breast Cancer Determined by Adeonsine Triphosphate Based Chemotherapy Response Assay / 한국유방암학회지

PURPOSE: Breast cancer is heterogeneous disease and the response to chemotherapeutic agents is also heterogeneous from patient to patient. Chemotherapy response assay is in vitro test that is performed to evaluate the degree of tumor growth inhibition by chemotherapy drugs. In this study, we performed the chemotherapy response assay using adenosine triphosphate (ATP-CRA) in breast cancer patients and assessed the clinical availability. METHODS: Sixty five breast cancer patients were enrolled in this study. Cancer cells were evenly divided and treated with commonly used chemotherapeutic drugs in breast cancer (doxorubicin, epirubicin, 5-fluorouracil, paclitaxel, docetaxel, vinorelbine, and gemcitabine). To verify in vitro ATP-CRA indirectly, we analyzed the correlation between cell death rate (CDR) of doxorubicin and epirubicin, and between doxorubicin and paclitaxel. We also analyzed the mean CDR of doxorubicin, epirubicin and paclitaxel by HER2 status. RESULTS: We could successfully perform the ATP-CRA in 60 patients (95.2%). In all cases, we can get the results within 7 days. The range of CDR was very wide, from 0 to more than 50%, except gemcitabine. Epirubicin showed the highest mean CDR (39.9%) and doxorubicin, paclitaxel in order. According to the chemosensitivity index, paclitaxel is the most frequently first-ranked and doxorubicin, epirubicin in order. Correlation coefficient between the cell death rate of doxorubicin and epirubicin is 0.4210 and 0.1299 between paclitaxel and doxorubicin. In HER2 positive group, mean CDR of paclitaxel, epirubicin and doxorubicin was higher than in HER2 negative group, even though epirubicin and doxorubicin were not statistically significant (p=0.018, p=0.114, p=0.311, respectively). CONCLUSION: ATP-CRA showed heterogeneous results in individual patients. ATP-CRA was successful and can be performed within short time period. According to our in vitro study, it showed similar results with in vivo study but for the clinical use, the prospective randomized controlled trial should be preceded.

Application of ATP-TCA in clinic / 国际肿瘤学杂志

ATP-tumor chemosensitivity assay(ATP-TCA)is a tumor chemosensitivity assay in vitro.Beacause of the special merites of A11P-TCA,it is seemed to be the most promising and effective method of tumor chemosensitivity in vitro.Application of ATP-TCA in clinic has got significant effects.ATP-TCA is a good method for individualized chemotherapy.

Chemosensitivity testing for human gastric cancer with short-term culture cells vs purified primary culture cells using the MTT assay / 中国癌症杂志

Purpose:To evaluate the feasibility, advantages and disadvantages of chemosensitivity test of human gastric cancer using MTT assay with short term culture tumor cells contaminated by nonmalignant cells compared with purifiedly primary culture tumor cells.Methods:Fifty nine fresh samples from patients with gastric cancer were obtained from operating rooms. Chemosensitivity results were provided by the MTT assay with short term culture cells. The primary culture cells were purified by means of a series of methods such as repeated cell attachment, differential trypsinization and natural purification which removed fibroblasts and other nonmalignant cells. The same MTT assay was conducted using purified cells. Chemosensitivity results between short term method and purification method were compared for seven antitumor drugs. Results:The success rate of short term method using the MTT assay for chemosensitivity testing was 81 4% (48 of 59 patients),and for the purification method it was 50.8(30 of 59 patients). The average was 20.2?9.5 days when primary cells were cultured into purified cancer cells. The optical density ( A ) value correlated directly with the the number of tumor cells with the two methods( P

Relationship between the sensitivity of human colon carcinoma cells to gefitinib and the expressions of Akt and MAPK / 解放军医学杂志

Objective To investigate the relationship between the sensitivity of colon carcinoma cells to epidermal growth factor receptor(EGFR) inhibitor,gefitinib,and the downstream proteins of EGFR.Methods The expressions of EGFR proteins of 6 colon carcinoma cell lines(Lovo,HCT116,HT29,LS174T,SW480 and SW620) were determined with immunocytochemistry staining.The inhibitory effects of gefitinib on the growth of colon carcinoma cells were assessed by MTT,and the expression levels of Akt and MAPK as well as their phosphorylated forms(p-Akt and p-MAPK) were assessed by Western blotting.Results EGFR protein expressed in all the Lovo,HCT116,HT29,LS174T and SW480 cells,and the highest expression was found in Lovo cells,but not in SW620 cells.Lovo cells showed the highest,HT29 and SW480 cells showed moderate,sensitivity to gefitinib,while the others showed more or less resistance to gefitinib.No significant difference was found between the growth inhibition and IC50 values among the 6 cell lines despite of being treated with fetal bovine serum or EGF.Akt protein existed in all the cell lines without notable difference.Lovo and SW620 cells showed the least of p-Akt expression(P

Relationship between antitumor activity of Gefitinib and epidermal growth factor receptor expression in human colorectal cancer cell lines / 解放军医学杂志

Objective To investigate the antitumor activity of Gefitinib, a selected epidermal growth factor receptor-tyrosine kinase inhibitor, on human colorectal cancer cell lines in vitro, and to explore the relationship between the inhibitory effect of Gefitinib on cancer cells and the expression of epidermal growth factor receptor (EGFR). Methods The growth inhibitory effects of Gefitinib, which expressed as the half growth inhibition dose IC50, on colorectal cancer cells were assessed by MTT assay. EGFR mRNA expression was detected by reverse transcriptional PCR (RT-PCR). Western blot was used to determine the expression of EGFR protein as well as its phosphorylated forms (p-EGFR). Results Gefitinib inhibited growth of all the six colorectal cancer cell lines in vitro with an IC50 range from 6.5 to 172.7?mol/L. Lovo cell line, with an IC50 value less than 10?mol/L, was the most sensitive one to Gefitinib, HT29 and SW480 were moderate sensitive to 10?mol/L