Objective To predict clinical chemotherapy sensitivity of primary non-small cell lung cancer（NSCLC） by methylthiazal （MTT） tumor chemosensitivity assay method in vitro and detection of multidrug resistance gene1 （MDR1）, and provide reference for clinical individualized treatment. Methods We selected 80 fresh primary NSCLC samples from NSCLC patients who underwent surgical resection in Zibo Central Hospital Affiliated to Binzhou Medical College between January 2009 and December 2011. There were 46 male patients and 34 female patients with their median age of 54 （29 to 81）years. Viable NSCLC cells obtained from malignant tissue were tested for their sensitivity to cisplatin （DDP）, gemcitabine （GEM）, docetaxe （DOC）, etoposide （VP-16） ,and vinorelbine （NVB） using MTT assay in vitro. Fluorescent quantitative reverse transcriptase-polymerase chain reaction （RT-PCR） was used to analysis the expression level of multidrug resistance gene1 （MDR1）. Results After exposure to antitumor drugs, morphologic changes, decrease of metabolic activity, and apoptosis were detected in NSCLC cells. MTT results showed that different individual cancer cells had different chemosensitivity to antitumor drugs, and cancer cells also had different chemosensitivity to different antitumor drugs. Inhibitory rates of cancer cells exposed to DOC, GEM, and VP-16 were significantly higher than those of cancer cells exposed to DDP and NVB （42.5%±9.5%, 40.5%±6.5%, 38.4%±7.6% versus 31.5%±8.5%，32.5%±7.8%, P＜0.05）.The positive rate of MDR1 in tumor tissues was 40.0% （32/80）. The expression of MDR1 was not associated with tumor histological type, degree of differentiation, lymph node metastasis and TNM stage. The expression of MDR1 was associated with resistance to NVB （χ2=5.209，P=0.022），GEM （χ2=4.769，P=0.029），VP-16 （χ2=4.596，P=0.032），and DDP（χ2=6.086，P=0.014）, but not associated with resistance to DOC（χ2=0.430，P=0.512）. Conclusion MTT chemosensitivity assay can effectively predict clinical chemotherapy sensitivity. Detection of MDR1, together with MTT chemosensitivity assay, can more accurately predict NSCLC chemosensitivity and be a guide for individualized chemotherapy of NSCLC.
ObjectiveTo investigate the primary culture method of human papillary thyroid carcinoma (PTC) cells for a long term and establish a monitoring and verification measures. MethodsPTC cells were isolated following routine procedures and cultured in the DMEM supplemented with 10% fetal bovine serum, glutamine, and 20 ng/ml epidermal growth factor (EGF). Thyroglobulin (Tg) and thyroperoxidase (TPO) in nutrient solution and specific antigen Tg expression of PTC cells cultured for different days were observed. ResultsThe PTC cells grew satisfactorily up to 45 days of incubation. Tg content in nutrient solution expressed the training period of a linear singular parabolic, achieved peak value (985.2 μg/L) at about 14 d. TPO had not been detected in nutrient solution. The Tg expressed positively by immunization fluorescent dyeing. ConclusionsPTC cells cultured in the present method can survive to over 45 days. A brief monitoring and evaluation systems of PTC cells has been established. This report prompts that cultured cells within 14 days maybe more suitable to gene research and provide alternative to the basic research of PTC events and features.
Objective To understand the effect of estradiol in different concentrations on proliferation of diverse mammary primary cells in vitro. Methods The primary cells of cancer tissue, the adjacent tissue to tumors and normal mammary tissue from patiens with breast cancer were obtained using collagenase digesting method. All the tissue samples were cultivated in vitro, and were given estradiol in different concentrations. The effect of estradiol on the proliferation of those primary cells was measured by MTT. Results Estradiol remarkedly promoted the proliferation of primary cells of cancer tissue and peritumor tissue in vitro, whose ER expression were positive. Whereas, the promotion effect of estradiol on the proliferation of normal mammary primary cells was relatively weak, and there was no correlation between the promotion effect with the expression of ER in cancer tissue. Conclusion The risks of occurrence and relapse of breast cancer would increase significantly when the concentration of estradiol is no less than 103 pmol/L in vivo.
ObjectiveTo establish a method of air-liquid interface culture and ciliary beat frequency measurement of mouse tracheal-bronchial epithelial cells to simulate the physiological function of airway epithelium.MethodsBALB/c mouse tracheal-bronchial epithelial cells were obtained by digestion with 1 mg/mL protease in cold temperature overnight, and the digestion time was optimized to ensure the quantity and viability of the obtained cells. After removing fibroblasts by differential velocity adhesion method, the cells were cultured into collagen coated Transwell inserts. Proliferating phase and air-liquid interface culture were promoted with different culture media.ResultsCell numbers obtained by cold protease overnight digestion for 12 h, 14 h and 16 h were (1.78±0.33)×105, (1.93±0.26)×105 and (2.01±0.28)×105, respectively. Cell viability by trypan blue staining were (96.86±0.25)%, (94.73±1.63)% and (86.87±5.95)%, respectively. Cells were 100% confluent in Transwell chamber after 1-week proliferation, and the ciliary beat frequency was observed under microscope after 2 - 3 weeks of air-liquid interface culture. The cilia structure was confirmed by hematoxylin-eosin staining, electron microscopy and immunofluorescence. Ciliary beat frequency of the cells obtained by this method was consistent with that of mouse trachea in vivo, which further demonstrated its capacity in simulating the physiological function of airway epithelium. ConclusionsThe separation and air-liquid interface culture system as well as the ciliary beat frequency measurement method established in this experiment is simple, stable, efficient and reliable, which establishes a substantial foundation for exploring the pathogenesis and treatment mechanism of airway diseases. It can also provide reference for the culture of epithelium in the airway of other species and/or other organs.