ABSTRACT
Aim To isolate cancer stem cells from human pan-creatic cancer cell line L3. 6pl and to identify their biological characteristics. Methods L3. 6pl cells were cultivated in com-mercial low adhesion plate with serum-free stem cell culture me-dium ( MEM/F12 1:1 ) supplemented with B27. The cancer stem cells reformed into floating spheres were isolated. The method of tumor sphere formation was used to isolate/enrich and characterize the cancer stem cells in pancreatic carcinoma cell line L3. 6pl. Cancer stem cell spheres were collected and sorted using magnetic cell sorting ( magnetic activated cell sorting, MACS) technology, with the cell surface markers of CD24 +CD44 + ESA+. Self-renewal and EMT-related oncogene expres-sion were measured with Western blot. Cancer stem cells differ-entiation potential and the expression of cancer stem cell related signs were checked with Immunofluorescence assay. To deter-mine tumorigenesis in vivo, Xenograft assay in NOD-SCID mice were performed respectively, then immunohistochemistry proto-oncogene c-Met and RON expression were also checked. West-ern blot was used to detect the changes of stemness relative tran-scriptional factors and epithelial markers expressed in spheres before and after differentiation. Drug resistance of pancreatic cancer stem cells to gemcitabine or paclitaxel was measured with MTT assay. Results CD24 + CD44 + ESA+ cells were signifi-cantly tumorigenic, and cultured in serum-free conditions to form spheroids, which had the characteristics of stem cells with self-renewal, EMT and drug-resistant capabilities, and had a posi-tive correlation with the c-Met, RON protein expression. Con-clusion Human pancreatic stem cells are successfully isolated, which provides a useful model for individualized therapy and e-valuation of the therapeutic efficacy for pancreatic cancer pa-tients.
ABSTRACT
BACKGROUND: There are many methods for studying the somatotype of adults or children. Among them, Heath-Carter somatotype method is a comprehensive evaluation method. Using this method, 10 items of anthropometric indicators are selected and 3 factors which could be gotten to represent relative content of body fat, growth degree of skeletal muscle and relative height and thinness of body (relative line degree), are calculated, respectively.OBJECTIVE: To analyze the rules and characteristics of somatotype development in Dong students from Hunan province, so as to supplement the essential data for physical anthropology.DESIGN, TIME AND SETTING: A cross-sectional investigation was performed at the Department of Biological Engineering, Huaihua College of Hunan Province in May 2006.PARTICIPANTS: A total of 989 Dong students (boys 492, girls 497), aged 7 to 17 years, were selected from the primary schools or middle schools of Tongdao Dong Nationality Autonomous Country in Hunan province and recruited into the present study. All the selected students were verified healthy by physical examinations at school. The subjects were divided into two groups by gender, and each group was divided into 11 subgroups according to the age.METHODS: By Heath-Carter somatotype method, 10 anthropological indexes were measured. Each indicator was measured twice and the average value was selected. The above-mentioned data were input into the computer to form a database. The following indicators were calculated in turn: 3 factors on somatotype, coordinate values of X and Y on somatotype chart, mean of dimensional distances from the average somatotype to all somatotypes in the sample, difference between the two somatotypes in three-dimensional space and frequency distribution of each somatotype.MAIN OUTCOME MEASURES: Height, body mass, upper arm circumference, calf circumference, intracondylar diameters of humerus and femur, skinfold of brachial triceps, subscapular skinfold, skinfold of anterosuperior iliac spine, skinfold of gastrocnemius muscle.RESULTS: All 989 students were included in the final analysis. The dynamic range of the average value of endomorphic factor was 1.2-1.9 and 1.3-4.1, and that of mesomorphic factor was 4.2-5.1 and 1.9-3.0, and that of ectomorphic factor was 2.8-3.7 and 2.6-3.9 for the boys and the girls, respectively. Statistical analysis revealed that there was significant difference in the average value of mesomorphic factor by gender in each age (P＜0.05-0.01). The average endomorphic factor of the girls was obviously higher than that of the boys in each age (P＜0.01) except 7 to 11 years old. The average ectomorphic factor of the girls was higher than that of the boys before 13 years old and it was contrary after 13 years old. There was significant difference of average ectomorphic factor between beys and girls only in 9,11 and 15 to 17 years old (P＜0.05-0.01). The results of t-test showed that there was significant difference for the somatotypes by gender in each age (P＜0.05-0.01). The distribution of girl somatotype demonstrated a dynamic process from 7 to 17 years old, which demonstrated balanced mesomorphy-ectomorphy changed to balanced ecdomorphy, ectomorphy-ectomorphy, balanced endomorphy-ectomorphy and balanced endomorphy in turn. But distribution of boy somatotype was relatively stable, and average somatotype was mesomorphy. Cluster analysis revealed that there was obvious difference of congenital somatotype between Dong students and other 9 populations. During puberty, the somatotype feature of Dong boys was more close to that of Korean boys (SAD=0.37), but which was more far to that of Han boys in the city (SAD=2.15). The somatotype feature of Dong girls was more close to that of Zhuang girls (SAD=0.71), but which was more far to that of Korean girls (SAD=2.35) and Miao girls (SAD=2.10).CONCLUSION: The congenital somatotype of Dong students has its unique characteristics. There is obvious difference in somatotype characteristics between boys and girls.
ABSTRACT
Jingzhaotoxin-V(JZTX-V) isolated from the venom of the spider Chilobrachys jingzhao is a novel potent inhibitor that acts on tetrodotoxin-resistant and tetrodotoxin-sensitive sodium channels in adult rat dorsal root ganglion(DRG) neurons. It is a 29-residue polypeptide toxin including three disulfide bridges. To investigate the structure-function relationship of the toxin, a mutant of JZTX-V in which Arg20 was substituted by Ala, was synthesized by solid-phase chemistry method with Fmoc-protected amino acids on the PS3 automated peptide synthesizer. The synthetic linear peptide was then purified by reversed-phase high performance liquid chromatography and oxidatively refolded under the optimal conditions. The refolded product was analyzed by matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry(MALDI-TOF MS) and electrophysiological experiments for its relative molecular weight and prohibitive activity of sodium channels respectively. The present findings show that the prohibitive effect of R20A-JZTX-V on TTX-S sodium channels in DRG neurons is almost the same as that of native JZTX-V, suggesting that Arg20 does not play any important role in inhibiting TTX-S sodium currents in DRG neurons. In contrast, the prohibitive level of R20A-JZTX-V on TTX-R sodium channels is reduced by at last 18.3 times, indicating that Arg20 is a key amino acid residue relative to the bioactivity of JZTX-V. It is presumed that the decrease in activity of R20A-JZTX-V is due to the changes of the property in the binding site in TTX-R sodium channels.