ABSTRACT
Objective To establish an animal model by placing one end of PICC in the hepatic portal vein of a beagle dog and leaving the other end out of its body.Methods Six Beagle dogs were given respiration anesthesia through orotracheal intubation.An incision was made through the right rectus abdominalis to locate the superior mesenteric vein (SMA) and the main hepatic portal vein.The left branch of SMA was separated and cut to put PICC into the main hepatic portal vein before being ligated and fixed.The other end of PICC was elicited through the right abdominal wall and passed beneath the skin to the back neck and fastened in case of movement.Results The anesthetic effect was good and all the operations were successful.The mean operation time was about an hour and the mean blood loss was about 15 ml.The incision healed 5-7 d after operation.Conclusion The establishment of the model can improve the effects of liver-targeting drugs,which can cut down the dosage,lower the cost of treatment and experiment and reduce the adverse effect of medicines.Through PICC,we can directly draw blood from the hepatic portal vein to measure the blood concentration before the first pass elimination.Then according to the concentration,we can calculate the absorption rate in the gastrointestinal tract,which can facilitate related experimental studies.
ABSTRACT
Objective To knockout the MATP gene of mouse melanoma cell line B16F10 using CRISPR/Cas9 system,and to lay foundation for the functional study of MATP gene.Methods Specific primers of MATP were designed according to the report in http://crispr.mit.edu/ website.The primers were linked to pCAS9/gRNA1 vector.Then the positive vector was transfected into mouse melanoma B16F10 cells,and monoclonal cell lines were obtained by the infinite dilution method.After the genomes of different monoclonal cell lines were extracted and sequenced,the cell lines with MATP gene cleavage were screened,and the expression of MATP in these cell lines was verified by Western-blot analysis.Results Three MATP gene knockout cell lines were successfully obtained.The western-blot results showed that the cell lines did not express MATP protein.Conclusions The knockout of MATP gene in B16F10 cell line can be successfully achieved using the pCAS9/gRNA1 vector.
ABSTRACT
Objective To analyze the effect of Noggin silencing on the BMP and Wnt signaling pathways in hair follicle development.Methods The expression of BMP-2, BMP-4, BMPR-IA, BMP-6, BMP-7, LEF-1 andβ-catenin in Noggin silencing MC3T3-E1 stable cell line was detected by RT-PCR and western blot.Results RT-PCR results showed that the expressions of five genes in BMP signaling pathway were all significantly influenced by Noggin silencing, the ex-pressions of BMP-2 (P<0.001), BMP-4 (P<0.01), BMP-6 (P<0.001) and BMP-7 (P<0.001) were all increased and the expression of BMPR-IA (P<0.01) was decreased.While the expressions of the two genes LEF-1 (P<0.001) and β-catenin ( P<0.001) in Wnt signaling pathway were significantly decreased.Western blot results showed that the ex-pressions of these proteins in the two signaling pathways were also affected.The expressions of BMP-2 (P<0.05), BMP-4 (P<0.05), BMP-6 (P<0.05) and BMP-7 (P<0.05) were all increased, while the expressions of BMPR-IA (P<0.05), LEF-1 (P<0.01) andβ-catenin (P<0.001) were decreased.Conclusions There may be a negative feedback regulation of Noggin on the BMP signaling pathway in vitro, but a positive feedback regulation on the Wnt signaling pathway in vitro.It provides certain evidence for studies on the effect of Noggin gene on BMP and Wnt signaling pathways in vivo. There may be an interaction between hair follicle development-related signaling pathways, which still needs further experi-ments to prove.