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Objective To establish a non-venous bypass orthotopic liver transplantation model in Bama miniature pigs with high repeatability and stability. Methods Twelve Bama miniature pigs were randomly divided into the donor group (n=6) and recipient group (n=6). Pigs underwent non-venous bypass orthotopic liver transplantation. The time of anhepatic phase during operation was shortened, blood pressure during anhepatic phase was stably maintained, and management of anesthesia and body fluid during operation were strengthened. The operation time, anhepatic phase and survival status of the recipients were observed and recorded. The intraoperative heart rate, mean arterial pressure (MAP) and changes in arterial blood gas analysis were monitored. The perioperative liver function was evaluated. Results Among 6 Bama miniature pigs, 1 died from transplantation failure intraoperatively. The operation time of the remaining 5 pigs was (247±27) min and the time of anhepatic phase was (46±4) min. Three animals survived for more than 2 weeks. Compared with the preanhepatic phase, the heart rate of the animals was significantly faster, MAP was considerably reduced to (46±6) mmHg, blood pH value, base excess (BE) and HCO3- level were all significantly decreased and serum level of K+ was significantly elevated during the anhepatic phase (all P < 0.05). In the neohepatic phase, MAP of Bama miniature pigs was significantly increased, heart rate was dramatically slower.Blood pH value, BE, HCO3- level were significantly increased and serum level of K+ was significantly declined (all P < 0.05). During abdominal closure, MAP, blood gas indexes and serum level of K+ were almost recovered to those in the preanhepatic phase. Compared with preoperative levels, the levels of alanine aminotransferase(ALT), aspartate aminotransferase(AST), lactate dehydrogenase(LDH)and alkaline phosphatase(ALP)were significantly increased after operation (all P < 0.05), the change in AST was the most obvious, and it gradually decreased at postoperative 2 d. The level of γ-gutamyl transferase(GGT) did not significantly elevated. The level of total bilirubin (TB) was evidently elevated at postoperative 5 d. Compared with the preoperative levels, the levels of total protein (TP) and albumin (ALB) were significantly decreased after operation (both P < 0.05), and began to gradually increase at postoperative 1 d. Conclusions The non-venous bypass orthotopic liver transplantation model of Bama miniature pig is convenient, with highly reproducible and survival rate, which can be utilized as a standardized liver transplantation model.
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OBJECTIVE： To compare low-polarity volatile constituents in supercritical CO2 extract from the roots and stem of Ilex asprella and its effects on the proliferation of IEC-6 in vitro， and to provide reference for making full use of wild resources of I. asprella and expanding its medicinal parts. METHODS： The low-polarity volatile constituents were extracted from the root and stem of I. asprella with supercritical fluid CO2 extraction（SFE-CO2）. The chemical constituents were analyzed by GC-MS. IEC-6 cells were treated with different concentrations of supercritical CO2 extracts （0， 1， 5， 10， 20， 40， 60， 80， 100 μg/mL） from roots or stems of I. asprella. MTT assay was used to detect the relative viability， and cell proliferation curve was drawn and EC50 of each extract were calculated. RESULTS： Sixty-two and forty-six low-polarity volatile constituents were identified from supercritical CO2 extract in the roots and stem of I. asprella with GC-MS； there were 24 common constituents totally， mainly including pelargonic acid（14.18％ and 6.14％），octanoic acid（10.59％ and 4.35％），hexanoic acid（8.63％ and 10.86％），paeonol（7.79％ and 6.00％），2-methyl-3-phenyl-propanal（6.3％ and 0.58％），acetic acid（1.72％ and 33.77％） in root and stem， respectively. The results of cell culture in vitro showed that when the concentration of supercritical CO2 extract from the roots and stems of I. asprella was lower （≤60 μg/mL）， it could significantly promote the proliferation of IEC-6 cells and their EC50 were 16.35， 20.20 μg/mL， respectively； when the concentration of the extract was higher （≥80 μg/mL）， it showed cytotoxicity and inhibited the proliferation of IEC-6 cells. CONCLUSIONS： There are similar species of volatile constituents in roots and stems of I. asprella and similar in vitro bioactivity of the supercritical CO2 extracts to IEC-6 cells. The short-chain fatty acids may be the active ingredient to promote cell proliferation， while paeonol may be the cytotoxic active ingredient.
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OBJECTIVE:To extract and identify the volatile components of the roots of Polyalthia nemoralis,and to evaluate its biological activity in vitro. METHODS:Supercritical CO2 extraction (SFE-CO2) was performed to extract volatile compounds from the roots of P. nemoralis,and the volatile components were separated and determined by GC-MS. Human hepatic cancer Huh-7 cells were cultured with 0(blank control),5,10,20,30,40 and 50 μg(medicinal material)/ml extract for 24 h,and then MTT assay was used to detect the inhibitory effect of the extracts on cells. Relative cell viability and IC50 were calculated. The an-ti-bacterial activities of extract to Staphylococcus aureus,Methicillin-resistant Staphylococcus aureus,Enterococcus faecalis and oth-er strain were determined by Kirby-bauer method and broth dilution method. RESULTS:Forty compounds were identified from the SFE-CO2 extracts from the roots of P. nemoralis. The main constituents were(Z,Z,Z)-9,12,15-octa-decatrien-1-ol,glycerin,cinna-maldehyde,n-hexadecanoic acid and eugenol. Compared with blank control group,SFE-CO2 extracts from the roots of P. nemoralis 5 μg(medicinal material)/ml and above showed significant inhibitory effect on cell growth(P<0.05),and the inhibitory effect was strengthened as the concentration of extracts increased,IC50 values was 5.2 μg(medicinal material)/ml. However,the supercritical extract didn't showed antibacterial activity against three microorganisms in 2 kinds of antibacterial tests. CONCLUSIONS:SFE-CO2 and GC-MS method can effectively extract and identify volatile components of the roots of P. nemoralis,and supercritical extracts can inhibit the viability of cells but have no antibacterial activity.