AIM:To improve the outcome of orthotopic transplantation in a mouse model,we used an absorbable gelatin sponge(AGS) in nude mice to establish an orthotopic implantation tumor model.METHODS:MHCC-97L hepatocellular carc...AIM:To improve the outcome of orthotopic transplantation in a mouse model,we used an absorbable gelatin sponge(AGS) in nude mice to establish an orthotopic implantation tumor model.METHODS:MHCC-97L hepatocellular carcinoma(HCC)cells stably expressing the luciferase gene were injected into the subcutaneous region of nude mice.One week later,the ectopic tumors were harvested and transplanted into the left liver lobe of nude mice.The AGS was used to establish the nude mouse orthotopic implantation tumor model.The tumor suppressor gene,paired box gene 5(PAX5),which is a tumor suppressor in HCC,was transfected into HCC cells to validate the model.Tumor growth was measured by bioluminescence imaging technology.Semi-quantitative reverse transcription polymerase chain reaction(RT-PCR) and histopathology were used to confirm the tumorigenicity of the implanted tumor from the MHCC-97L cell line.RESULTS:We successfully developed an orthotopic transplantation tumor model in nude mice with the use of an AGS.The success rate of tumor transplantation was improved from 60% in the control group to 100% in the experimental group using AGS.The detection of fluorescent signals showed that tumors grew in all live nude mice.The mice were divided into 3 groups:AGS-,AGS+/PAX5-and AGS+/PAX5 +.Tumor size was significantly smaller in PAX5 transfected nude mice compared to control mice(P < 0.0001).These fluorescent signal results were consistent with observations made during surgery.Pathologic examination further confirmed that the tissues from the ectopic tumor were HCC.Results from RT-PCR proved that the HCC originated from MHCC-97L cells.CONCLUSION:Using an AGS is a convenient and efficient way of establishing an indirect orthotopic liver transplantation tumor model with a high success rate.展开更多
AIM： To observe the curative effect of galactosylated chitosan （GC）/5-fluorouracil （5-FU） nanoparticles in liver caner mice and its side effects. METHODS： The GC/5-FU nanoparticle is a nanomate- rial made by cou...AIM： To observe the curative effect of galactosylated chitosan （GC）/5-fluorouracil （5-FU） nanoparticles in liver caner mice and its side effects. METHODS： The GC/5-FU nanoparticle is a nanomate- rial made by coupling GC and 5-FU. The release experiment was performed in vitro. The orthotropic liver cancer mouse models were established and divided into control, GC, 5-FU and GC/5-FU groups. Mice in the control and GC group received an intravenous injection of 200 μL saline and GC, respectively. Mice in the 5-FU and GC/5-FU groups received 200 μL （containing 0.371 mg 5-FU） 5-FU and GC/5-FU, respectively. The tumor weight and survival time were observed. The cell cycle and apoptosis in tumor tissues were monitored by flow cytometry. The expression of p53, Bax, Bcl-2, caspase-3 and poly adenosine 50-diphosphate-ribose polymerase 1 （PARP-1） was detected by immunohistochemistry, reverse transcription-polymerase chain reaction and Western blot. The serum blood biochemical parameters and cytotoxic activity of natural killer （NK） cell and cy- totoxicity T lymphocyte （CTL） were measured. RESULTS： The GC/5-FU nanoparticle is a sustained release system. The drug loading was 6.12% ± 1.36%, the encapsulation efficiency was 81.82% ± 5.32%, and the Zeta potential was 10.34 ± 1.43 mV. The tu- mor weight in the GC/5-FU group （0.4361±0.1153 g vs 1.5801 ± 0.2821 g, P 〈 0.001） and the 5-FU （0.7932±0.1283 g vs 1.5801 ±0.2821 g, P 〈 0.001） was sig- nificantly lower than that in the control group; GC/5- FU treatment can significantly lower the tumor weight （0.4361± 0.1153 g vs 0.7932±0.1283 g, P 〈 0.001）, and the longest median survival time was seen in the GC/5-FU group, compared with the control （12 d vs 30 d, P 〈 0.001）, GC （13 d vs 30 d, P 〈 0.001） and 5-FU groups （17 d vs 30 d, P 〈 0.001）. Flow cytom- etry revealed that compared with the control, GC/5- FU caused a higher rate of G0-G1 arrest （52.79% ± 13.42% vs 23.92%±9.09%, P = 0.014 ） and apopto- sis （2.55% ±1.10% vs 11.13% ±11.73%, P 〈 0.001） in hepatic cancer cells. Analysis of the apoptosis path- ways showed that GC/5-FU upregulated the expression of p53 at both the protein and the mRNA levels, which in turn lowered the ratio of Bcl-2lBax expression; this led to the release of cytochrome C into the cytosol from the mitochondria and the subsequent activation of caspase-3. Upregulation of caspase-3 expression de- creased the PARP-1 at both the mRNA and the protein levels, which contributed to apoptosis. 5-FU increased the levels of aspartate aminotransferase and alanine aminotransferase, and decreased the numbers of platelet, white blood cell and lymphocyte and cytotoxic activities of CTL and NK cells, however, there were no such side effects in the GC/5-FU group. CONCLUSION： GC/5-FU nanoparticles can significant- ly inhibit the growth of liver cancer in mice via the p53 apoptosis pathway, and relieve the side effects and im- munosuppression of 5-FU.展开更多
基金Supported by National Natural Science Foundation of China, No.81201963Inner Mongolia Natural Science Foundation of China,No.2010MS1123
文摘AIM:To improve the outcome of orthotopic transplantation in a mouse model,we used an absorbable gelatin sponge(AGS) in nude mice to establish an orthotopic implantation tumor model.METHODS:MHCC-97L hepatocellular carcinoma(HCC)cells stably expressing the luciferase gene were injected into the subcutaneous region of nude mice.One week later,the ectopic tumors were harvested and transplanted into the left liver lobe of nude mice.The AGS was used to establish the nude mouse orthotopic implantation tumor model.The tumor suppressor gene,paired box gene 5(PAX5),which is a tumor suppressor in HCC,was transfected into HCC cells to validate the model.Tumor growth was measured by bioluminescence imaging technology.Semi-quantitative reverse transcription polymerase chain reaction(RT-PCR) and histopathology were used to confirm the tumorigenicity of the implanted tumor from the MHCC-97L cell line.RESULTS:We successfully developed an orthotopic transplantation tumor model in nude mice with the use of an AGS.The success rate of tumor transplantation was improved from 60% in the control group to 100% in the experimental group using AGS.The detection of fluorescent signals showed that tumors grew in all live nude mice.The mice were divided into 3 groups:AGS-,AGS+/PAX5-and AGS+/PAX5 +.Tumor size was significantly smaller in PAX5 transfected nude mice compared to control mice(P < 0.0001).These fluorescent signal results were consistent with observations made during surgery.Pathologic examination further confirmed that the tissues from the ectopic tumor were HCC.Results from RT-PCR proved that the HCC originated from MHCC-97L cells.CONCLUSION:Using an AGS is a convenient and efficient way of establishing an indirect orthotopic liver transplantation tumor model with a high success rate.
基金Supported by Natural Science Foundation of Shanghai,No.09ZR1424700 and 114119a4700Minhang District Natural Science Foundation of Shanghai,No. 2009MHZ085grants from Minhang District Public Health Bureau of Shanghai,No.2009MW28
文摘AIM： To observe the curative effect of galactosylated chitosan （GC）/5-fluorouracil （5-FU） nanoparticles in liver caner mice and its side effects. METHODS： The GC/5-FU nanoparticle is a nanomate- rial made by coupling GC and 5-FU. The release experiment was performed in vitro. The orthotropic liver cancer mouse models were established and divided into control, GC, 5-FU and GC/5-FU groups. Mice in the control and GC group received an intravenous injection of 200 μL saline and GC, respectively. Mice in the 5-FU and GC/5-FU groups received 200 μL （containing 0.371 mg 5-FU） 5-FU and GC/5-FU, respectively. The tumor weight and survival time were observed. The cell cycle and apoptosis in tumor tissues were monitored by flow cytometry. The expression of p53, Bax, Bcl-2, caspase-3 and poly adenosine 50-diphosphate-ribose polymerase 1 （PARP-1） was detected by immunohistochemistry, reverse transcription-polymerase chain reaction and Western blot. The serum blood biochemical parameters and cytotoxic activity of natural killer （NK） cell and cy- totoxicity T lymphocyte （CTL） were measured. RESULTS： The GC/5-FU nanoparticle is a sustained release system. The drug loading was 6.12% ± 1.36%, the encapsulation efficiency was 81.82% ± 5.32%, and the Zeta potential was 10.34 ± 1.43 mV. The tu- mor weight in the GC/5-FU group （0.4361±0.1153 g vs 1.5801 ± 0.2821 g, P 〈 0.001） and the 5-FU （0.7932±0.1283 g vs 1.5801 ±0.2821 g, P 〈 0.001） was sig- nificantly lower than that in the control group; GC/5- FU treatment can significantly lower the tumor weight （0.4361± 0.1153 g vs 0.7932±0.1283 g, P 〈 0.001）, and the longest median survival time was seen in the GC/5-FU group, compared with the control （12 d vs 30 d, P 〈 0.001）, GC （13 d vs 30 d, P 〈 0.001） and 5-FU groups （17 d vs 30 d, P 〈 0.001）. Flow cytom- etry revealed that compared with the control, GC/5- FU caused a higher rate of G0-G1 arrest （52.79% ± 13.42% vs 23.92%±9.09%, P = 0.014 ） and apopto- sis （2.55% ±1.10% vs 11.13% ±11.73%, P 〈 0.001） in hepatic cancer cells. Analysis of the apoptosis path- ways showed that GC/5-FU upregulated the expression of p53 at both the protein and the mRNA levels, which in turn lowered the ratio of Bcl-2lBax expression; this led to the release of cytochrome C into the cytosol from the mitochondria and the subsequent activation of caspase-3. Upregulation of caspase-3 expression de- creased the PARP-1 at both the mRNA and the protein levels, which contributed to apoptosis. 5-FU increased the levels of aspartate aminotransferase and alanine aminotransferase, and decreased the numbers of platelet, white blood cell and lymphocyte and cytotoxic activities of CTL and NK cells, however, there were no such side effects in the GC/5-FU group. CONCLUSION： GC/5-FU nanoparticles can significant- ly inhibit the growth of liver cancer in mice via the p53 apoptosis pathway, and relieve the side effects and im- munosuppression of 5-FU.
