AIM: To investigate anti-tumor activities and apoptosis-regulated mechanisms of bufalin in the orthotopic transplantation tumor model of human hepatocellular carcinoma in nude mice.METHODS: BEL-7402 cells of human hep...AIM: To investigate anti-tumor activities and apoptosis-regulated mechanisms of bufalin in the orthotopic transplantation tumor model of human hepatocellular carcinoma in nude mice.METHODS: BEL-7402 cells of human hepatocellular carcinoma were inoculated to form subcutaneous tumors, and were implanted into the liver to establish orthotopic transplantation tumor models of human hepatocellular carcinoma in nude mice. Seventy-five animals were randomized divided into five groups (n = 15). Bufalin was injected intraperitoneally into three groups at doses of 1.5 mg/kg (BF1), 1 mg/kg (BF2) and 0.5 mg/kg (BF3) for d 15-24, respectively. The NS group was injected an equal volume of saline as above and adriamycin was injected intraperitoneally into the ADM group at a dose of 8.0 mg/kg for d 15. Ten mice in each group were killed at d 25 and the survival time in each group was calculated. We also observed the morphologic alterations in the myocardium, brain, liver, kidney and tumor tissues by pathology and electron microscopy, measured the apoptotic rate by TUNEL staining method, and detected the expression of apoptosis-regulated genes bcl-2 and bax by immunohistochemical staining and RT-PCR in tumor tissues. RESULTS: The tumor volumes in each group of bufalin were reduced significantly (35.21 ± 12.51 vs 170.39 ± 25.29; 49.83 ± 11.46 vs 170.39 ± 25.29; 83.99 ± 24.63 vs 170.39 ± 25.29, P < 0.01, respectively), and the survival times were prolonged in group BF1-2 (31.8 ± 4.2 vs 23.4 ± 2.1 and 29.4 ± 3.4 vs 23.4 ± 2.1, P < 0.05, respectively), and necrosis was mainly in severe or moderate degree in group BF1-2. No morphologicalchanges were detected in the myocardium, brain, liver and kidney tissues. Apoptotic characteristics could be seen in group BF1-2. The positive rates of bcl-2 and bax protein expression of each group by immunohistochemical staining were 10.0%, 10.0%, 20.0%, 10.0% and 20.0%; 90.0%, 80.0%, 80.0%, 40.0% and 30.0%, respectively. Loss of expression of bcl-2 mRNA in each group was to be found and the density of bax mRNA was increased progressively with increase of dose of bufalin by RT-PCR. CONCLUSION: Bufalin has significant anti-tumor activities in the orthotopic transplantation tumor model of human hepatocellular carcinoma in nude mice with no marked toxicity and was able to induce apoptosis of transplanted tumor cells. This apoptosis may be mediated mainly via up-regulating the expression of apoptosis-regulated gene bax, which may be involved in its anti-tumor mechanism of bufalin.展开更多
Bufalin is one of the main pharmacological and toxicological components of Venenum Bufonis and many traditional Chinese medicine preparations.The cardiotoxicity clearly limits its application to patients living in cou...Bufalin is one of the main pharmacological and toxicological components of Venenum Bufonis and many traditional Chinese medicine preparations.The cardiotoxicity clearly limits its application to patients living in countries.Hence,an investigation of its toxicological mechanism is helpful for new drug development and treatment of the related clinical adverse reactions.We investigate the cardiotoxicity of bufalin using human induced pluripotent stem cells-derived cardiomyocytes(hiPSC-CMs)(0.003–0.1μmol·L–1),human induced pluripotent stem cells-derived cardiomyocytes(hiPSC-CMs)(0.03–0.3μmol·L–1)and eight human cardiac ion channel currents(0.01–100μmol·L–1)combined with an impedance-based bioanalytical and patch clamp method.Biphasic effect of bufalin on the contractility in hiPSC-CMs,which has been shown to strengthen myocardial contractility,accelerate conduction,and increase beating rate at the earlier stage of administration,whereas weakened myocardial contractility,abolished conduction,and ceased beating rate at the later stage of administration.Bufalin decreased the action potential duration(Action potential duration at 30%,50%and 90%repolarization),cardiac action potential amplitude,and maximal depolarization rate and depolarized the resting membrane potential of hiPSC-CMs.Spontaneous beating rates of hiPSC-CMs were markedly increased at 0.03μmol·L–1,while were weakened at 0.3μmol·L–1 after application.Bufalin blocks INav1.5 in a concentration-dependent manner with half maximal inhibitory concentration of 74.5μmol·L–1.Bufalin respectively increased the late sodium current and Na+-Ca2+exchange current with a concentration for 50%of maximal effect of 2.48 and 66.06μmol·L–1 in hiPSC-CMs.Whereas,bufalin showed no significant effects on other cardiac ion channel currents.The enhancement of the late sodium current is one of the main mechanism for cardiotoxicity of bufalin.展开更多
OBJECTIVE To determine the characterization,anti-tumor efficacy and pharmacokinetics of bufalin-loaded PEGylated liposomes compared with bufalin entity.METHODS Bufalin-loaded PEGylated liposomes and bufalin-loaded lip...OBJECTIVE To determine the characterization,anti-tumor efficacy and pharmacokinetics of bufalin-loaded PEGylated liposomes compared with bufalin entity.METHODS Bufalin-loaded PEGylated liposomes and bufalin-loaded liposomes were prepared reproducibly with homogeneous particle size by the combination of thin film evaporation method and high pressure homogenization method.The particle size and zeta potential of the liposomes were determined by dynamic light scattering technique.The direct imaging of morphology of liposomes was charactered by transmission electron microscope.The content of bufalin in liposomes was analysed by HPLC method.The entrapment efficiency and the particle size was applied to assess the stability profile,after storage at 4℃ on day 0,7,15,30 and 90.The in-vitro release behaviours of bufalin from liposomes were conducted using dialysis bag technique at 37℃.In-vitro cytotoxicity studies were carried out using MTT[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide]assay on several kinds of tumor cel lines including SW620,PC-3,MDA-MB-231,A549,U251,U87 and HepG2.In-vivo pharmacokinetic study of bufalin liposomes was evaluated by HPLC method.RESULTS Their mean particle sizes were 127.6 nm and 155.0 nm,mean zeta potentials were 2.24 m V and-18.5 m V,entrapment efficiencies were 76.31%and 78.40%,respectively.In-vitro release profile revealed that the release of bufalin in bufalin-loaded PEGylated liposomes was slower than that of bufalin-loaded liposomes.The cytotoxicity of blank liposomes has been found within acceptable range,whereas bufalin-loaded PEGylated liposomes showed enhanced cytotoxicity to U251 cells compared with bufalin entity.In-vivo pharmacokinetics indicated that bufalinloaded PEGylated liposomes could extend eliminate half-life time of bufalin in plasma in rats.CONCLUSION The results suggested that bufalin-loaded PEGylated liposomes improved the solubility and increased the drug concentration in plasma.展开更多
OBJECTIVE: To probe insights into the reversal effect of bufalin on vincristine-acquired multidrug resistance(MDR) in human leukemia cell line K562/VCR.METHODS: Proliferative inhibition rate and the reversal index(RI)...OBJECTIVE: To probe insights into the reversal effect of bufalin on vincristine-acquired multidrug resistance(MDR) in human leukemia cell line K562/VCR.METHODS: Proliferative inhibition rate and the reversal index(RI) of bufalin were determined by Methyl thiazolyl tetrazolium assay. The uptake of Adriamycin(ADM) in K562/VCR cells, cell cycle and apoptosis rate were determined by flow cytometry(FCM). Cell morphologic changes were observed with Wright-Giemsa staining. The expression of P-glycoprotein(P-gp), multidrug-associated protein-1(MRP1), Bcl-x L and Bax protein were measured by immunocytochemistry.RESULTS: The human leukemia multidrug resistant K562/VCR cells showed no cross-resistance to bufalin. The RIs of bufalin at concentrations of 0.0002,0.001 and 0.005 μmol/L were 4.85, 6.94 and 14.77,respectively. Preincubation of 0.001 μmol/L bufalin for 2 h could increase intracellular ADM fluorescence intensity to 28.07%(P<0.05) and down-regulate MRP1 expression simultaneously, but no remarkable effect was found on P-gp protein. Cell cycle analysis indicated increased apoptosis rate and apparent decreased G2/M phase proportion after treatment with bufalin. When exposed to 0.01μmol/L bufalin, typical morphological changes of apoptosis could be observed. Down-regulation of Bcl-x L and up-regulation of Bax expression in K562/VCR cells could be detected by immunocytochemistry.CONCLUSION: Bufalin could partly reverse the MDR of K562/VCR cells, with a possible mechanism of down-regulating MRP1 expression and activating apoptosis pathway by altering Bcl-x L/Bax ratio.展开更多
Objective: Bufalin is an effective drug for the treatment of liver cancer. But its high toxicity, poor watersolubility, fast metabolism and short elimination half-life limit its use in tumor treatment. How to make the...Objective: Bufalin is an effective drug for the treatment of liver cancer. But its high toxicity, poor watersolubility, fast metabolism and short elimination half-life limit its use in tumor treatment. How to make the drug accumulate in the tumor and reduce side effects while maintaining its efficacy are urgent problems to be solved. The goal of this study is to solve these problems.Methods: A copolymer with tunable poly-N-isopropylacrylamide and polylactic acid was designed and synthesized. The corresponding dual targeting immunomicelles(DTIs) loaded with bufalin(DTIs–BF)were synthesized by copolymer self-assembly in an aqueous solution. The size and structure of DTIs–BF were determined by ZetaSizer Nano-ZS and transmission electron microscopy. Then, its temperature sensitivity, serum stability, critical micelle concentration(CMC), entrapment efficiency(EE), drug release and non-cytotoxicity of blank block copolymer micelles(BCMs) were evaluated. Next, the effects of DTIs–BF on cellular uptake, cytotoxicity, and tumor cell inhibition were evaluated. Finally, the accumulation of DTIs–fluorescein isothiocyanate(FITC) and the in vivo anti-tumor effect were observed using an interactive video information system.Results: DTIs–BF had a small size, spherical shape, good temperature sensitivity, high serum stability, low CMC, high EE, and slow drug release. The blank BCMs had very low cytotoxicity. Compared with free bufalin, the in vitro cellular internalization and cytotoxicity of DTIs–BF against SMMC-7721 cells were significantly enhanced, and the effects were obviously better at 40 ℃ than 37 ℃. In addition, the therapeutic effect on SMMC-7721 cells was further enhanced by the programmed cell death specifically caused by bufalin. When DTIs–FITC were injected intravenously in BALB/c nude mice bearing liver cancer,the accumulation of FITC was significantly increased in tumors.Conclusion: DTIs–BF is a potentially effective nano-formulation and has broad prospects in the clinical treatment of liver cancer.展开更多
Objective: To investigate anti-tumor effect of bufalin on the orthotopic transplantation tumor model of human hepatocellular carcinoma in nude mice. Methods: BEL-7402 cells of human hepatocellular carcinoma were inocu...Objective: To investigate anti-tumor effect of bufalin on the orthotopic transplantation tumor model of human hepatocellular carcinoma in nude mice. Methods: BEL-7402 cells of human hepatocellular carcinoma were inoculated to form subcutaneous tumors in nude mice by subcutaneous injection. Then the subcutaneous tumors were implanted into the liver of nude mice, and the orthotopic transplantation tumor models of human hepatocellular carcinoma were established. Seventy-five models were randomized into 5 groups ( n = 15) . Bufalin was injected intraperitoneally into the 3 groups at dose of 1.5,1 and 0.5 mg/kg for day 15 - 24, respectively. NS group were injected equal volume saline as above and adriamycin were injected intraperitoneally into ADM group at dose of 8.0 mg/kg for day 15. Ten mice in each group were killed at day 25 and detected on morphological and ultrastructural changes in myocardium, brain, liver, kidney and tumor tissues by pathology and electron microscope. The survival time in each group were observed. Results: The tumor volumes in each group of bufalin were reduced significantly compared with NS group (P < 0.01), the survival time were prolonged in group Bu 1 and Bu 2 compared with NS group ( P < 0.05), and tumor tissues were mainly necrosis in severe or moderate degree in Bu 1, Bu 2 groups, and mild degree or moderate degree in Bu 3 group. No morphological changes were detected in myocardium, brain, liver and kidney tissues, respectively. Apoptotic characteristics could be seen in tumor tissues of group Bu 1 and group Bu 2. Conclusion: Bufalin has significant anti-tumor effects on the orthotopic transplantation tumor model of human hepatocellular carcinoma in nude mice without marked toxicity. To guide cell apoptosis may be one of its anti-tumor mechanism of bufalin.展开更多
The biotransformation of bufalin by cell suspension cultures of Platycodon grandifiorus was investigated and two new biotransformed products were obtained, which was 3-epi-telocinobufagln and 3-epi-bufalin-3-O-β-D-gl...The biotransformation of bufalin by cell suspension cultures of Platycodon grandifiorus was investigated and two new biotransformed products were obtained, which was 3-epi-telocinobufagln and 3-epi-bufalin-3-O-β-D-glucoside.展开更多
OBJECTIVE Toad venom(Venenum Bufonis)isalways used for analgesia in China from ancient to modern times,but the effective component of it remains unclear.METHODS In the present study,we investigated the anti-nociceptiv...OBJECTIVE Toad venom(Venenum Bufonis)isalways used for analgesia in China from ancient to modern times,but the effective component of it remains unclear.METHODS In the present study,we investigated the anti-nociceptive effect and the underlying mechanism ofbufalin,an active ingredient fromtoad venom by animal behavior,patch clamp and calcium imaging.RESULTS Bufalin could significantly relieve formalin-induced spontaneous flinching and licking response as well as carrageenan-induced mechanical and thermal hyperalgesia.Using the whole-cel patch-clamp recording,bufalincaused remarkable suppressive effect on the peak currents of Na+channels in dorsal root ganglion neuroblastoma ND7-23 cel line in a U-shaped dependent manner.In addition,bufalinprompted the voltage-dependent activationand caused a negative shift of the fast-state inactivation of Na+channels.However,bufalin produced insignificant effect not onlyon voltage-dependent Kv4.2,Kv4.3 and BK channels,but also on the capsaicin induced Ca2+influx.CONCLUSION The present results indicate bufalin is capable of producing remarkable anti-nociceptive effects whichis probably ascribed to its specific modulation of voltage-gated Na+channels.展开更多
Objective Bufalin,the main active anti-tumor monomer of toad venom,is crucial in cancer treatment.However,intrinsic issues,such as poor solubility and systematic toxicity,have considerably mitigated its anticancer fun...Objective Bufalin,the main active anti-tumor monomer of toad venom,is crucial in cancer treatment.However,intrinsic issues,such as poor solubility and systematic toxicity,have considerably mitigated its anticancer functions and caused unwanted side effects.It is essential to develop innovative targeting systems to precisely and efficiently deliver anticancer drugs to achieve satisfying therapeutic efficiency.Methods This work established a novel and more efficient system for simultaneously detecting and killing colorectal cancer cells.The proposed method designed two allosteric probes,a report probe and a recognize probe.The method exhibited high sensitivity towards cell detection via the recognizing probe identifying target cancer cells and the report probe’s signal report.Combining bufalin and fluorouracil endowed better tumor cell inhibition.Results We observed significantly enhanced fluorescence dots surrounding the HCT-116 cell membranes.No fluorescence increments in the other three cells were identified,indicating that the established liposome complex could specifically bind with target cells.In addition,the best ratio of bufalin to fluorouracil was 0.15 and 0.5,respectively.This improved the anti-tumor effects and achieved more than 60%tumor cell inhibition.Conclusion This method will provide new opportunities for intracellular biomolecule detection and targeted cancer cell therapy.展开更多
Pancreatic cancer is one of the most lethal cancers worldwide and is characterized by a poor prognosis.1,2 Due to its aggressive nature and lack of early symptoms,most patients are diagnosed at an advanced stage,and c...Pancreatic cancer is one of the most lethal cancers worldwide and is characterized by a poor prognosis.1,2 Due to its aggressive nature and lack of early symptoms,most patients are diagnosed at an advanced stage,and chemotherapy is the optimal option.3,4 Epidemiology,the incidence rate of pancreatic cancer has increased in the last two decades,from 196,000 patients in 1990 to 441,000 in 2017.Based on 2020 global cancer statistics,the annual cases of pancreatic cancer have increased to 195,773.4 Unfortunately,patients demonstrate resistance to chemotherapy,and only some of them benefit from current therapeutic strategies.展开更多
Objective:Esophageal squamous cell carcinoma(ESCC),a commonly encountered malignant tumor in the gastrointestinal tract,poses a significant health burden.Bufalin,a pharmacologically active molecule,has been shown to e...Objective:Esophageal squamous cell carcinoma(ESCC),a commonly encountered malignant tumor in the gastrointestinal tract,poses a significant health burden.Bufalin,a pharmacologically active molecule,has been shown to exhibit antitumor activity against various types of cancers.This study investigates the molecular mechanism underpinning the effects of bufalin on ESCCs.Materials and Methods:The impact of bufalin on the proliferation and migration of ESCC cells was evaluated through the utilization of the Cell Counting Kit-8(CCK-8),scratch assay,and transwell assay.In addition,ribonucleic acid(RNA)sequencing was performed to identify genes that were abnormally expressed in response to bufalin treatment.Western blotting was utilized to ascertain the expression levels of protein arginine methyltransferase-6(PRMT6),phosphorylated AKT(p-AKT),and phosphorylated mammalian target of rapamycin(p-m TOR).Cell transfection was then performed to observe the rescue effect of PRMT6 on bufalin.Results:Bufalin displayed a significant time-dependent inhibition of the proliferation,migration,and invasion of ECA109 cells.An RNA sequencing(RNA-seq)analysis revealed that PRMT6 expression was downregulated in the cells treated with bufalin.PRMT6 promoted the proliferation,migration,and invasive potential of ECA109 cells.The overexpression of PRMT6 boosted p-AKT and p-m TOR levels in ECA109 cells and reversed bufalin inhibition.Conclusions:These findings indicate that bufalin exerts its inhibitory effects on ESCCs through the PRMT6/AKT/m TOR signaling pathway.These findings lay the groundwork for bufalin as a promising therapeutic candidate for the treatment of ESCC.展开更多
文摘AIM: To investigate anti-tumor activities and apoptosis-regulated mechanisms of bufalin in the orthotopic transplantation tumor model of human hepatocellular carcinoma in nude mice.METHODS: BEL-7402 cells of human hepatocellular carcinoma were inoculated to form subcutaneous tumors, and were implanted into the liver to establish orthotopic transplantation tumor models of human hepatocellular carcinoma in nude mice. Seventy-five animals were randomized divided into five groups (n = 15). Bufalin was injected intraperitoneally into three groups at doses of 1.5 mg/kg (BF1), 1 mg/kg (BF2) and 0.5 mg/kg (BF3) for d 15-24, respectively. The NS group was injected an equal volume of saline as above and adriamycin was injected intraperitoneally into the ADM group at a dose of 8.0 mg/kg for d 15. Ten mice in each group were killed at d 25 and the survival time in each group was calculated. We also observed the morphologic alterations in the myocardium, brain, liver, kidney and tumor tissues by pathology and electron microscopy, measured the apoptotic rate by TUNEL staining method, and detected the expression of apoptosis-regulated genes bcl-2 and bax by immunohistochemical staining and RT-PCR in tumor tissues. RESULTS: The tumor volumes in each group of bufalin were reduced significantly (35.21 ± 12.51 vs 170.39 ± 25.29; 49.83 ± 11.46 vs 170.39 ± 25.29; 83.99 ± 24.63 vs 170.39 ± 25.29, P < 0.01, respectively), and the survival times were prolonged in group BF1-2 (31.8 ± 4.2 vs 23.4 ± 2.1 and 29.4 ± 3.4 vs 23.4 ± 2.1, P < 0.05, respectively), and necrosis was mainly in severe or moderate degree in group BF1-2. No morphologicalchanges were detected in the myocardium, brain, liver and kidney tissues. Apoptotic characteristics could be seen in group BF1-2. The positive rates of bcl-2 and bax protein expression of each group by immunohistochemical staining were 10.0%, 10.0%, 20.0%, 10.0% and 20.0%; 90.0%, 80.0%, 80.0%, 40.0% and 30.0%, respectively. Loss of expression of bcl-2 mRNA in each group was to be found and the density of bax mRNA was increased progressively with increase of dose of bufalin by RT-PCR. CONCLUSION: Bufalin has significant anti-tumor activities in the orthotopic transplantation tumor model of human hepatocellular carcinoma in nude mice with no marked toxicity and was able to induce apoptosis of transplanted tumor cells. This apoptosis may be mediated mainly via up-regulating the expression of apoptosis-regulated gene bax, which may be involved in its anti-tumor mechanism of bufalin.
基金supported by the National Foundation of Science and Technology(2018ZX09201017-008)。
文摘Bufalin is one of the main pharmacological and toxicological components of Venenum Bufonis and many traditional Chinese medicine preparations.The cardiotoxicity clearly limits its application to patients living in countries.Hence,an investigation of its toxicological mechanism is helpful for new drug development and treatment of the related clinical adverse reactions.We investigate the cardiotoxicity of bufalin using human induced pluripotent stem cells-derived cardiomyocytes(hiPSC-CMs)(0.003–0.1μmol·L–1),human induced pluripotent stem cells-derived cardiomyocytes(hiPSC-CMs)(0.03–0.3μmol·L–1)and eight human cardiac ion channel currents(0.01–100μmol·L–1)combined with an impedance-based bioanalytical and patch clamp method.Biphasic effect of bufalin on the contractility in hiPSC-CMs,which has been shown to strengthen myocardial contractility,accelerate conduction,and increase beating rate at the earlier stage of administration,whereas weakened myocardial contractility,abolished conduction,and ceased beating rate at the later stage of administration.Bufalin decreased the action potential duration(Action potential duration at 30%,50%and 90%repolarization),cardiac action potential amplitude,and maximal depolarization rate and depolarized the resting membrane potential of hiPSC-CMs.Spontaneous beating rates of hiPSC-CMs were markedly increased at 0.03μmol·L–1,while were weakened at 0.3μmol·L–1 after application.Bufalin blocks INav1.5 in a concentration-dependent manner with half maximal inhibitory concentration of 74.5μmol·L–1.Bufalin respectively increased the late sodium current and Na+-Ca2+exchange current with a concentration for 50%of maximal effect of 2.48 and 66.06μmol·L–1 in hiPSC-CMs.Whereas,bufalin showed no significant effects on other cardiac ion channel currents.The enhancement of the late sodium current is one of the main mechanism for cardiotoxicity of bufalin.
基金Supported by Overall Innovation Plan Projects of Science and Technology of Shaanxi Province in China(2015KTZDSF02-01-02)
文摘OBJECTIVE To determine the characterization,anti-tumor efficacy and pharmacokinetics of bufalin-loaded PEGylated liposomes compared with bufalin entity.METHODS Bufalin-loaded PEGylated liposomes and bufalin-loaded liposomes were prepared reproducibly with homogeneous particle size by the combination of thin film evaporation method and high pressure homogenization method.The particle size and zeta potential of the liposomes were determined by dynamic light scattering technique.The direct imaging of morphology of liposomes was charactered by transmission electron microscope.The content of bufalin in liposomes was analysed by HPLC method.The entrapment efficiency and the particle size was applied to assess the stability profile,after storage at 4℃ on day 0,7,15,30 and 90.The in-vitro release behaviours of bufalin from liposomes were conducted using dialysis bag technique at 37℃.In-vitro cytotoxicity studies were carried out using MTT[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide]assay on several kinds of tumor cel lines including SW620,PC-3,MDA-MB-231,A549,U251,U87 and HepG2.In-vivo pharmacokinetic study of bufalin liposomes was evaluated by HPLC method.RESULTS Their mean particle sizes were 127.6 nm and 155.0 nm,mean zeta potentials were 2.24 m V and-18.5 m V,entrapment efficiencies were 76.31%and 78.40%,respectively.In-vitro release profile revealed that the release of bufalin in bufalin-loaded PEGylated liposomes was slower than that of bufalin-loaded liposomes.The cytotoxicity of blank liposomes has been found within acceptable range,whereas bufalin-loaded PEGylated liposomes showed enhanced cytotoxicity to U251 cells compared with bufalin entity.In-vivo pharmacokinetics indicated that bufalinloaded PEGylated liposomes could extend eliminate half-life time of bufalin in plasma in rats.CONCLUSION The results suggested that bufalin-loaded PEGylated liposomes improved the solubility and increased the drug concentration in plasma.
基金Shanghai Municipal Health Bureau:Traditional Chinese Medicine in Treating with Advanced Hepatocellular Carcinoma(No.ZYSNXD-CC-ZDYJ032)
文摘OBJECTIVE: To probe insights into the reversal effect of bufalin on vincristine-acquired multidrug resistance(MDR) in human leukemia cell line K562/VCR.METHODS: Proliferative inhibition rate and the reversal index(RI) of bufalin were determined by Methyl thiazolyl tetrazolium assay. The uptake of Adriamycin(ADM) in K562/VCR cells, cell cycle and apoptosis rate were determined by flow cytometry(FCM). Cell morphologic changes were observed with Wright-Giemsa staining. The expression of P-glycoprotein(P-gp), multidrug-associated protein-1(MRP1), Bcl-x L and Bax protein were measured by immunocytochemistry.RESULTS: The human leukemia multidrug resistant K562/VCR cells showed no cross-resistance to bufalin. The RIs of bufalin at concentrations of 0.0002,0.001 and 0.005 μmol/L were 4.85, 6.94 and 14.77,respectively. Preincubation of 0.001 μmol/L bufalin for 2 h could increase intracellular ADM fluorescence intensity to 28.07%(P<0.05) and down-regulate MRP1 expression simultaneously, but no remarkable effect was found on P-gp protein. Cell cycle analysis indicated increased apoptosis rate and apparent decreased G2/M phase proportion after treatment with bufalin. When exposed to 0.01μmol/L bufalin, typical morphological changes of apoptosis could be observed. Down-regulation of Bcl-x L and up-regulation of Bax expression in K562/VCR cells could be detected by immunocytochemistry.CONCLUSION: Bufalin could partly reverse the MDR of K562/VCR cells, with a possible mechanism of down-regulating MRP1 expression and activating apoptosis pathway by altering Bcl-x L/Bax ratio.
基金financially supported by the National Natural Science Foundation of China (81673739)the National Key Research and Development Program of China (No.2018YFC1705305)+2 种基金Shanghai Development Office of Traditional Chinese Medicine (No. ZY[2018-2020]-FWTX-4010)Shanghai “13th Five-year Plan” Clinical Key Department—Department of Dermatology of Traditional Chinese Medicine (No. shslczdzk05001)Clinical Research Plan of SHDC (No.SHDC2020CR3097B)。
文摘Objective: Bufalin is an effective drug for the treatment of liver cancer. But its high toxicity, poor watersolubility, fast metabolism and short elimination half-life limit its use in tumor treatment. How to make the drug accumulate in the tumor and reduce side effects while maintaining its efficacy are urgent problems to be solved. The goal of this study is to solve these problems.Methods: A copolymer with tunable poly-N-isopropylacrylamide and polylactic acid was designed and synthesized. The corresponding dual targeting immunomicelles(DTIs) loaded with bufalin(DTIs–BF)were synthesized by copolymer self-assembly in an aqueous solution. The size and structure of DTIs–BF were determined by ZetaSizer Nano-ZS and transmission electron microscopy. Then, its temperature sensitivity, serum stability, critical micelle concentration(CMC), entrapment efficiency(EE), drug release and non-cytotoxicity of blank block copolymer micelles(BCMs) were evaluated. Next, the effects of DTIs–BF on cellular uptake, cytotoxicity, and tumor cell inhibition were evaluated. Finally, the accumulation of DTIs–fluorescein isothiocyanate(FITC) and the in vivo anti-tumor effect were observed using an interactive video information system.Results: DTIs–BF had a small size, spherical shape, good temperature sensitivity, high serum stability, low CMC, high EE, and slow drug release. The blank BCMs had very low cytotoxicity. Compared with free bufalin, the in vitro cellular internalization and cytotoxicity of DTIs–BF against SMMC-7721 cells were significantly enhanced, and the effects were obviously better at 40 ℃ than 37 ℃. In addition, the therapeutic effect on SMMC-7721 cells was further enhanced by the programmed cell death specifically caused by bufalin. When DTIs–FITC were injected intravenously in BALB/c nude mice bearing liver cancer,the accumulation of FITC was significantly increased in tumors.Conclusion: DTIs–BF is a potentially effective nano-formulation and has broad prospects in the clinical treatment of liver cancer.
基金Supported by National Natural Science Foundation of China (No.30200364)
文摘Objective: To investigate anti-tumor effect of bufalin on the orthotopic transplantation tumor model of human hepatocellular carcinoma in nude mice. Methods: BEL-7402 cells of human hepatocellular carcinoma were inoculated to form subcutaneous tumors in nude mice by subcutaneous injection. Then the subcutaneous tumors were implanted into the liver of nude mice, and the orthotopic transplantation tumor models of human hepatocellular carcinoma were established. Seventy-five models were randomized into 5 groups ( n = 15) . Bufalin was injected intraperitoneally into the 3 groups at dose of 1.5,1 and 0.5 mg/kg for day 15 - 24, respectively. NS group were injected equal volume saline as above and adriamycin were injected intraperitoneally into ADM group at dose of 8.0 mg/kg for day 15. Ten mice in each group were killed at day 25 and detected on morphological and ultrastructural changes in myocardium, brain, liver, kidney and tumor tissues by pathology and electron microscope. The survival time in each group were observed. Results: The tumor volumes in each group of bufalin were reduced significantly compared with NS group (P < 0.01), the survival time were prolonged in group Bu 1 and Bu 2 compared with NS group ( P < 0.05), and tumor tissues were mainly necrosis in severe or moderate degree in Bu 1, Bu 2 groups, and mild degree or moderate degree in Bu 3 group. No morphological changes were detected in myocardium, brain, liver and kidney tissues, respectively. Apoptotic characteristics could be seen in tumor tissues of group Bu 1 and group Bu 2. Conclusion: Bufalin has significant anti-tumor effects on the orthotopic transplantation tumor model of human hepatocellular carcinoma in nude mice without marked toxicity. To guide cell apoptosis may be one of its anti-tumor mechanism of bufalin.
文摘The biotransformation of bufalin by cell suspension cultures of Platycodon grandifiorus was investigated and two new biotransformed products were obtained, which was 3-epi-telocinobufagln and 3-epi-bufalin-3-O-β-D-glucoside.
基金The project supported by Innovation Program of Shanghai Municipal Education Commission(15ZZ063)by Research Project of Putuo Hospital,Shanghai University of Traditional Chinese Medicine(2014YJ002)
文摘OBJECTIVE Toad venom(Venenum Bufonis)isalways used for analgesia in China from ancient to modern times,but the effective component of it remains unclear.METHODS In the present study,we investigated the anti-nociceptive effect and the underlying mechanism ofbufalin,an active ingredient fromtoad venom by animal behavior,patch clamp and calcium imaging.RESULTS Bufalin could significantly relieve formalin-induced spontaneous flinching and licking response as well as carrageenan-induced mechanical and thermal hyperalgesia.Using the whole-cel patch-clamp recording,bufalincaused remarkable suppressive effect on the peak currents of Na+channels in dorsal root ganglion neuroblastoma ND7-23 cel line in a U-shaped dependent manner.In addition,bufalinprompted the voltage-dependent activationand caused a negative shift of the fast-state inactivation of Na+channels.However,bufalin produced insignificant effect not onlyon voltage-dependent Kv4.2,Kv4.3 and BK channels,but also on the capsaicin induced Ca2+influx.CONCLUSION The present results indicate bufalin is capable of producing remarkable anti-nociceptive effects whichis probably ascribed to its specific modulation of voltage-gated Na+channels.
基金Supported by grants from the Medical Guidance Project of Shanghai Science and Technology Commission(No.19401935200)the Budget Internal Medicine Research Project of Shanghai University of Traditional Chinese Medicine(No.2020LK065).
文摘Objective Bufalin,the main active anti-tumor monomer of toad venom,is crucial in cancer treatment.However,intrinsic issues,such as poor solubility and systematic toxicity,have considerably mitigated its anticancer functions and caused unwanted side effects.It is essential to develop innovative targeting systems to precisely and efficiently deliver anticancer drugs to achieve satisfying therapeutic efficiency.Methods This work established a novel and more efficient system for simultaneously detecting and killing colorectal cancer cells.The proposed method designed two allosteric probes,a report probe and a recognize probe.The method exhibited high sensitivity towards cell detection via the recognizing probe identifying target cancer cells and the report probe’s signal report.Combining bufalin and fluorouracil endowed better tumor cell inhibition.Results We observed significantly enhanced fluorescence dots surrounding the HCT-116 cell membranes.No fluorescence increments in the other three cells were identified,indicating that the established liposome complex could specifically bind with target cells.In addition,the best ratio of bufalin to fluorouracil was 0.15 and 0.5,respectively.This improved the anti-tumor effects and achieved more than 60%tumor cell inhibition.Conclusion This method will provide new opportunities for intracellular biomolecule detection and targeted cancer cell therapy.
基金supported by the National Natural Science Foundation of China(No.82405167)the Shenzhen Science and Technology Program(Guangdong,China)(No.GJHZ20220913143005010)the fundamental research project of the Shenzhen Science and Technology Innovation Commission(No.20231120113324002 and No.RCBS20231211090635056).
文摘Pancreatic cancer is one of the most lethal cancers worldwide and is characterized by a poor prognosis.1,2 Due to its aggressive nature and lack of early symptoms,most patients are diagnosed at an advanced stage,and chemotherapy is the optimal option.3,4 Epidemiology,the incidence rate of pancreatic cancer has increased in the last two decades,from 196,000 patients in 1990 to 441,000 in 2017.Based on 2020 global cancer statistics,the annual cases of pancreatic cancer have increased to 195,773.4 Unfortunately,patients demonstrate resistance to chemotherapy,and only some of them benefit from current therapeutic strategies.
基金supported by funding from the Priority Academic Program Development of Jiangsu Higher Education Institutions(PAPD)the National Nature Science Foundation of China(81703557)。
文摘Objective:Esophageal squamous cell carcinoma(ESCC),a commonly encountered malignant tumor in the gastrointestinal tract,poses a significant health burden.Bufalin,a pharmacologically active molecule,has been shown to exhibit antitumor activity against various types of cancers.This study investigates the molecular mechanism underpinning the effects of bufalin on ESCCs.Materials and Methods:The impact of bufalin on the proliferation and migration of ESCC cells was evaluated through the utilization of the Cell Counting Kit-8(CCK-8),scratch assay,and transwell assay.In addition,ribonucleic acid(RNA)sequencing was performed to identify genes that were abnormally expressed in response to bufalin treatment.Western blotting was utilized to ascertain the expression levels of protein arginine methyltransferase-6(PRMT6),phosphorylated AKT(p-AKT),and phosphorylated mammalian target of rapamycin(p-m TOR).Cell transfection was then performed to observe the rescue effect of PRMT6 on bufalin.Results:Bufalin displayed a significant time-dependent inhibition of the proliferation,migration,and invasion of ECA109 cells.An RNA sequencing(RNA-seq)analysis revealed that PRMT6 expression was downregulated in the cells treated with bufalin.PRMT6 promoted the proliferation,migration,and invasive potential of ECA109 cells.The overexpression of PRMT6 boosted p-AKT and p-m TOR levels in ECA109 cells and reversed bufalin inhibition.Conclusions:These findings indicate that bufalin exerts its inhibitory effects on ESCCs through the PRMT6/AKT/m TOR signaling pathway.These findings lay the groundwork for bufalin as a promising therapeutic candidate for the treatment of ESCC.
