In this study,37 derivatives of phorbol esters were synthesized and their anti-HIV-1 activities evaluated,building upon our previous synthesis of 51 phorbol derivatives.12-Para-electron-acceptor-trans-cinnamoyl-13-dec...In this study,37 derivatives of phorbol esters were synthesized and their anti-HIV-1 activities evaluated,building upon our previous synthesis of 51 phorbol derivatives.12-Para-electron-acceptor-trans-cinnamoyl-13-decanoyl phorbol derivatives stood out,demonstrating remarkable anti-HIV-1 activities and inhibitory effects on syncytia formation.These derivatives exhibited a higher safety index compared with the positive control drug.Among them,12-(trans-4-fluorocinnamoyl)-13-decanoyl phorbol,designated as compound 3c,exhibited the most potent anti-HIV-1 activity(EC_(50)2.9 nmol·L^(−1),CC50/EC_(50)11117.24)and significantly inhibited the formation of syncytium(EC_(50)7.0 nmol·L^(−1),CC50/EC_(50)4891.43).Moreover,compound 3c is hypothesized to act both as an HIV-1 entry inhibitor and as an HIV-1 reverse transcriptase inhibitor.Isothermal titration calorimetry and molecular docking studies indicated that compound 3c may also function as a natural activator of protein kinase C(PKC).Therefore,compound 3c emerges as a potential candidate for developing new anti-HIV drugs.展开更多
Phorbol esters are recognized for their dual role as anti-HIV-1 agents and as activators of protein kinase C(PKC).The efficacy of phorbol esters in binding with PKC is attributed to the presence of oxygen groups at po...Phorbol esters are recognized for their dual role as anti-HIV-1 agents and as activators of protein kinase C(PKC).The efficacy of phorbol esters in binding with PKC is attributed to the presence of oxygen groups at positions C20,C3/C4,and C9 of phorbol.Concurrently,the lipids located at positions C12/C13 are essential for both the anti-HIV-1 activity and the formation of the PKC-ligand complex.The influence of the cyclopropane ring at positions C13 and C14 in phorbol derivatives on their anti-HIV-1 activity requires further exploration.This research entailed the hydrolysis of phorbol,producing seco-cyclic phorbol derivatives.The anti-HIV-1 efficacy of these derivatives was assessed,and the affinity constant(Kd)for PKC-δprotein of selected seco-cyclic phorbol derivatives was determined through isothermal titration calorimetry.The findings suggest that the chemical modification of cyclopropanols could affect both the anti-HIV-1 activity and the PKC binding affinity.Remarkably,compound S11,with an EC_(50) of 0.27μmol·L^(−1) and a CC_(50) of 153.92μmol·L^(−1),demonstrated a potent inhibitory effect on the intermediate products of HIV-1 reverse transcription(ssDNA and 2LTR),likely acting at the viral entry stage,yet showed no affinity for the PKC-δprotein.These results position compound S11 as a potential candidate for further preclinical investigation and for studies aimed at elucidating the pharmacological mechanism underlying its anti-HIV-1 activity.展开更多
To investigate the phytochemical constituents of aboveground parts of Croton tiglium L. （family Euphorbiaceae）, its ingredients were isolated by repeated chromatography on silica gel, Sephadex LH-20, ODS and prepara...To investigate the phytochemical constituents of aboveground parts of Croton tiglium L. （family Euphorbiaceae）, its ingredients were isolated by repeated chromatography on silica gel, Sephadex LH-20, ODS and preparative HPLC. Their structures were identified based on 1D, 2D NMR and mass spectral analysis. A total of 10 phorbol esters were obtained. Among them, compound 1 （12-O-（2-methyl）butyryl-4ct-deoxyphorbol-β-isobutyrate） and compound 10 （20-formyl-4a-deoxyphorbol- β-acetate） were two new compounds, and the other eight were known compounds.展开更多
The effects of phorbol-12,13-dibuterate （PDBu） on total sodium current （INa-total）, tetrodotoxin-resistant sodium current （INa-TFXr）, 4-AP-sensitive potassium current （IA） and TEA-sensitive potassium current ...The effects of phorbol-12,13-dibuterate （PDBu） on total sodium current （INa-total）, tetrodotoxin-resistant sodium current （INa-TFXr）, 4-AP-sensitive potassium current （IA） and TEA-sensitive potassium current （IK） in trigeminal ganglion （TG） neurons were investigated. Whole-cell patch clamp techniques were used to record ion currents in cultured TG neurons of rats. Results revealed that 0.5μmol/L PDBu reduced the amplitude of INa-total by （38.3±4.5）% （n=6, P〈0.05）, but neither the G-V curve （control： V0.5 =-17.1±4.3 mV, k=7.4±1.3; PDBu： V0.5=-15.9±5.9 mV, k=5.9±1.4; n=6, P〉0.05） nor the inactivation rate constant （control： 3.6±0.9 ms; PDBu： 3.6±0.8 ms; n=6, P〉0.05） was altered. 0.5 μmol/L PDBu could significantly increase the amplitude of INa-TFXr by （37.2± 3.2）% （n=9, P〈0.05） without affecting the G-V curve （control： V0.5=-14.7±6.0 mV, k=6.9± 1.4; PDBu： V0.5=- 11.1±5.3 mV, k=8.1± 1.5; n=5, P〉0.05 ） or the inactivation rate constant （control： 4.6±0.6 ms; PDBu： 4.2±0.5 ms; n=5, P〉0.05）. 0.5 μmol/L PDBu inhibited IK by （15.6±5.0） % （n=16, P〈0.05）, and V0.5 was significantly altered from - 4.7±1.4 mV to -7.9 ±1.8 mV （n=16, P〈0.05）. IA was not significantly affected by PDBu, 0.5μmol/L PDBu decreased IA by only （0.3±3.2）% （n=5, P〉0.05）. It was concluded that PDBu inhibited INa-total ：.but enhanced INa-TFXr, and inhibited IK without affecting IA. These data suggested that the activation of PKC pathway could exert the actions.展开更多
The deoiled Jatropha curcas meal by hexane extraction was detoxified phorbol esters by two different methods. These two methods were alkali in methanol and only ethanol washing. After both treatments, the PEs （phorbo...The deoiled Jatropha curcas meal by hexane extraction was detoxified phorbol esters by two different methods. These two methods were alkali in methanol and only ethanol washing. After both treatments, the PEs （phorbol esters） was decreased by 100%. The crude protein in detoxified meal of alkali in methanol washing was less amount than only ethanol washing. The result showed that treatment by only ethanol washing was a promising way to detoxify deoiled Jatropha curcas meal for animal feeds in industrial scale.展开更多
Dopamine （DA） exposure at a dose of 100 pmol/L for 24 hours causes oxidative stress in SH-SY5Y cells with induction of neuronal differentiation by retinoic acid （RA,10 pmol/L,72 hours） followed by phorbol ester 12...Dopamine （DA） exposure at a dose of 100 pmol/L for 24 hours causes oxidative stress in SH-SY5Y cells with induction of neuronal differentiation by retinoic acid （RA,10 pmol/L,72 hours） followed by phorbol ester 12-O-tetradecanoyl-phorbol-13-acetate （TPA,80 nmol/L,72 hours）. However,it remains unclear whether the alteration of phenotype observed in response to oxidative stress is associated with protein regulation in this cellular model for Parkinson＇s disease. The present study detected protein regulation affected by oxidative stress at a proteomic level：selection of differentially altered proteins using two dimensional difference in-gel electrophoresis and identification of these proteins using matrix assisted laser desorption/ionization time-of-flight mass spectrometry. The results demonstrated significant alterations in expression of six proteins in SH-SY5Y cells following the differentiation and fourteen proteins in the differentiated cells following the exposure,exemplified by an increase of tubulin alpha1 in the former but a decrease of tubulin alpha-ubiquitous chain in the latter. These results suggest that two potentially specific but relevant patterns of proteomic change may be produced in SH-SY5Y cells with the induction of differentiation by RA followed by TPA,and in the differentiated cells after DA exposure.展开更多
Objective:To compare the protein profile of culture supernatants in stimulated and unstimulated human fibroblasts to find some proteins indicating the presence of fibroblasts and their activation status.Methods:Dermal...Objective:To compare the protein profile of culture supernatants in stimulated and unstimulated human fibroblasts to find some proteins indicating the presence of fibroblasts and their activation status.Methods:Dermal fibroblasts were stimulated with phorbol 12-myristate 13-acetate(PMA)/ionomycine for 72 h.MTT assay was done to determine cell viability and A/E fluorescent staining was used to evaluate the cell death pattern.Protein analysis was performed by gradient SDS polyacrylamide gel electrophoresis 8%–16%.Results:The supernatant of 24 h cultured both stimulated and unstimulated fibroblasts showed two bands in SDS-PAGE analysis with relative molecular weights of 8.59 and78.8 k Da.These bands density was decreased during the next 48 h in unstimulated cells while their expression was continued in PMA or PMA/ionomycine stimulated cells and a new 85.3 k Da band was appeared in unstimulated and 72 h PMA stimulated cells.Moreover,we found another seven small size(10–19.5 k Da) proteins in supernatants of48 h and 72 h unstimulated but not in PMA or PMA/Ionomycine stimulated fibroblasts.Most of these proteins expression were down regulated following fibroblast activation.This down-regulation is consistent with our finding that PMA or PMA/ionomycine stimulated cells exhibited a significant level of apoptosis cell death.Conclusions:Human fibroblasts produce some small to intermediate sized proteins with specific SDS-PAGE profile upon cell activation.Most of these proteins can be excreted in urine and can be immunogen theoretically so this data provided a reliable clue for fibrosis biomarker screening based on designation of an appropriated immunoassay.展开更多
We tried to clarify the role of oxygen radicals released from granulocytes stimulated byphorbol myristate acetate(PMA) in rat acute lung injury. It was found that DNA strand-breakdamage(DSBD) in peripheral white blood...We tried to clarify the role of oxygen radicals released from granulocytes stimulated byphorbol myristate acetate(PMA) in rat acute lung injury. It was found that DNA strand-breakdamage(DSBD) in peripheral white blood cells (WBC) was significantly increased 40 min after injec-tion of PMA. DSBD in lung tissue of rats treated with PMA was also markedly increased comparedwith the controls. The PMA-treated rats showed significantly higher lipid-peroxide (LPO) level inplasma and lung tissue hemogenate than the controls did. These results suggest that determination ofDSBD, a simple and sensitive indicator for oxygen radical damaging, might be useful in thediagnosis of adult respiratory distress syndrome (ARDS), when it is used together with themeasurement of plasma LPO.展开更多
To investigate the correlation between the activity of kinases in the growth factor signal transduction pathway and the development of resistance of breast cancer to tamoxifen, reporter gene regulated by the regulat...To investigate the correlation between the activity of kinases in the growth factor signal transduction pathway and the development of resistance of breast cancer to tamoxifen, reporter gene regulated by the regulating fragment of CCD1 was transfected into the MCF 7 cells, and the influence of tamoxifen on the reporter gene expression was examined under different conditions of TPA treatment. Our results showed that the reporter gene expression was inhibited by tamoxifen and promoted by TPA. Furthermore, tamoxifen exerts an agonist effect on the reporter gene expression when the cells was treated by TPA previously for 12 h. It is concluded that TPA could induce estrogen like effect of tamoxifen on estrogen receptor positive breast cancer cells and it may be one of the mechanisms responsible for the development of tamoxifen resistance.展开更多
Relaxin is known to inhibit cardiac fibrosis. However, it is unclear whether relaxin could regulate the effects of Phorbol 12-myristate 13-acetate （PMA, PKC activator） on cardiac fibrosis. So the influence of relaxi...Relaxin is known to inhibit cardiac fibrosis. However, it is unclear whether relaxin could regulate the effects of Phorbol 12-myristate 13-acetate （PMA, PKC activator） on cardiac fibrosis. So the influence of relaxin on the cell proliferation and collagen expression induced by PMA in cultured cardiac fibroblasts was studied. It showed that PMA significantly increased cardiac fibroblasts proliferation, Type I pro-collagen protein expression, Type I pro-collagen mRNA expression, and rhRLX absolutely significantly decreased PMA induced effects on cardiac fibroblasts proliferation and Type I pro-collagen expressions, indicating that relaxin could inhibit cardiac fibrosis induced by PMA.展开更多
Cardiomyocytes isolated from neonatal rats were treated with phorboll 2-myristatel 3-acetate (PMA ) ranging from 10(-11)to 10-7 mol/L for 20 min, causing cytosol protein kinase A (PKA) activity to decrease while parti...Cardiomyocytes isolated from neonatal rats were treated with phorboll 2-myristatel 3-acetate (PMA ) ranging from 10(-11)to 10-7 mol/L for 20 min, causing cytosol protein kinase A (PKA) activity to decrease while particulate PKA activity increase in a concentration-dependent manner. The change of PKA activity induced by PMA was abolished completely by pretreatment of polymyxin B or depletion of protein kinase C (PKC). Type II PKA activity in particulate fraction was enhanced remarkably, while that of type I PKA was not altered when the cells were treated with 100 nmol/L PMA. The results suggested that subcellular distribution and activity of PKA in cardiomyocytes may be regulated by PKC.展开更多
Objective:To study the effect of arsenic trioxide (As2O3) combined with phorbol ester (PMA) on the proliferation of acute promyelocytic leukemia cell line Kasumi-1.Methods:Acute promyelocytic leukemia cell lines Kasum...Objective:To study the effect of arsenic trioxide (As2O3) combined with phorbol ester (PMA) on the proliferation of acute promyelocytic leukemia cell line Kasumi-1.Methods:Acute promyelocytic leukemia cell lines Kasumi-1 were cultured and randomly divided into control group (treated with the RPMI1640 medium without drugs or serum), As2O3 group (treated with serum-free RPMI1640 medium containing 20 μmol/L As2O3), PMA group (treated with serum-free RPMI1640 medium containing 160 nmol/L PMA) and As2O3+ PMA group (treated with serum-free RPMI1640 medium containing 20 μmol/L As2O3 and 160 nmol/L PMA). After treatment, the cell proliferation activity, cell cycle ratio and the protein expression of related genes were measured.Results: 12 h, 24 h and 48 h after treatment, the cell proliferation activity of As2O3 group, PMA group and As2O3+PMA group were significantly lower than that of control group, and the cell proliferation activity of As2O3+PMA group was significantly lower than that of As2O3 group and PMA group;48 h after treatment, the G1 phase and S phase ratio as well as CDK1 and CyclinB1 expression of As2O3 group, PMA group and As2O3+PMA group were significantly lower than those of control group while the G2 phase ratio as well as Bax, Caspase-3, Caspase-9, p-Chk1 and p-Cdc25C9 expression were significantly higher than those of control group;the G1 phase and S phase ratio as well as CDK1 and CyclinB1 expression of As2O3+PMA group were significantly lower than those of As2O3 group and PMA group while the G2 phase ratio as well as Bax, Caspase-3, Caspase-9, p-Chk1 and p-Cdc25C9 expression was significantly higher than those of As2O3 group and PMA group.Conclusion:As2O3 combined with PMA can inhibit the proliferation of acute promyelocytic leukemia cell line Kasumi-1 by inducing apoptosis and blocking cell cycle.展开更多
We have accomplished the efficient total synthesis of(+)-phorbol in 20 steps from inexpensive(+)-car-vone.This concise synthesis is partly due to the use of two pentamethyldisilyl(PMDS)groups as the masked hydroxyl gr...We have accomplished the efficient total synthesis of(+)-phorbol in 20 steps from inexpensive(+)-car-vone.This concise synthesis is partly due to the use of two pentamethyldisilyl(PMDS)groups as the masked hydroxyl groups.The significant advantages of the PMDS group over other silyl groups have been firmly demonstrated.This total synthesis features a Shapiro reaction that links two fragments possessing a unique PMDS group.This is followed by a Tamao-Fleming oxidation and a subsequent ring-closing metathesis reaction to construct the 5/7/6/3 tetra-cyclic skeleton.展开更多
Protein kinase C with a molecular weight of 82 kD has been purified to electrophoresis homogenous from rat brain through a series of chromatography cohmns including DE-52, Sepharose G-200 and phenyl-Sepharose. The enz...Protein kinase C with a molecular weight of 82 kD has been purified to electrophoresis homogenous from rat brain through a series of chromatography cohmns including DE-52, Sepharose G-200 and phenyl-Sepharose. The enzyme possessed autophosphorylation activity. Yuanhuacin A inhibited the ~3H-phorbol-12, 13-dibutyrate (~3H-PdBu) binding of PKC with an IC_(50) value of 1.48±0.287×10^(-8) mol/L when the concentration of ~3H-PdBu was 1.5×10^(-9) mol/L (K_i=1.2×10^(-8) mol/L). Yuanhuacin A inhibited the PdBu-stimulated PKC activity in the catalysis of the phosphorylation of Histone Ⅲ-S with an IC_(50) of 2.82±0.37×10^(-9) mol/L (PdBu=10^(-6) mol/L), while it had no effect on the basal and Ca^(2+)-stimulated PKC activity in the same assay system. This result suggests that Yuanhuacin A is a selective antagonist of the phorbol ester receptor in protein kinase C.展开更多
Activator protein-1(AP-1)transcription factor is a key component of many signal transduction pathways involved in the regulation of cellular processes and controls rapid responses of mammalian cells when exposed to th...Activator protein-1(AP-1)transcription factor is a key component of many signal transduction pathways involved in the regulation of cellular processes and controls rapid responses of mammalian cells when exposed to the variety of stimulus.The phorbol 12-myristate 13-acetate and Forskolin(Fo)are well-known kinase activators/stimulators of Protein Kinase C(PKC)and Protein Kinase A(PKA)respectively.Importantly,these kinases are found to be present in transitional points of many cell signaling pathways,especially those involved in proliferation.The stimulating effect of PKC and PKA on the expression of AP-1 factors in MCF-7 breast cell proliferation is not well characterized.Hence,the role of PKC by PMA treatment and the role of PKA by using Fo in MCF-7 cells is investigated.Where,cells treated with PMA showed increased cell proliferation,while Fo had no effect,but inhibited the PMA induced proliferation.The RT-PCR results showed the PMA induced c-Jun,c-Fos and Fra-1 expressions compared to control and Fo.However,Fo in combination with PMA,inhibit the PMA induced above mRNA expressions where Fo alone has no effect.Western blot studies validated the c-Jun expressions in PMA treated MCF-7 cells.Further,PMA increases the mRNA expression of Cyclin-E1,Cyclin-D1,and CDK-4,whereas Fo decreases their expressions.Thus,mitogenic effect of PMA and inhibitory action of Fo on MCF-7 cells is probably enhanced via activation of AP-1 factors and concomitant action of cell cycle regulators in the downstream singling cascade.展开更多
基金supported by the National Natural Science Foundation of China(Nos.81202882,82060670)Suzhou Science and Technology Planning Project in Jiangsu Province of China(No.SNG2021022)+1 种基金the Priority Academic Program Development of the Jiangsu Higher Education Institutes,China(PAPD)and the Project of Innovative Research Team of Yunnan Province(No.202005AE160005).
文摘In this study,37 derivatives of phorbol esters were synthesized and their anti-HIV-1 activities evaluated,building upon our previous synthesis of 51 phorbol derivatives.12-Para-electron-acceptor-trans-cinnamoyl-13-decanoyl phorbol derivatives stood out,demonstrating remarkable anti-HIV-1 activities and inhibitory effects on syncytia formation.These derivatives exhibited a higher safety index compared with the positive control drug.Among them,12-(trans-4-fluorocinnamoyl)-13-decanoyl phorbol,designated as compound 3c,exhibited the most potent anti-HIV-1 activity(EC_(50)2.9 nmol·L^(−1),CC50/EC_(50)11117.24)and significantly inhibited the formation of syncytium(EC_(50)7.0 nmol·L^(−1),CC50/EC_(50)4891.43).Moreover,compound 3c is hypothesized to act both as an HIV-1 entry inhibitor and as an HIV-1 reverse transcriptase inhibitor.Isothermal titration calorimetry and molecular docking studies indicated that compound 3c may also function as a natural activator of protein kinase C(PKC).Therefore,compound 3c emerges as a potential candidate for developing new anti-HIV drugs.
基金supported by the National Natural Science Foundation of China(Nos.81202882 and 82060670)the Suzhou Science and Technology Planning Project in Jiangsu Province of China(No.SNG2021022)+1 种基金the Priority Academic Program Development of the Jiangsu Higher Education Institutes,China(PAPD)Project of Innovative Research Team of Yunnan Province(No.202005AE160005)。
文摘Phorbol esters are recognized for their dual role as anti-HIV-1 agents and as activators of protein kinase C(PKC).The efficacy of phorbol esters in binding with PKC is attributed to the presence of oxygen groups at positions C20,C3/C4,and C9 of phorbol.Concurrently,the lipids located at positions C12/C13 are essential for both the anti-HIV-1 activity and the formation of the PKC-ligand complex.The influence of the cyclopropane ring at positions C13 and C14 in phorbol derivatives on their anti-HIV-1 activity requires further exploration.This research entailed the hydrolysis of phorbol,producing seco-cyclic phorbol derivatives.The anti-HIV-1 efficacy of these derivatives was assessed,and the affinity constant(Kd)for PKC-δprotein of selected seco-cyclic phorbol derivatives was determined through isothermal titration calorimetry.The findings suggest that the chemical modification of cyclopropanols could affect both the anti-HIV-1 activity and the PKC binding affinity.Remarkably,compound S11,with an EC_(50) of 0.27μmol·L^(−1) and a CC_(50) of 153.92μmol·L^(−1),demonstrated a potent inhibitory effect on the intermediate products of HIV-1 reverse transcription(ssDNA and 2LTR),likely acting at the viral entry stage,yet showed no affinity for the PKC-δprotein.These results position compound S11 as a potential candidate for further preclinical investigation and for studies aimed at elucidating the pharmacological mechanism underlying its anti-HIV-1 activity.
基金National Natural Science Foundation of China(Grant No.81172943)
文摘To investigate the phytochemical constituents of aboveground parts of Croton tiglium L. （family Euphorbiaceae）, its ingredients were isolated by repeated chromatography on silica gel, Sephadex LH-20, ODS and preparative HPLC. Their structures were identified based on 1D, 2D NMR and mass spectral analysis. A total of 10 phorbol esters were obtained. Among them, compound 1 （12-O-（2-methyl）butyryl-4ct-deoxyphorbol-β-isobutyrate） and compound 10 （20-formyl-4a-deoxyphorbol- β-acetate） were two new compounds, and the other eight were known compounds.
基金This project was supported by a grant from National Natu-ral Sciences Foundation of China (No. 30271500).
文摘The effects of phorbol-12,13-dibuterate （PDBu） on total sodium current （INa-total）, tetrodotoxin-resistant sodium current （INa-TFXr）, 4-AP-sensitive potassium current （IA） and TEA-sensitive potassium current （IK） in trigeminal ganglion （TG） neurons were investigated. Whole-cell patch clamp techniques were used to record ion currents in cultured TG neurons of rats. Results revealed that 0.5μmol/L PDBu reduced the amplitude of INa-total by （38.3±4.5）% （n=6, P〈0.05）, but neither the G-V curve （control： V0.5 =-17.1±4.3 mV, k=7.4±1.3; PDBu： V0.5=-15.9±5.9 mV, k=5.9±1.4; n=6, P〉0.05） nor the inactivation rate constant （control： 3.6±0.9 ms; PDBu： 3.6±0.8 ms; n=6, P〉0.05） was altered. 0.5 μmol/L PDBu could significantly increase the amplitude of INa-TFXr by （37.2± 3.2）% （n=9, P〈0.05） without affecting the G-V curve （control： V0.5=-14.7±6.0 mV, k=6.9± 1.4; PDBu： V0.5=- 11.1±5.3 mV, k=8.1± 1.5; n=5, P〉0.05 ） or the inactivation rate constant （control： 4.6±0.6 ms; PDBu： 4.2±0.5 ms; n=5, P〉0.05）. 0.5 μmol/L PDBu inhibited IK by （15.6±5.0） % （n=16, P〈0.05）, and V0.5 was significantly altered from - 4.7±1.4 mV to -7.9 ±1.8 mV （n=16, P〈0.05）. IA was not significantly affected by PDBu, 0.5μmol/L PDBu decreased IA by only （0.3±3.2）% （n=5, P〉0.05）. It was concluded that PDBu inhibited INa-total ：.but enhanced INa-TFXr, and inhibited IK without affecting IA. These data suggested that the activation of PKC pathway could exert the actions.
文摘The deoiled Jatropha curcas meal by hexane extraction was detoxified phorbol esters by two different methods. These two methods were alkali in methanol and only ethanol washing. After both treatments, the PEs （phorbol esters） was decreased by 100%. The crude protein in detoxified meal of alkali in methanol washing was less amount than only ethanol washing. The result showed that treatment by only ethanol washing was a promising way to detoxify deoiled Jatropha curcas meal for animal feeds in industrial scale.
基金the Science and Technology Development Program of Jilin Province, No. 200505200the Distinguished Professor Foundation of Jilin University, No. 450011011204
文摘Dopamine （DA） exposure at a dose of 100 pmol/L for 24 hours causes oxidative stress in SH-SY5Y cells with induction of neuronal differentiation by retinoic acid （RA,10 pmol/L,72 hours） followed by phorbol ester 12-O-tetradecanoyl-phorbol-13-acetate （TPA,80 nmol/L,72 hours）. However,it remains unclear whether the alteration of phenotype observed in response to oxidative stress is associated with protein regulation in this cellular model for Parkinson＇s disease. The present study detected protein regulation affected by oxidative stress at a proteomic level：selection of differentially altered proteins using two dimensional difference in-gel electrophoresis and identification of these proteins using matrix assisted laser desorption/ionization time-of-flight mass spectrometry. The results demonstrated significant alterations in expression of six proteins in SH-SY5Y cells following the differentiation and fourteen proteins in the differentiated cells following the exposure,exemplified by an increase of tubulin alpha1 in the former but a decrease of tubulin alpha-ubiquitous chain in the latter. These results suggest that two potentially specific but relevant patterns of proteomic change may be produced in SH-SY5Y cells with the induction of differentiation by RA followed by TPA,and in the differentiated cells after DA exposure.
基金Supported by the Research and Technology Council of Babol University of Medical Sciences with Grant No.9031029
文摘Objective:To compare the protein profile of culture supernatants in stimulated and unstimulated human fibroblasts to find some proteins indicating the presence of fibroblasts and their activation status.Methods:Dermal fibroblasts were stimulated with phorbol 12-myristate 13-acetate(PMA)/ionomycine for 72 h.MTT assay was done to determine cell viability and A/E fluorescent staining was used to evaluate the cell death pattern.Protein analysis was performed by gradient SDS polyacrylamide gel electrophoresis 8%–16%.Results:The supernatant of 24 h cultured both stimulated and unstimulated fibroblasts showed two bands in SDS-PAGE analysis with relative molecular weights of 8.59 and78.8 k Da.These bands density was decreased during the next 48 h in unstimulated cells while their expression was continued in PMA or PMA/ionomycine stimulated cells and a new 85.3 k Da band was appeared in unstimulated and 72 h PMA stimulated cells.Moreover,we found another seven small size(10–19.5 k Da) proteins in supernatants of48 h and 72 h unstimulated but not in PMA or PMA/Ionomycine stimulated fibroblasts.Most of these proteins expression were down regulated following fibroblast activation.This down-regulation is consistent with our finding that PMA or PMA/ionomycine stimulated cells exhibited a significant level of apoptosis cell death.Conclusions:Human fibroblasts produce some small to intermediate sized proteins with specific SDS-PAGE profile upon cell activation.Most of these proteins can be excreted in urine and can be immunogen theoretically so this data provided a reliable clue for fibrosis biomarker screening based on designation of an appropriated immunoassay.
文摘We tried to clarify the role of oxygen radicals released from granulocytes stimulated byphorbol myristate acetate(PMA) in rat acute lung injury. It was found that DNA strand-breakdamage(DSBD) in peripheral white blood cells (WBC) was significantly increased 40 min after injec-tion of PMA. DSBD in lung tissue of rats treated with PMA was also markedly increased comparedwith the controls. The PMA-treated rats showed significantly higher lipid-peroxide (LPO) level inplasma and lung tissue hemogenate than the controls did. These results suggest that determination ofDSBD, a simple and sensitive indicator for oxygen radical damaging, might be useful in thediagnosis of adult respiratory distress syndrome (ARDS), when it is used together with themeasurement of plasma LPO.
基金This project was supported by a grant of the National Nat-ural Sciences Foundation of China (No.39770 72 4)
文摘To investigate the correlation between the activity of kinases in the growth factor signal transduction pathway and the development of resistance of breast cancer to tamoxifen, reporter gene regulated by the regulating fragment of CCD1 was transfected into the MCF 7 cells, and the influence of tamoxifen on the reporter gene expression was examined under different conditions of TPA treatment. Our results showed that the reporter gene expression was inhibited by tamoxifen and promoted by TPA. Furthermore, tamoxifen exerts an agonist effect on the reporter gene expression when the cells was treated by TPA previously for 12 h. It is concluded that TPA could induce estrogen like effect of tamoxifen on estrogen receptor positive breast cancer cells and it may be one of the mechanisms responsible for the development of tamoxifen resistance.
基金This work was supported by the National Natural Science Foundation of China(Grant NoS.81100169).
文摘Relaxin is known to inhibit cardiac fibrosis. However, it is unclear whether relaxin could regulate the effects of Phorbol 12-myristate 13-acetate （PMA, PKC activator） on cardiac fibrosis. So the influence of relaxin on the cell proliferation and collagen expression induced by PMA in cultured cardiac fibroblasts was studied. It showed that PMA significantly increased cardiac fibroblasts proliferation, Type I pro-collagen protein expression, Type I pro-collagen mRNA expression, and rhRLX absolutely significantly decreased PMA induced effects on cardiac fibroblasts proliferation and Type I pro-collagen expressions, indicating that relaxin could inhibit cardiac fibrosis induced by PMA.
文摘Cardiomyocytes isolated from neonatal rats were treated with phorboll 2-myristatel 3-acetate (PMA ) ranging from 10(-11)to 10-7 mol/L for 20 min, causing cytosol protein kinase A (PKA) activity to decrease while particulate PKA activity increase in a concentration-dependent manner. The change of PKA activity induced by PMA was abolished completely by pretreatment of polymyxin B or depletion of protein kinase C (PKC). Type II PKA activity in particulate fraction was enhanced remarkably, while that of type I PKA was not altered when the cells were treated with 100 nmol/L PMA. The results suggested that subcellular distribution and activity of PKA in cardiomyocytes may be regulated by PKC.
文摘Objective:To study the effect of arsenic trioxide (As2O3) combined with phorbol ester (PMA) on the proliferation of acute promyelocytic leukemia cell line Kasumi-1.Methods:Acute promyelocytic leukemia cell lines Kasumi-1 were cultured and randomly divided into control group (treated with the RPMI1640 medium without drugs or serum), As2O3 group (treated with serum-free RPMI1640 medium containing 20 μmol/L As2O3), PMA group (treated with serum-free RPMI1640 medium containing 160 nmol/L PMA) and As2O3+ PMA group (treated with serum-free RPMI1640 medium containing 20 μmol/L As2O3 and 160 nmol/L PMA). After treatment, the cell proliferation activity, cell cycle ratio and the protein expression of related genes were measured.Results: 12 h, 24 h and 48 h after treatment, the cell proliferation activity of As2O3 group, PMA group and As2O3+PMA group were significantly lower than that of control group, and the cell proliferation activity of As2O3+PMA group was significantly lower than that of As2O3 group and PMA group;48 h after treatment, the G1 phase and S phase ratio as well as CDK1 and CyclinB1 expression of As2O3 group, PMA group and As2O3+PMA group were significantly lower than those of control group while the G2 phase ratio as well as Bax, Caspase-3, Caspase-9, p-Chk1 and p-Cdc25C9 expression were significantly higher than those of control group;the G1 phase and S phase ratio as well as CDK1 and CyclinB1 expression of As2O3+PMA group were significantly lower than those of As2O3 group and PMA group while the G2 phase ratio as well as Bax, Caspase-3, Caspase-9, p-Chk1 and p-Cdc25C9 expression was significantly higher than those of As2O3 group and PMA group.Conclusion:As2O3 combined with PMA can inhibit the proliferation of acute promyelocytic leukemia cell line Kasumi-1 by inducing apoptosis and blocking cell cycle.
基金supported by the National Natural Science Foundation of China(grant nos.21925101 and 22331001)the Yunnan Provincial Science and Technology Project at the Southwest United Graduate School(grant no.202302AP370004).
文摘We have accomplished the efficient total synthesis of(+)-phorbol in 20 steps from inexpensive(+)-car-vone.This concise synthesis is partly due to the use of two pentamethyldisilyl(PMDS)groups as the masked hydroxyl groups.The significant advantages of the PMDS group over other silyl groups have been firmly demonstrated.This total synthesis features a Shapiro reaction that links two fragments possessing a unique PMDS group.This is followed by a Tamao-Fleming oxidation and a subsequent ring-closing metathesis reaction to construct the 5/7/6/3 tetra-cyclic skeleton.
文摘Protein kinase C with a molecular weight of 82 kD has been purified to electrophoresis homogenous from rat brain through a series of chromatography cohmns including DE-52, Sepharose G-200 and phenyl-Sepharose. The enzyme possessed autophosphorylation activity. Yuanhuacin A inhibited the ~3H-phorbol-12, 13-dibutyrate (~3H-PdBu) binding of PKC with an IC_(50) value of 1.48±0.287×10^(-8) mol/L when the concentration of ~3H-PdBu was 1.5×10^(-9) mol/L (K_i=1.2×10^(-8) mol/L). Yuanhuacin A inhibited the PdBu-stimulated PKC activity in the catalysis of the phosphorylation of Histone Ⅲ-S with an IC_(50) of 2.82±0.37×10^(-9) mol/L (PdBu=10^(-6) mol/L), while it had no effect on the basal and Ca^(2+)-stimulated PKC activity in the same assay system. This result suggests that Yuanhuacin A is a selective antagonist of the phorbol ester receptor in protein kinase C.
基金Grants Commission(UGC),New Delhi for providing Major Research Project financial Grant(F.No.34-250/2008(SR))to SCS.
文摘Activator protein-1(AP-1)transcription factor is a key component of many signal transduction pathways involved in the regulation of cellular processes and controls rapid responses of mammalian cells when exposed to the variety of stimulus.The phorbol 12-myristate 13-acetate and Forskolin(Fo)are well-known kinase activators/stimulators of Protein Kinase C(PKC)and Protein Kinase A(PKA)respectively.Importantly,these kinases are found to be present in transitional points of many cell signaling pathways,especially those involved in proliferation.The stimulating effect of PKC and PKA on the expression of AP-1 factors in MCF-7 breast cell proliferation is not well characterized.Hence,the role of PKC by PMA treatment and the role of PKA by using Fo in MCF-7 cells is investigated.Where,cells treated with PMA showed increased cell proliferation,while Fo had no effect,but inhibited the PMA induced proliferation.The RT-PCR results showed the PMA induced c-Jun,c-Fos and Fra-1 expressions compared to control and Fo.However,Fo in combination with PMA,inhibit the PMA induced above mRNA expressions where Fo alone has no effect.Western blot studies validated the c-Jun expressions in PMA treated MCF-7 cells.Further,PMA increases the mRNA expression of Cyclin-E1,Cyclin-D1,and CDK-4,whereas Fo decreases their expressions.Thus,mitogenic effect of PMA and inhibitory action of Fo on MCF-7 cells is probably enhanced via activation of AP-1 factors and concomitant action of cell cycle regulators in the downstream singling cascade.
