In vitro and In vivo antimycobacterial, hepatoprotective ...
Evaluation of the novel nanocell (in vitro/in vivo)
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Transcript of Evaluation of the novel nanocell (in vitro/in vivo)
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Evaluation of the novel nanocell(in vitro/in vivo)Dr.Ramraj PantheeChulabhorn Research Institute
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OutlineIntroduction In vitro studiesIn vivo studiesSummary
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IntroductionNovel nanocellConfocal microscope In vitro Cell culture studies Co- culture studiesIn vivo Nude mice tumor studies
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The novel nanocellstructureFTY720Doxorubisin+5FU(D+5F)PLGA conjugation (Both)(F) Nanocell
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The novel nanocell Function24 13 NanocellTumor cellBVs shutdownNc into tumor cell
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Confocal microscopeStructure and function Laser ray 3 dimensional picture Computerized device
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In vitro studies controls
Vehicle is phosphate buffer solution(PBS)FTY720
Doxo +5FUDoxorubicin5FU
Dox+FTYDoxv
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Cell culture studies+Liver cancer cellsCalf serum culture medium24 hours5%Cells + Drugs Viable rateDrugs action on liver cancer cells Survival
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Cont. Cell culture studies
Plated for 24 hrs,Doxorubicin +5FU Conjugated mixed for 48hrs.Evaluation: Trypan blue exclusion method. Ec 50 calculation curve fittingCellcultureTrypan blue cell viabilityEc50 - 50% test cells effectiveby drugs
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Tumor-endothelium co-culture studiesLiver cancer cells with GFP Expre..Endothelial cellsMatrigelmatrixParaformaldehyde(4%)Incubation- propidium IodideExposed to different treatmentLiver cancer cellEndothelial cell
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Microscopic studies of cellControl 30 hFTY720DOXO+5FU Modified picture from: Nature vol. 436 July 2005 S.sengupta et.al.. Control 12 hNanocell 12hNanocell 30hNanocell in 12h action ,BV Nanocell in 30h tumor
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Microscopic studies (cont.)Yellow stained Liver cancer cellsRed stained - endothelial cellsDoxorubicin +5FU Tumor reducedFTY720 Vascular shutdownNanocell in 12hours little effectNanocell in 30hours complete tumor reduced
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Stereological quantification of co-culture
Vascular component:VEHICLE 12hr-no changeIn 30hrs. Vascular changed Model picture from Nature vol.436 2005------ 12 h--- -------30 h---------VehicleNanocellVehicle
FTY7205Fluouracil +Doxo
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Stereological quantification of co-cultureTumor component:Vehicle 12hrs no change
Doxo+5FU 30hrsModel picture from Nature vol.436 2005VehicleNanocellvehicleFTY720Doxo+5FTumor component
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Physicochemical release kinetic of drugsDoxorubicin + 5FU slow releaseFTY720 rapid releaseFTY720Doxorubicin+5FU Model picture from Nature vol. 436 2005
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In vivo nanocell studiesNude mouse
Nude mouseHigh sensitive to antigenWell manifestation of tumor
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In vivo nanocells (cont.) in vivo tumor studiesGFP expressionLiver cancer cellsImplanted male nude miceTumor vol. 50mm3Treatment started with 100 l ivFTY720 anddoxorbicin+5FUAnimals were killed in periods Necropsies tumor - histopathology
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In vivo NanocellNude mouse tumorLiver cancer cells injected into sub -cutaneous tissue Tumor 50mm3 in size
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Tumor studies
Size of tumor with different controls Modified picture from Nature vol.436,july 2005 S. sengupta et.al.D+5F+FF5FND+FV
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Effects of treatment
SurvivalToxicityTraditional chemotherapy 30days surviveNanocell combined 65 days surviveLess toxicWBC count show - normal Without any treatment20days survive Source-Nature:436:2005 S.Sengupta et.al
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Tissue distribution studiesNC + Fluorescence dyeInjected miceMice were killed 5, 10, 24 hrs. after injectionOrgans collection liver, lungs, spleen, tumor and serum after necropsedWt. organs and fluorescence extracted
Purpose - Distribution of drugs into the tissues Result Nanocells were detected within 5 hours
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SummaryA novel nanocell therapy Slow release Target specific Less toxic Cytotoxic Lifespan