Classification
Epithelialtissueisdividedintocoveringepithelium,glandularepithelium,sensoryepithelium,reproductiveepithelium,myoepithelial,etc.Theepitheliumusuallyreferstothecoveringepithelium.
Epithelialcellscanbedividedintotwotypesaccordingtothenumberofcelllayers,monolayerandmulti-layer,andcolumnarandsquamousaccordingtotheirshape.
Columnarepithelialcellae: maxime distribuuntur in thenasalcatas, nasopharynx, organa, pulmones, ventris, intestina, cervix, endometrium et fallopiantubes.
Squamous epithelialcellae: cutis totum corpus, os, fauces, pars thenasopharynx, gula, allofvagina, et cervix.
Munus
Theepithelialcellsintheouterlayeroftheskinaregenerallykeratinized,whichcanprotectandabsorb.
Theepithelialcellsinthecavityarehighlydifferentiatedandhavethefunctionsofsecretion,excretionandabsorption.
Theepithelialcellsonthesurfaceofdigestiveorgans,lungsandotherorgansplayanimportantroleindigestion,absorption,andexchangeofoxygen.
Culture
I) Epidermalcellculture
1.Materials:Takesmallpiecesofsurgicalskingraftsorsurgicalresidualskin,preferablywiththinkeratinizationlayer,andbetterskinforprematureabortioninfants,cutintosmallpiecesof0.5to1squarecentimeters.
2.EDTAtreatment:primum putitin0.02%EDTAatroomtemperatefor5minutes.
3.Colddigestion:Changeinto0.25% trypsinandplaceat4°Covernight.
4.Separation:Takeouttheskinandseparatetheepidermisfromthedermiswithvascularforcepsortweezers.
5. Warmdigestion: Accipe epidermis et treatiseparate, cutintosmallerpieces forfices, putitinanew0.25% trypsin, et thendigestitat37°Cfor30-60minutes.
6.Useapipettetogentlypipetterepeatedlytomakeacellsuspension.
7.Culturesolution:Afterfilteringthrough80meshstainlesssteelgauze,centrifugeatlowspeed,suckthesupernatant,directlyaddEaglesolutionand20%calfserumtomakecellsuspension,inoculateitintodishes,andcultureinCO2incubator.
II) Ubera
Directculturemethod:(suitableforculturingsofttissuewithlessfiber)
1.InacontainercontainingasmallamountofculturesolutionorHankssolution,useasharpbladetorepeatedlycutthetissueintopieces.
2.Pourthetissuefragmentsandliquidintothecentrifugetube,addalittleculturemedium,pipetteforawhilewithapipette,andplacethetesttuberackfor3to5minutes.Aspiratetheupperlayeroffluidtoexcludethenon-breastcellpart.Repeat2to3times.
3.Afterthelasttreatment,addculturesolutiontothesedimentationtubeandpipettealittlewithapipettetoresuspendthesediment.Withoutwaitingforthecellmasstodrop,filteritintoanothertubethrough3to4layersofsterilegauze.
4.Adjusttheappropriatedensityandinoculateitintoacultureflaskforcultivation.
Collagenasedigestionmethod:(suitableforprocessinghardtissueswithmorefibers)
Theprocessisthesame culturae alia.
III) Cultureofgastricepithelialcells
1.Collection:Takeasmallamountofmucosainthedistalnon-lesionareaofgastricspecimensfromgastriculcerorgastriccancer.
2.Cleaning:Peeloff the lowermucosa withablunttoolandcutitintoasizeof1mm.
3.Digestion:DigestionintypeIcollagenaseandhyaluronidaseat37°Cfor80minutes.
4.Centrifugation:CollectthecellsuspensionandrinsetwicewithHankssolutionafter800rpm/centrifugation.
5.Inoculation:Afterthelastcentrifugation,addacompletemediumcontaining1%to2%fetalbovineserumandinoculateitintoacultureplatewithdifferentnumbersofwells.Theamountofinoculationdependsonthepurposeoftheexperiment.
IV) Hepatocyteculture
Firstgenerationtissueblockculture:Takefreshliver,firststripoffthefibrouscomponentssuchascapsuleandbloodvessels,andcuttheliverinto1mmwithaknifeorscissors3Thesmallpiecesontheleftandrightsidesarecultivatedonthewall.
5) Endothelialcellculture
1.Takethefreshumbilicalcordafterdelivery.Ifnotcultivatedimmediately,itcanbestoredat4°C,butitshouldnotexceed12hours.Cutasectionof10-15cmlongaseptically.Otherlargebloodvessels,suchasembryosandlarvae,canalsobeusedforculture.
2.First,useathree-waysyringetoabsorbthewarmPBSsolutionandinjectitintotheveinoftheumbilicalcordtowashawaytheresidualblood.Theinjectionportshouldbeligatedwithastringtopreventliquidreflux.
3.Clamponeendoftheumbilicalcordwithavascularclamp,andslowlyinjectcollagenasewithafinalconcentrationof0.1%intotheumbilicalveinfromtheotherend.Afterliquidappearsattheend,itisligatedtofillthebloodvessel.Theinjectionportshouldalsobeligatedtopreventliquidreflux.Digestfor3-10minutes.
4.Aspiratethedigestionsolutioncontainingendothelialcellsandpouritintoacentrifugetube.Inordertogetmorecells,injectwarmPBStorinse2or3timestocompletelyremovetheremainingcells,andthenpouritintothecentrifugetubeandcentrifuge.
5.Aspiratethesupernatant,add1640culturemediumtomakeacellsuspension,inoculateitintoaflaskanddishforculture,whenitgoeswell,thecellscangrowintoamonolayerwithin2to3days.
6) Capillaryendothelialcellculture
Preparationoftumorconditionedmedium;
1.S-180solidsarcomatissuefromC3Hmicewastaken.
2.Trypsindigestion,15mlofDulbecco'smodificatum est sculturemedium (continens 10% calfserum), inoculatum inoculatum-75Falconcultureflassforculture.
3.Whenthecellgrowthisclosetocompleteconfluence,theculturemediumiscollectedtomakeaconditionedmedium;afteranewmediumcontaining10%calfserumisused,itisalsocollectedeverytwodaystoobtainalargeamountofconditionedculture.
4.After4000revolutiones/centrifugae, itis filteredthrougha0.22μmmicroporousmembraneus-storedat-20°Cforlateruse