Imethylene blue kunye ne-nitric oxide: i-NOS, i-cGMP, imiqondiso yemithambo yegazi

Umbuzo uvakala ulula: ngaba imethylene blue iyayinqanda initric oxide? Elabhoratri impendulo yahlulahlulwa ibe yimibuzo emine eyahlukileyo, yaye ngamnye unovavanyo olwahlukileyo. Ukuzibhidanisa sesona sizathu sibangela ukuba okubhaliweyo ngemethylene blue kunye ne-nitric oxide kuvakale kuqinisekile kodwa kungacaci.

Cinga ngomqondiso we-nitric oxide njengekhonkco elinamalungu amane. Imfuza ikhutshelwa kwi-RNA ukuze kwenziwe initric oxide synthase. Ienzayim iguqula iarginine ibe yinitric oxide. Le gesi isasazeka iye kwindawo ekujoliswe kuyo ngaphambi kokuba isabele kwenye into. Iguanylate cyclase enyibilikayo iyayibona ize yenze iGMP eyisiklikhi. Iziphumo zemethylene blue kuwo onke la malungu zipapashiwe, kodwa ubungqina buvela kwiinkqubo zovavanyo ezahlukeneyo, kumazinga oxinaniso ahlukeneyo, nakwiindidi ezahlukeneyo zeenzayim. Isiphumo kwelinye ilungu asingobungqina bokusebenza kwelinye.

Ikhonkco eliqhelekileyo, lichazwe ngokucacileyo

Kudonga lomthambo wegazi, ulandelelwano oluqhelekileyo luhamba ngolu hlobo. Isivuseleli sisebenza kwiseli ye-endothelium. Initric oxide synthase ye-endothelium, ebizwa ngokuba yi-eNOS okanye yi-NOS3, yenza isixa esincinci se-nitric oxide ixesha elifutshane. Le gesi isasazeka iye kwiseli ekufutshane yesihlunu esigudileyo, ibopheleleke kwiheme enentsimbi ekwimo encitshisiweyo yeguanylate cyclase enyibilikayo, ize ikhawulezise ukuguqulwa kwe-GTP ibe yiGMP eyisiklikhi. IGMP eyisiklikhi ivuselela iprotein kinase G, indlela esetyenziswa ngayo ikhalsiyam iyatshintsha, size isihlunu siphumle.

Kukho ikhonkco lesibini ekufanele ligcinwe lahlukile. Ngexesha losulelo okanye lokuvezwa kwiicytokine, iiseli zikhuphela umyalezo wemfuza kwenye ienzayim, initric oxide synthase evuselelwayo okanye i-iNOS, ekwabizwa ngokuba yi-NOS2. Ivelisa ukuhamba kwe-nitric oxide okukhulu nokuhlala ixesha elide kunoko kwe-eNOS. Uhlobo lwesithathu, i-NOS yeeseli zemithambo-luvo okanye i-nNOS, lusebenza kwenye imeko. Xa iphepha lophando lisithi initric oxide synthase ngaphandle kokuchaza uhlobo lwayo, yima uze ukhangele ukuba loluphi olulinganisiweyo. Isishwankathelo sendlela imethylene blue ehambisa ngayo iielektroni senza ingongoma efanayo ngeendawo ekujoliswe kuzo ze-redox ngokubanzi: inkqubo yokudlulisela ayenzi nto iyodwa, yaye iqabane elikwisiphelo ngasinye limisela isiphumo.

IGMP eyisiklikhi elinganisiweyo ikwalufundo oludibanisa iinkqubo ezininzi, hayi uvavanyo oluthe ngqo lwe-enzayim. Inqanaba layo libonisa ukwenziwa kwayo yicyclase kuthatyathwe ukuchithwa kwayo ziiphosphodiesterase, ingakumbi i-PDE5 kwisihlunu esigudileyo semithambo yegazi. Ukuhla kweGMP eyisiklikhi kuthetha ke ukuba umqondiso omncinci ufike kwisamkeli. Ngokwayo, ayichazi ukuba leliphi ilungu elingasebenzanga.

Ilungu 1: kwenziwa ienzayim ye-iNOS encinci

Kwiimacrophage zeempuku ezincinci ezivuselelwe ngelipopolysaccharide yeebhaktheriya, nge-interferon gamma, okanye ngazo zombini, imethylene blue yathintela ukunyuka okulindelekileyo kweproteni ye-iNOS yaza yayicinezela kakhulu i-RNA engumthunywa ye-iNOS kuvavanyo lokukhutshelwa kwemfuza kwi-RNA kwiimacrophage. Eyona nto inika ulwazi koko kungazange kutshintshe. Ukuchithwa kwesithinteli i-I kappa B alpha, ukongezwa kwephosphate kwi-NF kappa B nakwi-STAT1, nokungena kwezo zinto kwinucleus, konke kwaqhubeka ubukhulu becala kungatshintshanga. Into eyehlayo yayikukubophelela kwazo kwi-DNA, okuqinisekiswe luvavanyo lwechromatin immunoprecipitation olwalubonisa ukuncipha kokubophelela kummandla okhuthaza ukukhutshelwa kwemfuza ye-iNOS.

Eso sisiphumo solawulo lwemfuza esenzeka ngaphambi kokumiselwa kobuninzi be-enzayim, hayi ukuthintelwa okukhawulezileyo kwe-enzayim esele ikho. Sisebenza ekudluliselweni kwemiqondiso yokudumba, apho izinto ezilawula ukukhutshelwa kwemfuza kufuneka zibopheleleke kummandla okhuthazayo ngaphambi kokuba kuvele iproteni. Asithethi nto ngokuthe ngqo nge-eNOS ehlala ikhona kumthambo ongavuselelwanga, yaye kufuneka sichazwe ngaloo ndlela.

Ilungu 2: ienzayim ye-NOS esele ikho isebenza kancinci

Uphononongo kwiimpuku ezinkulu lwavavanya inqanaba elahlukileyo. Iimpuku ezinkulu ezingamaduna ezisezingqondweni zafumana ilipopolysaccharide ngokuyifakwa emthanjeni, zaza emva kweeyure ezintathu zafumana isisombululo setyuwa okanye imethylene blue eyi-3 mg per kg emthanjeni. Umsebenzi we-nitric oxide synthase kwindawo ephakathi nesezantsi ye-hypothalamus walinganiswa ngokuguqulwa kwearginine ephawuliweyo ibe yicitrulline kuphononongo lomsebenzi we-NOS kwi-hypothalamus. Uxinzelelo kunye nesisombululo setyuwa kuphela lwavuselela umsebenzi othile ongaxhomekekanga kwikhalsiyam, yaye ilipopolysaccharide yawunyusa wada wamalunga namaxesha alishumi enqanaba leqela lolawulo. Imethylene blue yawuphelisa omabini la macandelo avuselelweyo. Umsebenzi osoloko ukhona oxhomekeke kwikhalsiyam awuzange utshintshe kwezo zicubu ngeso sixa nangelo xesha.

Sifunde eso siphumo kanye njengoko sibhaliwe: umsebenzi olinganisiweyo wohlobo lwe-iNOS wacinezelwa phantsi kwezo meko. Ayingomlinganiselo wokubonakaliswa kwemfuza, yaye ayibobungqina bokhetho olusisigxina lwe-iNOS endaweni ye-eNOS okanye i-nNOS. Uvavanyo lwe-enzayim esulungekisiweyo lulwenza lucace ngakumbi olu lumkiso. Kuvavanyo oluthe ngqo lwe-nitric oxide synthase esulungekisiweyo, imethylene blue yathintela ngokupheleleyo ukwenziwa kwecitrulline kwi-30 micromolar, ngoxinaniso oluthintela isiqingatha lomsebenzi olusondele kwi-5.3 micromolar xa kungekho superoxide dismutase nakwi-9.2 micromolar xa ikhona, njengoko kuchazwe kuthelekiso lwe-NOS esulungekisiweyo kunye neguanylate cyclase. Uphononongo olwahlukileyo lwe-nNOS esulungekisiweyo lwabonisa indima yokufakwa kweheme ebopheleleke kwi-enzayim kwimo ye-oxidation. Isifundo esibalulekileyo sesokuba ukuthintelwa kwe-NOS ngokuthe ngqo kuyenzeka kwiinkqubo ezibekwe zodwa, kodwa isiphumo se-hypothalamus asinakwandiswa sibe libango elibanzi lokuba imethylene blue ayichaphazeli zonke ii-NOS ezisoloko zikhona naphi na.

Ilungu 3: initric oxide ilahleka ngaphambi kokuba ifike

Obona bungqina bucacileyo bokuba enye into ngaphandle kokuthintelwa kwe-enzayim ingachaza ukuhla kweGMP eyisiklikhi buvela kwiiseli zesihlunu esigudileyo somthambo wemiphunga yomvundla ezikhuliswe elabhoratri. Uyilo lovavanyo lwayidlula ngokupheleleyo i-NOS eveliswa ngaphakathi ngokongeza izinto ezikhupha initric oxide ngokuthe ngqo, lwaza lwabuza ukuba imethylene blue isayicinezela na impendulo yeGMP eyisiklikhi kuphononongo lwe-superoxide kwisihlunu esigudileyo semithambo yegazi.

Izinto ezintathu ezikhupha initric oxide zavavanywa ukusuka kwi-1 ukuya kwi-100 micromolar: i-S nitroso L cysteine, isodium nitroprusside, kunye nesakhiwo esifana nazo kodwa esizinzileyo nesikhutshwe iqela le-amino, esibizwa ngokuba yi-MPANO. Zonke zayinyusa iGMP eyisiklikhi phakathi kwamaxesha amalunga ne-1.5 kunye ne-12. Imethylene blue eyi-10 micromolar yathintela ezimbini zokuqala ngepesenti eziyi-51 ukuya kwi-100, yaza yayishiya ingatshintshanga impendulo ye-MPANO. Ukuba imethylene blue ibithintela nje indawo yecyclase apho kuqhutywa khona ukuguqulwa kwezinto, bekuya kuba nzima ukuchaza loo mahluko phakathi kwezinto ezikhupha initric oxide, kuba zontathu ziphelela kwi-enzayim enye ekwinqanaba elilandelayo.

Emva koko iimvavanyo ezintathu zolawulo zanxulumanisa le ndlela yokusebenza ne-superoxide. Okokuqala, ukuvelisa isuperoxide ngaphandle kweeseli kusetyenziswa ixanthine kunye nexanthine oxidase kwavelisa ipateni efanayo: i-CYSNO nenitroprusside zathintelwa, i-MPANO ayachaphazeleka. Okwesibini, imethylene blue yabangela ukuba iiseli zesihlunu esigudileyo zivelise isuperoxide ngokuxhomekeke kwisixa nakwixesha, nto leyo eyalinganiswa ngokuncitshiswa kwecytochrome c. Okwesithathu, isuperoxide dismutase yakuthintela ngokupheleleyo oko kuthintelwa komqondiso, ngoxa icatalase ingazange ikwenze oko. Oku kwalatha kwi-superoxide kunakwihydrogen peroxide njengeyona nto ibangela isiphumo ngokuthe ngqo kuloo nkqubo yovavanyo.

Kufanelekile ukuchaza uhlobo lovavanyo lokukhulisa iiseli kunye, kuba lubonisa ukuba isiphumo senzeka phi. Iiseli ze-endothelium ezikwiiglasi ezincinci zokugquma zakhuliswa kunye neeseli zesihlunu esigudileyo okwethutyana, ukuze into ekhutshwa yi-endothelium kwimeko yayo esisiseko inyuse iGMP eyisiklikhi yesihlunu esigudileyo kathathu ukuya kasibhozo. Ukunyanga kuqala naliphi kula maqela eeseli ngemethylene blue kwawunciphisa loo mqondiso, yaye isuperoxide dismutase yasibuyisela ngokuyinxenye isiphumo sokunyanga kuqala i-endothelium. Ngamanye amazwi, akuyomfuneko ukuba imethylene blue isebenze ngaphakathi kwiseli yesihlunu esigudileyo esamkela umqondiso. Inokunciphisa umqondiso ngokususa umthunywa endleleni. Isikhokelo sokufunda uphando ngemethylene blue sikuthatha oku njengomgaqo jikelele: chaza indawo apho ungenelelo lwenzeke khona ngaphambi kokuchaza imolekyuli ekujoliswe kuyo.

Ukubiza oku ngokuthintelwa kwe-NOS bekuya kuba yimpazamo. Akukho nyathelo le-synthase elalifuneka ukuze izinto ezikhupha initric oxide zisebenze, kodwa umqondiso wehla nangona kunjalo.

Ilungu 4: isamkeli ngokwaso siphendula kancinci

Iziphumo ezithe ngqo kuguanylate cyclase enyibilikayo zikhona, kodwa amazinga oxinaniso abalulekile. Kuthelekiso lwe-enzayim esulungekisiweyo olukhankanywe ngasentla, icyclase evuselelwe yi-nitric oxide yayifuna malunga ne-60 micromolar yemethylene blue ukuze kuthintelwe isiqingatha somsebenzi wayo, xa kuthelekiswa nokuthintelwa kwesiqingatha se-NOS kufutshane ne-5 ukuya kwi-9 micromolar, yaye nokuba yi-1 millimolar yavelisa kuphela ukuthintelwa okungaphelelanga kumgangatho wayo ophezulu. Lowo ngumahluko omalunga namaxesha amathandathu ukuya kwalishumi elinanye. Uvavanyo lweeseli olunoxinaniso oluphantsi kwinqanaba le-micromolar olunciphisa iGMP eyisiklikhi ngoko alunakuchazwa njengokuthintelwa okukhethayo kwecyclase ngenxa nje yenani leGMP eyisiklikhi.

Uphononongo lwe-redox lwamva longeza isilumkiso sesibini: imethylene blue yenza icyclase evuselelwe yi-nitric oxide ingasebenzi ngendlela eyahlukileyo kukufakwa nje kwentsimbi yeheme kwimo ye-oxidation ngumoya. Ngoko isamkeli sinokuphazamiseka, kodwa indlela yemichiza ayikaqinisekiswa, yaye umsantsa phakathi kwamazinga oxinaniso uthetha ukuba iziphumo ze-superoxide ne-NOS kufuneka kuqala zihlolwe zize zikhutshelwe ngaphandle xa ukuvezwa kukwinqanaba eliphantsi le-micromolar. Kulapha ke apho kufuneka kungene khona uvavanyo lolawulo lwephosphodiesterase. Ngaphandle kokuba ukuchithwa kwayo kulinganisiwe okanye kuthintelwe, ukuncipha kokuqokelelana kweGMP eyisiklikhi kuyahambelana nokwenziwa okuncinci, ukuchithwa okukhawulezayo, okanye zombini.

Kutheni oku kungenakushwankathelwa kube libango elinye ngoxinzelelo lwegazi

Ilungu ngalinye laqinisekiswa kwinkqubo eyahlukileyo. Ukukhutshelwa kwemfuza kwi-RNA kwiimacrophage akufani nomsebenzi we-enzayim kwi-hypothalamus. Umsebenzi we-hypothalamus emva kwelipopolysaccharide awufani nemeko ye-endothelium engavuselelwanga. Into ekhutshwa yi-endothelium kwimeko esisiseko xa iiseli zikhuliswa kunye ayifani nokwandiswa kwemithambo yegazi okuvuselelwa sisivuseleli. Iiseli zemithambo yemiphunga ezikhuliswe elabhoratri ezikwi-10 micromolar yemethylene blue aziyiyo inkqubo yokuhamba kwegazi yomntu. Ukuzidibanisa zibe yintetho ethi imethylene blue inyusa uxinzelelo lwegazi ngokuthintela initric oxide kulahla kanye loo mahluko iimvavanyo zolawulo ezazenzelwe ukuwugcina.

Izinto ezintathu ekufuneka zihlolwe zigcina ingcaciso ithembekile. Okokuqala, chaza uhlobo lwe-enzayim: i-eNOS kwimeko esisiseko yokuxinana kwemithambo yegazi, i-iNOS ekuhambeni kwe-nitric oxide okuvuselelwa kukudumba, ne-nNOS kwiimeko zemithambo-luvo nakwezinye zeenzayim ezisulungekisiweyo. Okwesibini, chaza indlela umqondiso we-nitric oxide ohanjiswa ngayo: ukwenziwa ngaphakathi, ukukhutshwa yi-endothelium kwimeko esisiseko, okanye into ethile eyikhuphayo efana ne-CYSNO, initroprusside, okanye i-MPANO, kuba le nto izinzileyo yayimelana nemethylene blue ngoxa ezo zingazinzanga zingazange zimelane nayo. Okwesithathu, chaza uxinaniso nokubuyiselwa komqondiso: ukubuyiselwa kwawo yi-superoxide dismutase kubonisa indima ye-superoxide, ukusilela kwecatalase kunciphisa amathuba okuba ihydrogen peroxide ibe ngunobangela kuloo nkqubo yovavanyo, ukulinganisa nge-xanthine oxidase kubonisa ukuba oko kwanele ukuvelisa isiphumo, yaye iimvavanyo zeenzayim ezisulungekisiweyo zahlula iziphumo ezithe ngqo kumsebenzi wokuguqula izinto ekulahlekeni komqondiso endleleni.

Isishwankathelo esifutshane sesi. Imethylene blue ifanele iqondwe njengento ephazamisa kwiindawo ezininzi ukudluliselwa komqondiso ukusuka kwi-nitric oxide ukuya kwiGMP eyisiklikhi, kunokuba iqondwe njengesithinteli esikhetha icyclase kuphela. Ngokuxhomekeke kwinkqubo yovavanyo nakuxinaniso, inokunciphisa ukubophelela kwezinto ezilawula ukukhutshelwa kwemfuza ye-iNOS kwi-RNA, ithintele ngokuthe ngqo ienzayim ze-NOS, inciphise ukufumaneka kwe-nitric oxide ngokuvelisa isuperoxide, ize iphazamise icyclase evuselelwe yi-nitric oxide. Indima yokudluliselwa kwemiqondiso ngeGMP eyisiklikhi kwizifundo zemithambo yegazi kunye nothelekiso lwezinto ezikhupha initric oxide ezisetyenziswa kuvavanyo lweeseli nganye ixoxa ngokweenkcukacha ngesinye sezo ndlela, yaye iindlela ezisetyenziselwa ukwahlula iziphumo ze-redox kwiziphumo zeenzayim zichaza isizathu sokuba isuperoxide dismutase kunye necatalase kunye novavanyo lolawulo ngento ezinzileyo ekhupha initric oxide zivele rhoqo kangaka kolu phando.

Ixabiso eliphantsi leGMP eyisiklikhi emva kwemethylene blue libonisa ikhonkco eliqhawukileyo. Ziimvavanyo zolawulo kuphela ezichaza ukuba leliphi ilungu eliqhawukileyo.