Methylene blue, Nrf2, da samuwar sabbin mitochondria: abin da aka auna da abin da har yanzu ake hasashe

Ana yawan samun wata jimla a rubuce-rubucen da suka shafi methylene blue: yana kunna Nrf2 kuma yana gina sabbin mitochondria. Jimlar tana kama da ikirari guda ɗaya, amma ta ƙunshi ra'ayoyi daban-daban guda biyu, waɗanda aka yi nazarinsu a tsarin gwaji daban-daban, tare da nau'ikan hujjoji daban-daban. Na farko hanyar isar da sigina ce, wani na'urar ganowa da ke kunna kwayoyin gado na kariya. Na biyu shirin gini ne, inda tantanin halitta ke ƙera ƙarin kayan aikin mitochondria. Sigina na iya tayar da aikin gini, amma nuna cewa siginar ta tashi ba daidai yake da nuna cewa an kammala ginin ba, kuma nuna ƙarin kayan gini ba daidai yake da nuna ingantacciyar samar da makamashi ba. Wannan labarin ya ware waɗannan abubuwa uku daga juna.

Ra'ayin Nrf2: na'urar ganowa, ba garkuwa ba

Nrf2 (nuclear factor erythroid 2-related factor 2) furotin ne mai sarrafa kwafin bayanan kwayoyin gado, wanda aikinsa shi ne kunna kwayoyin gado. A yanayin da babu wani abin tayar da martani, wani furotin mai ganowa da ake kira Keap1 yana riƙe shi a cytoplasm, sannan ya miƙa shi ga tsarin zubar da abubuwa na tantanin halitta don ci gaba da lalata shi. Idan sinadaran da ke haifar da oxidation ko electrophiles suka canza cysteines masu saurin amsawa da ke kan Keap1, wannan zubarwar tana tsayawa. Sabon Nrf2 da aka ƙera yana taruwa, ya shiga tsakiyar tantanin halitta, ya haɗu da ƙananan furotin na Maf, sannan ya ɗaure da jerin DNA da ake kira wuraren amsa na kariya daga oxidation. Wannan ɗaurin yana kunna jerin kwayoyin gado na kariya.

Ma'aunan da aka fi amincewa da su na wannan hanyar takamaimai ne. NQO1 (NAD(P)H quinone dehydrogenase 1) da TrxR1 (thioredoxin reductase 1) suna daga cikin kwayoyin gado da Nrf2 ke nufi waɗanda aka fi dogaro da su. Ana iya tayar da HO-1 (heme oxygenase-1) sosai, amma shi ma yana amsa wa wasu masu sarrafawa. Enzymes kamar SOD da catalase kayan aikin kariya ne daga oxidation, amma furotin masu sarrafa kwafin bayanan kwayoyin gado da yawa suna sarrafa matakansu, don haka ƙaruwar SOD kaɗai ba ta tabbatar da cewa Nrf2 ne ya jawo ta ba. Hujjar da ta fi kusa da faruwar siginar ita ce taruwar Nrf2 a tsakiyar tantanin halitta, kunna na'urar rahoton ARE, ko tayar da kwayoyin gado da aka san Nrf2 ke nufi, wanda yake gushewa idan an cire Nrf2. Duk sauran abubuwan, kamar raguwar sinadaran da ke haifar da oxidation, ingantacciyar rayuwa yayin damuwa, ko ƙaruwar glutathione, sakamako ne da ke zuwa daga baya wanda hanyoyi da yawa za su iya haifarwa.

Sakamakon farko na methylene blue a wannan labarin ya dace da ɓangaren wannan ma'auni da ya fi kusa da siginar. Fibroblasts da aka fallasa wa methylene blue a matakan nanomolar sun nuna jinkirin tsufar tantanin halitta tare da ƙarin adadin complex IV, ƙarin amfani da oxygen, da tayar da enzymes na phase-2, TrxR1 da NQO1, kamar yadda aka ruwaito a binciken da ya fara danganta methylene blue da inganta mitochondria da jinkirta tsufar tantanin halitta. TrxR1 da NQO1 su ne ainihin kwayoyin gado da aka san Nrf2 ke nufi, waɗanda mai karatu mai lura zai so ya gani. Abin da binciken bai nuna ba shi ne faruwar siginar kanta: babu auna Nrf2 a tsakiyar tantanin halitta, babu na'urar rahoton ARE, babu gwajin kwatantawa da aka kawar da Nrf2. Saboda haka, fassarar da ta dace ita ce methylene blue yana tayar da kwayoyin gado da aka san Nrf2 ke nufi a wannan tsarin. Wannan ya yi daidai da kunna Nrf2, amma bai ware Nrf2 a matsayin sanadin ba. Tayar da kwayar gado da ake nufi mataki ɗaya ne bayan an kunna hanyar.

Ra'ayin samuwar sabbin mitochondria: shiri mai wuraren dubawa uku

Samuwar sabbin mitochondria shi ne shirin da tantanin halitta ke amfani da shi wajen ƙara ƙarfin aikinsa na mitochondria. Tsarin matakansa kamar yadda littattafan koyarwa suka bayyana yana bi ta wuraren dubawa uku. Da farko, mataimakin kunnawa PGC-1alpha yana kunna shirin. Na biyu, furotin masu sarrafa kwafin bayanan kwayoyin gado a tsakiyar tantanin halitta suna ƙarfafa bayyana kwayoyin gado na mitochondria, ciki har da TFAM, wanda ke kula da DNA na mitochondria kuma yake yin kwafinsa. Wani abin ruɗani shi ne cewa ana kiran ɗaya daga cikin waɗannan furotin NRF1, nuclear respiratory factor 1, wanda ya bambanta da Nrf2. Na uku, tantanin halitta yana samun ƙarin cikakkun bayanan gado na mitochondria da ƙarin kayan aikin samar da makamashi ta hanyar numfashi. Ana iya auna waɗannan ta adadin kwafin mtDNA, aikin citrate synthase, ko yawan tarin furotin na numfashi da aikinsu.

Kowane wurin dubawa yana bayar da ikirari mai rauni ko mai ƙarfi fiye da wanda ya gabace shi. Ƙaruwar RNA mai ɗaukar saƙo na PGC-1alpha ko TFAM tana nuna cewa an nemi fara shirin, ba cewa an kammala shi ba. Ƙarin mtDNA ko ƙarin furotin na complex IV yana nuna ƙarin kayan aiki, amma mitochondria da aka sake tsara su na iya nuna irin wannan yanayin. Ingantaccen aikin numfashi, ƙarin amfani da oxygen ko aikin complex, yana nuna cewa aiki ya canza, amma aiki na iya inganta ba tare da samuwar ko mitochondrion guda ɗaya ba. Ikirari mai ƙarfi na samuwar sabbin mitochondria yana haɗa abubuwa uku: mai sarrafawa, adadin kayan aiki, da aiki.

Binciken da aka yi don bin diddigin fibroblasts ya bi mafi yawan wannan jerin. A tantanin halittar ɗan adam na IMR90 da aka yi wa amfani da methylene blue mai 100 nanomolar, rabon NAD zuwa NADH ya ƙaru na ɗan lokaci cikin 15 mintuna, sannan ya koma matakin farko cikin kwana ɗaya. Bayan wannan siginar makamashi ta ɗan lokaci, an sami ƙara rukunin phosphate na ɗan lokaci ga na'urar gano makamashi AMPK, sannan tayar da PGC-1alpha da SURF1, wani furotin da ake buƙata don haɗa complex IV mai aiki. Aikin complex IV ya kusan ninka sau biyu, sinadaran da ke haifar da oxidation a tantanin halitta sun ragu da kusan kashi 28, kuma telomeres sun ci gaba da lalacewa a hankali. An bayyana cikakken jerin a takardar binciken da ke haɗa aikin redox na methylene blue da AMPK da samuwar sabbin mitochondria.

Wasu bayanai guda biyu a wannan takardar sun cancanci a kula da su domin su ne irin bayanan da ake yawan barin su. Na farko, salon kunnawar ya fi kasancewarta muhimmanci: methylene blue ya kunna AMPK na ɗan lokaci, yayin da maganin kwatantawa AICAR ya ci gaba da kunna shi na dogon lokaci, kuma mai kunnawa na ɗan lokaci ya jinkirta tsufar tantanin halitta da tasiri kusan sau takwas fiye da ɗayan. Saboda haka, ikirari irin na cewa "methylene blue yana kunna AMPK" bai cika ba; abin da ya dace a faɗa shi ne yana kunna AMPK na ɗan lokaci, kuma kwatantawar tana nuna cewa kunnawa na dogon lokaci ba ya yin aikin iri ɗaya. Na biyu, methylene blue bai canza zagayen rayuwar tantanin halitta ba. Wannan ya kawar da bayanin madadin da ya fi sauƙi: cewa tantanin halittar kawai suna rabuwa a hankali ne, don haka suka yi kama da matasa.

Reshen binciken beraye: ɗabi'a, bayanan gado, da wani abin tayarwa da ke kama da saɓani

Wata ƙungiya mai nazarin beraye masu matsakaicin shekaru ta faɗaɗa aikin gwajin tantanin halitta zuwa dabbobi. Bayan 60 kwanaki na ba da methylene blue ta baki, berayen da aka yi wa amfani da shi sun riƙe ƙarin juriyarsu ta jiki da ɗabi'ar binciken muhalli, kuma sun nuna ƙarancin ɗabi'a mai kama da damuwa fiye da tsofaffin berayen da ba a yi wa amfani da shi ba. A naman kwakwalwa, kwayoyin gado da tsufa ya danne, wato NRF1, TFAM, SOD2, da ɓangaren complex IV mai suna COX1, sun dawo kusa da matakan manyan beraye, kuma adadin kwafin mtDNA ya ƙaru. An ruwaito waɗannan sakamakon, wato ɗabi'a tare da masu sarrafawa da yawan cikakkun bayanan gado, a binciken beraye masu matsakaicin shekaru kan methylene blue da samuwar sabbin mitochondria a kwakwalwa.

Abin tayarwar da aka gabatar a matsayin hasashe shi ne ɓangaren da ya fi jan hankali, domin ya saɓa wa labarin mai sauƙi cewa methylene blue sinadarin kariya ne daga oxidation. Berayen da aka yi wa amfani da shi sun nuna ƙarin samar da nau'ikan oxygen masu saurin amsawa a mitochondria na kwakwalwa, musamman hydrogen peroxide. Hasashen marubutan shi ne: siginar peroxide mai sauƙi ita ce ke kunna hanyar Nrf2, wadda daga nan take ƙarfafa shirin samuwar sabbin mitochondria. A wannan tsari, methylene blue yana ƙarfafa oxidation a matakin tayarwa, sannan yana rage oxidation a matakin sakamako. Masana ilimin magunguna suna kiran wannan salo hormesis, inda ɗan ƙaramin damuwa yake tayar da kariya da ta fi damuwar kanta girma. Ba a gwada kai tsaye ko wannan ƙaruwar peroxide ce a zahiri take haifar da kunna Nrf2 ba; ita ce alaƙar da aka gabatar a matsayin hasashe, ba alaƙar da aka auna ba.

Wani bincike na haɗin gwiwa daga ƙungiyar ɗaya ya gwada ɓangaren aminci na wannan hasashe a ƙarƙashin yanayi masu ƙalubale. A mitochondria na kwakwalwa da aka ware, methylene blue ya ƙara samar da hydrogen peroxide a matakan da suka kai ƙarancin 50 nanomolar, amma matakan da ake amfani da su don jiyya ba su haifar da wata lalacewar DNA na mitochondria da aka iya ganowa ba. Sai a 100 micromolar, matakin da ya fi na jiyya nesa ba kusa ba, aka sami lalacewa a wasu zaɓaɓɓun ɓangarorin mtDNA. Haɗa sinadarin rini mai 20 micromolar da jan haske kuma ya samar da singlet oxygen wanda ya lalata mtDNA a wurare da yawa. Wannan shi ne sanannen halin sinadarin rinin na tayar da martani da haske, ba aikinsa na ilimin magunguna a duhu ba. Yin amfani da shi tun kafin fallasawa ma ya kare mitochondria na ɓangaren ƙasa na tsakiyar kwakwalwa daga raunukan mtDNA da rotenone ya haifar. An bayyana alaƙar adadi da martani da kuma gwaje-gwajen kwatantawa na cutar da kayan gado a binciken kan samar da peroxide da methylene blue ke yi ba tare da cutar da kayan gado ba.

Abin da aka auna da abin da ake hasashe: karanta zane

An zana hoton hanyar da ke ƙasa musamman don wannan labarin domin jaddada bambancin da rubutun ya riƙa yi. Akwatunan da ke da iyakoki masu layi cikakke matakai ne da ke da hujjar gwaji kai tsaye a rubuce-rubucen bincike kan methylene blue: zagayen redox da ke sauya NAD/NADH, ƙara rukunin phosphate ga AMPK na ɗan lokaci, tayar da PGC-1alpha da SURF1, ƙarin aikin complex IV, tayar da NQO1 da TrxR1, dawo da bayyana kwayoyin gado na samuwar sabbin mitochondria da adadin kwafin mtDNA a kwakwalwar beraye, da ƙaruwar peroxide ba tare da lalacewar mtDNA ba a adadin da ake amfani da shi don jiyya. Akwatunan da ke da iyakoki masu layi tsattsage su ne alaƙoƙin da aka gabatar a matsayin hasashe: Keap1 yana gano siginar peroxide, taruwar Nrf2 a tsakiyar tantanin halitta tana ƙarfafa kwayoyin gado na kariya, da kibiyar sanadi daga kunna Nrf2 zuwa shirin samuwar sabbin mitochondria. Waɗannan matakai masu layi tsattsage suna da ma'ana, sun yi daidai da bayanan gwajin, kuma suna cikin sanannen ilimin halitta na littattafan koyarwa. Amma a waɗannan takardu ba a auna su tare da gwaje-gwajen kwatantawa ba, wato Nrf2 a tsakiyar tantanin halitta, na'urorin rahoton ARE, da kawar da Nrf2, waɗanda za su ba su matsayin akwatuna masu layi cikakke.

Abubuwa uku masu ruɗani suna bayyana da zarar zanen yana gabanka. Na farko, NRF1 da Nrf2 furotin ne daban-daban masu sunaye kusan iri ɗaya; binciken berayen ya auna nuclear respiratory factor 1, mai sarrafa samuwar sabbin mitochondria, ba furotin mai sarrafa kwafin bayanan kwayoyin gado na kariya daga oxidation ba. Haɗa su a matsayin abu ɗaya yana mayar da dukan zanen zuwa kumburi guda. Na biyu, bayyana kwayoyin gado farkon labarin ne, ba ƙarshensa ba; dawo da saƙonnin TFAM ko COX1 yana goyon bayan buƙatar samuwar sabbin mitochondria, yayin da adadin kwafin mtDNA da aikin complex IV su ne hujjojin cewa an biya buƙatar. Na uku, sakamakon kariya daga oxidation ba ya tabbatar da hanyar Nrf2; raguwar sinadaran da ke haifar da oxidation da kashi 28 a fibroblasts da kariya daga rotenone a beraye sakamako ne na gaske da ke zuwa daga baya, amma hanyoyi da dama suna haɗuwa wajen samar da su.

Abin da zai sauya akwatin mai layi tsattsage zuwa mai layi cikakke

Gwaje-gwajen da suka rage sanannu ne. Auna furotin Nrf2 a tsakiyar tantanin halitta bayan fallasa shi ga methylene blue, tare da jadawalin lokaci da ya dace da siginar AMPK ta ɗan lokaci, zai gwada ko rassan biyu suna farawa tare da gaske, kamar yadda hasashen kunnawa a lokaci ɗaya yake ikirari. Na'urar rahoto da ARE ke sarrafawa za ta tabbatar da aikin kwafin bayanan kwayoyin gado maimakon hasashensa daga kwayoyin gado da ake nufi. Maimaita ma'aunan samuwar sabbin mitochondria bayan an rage Nrf2 ko an kawar da shi zai gwada kibiyar sanadi kai tsaye: idan tayar da complex IV da ƙaruwar mtDNA suka ci gaba ba tare da Nrf2 ba, alaƙar da aka gabatar ba daidai ba ce, kuma reshen AMPK shi kaɗai yake haifar da sakamakon. A ɓangaren dabbobi, binciken ba da sinadarin na dogon lokaci tare da wuraren dubawa guda uku iri ɗaya, wato mai sarrafawa, adadin kayan aiki, da aiki, zai gwada ko shirin yana ci gaba da kasancewa a kunne ko yana daidaitawa da yanayin. Har sai an yi haka, kunna hanyoyin makamashi da kariya a lokaci ɗaya zai ci gaba da kasancewa abin da marubutansa suka kira shi: wani abu da zai iya zama muhimmi, ba wani tsarin aiki da aka tabbatar ba.

Masu karatu da suka zo daga fagen ilimin magunguna mafi faɗi na iya son bayanan da ke kewaye da batun: yadda methylene blue ke safarar electrons a matakin mitochondria, sauran abubuwan da yake yi wa mitochondria da makamashin tantanin halitta, inda sakamakon tsufar tantanin halitta yake a binciken tsufa, yadda ikirarin samuwar sabbin mitochondria yake da alaƙa da aikin fahimta, da tarihin aminci wanda yake iyakance duk wata tattaunawa kan adadin amfani. Kowannensu ra'ayi ne daban mai nasa hujjoji, kuma kowannensu ya cancanci irin wannan warewar abin da aka auna daga abin da ake hasashe da wannan labarin ya yi wa Nrf2.