Te ine i ngā momo hāora hohe i ngā whakamātautau puru metilīni

E rua ngā taiwhanga e whakahaere ana i tā tēnā, i tā tēnā e kī nei he whakamātautau ārai hāoratanga mō te puru metilīni. E pūrongo ana tētahi ka tāmi te waikano i ngā momo hāora hohe. E pūrongo ana tērā atu ka whakaputa kē i aua momo. E whakaatu ana ngā taiwhanga e rua i ngā ānau mārama o te urupare ki te rahinga. He pono ngā taiwhanga e rua. Ehara te rerekētanga i te tinihanga, i te kore mōhio rānei. Ko te take kē, i ine ngā rōpū e rua i ngā mea rerekē, i raro i ngā māramatanga rerekē, mā ngā pūoko e whakautu ana i ngā pātai rerekē, kātahi ka whakaahua i ngā whakautu e rua mā te kupu kotahi: ROS.

He rāpoi ngota uaua te puru metilīni ki te ine tika, nā te mea ka whai wāhi ki te matū o te pūnaha e whakaarotia ana māna e mātakitaki. I te pōuri, ka kawe ia i ngā irahiko i waenga i ngā kaituku me ngā kaiwhiwhi, ā, ka hoki ki tōna āhua tīmatanga, kua rite ki te huri anō i te huringa. I raro i te rama, ka noho hei matū whakahohe mā te aho, ka tae ki tētahi āhua torutoru kua whakaohohia. Mai i taua āhua ka taea e ia te tuku pūngao ki te hāora kia puta ai te hāora takitahi, te whakawhiti irahiko rānei kia puta ai ngā tuwhena me te perōkai hauwai. Ka taea e te waikano tonu te whakahāora, te whakaheke, te whakapōuri i te kōrekoreko, te huna ā-whatu rānei i tētahi pūoko ka whakaurua ki taua pūnaha. Nō reira, ehara rawa te pātai tuatahi mō tētahi pānuitanga ROS i te pātai he aha te tau. Ko te pātai kē, he aha rawa tā te whakamātautau e kite ai, ā, he aha tā te waikano e mahi ai ki taua whakamātautau.

Amplex Red: he whakamātautau perōkai hauwai kua whakatikatikahia

Ko te whakakotahitanga puru metilīni me Amplex Red te takirua e tino taea ana te tautoko i roto i ēnei tuhinga rangahau, ina tika te whakarite. He tauwhāiti te tauhohenga matū: ka whakamahia e te peroxidase horseradish te perōkai hauwai hei whakahāora i te tïpako Amplex kia puta ai te hua kōrekoreko, arā, te resorufin, me te ōwehenga matū e tata ana ki te kotahi ki te kotahi. Ka huri te perōkai o te tauira hei kōrekoreko i te pūrere kite. Kāore tētahi atu o te whānau ROS tikanga e kōkiri tika ana i taua tauhohenga.

Ko te painga matua ko te whakatikatika ine. Ka whakamātau tētahi kaimahi tūpato i ngā paerewa perōkai e mōhiotia ana i roto i te wairewa ārai pH kotahi, me ngā kukūtanga peroxidase me te tïpako kotahi, te wā whakatipu kotahi, me ngā tautuhinga whatu kotahi, kātahi ka pānui i ngā tauira mā te whakataurite ki taua ānau. I te rangahau mō ngā mitokōria roro kua wehea e tūāpapa ana i tēnei tikanga, i pēnei tonu ngā kaituhi: i tāpirihia ngā rahinga perōkai e mōhiotia ana i te mutunga o ia whakamātautau. Ka pūrongotia ngā tau hei reiti, ki te pikomōra ia meneti mō ia mirikarama pūmua, kaua hei waeine kōrekoreko noa. Koinei te mea e huri ai te kanapa hei inenga.

E toru ngā taipitopito o taua rangahau e tika ana kia whāia. Tuatahi, i whakaū ngā kaituhi i te tohu mā te catalase, e whakangaro ana i te perōkai. I mutu te pikinga kōrekoreko i whakaaweawehia e te puru metilīni i te tāpiritanga o te catalase, e whakaū ana he perōkai te pūtake o te tohu, ehara i te tohu horihori ā-whatu. Tuarua, i whakamātau tika rātou i te whakamarama mā te kati i te kurupae whakaoho kia whiwhi ai ngā tauira i te kotahi hauwaru o te rahinga aho noa, ā, e iwa ōrau noa iho te hekenga o te tohu. Neke atu i te iwa tekau ōrau o te perōkai i pūrongotia e rātou kāore i whakawhirinaki ki te aho ine. Tuatoru, i ine motuhake rātou i te whakakorenga perōkai mā tētahi kawehiko, ā, i kitea ka whakapiki te waikano i te whakaputanga, ka whakaroa hoki i te whakawāteatanga. Mehemea i pūrongo te pūoko i te kōrekoreko toenga anake, kua whakahanumihia aua pānga e rua.

He āhuatanga tonu e hapa ai te whakamātautau, ā, ka pā te puru metilīni ki ētahi o aua āhuatanga. Ka ngongo whānui te waikano i roto i te rohe whero kotahi e tuku aho ai te resorufin. Nō reira, i ngā kukūtanga waikano tiketike, ka taea e te tauira te huna i tōna ake tohu mā ngā pānga tātari o roto. Nā reira me rite te kukūtanga waikano o ngā paerewa ki tō ngā tauira. Ka taea hoki te Amplex Red te whakahāora mā te aho i a ia anō, ā, ka taea e tōna hua, te resorufin, te uru ki ngā huringa hāora-whakaheke mā te aho e whakaputa ana i te superōkai me te perōkai. He take nui tēnei ina whakamāramatia hoki tētahi matū whakahohe mā te aho i roto i te whakamātautau. Ko te whakatikatika whaihua, me whakamārama tuatahi ngā tauira puru metilīni, kātahi ka whakamātau i ngā wāhanga tauira ki ngā matū Amplex i te pōuri, kaua e whakamārama tahi i te waikano me te pūoko. Hei whakamutunga, ehara te mahi a te superoxide dismutase i te tāmi noa i tētahi tohu Amplex: ka taea e te huringa o te superōkai hei perōkai te whakapiki i taua tohu. Kāore tētahi mana whakahaere SOD e whakapiki ana i te kōrekoreko i hapa. Kua tautuhia kē e ia te superōkai hei momo kei mua ake i te ara tauhohenga.

DCFDA: he tohu hāoratanga kāore i tauwhāiti

Ko te takirua puru metilīni me DCFDA te pūtake o te nuinga o te rangirua. He matū tīmatanga te DCFH-DA mō te whakauru pūoko ki te pūtau. Ka tangohia e ngā esterase o roto i te pūtau ōna rōpū acetate, ka mahue ko te DCFH kāore e kōrekoreko, ā, mā te hāoratanga o te DCFH ka puta te DCF kōrekoreko. He roa te rārangi o ngā mea e whakahāora ana i a ia: ngā tuwhena hydroxyl, ngā matū takawaenga peroxidase me te heme, te hauota hauhā, te waikawa hypochlorous, me ngā tuwhena i ahu mai i te peroxynitrite. Kāore te perōkai hauwai anake e whakahāora tika i a ia i tētahi reiti whaihua; ko te tikanga me whai rino, pūmua heme, tauhohenga peroxidase rānei te kōrekoreko e whakawhirinaki ana ki te perōkai.

Ko te arohaehae e tika ana kia tohutorohia i ia wāhanga tikanga rangahau, i ahu mai i te tātari a Kalyanaraman me ōna hoa i te DCFH hei pūoko, e whakarāpopototia ana i te rangahau mitokōria roro i whakakore i te DCFDA mō tēnei take. I whakaatu rātou he pōturi te tauhohe a te waikano, kāore anō tōna ōwehenga matū kia tautuhia, kāore he whakatikatika ine whaimana, ā, ka tauhohe hoki ki ngā pūhui ehara i te ROS. Ka whakakaha hoki te pūoko i tāna e ine ai: mā te hāoratanga irahiko kotahi o te DCFH ka puta tētahi tuwhena DCF, ka tauhohe ki te hāora kia whakaputa ai i te superōkai, ka wehe-tauhohe ai taua superōkai hei perōkai anō. Nō reira, ko tētahi wāhanga o te matū e pūrongotia ana e te pānuitanga, nā taua pānuitanga tonu i hanga.

Ehara tēnei i te kī kāore he painga o te DCFDA. Ko te tikanga, he whakamātautau tirotiro kē, ehara i te tautuhi. Ko te pikinga o te kōrekoreko DCF e tohu ana kua piki te hāoratanga o te pūoko e whakawhirinaki ana ki ngā matū whakahāora. Kāore e tohu ana kua piki te perōkai, ā, kāore rawa e tohu ana kua piki te tapeke ROS, he rahinga kāore e taea e tētahi pūoko kotahi te ine. I pūrongo ngā rangahau puru metilīni o mua i whakawhirinaki ki te hāoratanga DCF i runga ake i te kotahi maikromōra i ngā pānga whakatairanga hāoratanga. I taua wā anō, i pūrongo ētahi atu pūnaha i te whakamarutanga i te mate pūtau nā te perōkai. Ka taea ēnei kitenga e rua te noho pono mō te waikano kotahi i ngā horopaki hāora-whakaheke rerekē, ā, kāore e taea e te hua DCF anake te whakaatu ko tēhea matū whakahāora i kawe i te tohu. Ko te hiranga o te rangahau mitokōria roro o mua, i whakakapia e ia tēnei whakamātautau tirotiro ki tētahi inenga perōkai kua whakatikatikahia, ā, i kitea tōtika te taumahatanga: ahakoa kotahi rau nanomōra noa te waikano, i piki maha whakareatanga te tukunga perōkai i ia tïpako me ia āhua hā pūtau i whakamātauria.

He mahanga anō tō DCFDA e pā motuhake ana ki te puru metilīni. He kaha te tae o te waikano, ā, he hohe ki ngā tauhohenga hāora-whakaheke. Nō reira ka taea e ia te whakapōuri i te kōrekoreko, te tinei i te tukunga aho, te whakahāora rānei i ngā matū o te pūoko me te kore whakawhirinaki ki te ROS pūtau. Ehara tētahi mana whakahaere kāore he pūtau, engari he waikano me te pūoko, i te whakapaipai kōwhiringa noa. Koia te whakamātautau e whakaatu ana mēnā ka tauhohe te waikano ki te pūoko.

Te hopu takahuri me te horomanga hāora: te tuakiri me te rere

Ka pūrongo ngā pūoko kōrekoreko kua puta tētahi mea e taea ana te whakahāora. Engari, mā te orooro takahuri irahiko me ngā matū hopu takahuri ka tata ake te pūrongo ki te tautuhi he aha taua mea. I whakamahia e te rangahau matarohia mō te matū aho o te puru metilīni te ESR hei kite i te tuwhena ascorbate i ahu mai i te waikano, te ascorbate me te aho. Kātahi ka whakaaturia te hanganga perōkai i ngā whakamātautau horomanga hāora, me te hanganga tuwhena hydroxyl nā te whakakōkī a te rino i ngā whakamātautau hopu takahuri. I whakautua e ia tikanga te pātai kāore i taea e ērā atu te whakautu: nā te ESR te tuakiri tuwhena, nā te horomanga hāora te rere tauhohenga kāore e whakawhirinaki ana ki tētahi pūoko kōrekoreko, ā, nā te whakakotahitanga i whakaatu e mahi tahi ana ngā ara hāora takitahi me ngā ara tuwhena i roto i te pūnaha kotahi kua whakamāramatia.

He rite tonu te pānui i taua hua o 1984 ki te pānui i tētahi tapanga whakatūpato. I te aroaro o te aho me tētahi matū whakaheke, i whakaputa te waikano i te perōkai. I te aroaro hoki o te rino, i whakaputa te pūnaha i te tuwhena hydroxyl, te momo tino tauhohe o taua huinga. I hono ngā kaituhi i tēnei ki te ascorbate o te karu, ā, i whakapae i te whakamahinga noa ki ngā karu me te kore āta whakaaro. Me aro ngā kaipānui o ēnei rā ki ngā āhuatanga: te aho me te matū whakaheke me te rino. Tangohia tētahi o ēnei, ka ngoikore te ara, ka ngaro rānei.

He take kē e whai wāhi ai ngā kawehiko hāora. Ka ine tētahi kawehiko momo Clark i te kohinga hāora e pau ana, e whakaū ana kei te whakapau hāora rāpoi ngota ngā tauhohenga hāora-whakaheke e kōkiritia ana e te waikano. Ka whakarato hoki i ngā raraunga tere tauhohenga kāore e taea e tētahi tohu horihori kōrekoreko te hanga. Kāore ia e tautuhi i te hua. Ka pau te hāora i te hanganga o te perōkai, te superōkai me te wai, me te memeha o te hāora takitahi. He rite te kore oti o te rere kāore he tuakiri ki te tuakiri kāore he rere. Nō reira ka whakakotahi ngā pepa rangahau pakari i te kawehiko ki tētahi whakamātautau perōkai kua whakatikatikahia, ki tētahi matū hopu rānei.

He aha ngā hua ārai hāoratanga e taupatupatu ai

Whakatakotoria ngā pānuitanga e toru ki te taha o tētahi ki tētahi, ā, ka kitea ko ngā taupatupatu o ngā tuhinga rangahau puru metilīni he rerekētanga tikanga, me ngā meka koiora e toru.

Tuatahi, he kaikawe irahiko te waikano, he pūtake perōkai hoki. Mā te whakatahuri i ngā irahiko i ngā wāhi hā pūtau e huri kē ai te hāora hei superōkai, ka taea e ia te whakaheke i tētahi ROS. I taua wā anō, ka whakaheke tōna āhua kua whakahekea i te hāora kia tika tonu te puta hei perōkai, ka whakapiki ai i tētahi atu. Nō reira, me tūmanako tētahi taiwhanga e mahi ana ki te matū hāora-whakaheke mitokōria me tōna mahi hei ara karo ki ngā whakautu e whakawhirinaki ana ki te pūoko: ka heke tētahi pānuitanga e aro ana ki te superōkai, engari ka piki tētahi pānuitanga perōkai i roto i te ipu ine kotahi.

Tuarua, i ngā rahinga iti, he tohu te perōkai, ehara i te kino anake. I kitea e te rangahau mitokōria roro kiore i whai mai i te mahi o mua te whakaputanga perōkai mai i te rima tekau nanomōra noa o te waikano, me te kore wharanga DNA mitokōria i ngā taumata haumanu, me te whakamarutanga i ngā wharanga nā te rotenone i muri i te maimoatanga tōmua. I te kotahi rau maikromōra anake, kei runga rawa ake i te awhe haumanu, ka tūkino te waikano i ētahi wāhanga DNA kua tīpakohia. Nā te rama whero me te rua tekau maikromōra waikano i whakaputa te hāora takitahi i tūkino i ngā wāhanga katoa i whakamātauria. Ko te tikanga e whakaarohia ana he hormesis: ka whakaoho te whakaputanga perōkai iti i ngā parepare ārai hāoratanga e hono ana ki te Nrf2, me te hanganga mitokōria. Nō reira, ka puta he pānuitanga whakatairanga hāoratanga i te wā poto i mua i tētahi hua ārai hāoratanga i te wā roa. Ka taea te rerekē o tētahi whakamātautau ka pānuitia i te kotahi hāora me tētahi ka pānuitia i te kotahi rā, nā te mea kua rerekē te koiora i waenga i aua wā.

Tuatoru, mā te aho tonu e whakatau ko tēhea rāpoi ngota e noho ana. Ka tango te whakamarama i ngā rōpū metilī i te waikano kia ahu atu ai ki te Azure B me ngā hua e pā ana. He nui noa te rama karu whakarahi i runga ake i tētahi rīhi hei whakahohe i a ia mā te aho. He whakamātautau rerekē ngā whakamātautau pōuri me ngā whakamātautau kua whakamāramatia. Me pērā hoki te titiro a te wāhanga mana whakahaere mō te whakamarama me te whakahaere waikano o ia kawa whakamātautau.

He huinga mana whakahaere iti rawa, me te pānui i ngā kerēme

Ko te whakataurite i raro nei te rauemi tohutoro taketake mō tēnei tuhinga. I mua i te whakaae ki tētahi hua ROS puru metilīni, tirohia ko ēhea rārangi i whakatutukihia e ngā kaituhi, ko ēhea hoki i mahue.

Ka whai mai tētahi ture pānui whaihua. Ko te Amplex me te peroxidase, kua whakatikatikahia, kua arowhaingia hoki ki te catalase, he taunakitanga mō te perōkai. He taunakitanga te DCFDA mō te hāoratanga pūoko kāore i tauwhāiti, kāore he mea kē atu. He taunakitanga te hopu takahuri me ngā mana whakahaere matū hopu tuwhena mō te tuakiri o ngā tuwhena. He taunakitanga te horomanga hāora mō te rere. Ko tētahi rerenga kōrero e kerēme ana i tua atu i ngā mea ka taea e tōna whakamātautau te tautoko, ahakoa te māia o te tuhi, he rerenga mō te pūoko, ehara i te rerenga mō te waikano. Ka whakawhānui ngā matapakinga e pā ana ki te whakawhirinaki ki te rahinga me te awhe hormesis, te rere irahiko mitokōria, te whakararuraru whakamātautau i ngā tauira kua waikanotia, me ngā tohu horihori nā te whakamarama karu whakarahi, i ia take nei. Ka whaimana tonu ia take ina whakakapia tōna tohutoro puriwāhi.