Te puru metilīni me te kai-whaiaro: AMPK, mTOR, me ngā whakamātautau pūtau io

Whakaarohia kei te whakahaere koe i tētahi whakamātautau pūtau, ka tāpiri i te puru metilīni, ā, ka kite i tētahi rārangi pūmua e kīia nei ko LC3-II e pōuri haere ana i tō pepa tā pūmua. He māmā te hiahia ki te kī ka “whakapiki te pūhui i te kai-whaiaro”, kātahi ka haere tonu. He wawe rawa tēnā whakamāoritanga o te kitenga. Ko te pōuri haere o te rārangi LC3-II pea he tohu kei te hanga te pūtau i ētahi atu mirumiru horoi, he tohu rānei kei te putu ngā mirumiru nā te ārainga o te hipanga whakakore. E anga ana ēnei whakamāramatanga e rua ki ngā ahunga tauaro, ā, kei ngā tuhinga rangahau mō te puru metilīni i ngā pūtau io he tauira taunakitanga o ia momo. Ka tirohia e tēnei tuhinga ngā mea i ine tūturu ai, whakamātautau atu, whakamātautau mai.

Ko te kai-whaiaro whānui, te momo i rangahaua i konei, te tukanga hangarua ā-tōpū a te pūtau. Ka tupu tētahi pūtea kiriuhi e kīia nei ko te phagophore huri noa i ngā pūmua me ngā wāhanga pūtau kua tawhito, ka kati hei pūtea kai-whaiaro, ka hanumi ki tētahi lysosome, ā, ka wāwāhia ōna utanga, ka tukuna kia whakamahia anō. Ka whakawhirinaki ngā pūtau io ki tēnei tukanga auau, nā te mea kāore rātou e whakawehe, ā, kāore e taea te whakaiti i te kukū o ngā wāhanga kua tūkinotia mā te whakawehe. Ina tū te hangarua, ka putu ngā para. Ina haere tonu, ka whakahokia mai e te pūtau ngā wae hanga i raro i te ahotea.

Ngā mea i whakaaturia e tētahi whakamātautau hippocampus

I whakamahia e te rangahau matua mō te whakakōkiri ngā pūtau hippocampus kiore HT22 i whakamamaetia mā te kore serum mō te 24 hāora. Mā tēnei āhuatanga ka tangohia ngā āhuatanga whakatipu me ngā tohu tautoko ora, ā, ka whakaoho i te mate pūtau kua whakaritea me te kai-whaiaro. I pūrongo ngā kairangahau nā te puru metilīni i whakaiti te mate pūtau i runga i te rahinga i hoatu, ā, i haere tahi tēnei whakamarutanga me ngā tohu o te kai-whaiaro whānui.

He tauwhāiti ngā kitenga o ngā tohu. I tīmata te whakaritenga anō o te whakakikoruatanga LC3B marara hei ira huri noa i te karihi i te takiwā o te 20 nM puru metilīni, ā, ko te urupare LC3-I me LC3-II kaha rawa i te takiwā o te 1 μM. I piki ngātahi a LC3B-I me LC3B-II i runga i te rahinga i hoatu me te roanga o te wā. Ko te mea nui, i neke atu te rōpū i te tā LC3 tū noa. I tāpiritia e rātou te 10 μM chloroquine, e aukati ana i te whakawātea a ngā lysosome, ā, i kitea he LC3-II teitei ake me te p62 kua pūmau ake i ngā pūtau i whiwhi puru metilīni. E tautoko ana taua tauira i te hurihanga e whakawhirinaki ana ki ngā lysosome, kaua i te putunga noa iho. I whakaaturia hoki nā te chloroquine i whakahoki whakamuri tētahi wāhanga o te pānga ārai mate pūtau kua whakaritea, e hono ana i te whakamarutanga ki te ara kai-whaiaro me te lysosome i tēnei tauira.

Ko te kitenga mō ngā tohu whakawhiti te wāhanga e tino hē ana te whakahua. I piki ngā tohu whakawhiti AMPK i te 12 me te 24 hāora i runga i te rahinga i hoatu, ko ngā inenga ko te AMPK whai phosphate me tōna ūnga whakararo, te ACC whai phosphate. Nā te pūārai AMPK Compound C, i hoatu i te 20 μM i ngā hāora 4 whakamutunga, i whakaiti te whakapiringa phosphate ki te ACC me te whakahohe LC3. Engari, kāore i kitea he pehanga o te mTOR whai phosphate. Nō reira, i ngā pūtau hippocampus e ahotea ana, ko te kitenga he whakakōkiri e hono ana ki te AMPK mā tētahi ara e āhua motuhake ana i te mTOR. I puta taua tauira anō i ngā kiore ora, i reira nā te maimoatanga puru metilīni i whakakōkiri ngā tohu kai-whaiaro whānui i te uho roro me te hippocampus.

Ehara te whakakōkiri i te rere tukanga kua oti

I konei tino whai take ai te wehewehe i ngā ariā. Ko te whakakōkiri te hanga a ngā pūnaha whakarunga i ētahi atu pūtea kai-whaiaro. Ko te rere tukanga te haere a ngā utanga i te ara katoa: te karapoti, te pakaritanga, te hanumi ki te lysosome, te wāwāhi, te hangarua. Ko ia pikinga LC3-II he kerēme whakakōkiri noa kia whakaū rā anō tētahi whakamātautau rere tukanga i te toenga o te haerenga.

Ka ine te whakamātautau rere tukanga paerewa i te LC3-II me te ārainga poto o ngā lysosome, ā, me te kore hoki o taua ārainga, pērā i te bafilomycin A1, i te chloroquine rānei. Mēnā e whakatere tūturu ana te maimoatanga i te hanga, mā te ārai i te whakangaro ka piki anō te LC3-II. Ka tāpiri tētahi pūrongo tohu takirua mCherry-GFP-LC3 i tētahi atu tirohanga: ka kōwhai te kanapa o ngā hanganga i mua i te hanumi ki te lysosome, ā, ka whero anake i muri i te tineinga o te tohu GFP e te waikawa. Nō reira, ka tautoko te nui ake o ngā ira whero-anake i te anga whakamua ki roto i ngā autolysosome waikawa. Ka taea e te hekenga o te pūmua p62 te whakaū i te hurihanga o ngā utanga, engari me mahara ka urupare hoki te tuhinga ira p62 ki te ahotea, ā, me tirotiro i te taumata mRNA.

E whakaatu ana te rārangi arowhai i raro nei i ia whakamātautau i whakamahia i ēnei tuhinga rangahau, ngā mea e inea ana, me ngā mea e kore e taea te whakaū mā taua whakamātautau anake. Ka taea e ngā kaipānui te whakamahi anō mō ngā kerēme kai-whaiaro whānui ā muri ake nei, tae atu ki ngā tohenga mō te noho kore kai me te roa o te ora e whakamahi ana i ēnei tohu.

Kia mahara ki tētahi mahanga kupu e āraitia ana e te ripanga. Ko te ACC whai phosphate te āhua kua āraia tōna mahi whākōkī. E whakaatu ana tōna pikinga i te putanga o te AMPK, ehara i te whakahohe ACC.

Me pēnei anō te tika o te rerenga “nā te ārainga o te kai-whaiaro whānui i whakakore te whakamarutanga pūtau io”. I te tauira HT22, ko tōna tikanga, nā te whakararu i te ara kai-whaiaro me te lysosome i whakakore te painga oranga a te puru metilīni i raro i te kore serum. E tautoko ana tēnei i te kai-whaiaro hei tukanga e tika ana, e whai wāhi ana rānei, i taua rīhi whakamātautau. Kāore e whakaū ana ko te kai-whaiaro anake te tikanga whakamaru a te pūhui, kāore hoki e whakaū ana mā te kai-whaiaro anake te chloroquine e mahi ai, ā, me whakararu ā-ira i te ATG5, ATG7, ULK1 rānei kia pakari ake ai te kerēme take-pānga.

E rua ngā ara ki te pūnaha kotahi

Ka āwhina tētahi mahere māmā. Ka whakapiki te ahotea pūngao i te AMPK, ka whakahohe te AMPK i te huinga ULK1, ā, ka tīmata te kai-whaiaro. Ka whakapiki ngā āhuatanga whakatipu me ngā taiora i te mTORC1, ka ārai te mTORC1 i te ULK1, ā, ka noho whakaweto te kai-whaiaro. I raro atu i te ULK1, ka tīmata te huinga Beclin-1 me VPS34 i te hanganga o te phagophore, ka whakapiritia he ngako ki te LC3-I kia huri hei LC3-II, ka kati te pūtea kai-whaiaro, ka hanumi ki te lysosome, ā, ka wāwāhia ngā utanga me te hurihanga o te p62.

Ka whakatakotoria e te hoahoa ia kitenga puru metilīni ki tēnei mahere: ko te kitenga hippocampus ki te taha o te peka AMPK, ko te kitenga pūtau tīpuna ki te taha o te peka mTOR.

Mā tēnei anga e whakatau te taupatupatu e āhua kitea ana ki te rangahau tuarua a Xie. I ngā pūtau tīpuna io kiore e whakarea ana, i kitea e tētahi whakamātautau motuhake nā te 5 μM puru metilīni i ārai te whakareatanga, i whakatairanga hoki i te noho ngū. I heke ngā pūtau whai tohu Ki67 i ngā rā 1 me 3, engari i mau tonu te whanaketanga o ngā pūtau kua ū ki te ara pūtau io. I reira, i haere te tikanga mā te pehanga o ngā tohu whakawhiti mTOR: i heke te mTOR katoa me te mTOR whai phosphate, i te taha o te whakaputanga iti ake o te p70S6K me te 4EBP1, me te whakahekenga o ngā cyclin E1, B1, D1, me D2. Ko te āhuatanga matua i kitea ko te noho ngū o ngā pūtau tīpuna, i honoa e ngā kaituhi ki te tōmuri o te kaumātuatanga pūtau, kaua ki te whakaoranga o ngā pūtau hippocampus e ahotea ana.

Kāore he taupatupatu i waenga i ngā pepa e rua. He pūnaha whakakotahi urupare a AMPK me mTOR e tairongo ana ki te horopaki, ehara i te pana pūmau. Ka taea e tētahi pūtau āhua rite ki te pūtau io, e ahotea ana, te whakamahi i te AMPK me te kore ārainga mTOR e taea te ine. Engari ka taea e tētahi pūtau tīpuna e akiakina ana e ngā matū whakawehe pūtau te urupare ki taua pūhui anō mā te whakaiti i te mahi a ngā pūnaha whakatipu e whakawhirinaki ana ki te mTOR. Mā te momo pūtau, te horopaki ahotea, te kukū, me te wā inenga e whakatau ko tēhea peka ka kaha ake. Koia hoki te take me uru ngā pānga pūngao mitokōria me ngā pānga tauhohenga hāora-whakahoki a te puru metilīni ki taua tirohanga: e ai ki tētahi arotake tikanga mahi mō te whakamarutanga pūtau io, ka mahi te pūhui hei kaikawe irahiko kē, ka whakamahi i ngā tohu whakawhiti ārai hāoratanga me te ārai mate pūtau kua whakaritea, ka ārai i te MAO me te NOS, ā, ka hono ki te whakahoutanga mitokōria mā te hanga hou me te kai-whaiaro. Kotahi noa te peka o te huinga pānga maha ko te kai-whaiaro, ehara i te kōrero katoa.

Ngā mea kāore e whakaaturia ana e ngā whakamātautau

Ka whakatauira te kore serum i te ahotea tautoko ora me te ahotea pākia i roto i tētahi rīhi. Kāore e whakatauira ana i te noho kore kai o te rauropi katoa: kei te wē whakatipu taketake tonu te kūhuka, ngā waikawa amino, me ngā tote, ā, kāore he whanonga kai. Kāore e taea te huri ngā kukū whakatipu pūtau mai i te 20 nM ki te 5 μM hei rahinga mā te tangata mā te tātai noa, nā te mea he rerekē te ngongo, te here ki ngā pūmua, te āhua hāora-whakahoki, te pākia, te putunga ki ngā kiko, me te roanga o te whakapā.

Kāore anō kia hoahoatia he whakamātautau tangata hei whakaatu i te rere tukanga kai-whaiaro i whakakōkiritia e te puru metilīni i ngā tūroro. Kei ngā rangahau pūtau, ngā rangahau kiko i waho i te tinana, me ngā rangahau kararehe tonu ngā taunakitanga tikanga mahi, ā, e kī tonu ana ngā arotake he tōmua noa ngā taunakitanga mō te whakawhiti ki te whakamahinga tangata. Nō reira, kāore ēnei rangahau e whakaatu ana ka tārua te puru metilīni i te noho kore kai, ka whakaroa i te oranga tangata mā te kai-whaiaro, ka whakaoti rānei i te rere tukanga kai-whaiaro i te roro tangata. Me whai taunakitanga haumanu motuhake ngā kerēme tārua noho kore kai me te roa o te ora, ā, kāore ngā whakamātautau puru metilīni ki te tangata mō ētahi atu whakamahinga e huri hei whakamātautau kai-whaiaro nā te hononga noa iho. Me pānui ngā rangahau mō ngā tauira ahotea taiora me te roa o te ora me te pupuri tonu i taua rohe.

Mō te pepa e whai ake nei ka pānuihia e koe, whakamahia te rārangi arowhai i runga i tōna raupapa. Tuatahi, pātai ko tēhea peka i panoni: AMPK, mTOR, kāore rānei tētahi. Kātahi ka pātai mēnā i haere tahi te kitenga LC3 me tētahi whakamātautau rere tukanga, i tū takitahi rānei. Kātahi ka tirotiro mēnā nā te ārai i te ara i whakakore te painga, ā, mā tēhea tikanga. Ko tētahi rangahau e whakautu ana i ēnei pātai e toru kua whai pūtake mō tētahi kerēme tikanga mahi pakari. Ko tētahi e whakautu ana i te mea tuatahi anake kua whakaatu noa i tētahi tohu whakaae, ā, kāore anō kia maherehia te toenga o te haerenga.