Theaflavin-regulated Imd Condensates Tswj Drosophila plab hnyuv Homeostasis thiab laus

Thov hu rauoscar.xiao@wecistanche.comyog xav paub ntxiv

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Cov tshuaj yej dub yog cov tshuaj yej haus ntau tshaj plaws hauv ntiaj teb thiab tau tshaj tawm tsis tu ncua kom muaj txiaj ntsig los tiv thaiv kev laus. Txawm li cas los xij, txawm tias theaflavins, ib hom ntawm cov yam ntxwv phytochemicals hauv cov tshuaj yej dub extract, koom nrog kev tswj kev laus thiab kev ua neej nyob hauv cov neeg siv khoom tseem tsis paub ntau. Hauv txoj kev tshawb no, peb pom tau tias theaflavins ua lub luag haujlwm tseem ceeb hauv kev tiv thaiv lub hnub nyoog ntawm plab hnyuv tawm thiab dysbiosis, yog li ncua kev laus hauv Drosophila. Mechanistically, theaflavins tswj cov condensate los ua ke ntawm Imd kom tsis zoo tswj lub overactivation ntawm Imd signals nyob rau hauv cov txiv hmab txiv ntoo yoov. Tsis tas li ntawd, theaflavins tiv thaiv DSS-induced colitis hauv nas, qhia tias theaflavins ua lub luag haujlwm hauv kev hloov kho plab hnyuv. Zuag qhia tag nrho, peb txoj kev tshawb fawb nthuav tawm cov txheej txheem molecular uas theaflavins tswj lub plab homeostasis yuav los ntawm kev tswj hwm Imd coalescence.

Thov nias ntawm no kom paub ntxiv

Taw qhia

Kev laus, tus cwj pwm los ntawm kev poob qis hauv kev ua haujlwm ntawm lub cev hauv tib neeg lub cev thiab kev pheej hmoo ntawm cov kab mob thiab kev tuag, cuam tshuam nrog kev puas tsuaj rau cov molecules, hlwb, thiab cov ntaub so ntswg (Alavez li al., 2011; Bartke li al., 2019; Engeet ib., 2017). Kev txheeb xyuas cov khoom siv ua haujlwm lossis cov tshuaj tiv thaiv kev puas tsuaj lom los ncua kev laus thiab kev ua neej ntev yog qhov tseem ceeb (Bar-ardo li al., 2017; Kapahi li al., 2017). Cov kev tshawb fawb yav dhau los tau tshaj tawm tias cov tshuaj yej dub, cov tshuaj yej siv ntau tshaj plaws hauv ntiaj teb, muaj cov haujlwm tseem ceeb uas ncua kev laus (Cameron li al.,2008; Fei li al.,2017; Kumar thiab Rizvi, 2017; Naumovski li al.,2019 ; Peng et al., 2009; Xiao et al., 2020). Kev noj zaub mov ntxiv nrog cov extracts ntawm cov tshuaj yej dub zoo txuas ntxiv lub neej ntawm cov tsiaj sim xws li cab (Feet al.,2017), txiv hmab txiv ntoo yoov(Peng li al.,2009; Site al.,2011), nas (Site al.,2011). ; Xiao et al., 2020), thiab nas (Kumar thiab Rizvi, 2017).

Nws yog qhov paub zoo tias cov tshuaj yej dub extracts feem ntau muaj ob hom phytochemicals, uas yog, catechins thiab theaflavins (TFs) (Cameron li al.,2008; Kondo li al.,2019; Li et al., 2013), tom kawg uas yog tsim los ntawm catechins los ntawm endogenous polyphenol oxidase thiab peroxidase thaum lub sij hawm oxidation txheej txheem nyob rau hauv dub tshuaj yej ntau lawm (Li li al., 2013). Cov kev tshawb fawb ntau tau txheeb xyuas lub luag haujlwm tiv thaiv kev laus ntawm cov catechins los ntawm kev txhim kho oxidative kev nyuaj siab thiab muaj hnub nyoog txuam nrog o thiab txo cov nqaij mos puas (Cameron li al., 2008; Niu et al., 2013; Peng et al., 2009; Si et al. .2011; Waaner et al., 2015). Txawm li cas los xij, cov kev tshawb fawb ua haujlwm ntawm kev koom tes ntawm TFs rau kev laus tau poob qis heev tom qab cov neeg ntawm catechins, vim tias nws nyuaj rau rho tawm cov nyiaj txaus ntawm TFs los ntawm cov tshuaj yej dub nplooj rau kev tshawb fawb kho mob (Takemoto thiab Takemoto, 2018). Tsis ntev los no, ob peb txoj hauv kev biosynthetic tau tsim los rau kev tsim khoom loj ntawm TFs (Takemoto thiab Takemoto, 2018), ua rau nws muaj peev xwm tshawb nrhiav cov txiaj ntsig lom neeg ntawm TFs thiab cov txheej txheem tswj hwm hauv qab.

Tsis ntev los no, cov ntaub ntawv ntau ntxiv tau hais txog lub luag haujlwm tseem ceeb ntawm txoj hnyuv hauv kev laus thiab kev ua neej nyob (Guo li al., 2014; Jji li al., 2019; Maynard thiab Weinkove, 2018; Rothenberg thiab Zhang, 2019; Salazar li al. , 2018). Lub plab hnyuv epithelium, uas ua raws li kev xaiv thaiv, tso cai rau kev nqus ntawm cov as-ham, ions, thiab dej thiab txwv tsis pub muaj kev sib cuag nrog cov khoom tsim kev puas tsuaj, suav nrog cov kab mob, cov khoom noj khoom haus antigens, thiab cov co toxins (Capo et al., 2019; Nicholson li al. , 2012; Ryu et al., 2010). Raws li cov ntaub so ntswg hloov pauv siab hauv cov tib neeg, lub plab hnyuv muab qhov chaw nyob zoo tshaj plaws rau cov kab mob symbiotic microorganisms, suav nrog cov xwm txheej ntuj anaerobic, cov khoom noj muaj txiaj ntsig, thiab qhov ntsuas kub thiab pH.

Daim duab 1. Theaflavins prolong Drosophila lub neej thiab txhim khu lub peev xwm nce toj

(A thiab B) Cov txiv hmab txiv ntoo yoov tau noj nrog cov khoom noj uas tau muab ntxiv nrog 1 mg / ml lossis 2.5 mg / mL theaflavins (TFs) lossis tsis muaj TFs li basal. Cov yoov tau pauv mus rau cov vials tshiab uas muaj cov xov xwm tshiab txhua hnub, thiab yoov tuag tau qhab nia thoob plaws hauv cov neeg laus lub neej, Survival curves (A) thiab txhais tau tias lub neej ntev (B) ntawm poj niam yoov tau txheeb xyuas thiab qhia.

(Cand D) Poj niam w18 tau noj nrog cov khoom noj Drosophila txheem (xa Basal) lossis nrog cov khoom noj ntxiv nrog 1mg / ml lossis 2.5mg / ETFs, raws li. Rau pawg yoov ywj pheej ntawm cov hnub nyoog uas tau teev tseg (10 hnub, 30 hnub, 40 hnub, thiab 50 hnub, raws li) tau raug rau lub tsev noj mov (C) thiab Fluorescein feeding (D). tus nqi, tsim niches nyob los ntawm cov kab mob tshwj xeeb hauv lub cev (Bonfini li al.,2016; Buchon li al., 2013; Nicholson li al., 2012). Lub caij no, cov kab mob symbiotic thiab lawv cov metabolites kuj ncaj qha lossis tsis ncaj qha cuam tshuam rau kev ua zaub mov; digestion thiab nqus; zog tshuav; tiv thaiv kev ua haujlwm; plab hnyuv loj hlob thiab maturation; thiab ntau lwm yam haujlwm tseem ceeb ntawm lub cev (Bonfini et al., 2016; Broderick, 2016; Maynard thiab VVeinkove, 2018; Nicholson et al., 2012). Kev sib raug zoo ntawm ob lub symbionts tuaj yeem tswj tau qhov kev ruaj ntseg thiab kev sib npaug ntawm txoj hnyuv micro-ecosystem (Bonfini li al.,2016; Brodeerick, 2016; Broderick li al., 2014; Guo et., 2014; May-nard thiab Weinkove, 2018; Nicholson et al., 2012).

cistanche tuaj yeem tiv thaiv kev laus

Ib txoj kev tshawb fawb tau pom tias qhov kev tiv thaiv kab mob hauv lub cev yog qhov tseem ceeb rau kev tswj cov plab hnyuv microbiota homeostasis thiab tau txais kev cawmdim los ntawm cov pob txha mus rau vertebrates (Guillouet al., 2016; Guo et al., 2014; Ryu et al., 2006, Vijay-2010; Kumar et al., 2010). Hauv Drosophila, ob txoj kev taw qhia tseem ceeb, uas yog txoj hauv kev hu xov tooj thiab kev tiv thaiv kab mob tsis muaj zog (Imd), tau koom nrog hauv cov lus teb hauv lub cev (Lu li al., 2020; Myllymakiet al., 2014; Valanne et al., 2011). Qhov tshwj xeeb tshaj yog, Drosophila Tus Xov Tooj txoj hauv kev sib koom nrog cov tsiaj nyeg interleukin -1 receptor (IL-1R) thiab MyD88-dependent Toll-like receptor (TLR) pathways, whereas Imd txoj kev zoo ib yam. mus rau cov qog necrosis factor receptor (TNFR) txoj hauv kev thiab TRIF-dependent TLR txoj hauv cov tsiaj (Imler, 2014; Myllymaki li al., 2014; Valanne et al., 2011). Ob txoj hauv kev no tswj kev qhia ntawm cov tshuaj tiv thaiv kab mob peptides (AMPs) los ntawm kev ua kom muaj NF-KB transcription yam (Imler, 2014; Myllymaki li al., 2014; Valanne et al., 2011).bioflavonoidsLub Imd signaling pathway yog nquag qhib los ntawm Gram-negative kab mob thiab ua rau kev qhia ntawm lwm cov txheej txheem AMPs, xws li Attacin, Cecropin, thiab Diptericin (Kleino thiab Silverman, 2014; Myllymaki li al., 2014). Kev nthuav qhia ntawm cov AMPs no yuav tsum muaj peptidoglycan recognition protein (PGRP)-LC receptor nyob rau hauv Imd txoj kev thiab lub teeb liab-dependent cleavage thiab nuclear translocation ntawm Relish, ib tug NF-kB tsev neeg transcription factor (Kleino thiab Silverman, 2014; Myllymaki li al., 2014). Ib txoj kev tshawb fawb tsis ntev los no tau pom tias kev tsim cov amyloid yuav tsum tau ua kom lub Drosophila Imd txoj hauv kev thaum pom cov kab mob peptidoglycans (Kleino li al., 2017). Txawm li cas los xij, seb qhov Imd condensate no puas koom nrog lwm cov txheej txheem lom neeg, xws li kev tu lub plab hnyuv homeostasis thiab kev tswj kev laus, tseem tsis tau paub ntau.

Nyob rau hauv txoj kev tshawb no, peb siv cov txiv hmab txiv ntoo yoov thiab nas ua tsiaj qauv los tshawb xyuas lub luag haujlwm ntawm TFs hauv kev tswj cov hnyuv thiab kev laus. Peb tau pom tias TFs ncua lub hnub nyoog-pib overproliferation ntawm cov kab mob hauv plab hnyuv, tiv thaiv dysbiosis ntawm plab microbiome, thiab tiv thaiv kev ua haujlwm ntawm Imd signaling pathway, yog li ncua lub neej ntawm Drosophila. Cov kev tshawb fawb txuas ntxiv tau qhia tias TFs tsis zoo ua rau Imd signaling tuaj yeem yog los ntawm kev thaiv cov condensation ntawm Imd es tsis yog nws ubiquitination. Ntxiv mus, peb pom tias TFs muaj txiaj ntsig zoo hauv kev tiv thaiv DSS-induced colitis hauv nas. Ua ke, peb qhov kev tshawb pom tau nthuav tawm lub luag haujlwm tseem ceeb ntawm TFs hauv kev hloov pauv tus cwj pwm Imd, uas tuaj yeem yog qhov tseem ceeb hauv lawv cov txiaj ntsig zoo rau txoj hnyuv homeostasis thiab lub neej ntev.

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Kev noj zaub mov ntxiv ntawm theaflavins txuas ntxiv Drosophila lub neej

Txhawm rau tshuaj xyuas seb theaflavins (TFs) puas cuam tshuam rau kev laus thiab kev ua neej nyob hauv Drosophila, peb tau tsa poj niam w'118 yoov nrog cov khoom noj khoom noj uas tau muab ntxiv nrog 1mg / mL lossis 2.5mg / ml TFs thiab ua qhov ntsuas lub neej raws li tau piav qhia dhau los (Ji li al Ib., 2019). yoov tau noj cov zaub mov ntxiv nrog qhov sib npaug ntawm TFs hnyav (H2O) tau suav tias yog kev tswj hwm. Raws li pom nyob rau hauv daim duab 1A, kev noj haus ntxiv ntawm qhov tsis tshua muaj siab ntawm TFs (1 mg / mL) me ntsis ntev lub txiv hmab txiv ntoo ya lub neej, thiab qhov siab ntawm TFs (2.5 mg / mL) muaj txiaj ntsig zoo rau kev ua neej ntev. Qhov tseem ceeb, kev noj qab haus huv mus sij hawm ntev ntawm qhov siab ntawm TFs (2.5 mg / mL) ua rau kwv yees li 8 hnub ntawm kev nce siab hauv qhov nruab nrab ntawm cov poj niam yoov (Daim duab 1B), qhia txog lub luag haujlwm muaj txiaj ntsig ntawm TFsin hauv kev tswj hwm lub neej ntev hauv Drosophila. Cov txiaj ntsig zoo sib xws tau txais thaum peb kuaj qhov muaj peev xwm ntawm TFs ntxiv ntawm kev ua neej nyob ntawm txiv neej w'*'8 yoov (Figures S1A thiab S1B).

Daim duab 2. Theaflavins tiv thaiv lub hnub nyoog pib lub plab tsis ua haujlwm thiab microbiota dysbiosis hauv Drosophila

(A thiab B) Kev poob ntawm txoj hnyuv tau raug soj ntsuam los ntawm Smurftest thaum muaj hnub nyoog 10 hnub, 30 hnub, 40 hnub, thiab 50 hnub ntawm poj niam w18 cov neeg laus. Piv txwv ntawm cov tsis-Smurf thiab Smurf tau qhia hauv (A). Cov feem pua ​​​​ntawm Smurf hauv cov pab pawg sib txawv tau txheeb xyuas thiab qhia hauv (B).

(C) Bacterialabundances cawm tau los ntawm qPCR ntawm 16S rRNA noob hauv poj niam w18files los ntawm ntau pawg qhia tau txheeb xyuas thiab pom.

(DF) Sequencing ntawm commensal bacterial 16S rRNA noob tau ua. Qhov feem ntawm cov kab mob taxa (D), daim ntawv qhia tshav kub ntawm lub ntsiab microbial taxa (F), thiab cov txheeb ze ntawm Gammaproteobacteria (F) tau txheeb xyuas thiab nthuav tawm.

Daim duab 2. Txuas ntxiv mus

(G thiab H) Lub plab tau muab tshem tawm los ntawm cov poj niam yoov uas tau noj nrog 2.5 mg / ml TFs lossis tsis muaj TFs raws li kev tswj hwm ntawm 10 hnub thiab 40 hnub ntawm hnub nyoog. Cov duab ntawm midgut slides tau ntes los ntawm fluorescence microscopy rau GFP signals (G). Kev txheeb xyuas txheeb cais qhia qhov feem pua ​​​​ntawm GFP-zoo hlwb yog qhia hauv (H).

(I) Lub plab los ntawm cov poj niam yoov tau raug sau thiab raug tshuaj tiv thaiv kab mob siv cov tshuaj tiv thaiv kab mob tiv thaiv phosphorylated Histone 3(pH3) Cov txiaj ntsig tau txheeb xyuas pH3- cov hlwb zoo ib lub plab tau txheeb xyuas thiab pom. Hauv B, C, H, thiab, cov ntaub ntawv raug tshuaj xyuas los ntawm ob tus tub ntxhais kawm qhov kev xeem thiab tau pom tias txhais tau tias ± SEM.NS, tsis tseem ceeb, * p<><0.001. see="" also="" figure="">

ETFs tau paub tias muaj qhov tsw qab, thiab cov khoom noj iab feem ntau zam los ntawm ob tus neeg siv khoom xws li cov txiv hmab txiv ntoo yoov thiab tsiaj txhu (Yamazaki li al., 2014; Yang et al., 2018). Peb mam li nrhiav kom paub meej seb TFs puas tuaj yeem cuam tshuam rau Drosophila noj zaub mov, vim tias cov kev tshawb fawb yav dhau los tau ua pov thawj tias kev txwv kev noj haus ua rau muaj kev ua neej nyob ntev ob leeg hauv vertebrates thiab invertebrates (Hudry li al., 2019; Kapahi li al., 2017; Moger-Reischer et al. ., 2020). Txhawm rau ua qhov no, peb tau ua thawj qhov kev ntsuam xyuas café, ib txoj hauv kev siv dav los ntsuas qhov kev noj zaub mov ncaj qha thiab raug los ntawm kev siv lub capillary feeder uas muaj cov kua nruab nrab, raws li tau piav qhia yav dhau los (Ja et al., 2007). Raws li pom hauv daim duab 1C thiab S1C, kev noj zaub mov ntxiv ntawm TFs (1 mg / mL thiab 2.5 mg / mL, feem) tsis tshua hloov cov zaub mov noj ntawm cov yoov kuaj. Txhawm rau kom paub meej ntxiv qhov no, peb mam li siv cov Fluorescein ua cov khoom noj (Rera et al.,2012; Wang et al.,2005) thiab ua qhov kev ntsuam xyuas kev noj zaub mov zoo (Danilov li al.,2015). Raws li pom nyob rau hauv daim duab 1D thiab S1D, peb ua tsis tau tejyam soj ntsuam kev sib raug zoo ntawm TFs supplementation thiab qib ntawm cov khoom noj. Ua ke, peb cov txiaj ntsig tau qhia tias lub luag haujlwm muaj txiaj ntsig ntawm TFs hauv kev ua neej ntev yuav tsis yog vim kev txwv kev noj haus hauv Drosophila.

Ntxiv TFs yog dispensable rau cuam tshuam Drosophila zog homeostasis

Nws tau raug pom zoo tias cov tshuaj yej dub polyphenols ua rau muaj txiaj ntsig zoo ntawm kev tiv thaiv kev rog los ntawm inhibiting lipid thiab saccharide digestion, nqus, thiab kev noj haus, yog li txo qis calorie kom tsawg (Cameron li al., 2008). Peb mam li nrhiav kom paub meej seb TFs-tswj LDrosophila lub neej ncua ntev yog vim muaj kev hloov pauv hauv kev nqus cov as-ham thiab lub zog cia. Txhawm rau ua qhov no, peb thawj zaug tshuaj xyuas qib ntawm triacylglyceride (TAG), ib qho ntawm feem ntau pom cov lipid metabolites (Tennessen li al., 2014), hauv poj niam yoov thoob plaws lawv lub neej raws li tau piav qhia dhau los (Fan et al., 2017).muab cistancheRaws li pom nyob rau hauv daim duab 1E, kev kho mob ntawm TFs tsis tshua muaj kev cuam tshuam tag nrho TAG abundance ntawm cov hnub nyoog sib txawv. Thaum peb tshawb pom ntxiv glycogen cov ntsiab lus thiab kom muaj nuj nqis ntawm cov carbohydrates (glucose thiab trehalose) los ntawm cov khoom siv Hexokinase (Li li al., 2018; Ten-nessen li al., 2014), cov txiaj ntsig tau pom tias tsis muaj kev hloov pauv ntawm cov qauv nrog lossis tsis muaj TFs kev kho mob (Figures 1F-1H). Nco ntsoov, cov txiaj ntsig zoo ib yam tau txais thaum peb tshuaj xyuas cov tshuaj metabolic phenotypes siv txiv neej w118 ua qauv tsiaj (Figures S1E-S1H). Ua ke, cov ntaub ntawv no tau qhia tias kev noj zaub mov ntxiv ntawm TFs yog siv tau rau kev tswj hwm ntawm carbohydrate thiab lub zog homeostasis thoob plaws lub neej ntawm Drosophila.

Peb tau tshawb xyuas ntxiv txog kinetics ntawm lub ntuj senescence ntawm Drosophila nyob rau hauv kev noj haus TFs cuam tshuam los ntawm kev ua ceev ceev iterative negative geotaxis (RING) kev ntsuam xyuas raws li yav tas los tau piav (Dilliane li al., 2017; Staats li al.,2018). Raws li pom nyob rau hauv daim duab 1l,1J, S1l, thiab S1J, nce indexes ntawm lub hnub nyoog (40 hnub) w1118 (ob leeg poj niam thiab txiv neej) nyob rau hauv TFs supplementation pab pawg neeg tau zoo heev (yuav luag 12 feem pua ​​​​nce hauv cov poj niam thiab ze li ntawm 18 feem pua ​​​​nce hauv cov txiv neej. ) piv nrog cov pab pawg tswj hwm, qhia tias TFs tuaj yeem txhim kho Drosophila nce toj thiab locomotor kev ua ub no.

Theaflavins tiv thaiv lub hnub nyoog pib lub plab to thiab microbiota dysbiosis

Cov kev tshawb fawb yav dhau los tau pom tias cov txiv hmab txiv ntoo ya locomotor kev ua ub no thiab lub neej muaj feem cuam tshuam rau tus tswv tsev noj qab haus huv, xws li poj niam txiv neej, noj zaub mov, hnub nyoog, thiab genotype (Caruso et al., 2013; Heintz and Mair, 2014). Tsis ntev los no, ntau cov ntaub ntawv pov thawj tau ua pov thawj tias khaws cia plab hnyuv Imd signaling homeostasis ameliorates plab kev ncaj ncees los ntawm kev tiv thaiv over-proliferation ntawm ISC thiab precursor hlwb thiab los ntawm kev ncua hnub nyoog-pib plab hnyuv epithelial dysfunction, uas nyob rau hauv lem ua tau zoo pab rau lub cev noj qab haus huv (Clark li al. , 2015; Guo et al., 2014). Hauv Drosophila, lub neej luv luv yog cuam tshuam nrog kev ua haujlwm tsis zoo ntawm txoj hnyuv epithelial barrier, thiab kev txhim kho ntawm kev ncaj ncees ntawm qhov teeb meem no thaum lub sij hawm kev laus ua rau lub neej txuas ntxiv (Clark et al., 2015; Guoet al., 2014; Jet al., 2019). Yog li, peb nrhiav kev txiav txim siab seb TFs hloov qhov kev ncaj ncees ntawm txoj hnyuv los ntawm kev ua Smurf assay, ib txoj hauv kev uas tau piav qhia yav dhau los (Clark li al., 2015; Rera et al., 2011). Raws li pom nyob rau hauv daim duab 2A, cov txiv hmab txiv ntoo sim yoov nyob rau hauv uas xiav dyes raug txwv rau lub plab zom mov raug xa mus rau raws li tsis-Smurf (sab laug vaj huam sib luag nyob rau hauv daim duab 2A). Yog tias cov xim xiav tau pom

Daim duab 3. Txuas ntxiv mus

(EG) Lub plab raug cais tawm ntawm poj niam w1118 cov ntaub ntawv pub nrog 2.5 mg / ml TFs lossis tsis muaj TFs raws li basal ntawm 10 hnub, 30 hnub, 40 hnub, thiab 50 hnub ntawm hnub nyoog, feem. Tag nrho RNA raug muab rho tawm thiab raug rau kev tshuaj ntsuam ART-PCR. MRNA qhia theem ntawm diptericin (F), attacin (F) thiab cecropin A1 (G) tau txheeb xyuas thiab pom. Hauv EG, cov ntaub ntawv raug tshuaj xyuas los ntawm ob tus tub-ntxhais kawm ntawv t-test thiab qhia tias txhais tau tias ± SEM.NS, tsis tseem ceeb, *p<><><0.001. see="" also="" figure="">

tawm ntawm lub plab mus rau hauv plab, cov tswv tau suav tias yog Smurf (txoj cai vaj huam sib luag hauv daim duab 2A). Raws li qhov kev tshawb pom yav dhau los (Clark et al.,2015), qhov feem ntawm Smurf maj mam nce thaum laus (Daim duab 2B). Interestingly, cov kev faib ua feem ntawm Smurf cov neeg laus tau poob qis heev nyob rau hauv TFs-fed pab pawg piv nrog rau cov hnub nyoog-match tswj ntawm ob tug poj niam (los ntawm 8.2 feem pua ​​​​mus rau 3.7 feem pua, los ntawm 19.4 feem pua ​​​​rau 15.6 feem pua ​​, thiab los ntawm 35.4 feem pua ​​​​rau 27.1 feem pua ​​​​rau cov pab pawg. Thaum muaj hnub nyoog 30 hnub, 40 hnub, thiab 50 hnub, raws li, Figure2B) thiab cov txiv neej laus (los ntawm 2.7 feem pua ​​​​rau 0.9 feem pua ​​​​, los ntawm 6.9 feem pua ​​​​rau 0.9 feem pua ​​, thiab los ntawm 8.3 feem pua ​​​​rau 2.7 feem pua ​​​​ntawm hnub nyoog 30 xyoo. hnub, 40 hnub, thiab 50 hnub, raws li, daim duab S2A), qhia tias TFs tuaj yeem tiv thaiv plab hnyuv tsis ua haujlwm ntawm cov txiv hmab txiv ntoo muaj hnub nyoog. Nco ntsoov, peb tau pom qhov txo qis ntawm Smurf feem pua ​​​​ntawm cov txiv neej (txo yuav luag 65.8 feem pua ​​, 86.9 feem pua ​​, thiab 67.4 feem pua ​​​​ntawm cov hnub nyoog 30 hnub, 40 hnub, thiab 50 hnub, feem) dua li cov hnub nyoog. -matched poj niam (txo los ntawm yuav luag 54.1 feem pua, 19.5 feem pua, thiab 23.3 feem pua ​​​​ntawm cov hnub nyoog 30 hnub, 40 hnub, thiab 50 hnub, raws li), qhia tau hais tias TFs yuav ua tau lub luag hauj lwm zoo hauv kev txo cov plab hnyuv to hauv txiv neej yoov.

Cov kev tshawb fawb yav dhau los tau qhia tias muaj hnub nyoog pib lub plab hnyuv tsis ua haujlwm feem ntau cuam tshuam nrog dysbiosis ntawm plab microbiota (Clark et al., 2015; Guo et al., 2014). Yog li, peb tau tshuaj xyuas cov kab mob hauv plab hnyuv commensal kab mob tom qab TFs supplementation. Peb thawj zaug siv universal primers thiab ua qhov kev sim PCR kom muaj nuj nqis los ntsuas cov kab mob 16S rRNA gene theem. Raws li pom nyob rau hauv daim duab 2C, nyob rau hauv cov laus txiav, tag nrho cov kab mob cov neeg los ntawm TFs kev cuam tshuam pab pawg pom ib tug striking txo (yuav luag 45 feem pua ​​-55 feem pua ​​) piv nrog rau cov tswj, qhia tias TFs supplementation tej zaum pre-vents microbiota expansion thaum laus. . Tom qab ntawd, peb tau ua cov metagenomics los tshuaj xyuas kom meej cov kev hloov pauv hauv cov hnyuv microbial zej zog tshwm sim los ntawm kev noj zaub mov ntxiv nrog TFs. Raws li pom nyob rau hauv daim duab 2D-2F, qhov feem ntawm Gammaproteobacteria, qhov nthuav dav uas tau pom tias muaj kev cuam tshuam nrog txoj hnyuv tsis ua haujlwm (Clark li al.,2015), tau pom tseeb txo qis los ntawm TFs supplementation hauv cov laus cov hnyuv ( yuav luag 7 feem pua ​​​​txo), implying tias TFs zoo pab rau lub homeostasis ntawm plab microbiota thaum laus hauv Drosophila.

Theaflavins txhim kho plab hnyuv proliferative homeostasis hauv Drosophila

We further examined the intestinal integrity of flies that were treated with or without TFs. First, we employed the widely-used P{Esg-Gal4}/P{Uasp-GFP};{Tub-Gal80*s}strain(referred to as esgts>GFP), uas cov kab mob hauv plab hnyuv (ISCs) thiab cov hlwb progenitor tau sau nrog GFP. Raws li pom nyob rau hauv Figures2G thiab 2H, tus naj npawb ntawm GFP-zoo hlwb hauv TFs cuam tshuam pab pawg thaum muaj hnub nyoog 40 hnub (20 lub plab raug suav) tsawg dua (ze li 29.7 feem pua) ntau dua li ntawm cov hnub nyoog sib xws (yuav luag 47.6 feem pua ​​thiab 19 lub siab tau suav), qhia tias TFs tuaj yeem cuam tshuam lub hnub nyoog pib tshaj tawm ntawm ISCs. Tsis tas li ntawd, peb tau ua qhov kev sim tshuaj tiv thaiv kab mob siv cov tshuaj tiv thaiv kab mob Phospho-Histone 3 (pH3), ib qho cim tshwj xeeb rau cov hlwb mitotic hauv cov ntaub so ntswg. Peb tau pom tsawg dua pH3- cov hlwb zoo hauv lub plab los ntawm cov neeg laus uas muaj hnub nyoog TFs ntxiv (tus lej nruab nrab yog 20.6) dua li ntawm cov kev tswj hwm (tus lej nruab nrab yog 39.5, Daim duab 2). Ua ke, peb cov txiaj ntsig tau qhia tias kev noj zaub mov ntxiv nrog TFs ua kom lub plab hnyuv loj hlob hauv homeostasis thiab tiv thaiv hnub nyoog pib microbiota dysplasia, yog li ncua kev laus hauv Drosophila.

Theaflavins tsis zoo hloov cov plab hnyuv Imd cov cim

Txhawm rau tshawb nrhiav cov txheej txheem hauv qab molecular uas TFs tau txais txiaj ntsig zoo rau Drosophila lub plab homeostasis, peb tau ua RNA-seq tsom xam siv cov hnub nyoog sib txuam plab hnyuv los ntawm poj niam w1118 cov neeg laus kho nrog lossis tsis muaj TFs. Fragments ib kilobase ib lab (FPKM) kev soj ntsuam qhia tias tag nrho cov qauv ntawm cov noob qhia tau zoo sib xws ntawm ob pawg kev sim (Daim duab 3A). Txawm li cas los xij, kev txheeb xyuas qhov sib txawv ntxiv, txawm li cas los xij, tau txheeb xyuas tag nrho ntawm 229 qhov sib txawv ntawm cov noob, ntawm uas 99 tau hloov kho thiab 126 tau raug txo qis (Daim duab 3B). Interestingly, gene ontology tsom xam ntawm cov noob no tau qhia tias txoj kev tiv thaiv kab mob tau hloov pauv los ntawm kev noj zaub mov ntxiv nrog TFs (Daim duab 3C).

Hauv Drosophila, txoj hauv kev tiv thaiv kab mob hauv lub cev tau raug pov thawj los ua lub luag haujlwm tseem ceeb hauv kev tswj lub hnub nyoog pib lub plab homeostasis thiab lub neej ntev (Clark et al, 2015; Guo et al, 2014). Tom qab ntawd peb tsom mus rau kev tshuaj xyuas cov qauv qhia ntawm cov tshuaj tiv thaiv kab mob peptides (AMPs) hauv qab ntawm cov kab mob hauv lub cev. Raws li qhia

Daim duab 4. Txuas ntxiv mus

(D) Purified Imd proteins tau incubated nrog ntau yam concentrations ntawm TFs (50 uM, 100 uM, thiab 200 uM, ntsig txog) ntawm chav tsev kub rau 5 min. Cov qauv tom qab ntawd raug rau fluorescence emission spectrum tsom xam siv lub wavelengths qhia (excitation ntawm 430 nm, emission ntawm 450 nm mus rau 550 nm nrog ib tug slit dav ntawm 5 nm). Ib qho piv txwv uas tsis muaj TFs tau siv los ua tus tswj lub hauv paus.

（E and F）Lysates of guts from NP11>Myc-Imd yoov tau npaj thiab incubated nrog TFs (100μM) ntawm chav tsev kub rau 30 min. SDD-AGE (sab saum toj vaj huam sib luag hauv E) thiab SDS-PAGE (qis vaj huam sib luag hauv E) tau ua los soj ntsuam Imd protein aggregates. Tubulin tau siv los ua kev tswj kev thauj khoom.cistanchKev tsom xam Densitometry tau ua kom muaj nuj nqis Imd condensates hauv cov qauv sib txawv, thiab cov txiaj ntsig tau pom hauv (F). (G) Tus neeg sawv cev chromatogram ntawm TFs qauv.

(H thiab kuv) Purified Flag-Imd proteins tau incubated nrog TFs, TF1, TF2a, TF2b, los yog TF3 (100 μM rau txhua tus qauv) los yog nrog ib tug sib npaug ntim ntawm tsis (tswj) ntawm chav tsev kub rau 30 min raws li qhia. Cov qauv tom qab ntawd raug rau SDD-AGE (sab saum toj vaj huam sib luag hauv H) thiab SDS-PAGE (qis vaj huam sib luag hauv H) kev ntsuam xyuas los txiav txim cov qauv sib sau ua ke ntawm Imd. Densitometry tsom xam kom muaj nuj nqis Imd condensates hauv cov qauv sib txawv tau ua thiab qhia hauv (). Hauv C, F, thiab kuv, cov ntaub ntawv raug tshuaj xyuas los ntawm ob tus tub ntxhais kawm qhov kev xeem thiab qhia tias txhais tau tias ± SEM.***p< 0.001.="" see="" also="" figure="">

Hauv daim duab 3D, AMP noob tswj hwm los ntawm txoj kev tiv thaiv kab mob tsis muaj zog (Imd), suav nrog diptericin A (DptA), dip-tericin B(DptB), attacin A (AttA), cecropin A1 (CecA1), cecropin A2 (CecA2), thiab cecropin C (Cece), tau txo qis (los ntawm 39.4 feem pua, 72.3 feem pua, 10.5 feem pua, 86.5 feem pua, 99.8 feem pua, thiab 36.1 feem pua, raws li) nyob rau hauv TFs-kho pab pawg piv nrog cov tswj pawg. Txhawm rau kom paub meej ntxiv cov txiaj ntsig tau los ntawm RNA-seq tsom xam, peb tau ua ntau qhov rov qab hloov pauv-polymerase cov tshuaj tiv thaiv kab mob (gRT-PCR) txhawm rau txhawm rau txheeb xyuas cov theem sib txheeb ntawm Imd-txog AMP cov noob hauv Drosophila lub plab kuaj tau ntawm plaub lub hnub nyoog sib txawv. suav nrog 10 hnub, 30 hnub, 40 hnub, thiab 50 hnub. Raws li pom hauv daim duab 3E-3G, mRNA qhia theem ntawm qee cov noob, suav nrog diptericin, attacin, thiab cecropin A1, tau txo qis (los ntawm yuav luag 35.9 feem pua ​​-66.6 feem pua) hauv TFs-kho cov laus. Sib sau ua ke, peb cov ntaub ntawv tau tshaj tawm tias TFs muaj peev xwm tiv thaiv kev ua haujlwm ntawm Imd signaling pathway hauv Drosophila lub plab hlwb thaum laus.

Theaflavins yog dispensable rau transcriptional kev cai ntawm txoj kev Imd los yog ubiquitination ntawm Imd

Peb tau nrhiav kev tshuaj xyuas kev koom tes ntawm TFs thiab Imd thiab ua qhov kev ntsuam xyuas nto plasmon resonance (SPR) raws li tau piav qhia yav dhau los (Lan li al., 2020). Raws li pom hauv daim duab 4A. TFs tau nthuav tawm cov kev sib txuas nruab nrab nrog cov purified Imd protein ntawm qhov kwv yees Kp tas li ntawm 8.078 uM. Tom qab ntawd peb yuav txiav txim siab li cas TFs tswj txoj hnyuv Imd signaling txoj kev. Raws li cov txiaj ntsig ntawm kev txheeb xyuas cov ntaub ntawv los ntawm RNA-seq cov ntaub ntawv, peb pom tias cov lus qhia theem ntawm cov kev cai tseem ceeb hauv Imd signaling pathway, xws li PGRP-LC, Imd, thiab Relish, tsis tshua muaj kev hloov pauv los ntawm TFs kev kho mob (Daim duab S3A. ). Peb tau sau cov kab mob plab ntxiv los ntawm yoov uas tau txais kev noj zaub mov ntxiv nrog lossis tsis muaj TFs ntawm ntau lub hnub nyoog thiab ua cov kev sim qRT-PCR tshwj xeeb txhawm rau txheeb xyuas cov qauv qhia ntawm cov noob no. Raws li pom hauv daim duab S3B, mRNA qib ntawm qee cov noob, suav nrog effete, faded, tak1, pgrp-Ica, uev1a, bendless, relish, imd, pgrp-sc2, tab2, ird5 Kenny, thiab diap2, tau zoo sib xws ntawm pawg tswj hwm. thiab TFs kev cuam tshuam pab pawg, tawm tswv yim tias TFs tsis koom nrog kev tswj hwm Imd cov cim ntawm qib transcriptional.

Tom ntej no, peb tau tshuaj xyuas Imd ubiquitination profiles vim cov kev tshawb fawb yav dhau los tau qhia tias ubigui-tination ntawm Imd yog qhov tseem ceeb rau kev xa cov teeb liab qis hauv txoj kev Imd (Myllymaki li al., 2014; Zhou et al., 2005). Peb thawj zaug tau ua ib qho kev ntsuam xyuas hauv vitro E1 thiab pom tias kev kho mob nrog TFs tsis muaj kev cuam tshuam rau cov tshuaj tiv thaiv enzymatic ntawm E1 thiab ubiquitin los ua conjugated E1 / Ub (Daim duab S4A). Ntxiv mus, peb kis tau Drosophila S2 hlwb nrog plasmids qhia Flag-tagged Imd ua ke nrog HA-tagged Ub thiab tom qab ntawd kho cov hlwb nrog lossis tsis muaj TFs. Ntxiv ubiquitination assays qhia tias TFs supplementation NW nyuam qhuav cuam tshuam cov theem ntawm ubiquitinated Imd nyob rau hauv kab lis kev cai hlwb (Figures S4B thiab S4C). Txhawm rau txiav txim siab txog kev sib raug zoo ntawm TFs thiab Imd ubiquitination hauv vivo, peb tau siv lub transgenic ya P {NP1-Gal4}/P{Uasp-Myc-Imd}; P{Tub-Gal80*} (hu ua NP1* s > Myc-Imd), nyob rau hauv uas Myc-tagged Imd protein tau nthuav tawm hauv plab hnyuv (Daim duab S4D). Peb qhov kev sim ubiquitination tau pom tias kev noj zaub mov ntxiv nrog TFs yog qhov siv tau rau kev hloov pauv ntawm ubiquitination ntawm Imd hauv plab cov ntaub so ntswg (Figures S4D thiab S4E). Ua ke, peb cov txiaj ntsig tau qhia tias TFs tsis zoo tswj hwm Drosophila Imd cov cim qhia los ntawm kev tsis cuam tshuam rau tus qauv ubiquitination ntawm Imd.

Theaflavins tiv thaiv Imd condensate los ua ke

Ib txoj kev tshawb fawb tsis ntev los no tau qhia tias kev sib sau ntawm Imd yog qhov tseem ceeb rau kev ua kom cov noob caj noob ces hauv txoj kev Imd (Kleino li al., 2017). Peb mam li nrhiav kev tshuaj xyuas seb TFs tswj cov txheej txheem ntawm

Daim duab 5. Txuas ntxiv mus

(F thiab G) Poj Niam w18 (hu ua imd ntxiv / ntxiv), imd' heterozygous thiab homozygous mutants (xa mus rau imd ntxiv /- thiab imd-/-, raws li) tau khaws cov khoom noj Drosophila (xa mus rau Basal) los yog kev noj zaub mov ntxiv nrog 2.5 mg / mL (hu ua TFs) thiab tom qab ntawd raug soj ntsuam txog kev ua neej nyob txoj sia nyob nkhaus (F) thiab txhais tau tias ntev (G) raug tshuaj xyuas thiab qhia.

Hauv thiab F, qhov ntsuas ntsuas ntsuas tau siv rau kev ntsuas ntsuas. Nyob rau hauv BE, thiab, ob-tailed Student's t-test tau siv rau kev txheeb xyuas txheeb cais, thiab cov ntaub ntawv yog sh raws li txhais tau tias ± SEM. NS, tsis tseem ceeb, *p<0.05, **p="" <="" 0.01,=""><0.001. see="" also="" figure="">

Imd aggregate tsim. Ua ntej, peb tau ntxuav Myc-tagged Imd proteins los ntawm kab lis kev cai S2 hlwb thiab incubated cov proteins uas sib txawv ntawm TFs (Daim duab 4B). Interestingly, peb pom tias TFs cim tau tiv thaiv kev sib sau ua ke ntawm Imd (txo 81.7 feem pua ​​-89.7 feem pua ​​, Daim duab 4B thiab 4C), raws li tau hais los ntawm peb cov kev ntsuam xyuas semidenaturing agarose gel electrophoresis (SDD-AGE) . Txhawm rau kom paub meej qhov kev tshawb pom no, peb tau ua ntxiv thioflavin-T binding assays thiab tau txais txiaj ntsig zoo ib yam (Daim duab 4D). Thaum kawg, peb tau tsa NP1ts > Myc-Imd transgenic yoov uas tau txais kev noj zaub mov ntxiv nrog lossis tsis muaj TFs thiab sau cov qauv plab los ntawm cov neeg laus. Qhov kev sim SDD-AGE tau qhia tias kev sib sau ntawm Imd tau txo qis heev los ntawm TFs hauv cov hnyuv (tso los ntawm 16.7 feem pua ​​-22 feem pua ​​, Daim duab 4E, thiab 4F).

Txhawm rau txiav txim siab li cas TFs pab txhawb rau kev tswj hwm Imd condensate sib dhos, peb raug TFs rau HPLCanal-ysis thiab pom tias lawv muaj plaub hom monomers (Daim duab 4G), uas yog, theaflavin (TF1), theaflavin 3-O-gallate ( TF2a), theaflavin 3'-O-gallate (TF2b), thiab theaflavin 3,3'-di-O-gallate (TF3), uas tau ua raws li cov ntaub ntawv dhau los (Li et al., 2013; Takemoto thiab Takemoto, 2018) . Tom qab ntawd, peb incubated txhua yam ntawm cov tshuaj no nrog purified Imdprotein thiab ua SDD-AGE thwmsim. Raws li pom nyob rau hauv daim duab 4H thiab 4l, kev kho mob ntawm tag nrho cov TFmonomers no ua rau muaj kev txo qis (los ntawm 59.5 feem pua, 60.9 feem pua, 58.4 feem pua ​​.58.2 feem pua. thiab 55.8 feem pua ​​rau TFs, TF1, TF2a, TF2b, thiab TF3, feem) cov theem imd aggregate. Sib sau ua ke, peb cov txiaj ntsig tau qhia tias TFs tsis zoo tswj hwm Drosophila Imd txoj kev taw qhia tej zaum los ntawm kev tswj hwm tus cwj pwm ntawm Imd condensates.

Theaflavins txuas Drosophila lifespan nyob rau hauv ib qho Imd-dependent yam

To examine whether TFs prolong Drosophila lifespan depending on their regulatory role in Imd, we specifically knocked down Imd in gut tissues using the transgenic flies P{NP1-gal4)PITub-gal80*}/P{Uasp-imd-IR(KK)} (referred to as Imd RNAi). As shown in Figures 5A and 5B, dietary supplementation of TFs extended the lifespan of controls (elevation of nearly 7.2 days in the mean lifespan), whereas prevention of Imd expression in guts apparently reversed the lifespan extension caused by TFs. Consistent results were obtained when we utilized male Imd RNAi and control flies to perform lifespan assays (Figures S5A and S5B). Further qRT-PCR assays showed that the mRNA expression levels of AMP genes at ages of 10 days, 30 days, 40 days, and 50 days, including diptericin, attacin, and cecropin A1.hardly changed in the gut tissues of Imd RNAi flies(Figures 5C-5E), whereas additional TFs caused reductions of 31%-72.3% in mRNA abundances of these genes in samples from control groups. To confirm these results from Imd RNAi flies, we first performed lifespan assays using w18(control, referred to as imd+/+), imd' heterozygous (referred to as imd+/-), and homozygous(referred as imd-/-)mutant flies. We found that TFs addition extended lifespan and increased mean life longevity of imd+/-flies (elevation of nearly 6.2 days, Figures 5F and 5G). However, this lifespan extension effect by TFs was almost abolished in imd-/-flies (Figures 5F and 5G). Next, we performed lifespan assays utilizing Relish RNAi(NP1's>Relish RNAi and control(NP1ts>plus) yoov. vim tias Relish yog qhov tseem ceeb ntawm kev hloov pauv ntawm lub luag haujlwm rau Imd-downstream AMP gene expressions(Myllymaki li al.,2014).Raws li qhia hauv Fiqures S5C thiab S5D. Kev noj zaub mov ntxiv ntawm TFs tau ua rau lub sijhawm ntev ntawm kev tswj hwm yoov, thiab qhov kev ncua ntev ntawm kev ua neej no tau tiv thaiv tau zoo hauv Relish RNAi yoov. Sib sau ua ke, peb cov ntaub ntawv tau qhia tias kev noj zaub mov ntxiv ntawm TFs ua rau muaj txiaj ntsig zoo rau Drosophila lifespan yuav nyob ntawm plab hnyuv Imd cov cim.

Txhawm rau pom seb qhov txuas ntxiv ntawm kev ua neej nyob los ntawm TFstill tshwm sim hauv keeb kwm ntawm axenic, peb tau ua qhov kev ntsuam xyuas lifespan siv w1118 cov poj niam nyob rau hauv axenic mob. Raws li pom hauv daim duab S5E thiab S5F, qhov sib ntxiv ntawm qhov sib txawv ntawm TFs (1 mg / mL thiab 2.5 mg / mL) tsis tshua muaj kev cuam tshuam rau kev sib piv ntawm kev tuag thiab txhais tau tias lub neej ntev ntawm cov neeg laus w18. Vim tias cov kab mob axenic tuaj yeem tiv thaiv kom tsis txhob muaj microbiota ntau, yog li txwv tsis pub Imd cov cim hauv kev txiav (Clarke al., 2015), peb cov ntaub ntawv qhia tias TFs ntev Drosophila lifespan yuav yog los ntawm kev hloov kho ntawm plab microbiota-tswj lub cev tiv thaiv kab mob.

Daim duab 6. Theaflavins alleviate DSS-induced colitis hauv CD-1 nas

(A) Kev kho TFs tsis cuam tshuam rau kev siv dej hauv ntau pawg CD-1 nas.

(BD) Gavage kev tswj hwm ntawm ntau qhov ntau ntawm TFs (1 mg / mL, 2.5 mg / mL, thiab 5mg / mL, raws li) ua kom txo qis ntawm txoj hnyuv thiab qhov loj ntawm tus po tshwm sim los ntawm DSS hauv CD-1 nas. Cov duab sawv cev (B) thiab statistical analyses ntawm txoj hnyuv (C) thiab spleen (D) tau pom. (E) Kev tswj hwm ntawm ntau qhov ntau ntawm TFs (1 mg / mL, 2.5 mg / ml, thiab 5mg / mL, raws li) tiv thaiv plab hnyuv tawm los ntawm DSS hauv CD-1 nas. (F thiab G) Colons tau sau los ntawm cov pab pawg nas. Tag Nrho RNA raug rho tawm thiab raug rau qRT-PCR kev soj ntsuam txhawm rau txheeb xyuas mRNA qhia theem ntawm TNFa(F) thiab IL-6 (G).

(HL) Colons los ntawm ntau pawg nas tau sau longitudinally thiab raug rau HE staining assay. Cov qauv raug kuaj xyuas los ntawm microscopy, thiab cov duab sawv cev ntawm cov qauv qhia (HL) tau txais thiab pom. Scale bar, 40 μm. (M-Q') SEM cov duab qhia cov microtrichomes ntawm cov nyuv los ntawm cov qauv qhia sib txawv. Scale bars, 50um (MQ) thiab 4um (M'-Q').

Hauv CG, cov ntaub ntawv raug tshuaj xyuas los ntawm ob tus tub ntxhais kawm qhov kev xeem thiab qhia tias txhais tau tias ± SEM.NS, tsis tseem ceeb, *p<><><0.001. see="" also="" figure="">

Theaflavins alleviate DSS-induced colitis hauv CD-1 nas

Txhawm rau tshawb xyuas ntxiv txog lub luag haujlwm ntawm TFs hauv kev tswj cov hnyuv homeostasis hauv cov tsiaj, CD-1 nas tau siv los ua kom mob plab los ntawm dextran sulfate sodium (DSS). Cov nas tsuag tau siv tshuaj intragastrically sib txawv ntawm TFs (1 g / L, 2.5 g / L, lossis 5 a / L) lossis dej raws li kev tswj hwm plaub lub lis piam, tom qab ntawd kho nrog 2 feem pua ​​​​DSS hauv dej haus rau 7 hnub. Txhawm rau saib seb TFs puas cuam tshuam rau qhov haus ntawm DSSacross ntau pawg nas, peb tau saib xyuas cov nas haus txhua hnub. Raws li pom nyob rau hauv Fiqure6A, TFs kev kho mob yog dispensable rau cuam tshuam kev haus dej haus nyob rau hauv DSS-induced colitis nas, DSS-kho nas qhia colitis syndrome nrog raws plab thiab / los yog hematochezia, uas tau tiv thaiv los ntawm koob tshuaj-dependent TFs kev kho mob (Figures S6A-S6E).cistanche AustraliaTsis tas li ntawd, kev cuam tshuam nrog TFs cawm tau qhov txo qis hauv txoj hnyuv loj los ntawm DSS thiab ua kom lub qhov loj ntawm tus po (Figures 6B-6D).

Tom qab ntawd peb siv dextran-4000-FITC (FITC-DX) los tshuaj xyuas lub plab hnyuv ua haujlwm ntawm cov nas no. Raws li pom nyob rau hauv daim duab 6E, nas los ntawm DSS-kho cov pab pawg pom muaj siab concentration ntawm FITC-DX nyob rau hauv cov ntshav kuaj, qhia lub plab to hauv cov tsiaj, Txawm li cas los, kev cuam tshuam nrog TFs ho tiv thaiv kev thauj ntawm FITC-DX los ntawm digestive kabmob mus rau hauv lub plab. circulatory system (txo 56.1 feem pua ​​-77.5 feem pua ​​, Daim duab 6E). Qhov tseem ceeb, kev tswj hwm ntawm TFs tsis hloov lub plab permeability ntawm nas nyob rau hauv ib txwm muaj mob (Daim duab S6F). Vim tias qhov mob ua rau lub luag haujlwm tseem ceeb hauv kev txhim kho colitis ntawm tus qauv nas, peb tau ua ntxiv qRT-PCR kev sim, qhia tias TFs kev kho mob txo qis IL-6 thiab TNF cov lus qhia hauv cov hnyuv ntawm DSS-kho nas (Figures 6F thiab 6G).

Txhawm rau txheeb xyuas ntxiv cov kev hloov pauv hauv txoj hnyuv, peb tau ua cov tshuaj hematoxylin-eosin (HE) staining. Raws li pom nyob rau hauv Figures6H-6L thiab S6G-S6J, DSS-induced hnyav epithelial puas tau zoo heev ameliorated los ntawm TFs kev kho mob raws li koob tshuaj. Kev kuaj xyuas electron microscopy imaging ntawm cov kab mob plab hnyuv ntxiv tau muab cov txiaj ntsig zoo ib yam (Figures 6M-6Q' thiab S6K-S6N'). Ua ke, peb qhov kev tshawb pom tau qhia tias TFs ua lub luag haujlwm muaj txiaj ntsig zoo hauv kev tiv thaiv plab hnyuv ua haujlwm hauv ob qho tib si kab thiab tsiaj txhu.

Kev sib tham

Nyob rau hauv txoj kev tshawb no, peb tau tshawb xyuas lub physiological functions ntawm TFs nyob rau hauv modulating plab hnyuv homeostasis thiab lub neej ntev thiab lub hauv paus molecular mechanisms. Peb tau pom tias kev noj zaub mov ntxiv nrog TFs muaj txiaj ntsig zoo rau kev tiv thaiv lub hnub nyoog pib dysbiosis ntawm plab hnyuv microbiota thiab qeeb ntawm plab epithelial tsis ua haujlwm, yog li ncua lub neej ntawm Drosophila. Peb cov kev tshawb fawb hauv vitro thiab hauv vivo tau tshaj tawm tias TFs yuav ua lub luag haujlwm tsis zoo hauv kev tswj hwm txoj kev taw qhia Imd los ntawm kev thaiv cov coalescence ntawm Imd. Ntxiv mus, peb pom tias DSS-induced colitis tuaj yeem txo tau zoo los ntawm TFs supplementation hauv CD-1 nas. Peb cov txiaj ntsig txhawb kev xav tias TFs tau txais txiaj ntsig zoo rau txoj hnyuv homeostasis nyob rau hauv ob qho tib si invertebrates thiab vertebrates.

Kev noj zaub mov ntxiv nrog theaflavins prolongs Drosophila lub neej

Kev haus tshuaj yej yog nrov thoob ntiaj teb thiab yog thib ob tsuas yog siv dej (Rothenberg thiab Zhang, 2019). Ntau cov kev tshawb fawb tau tsom mus rau kev ua haujlwm ntawm lub cev feem ntau los ntawm kev sim thiab kev soj ntsuam (Cameron li al.,2008; Niu li al.,2013; Peng li al.,2009; Rothenberg thiab Zhang.2019; Spindleret al.,2013; Takemoto thiab Takemoto, 2018; Unno et al., 2020; Zhou et al., 2020.Nws tau nthuav dav qhia tias tshuaj yej pab kev noj qab haus huv los ntawm kev txo qis lipid thiab los ntawm kev tiv thaiv kev rog, thiab cov ntsiab lus tseem ceeb hauv tshuaj yej yog catechins, tshwj xeeb tshaj yog epigallocatechin gallate (EGCG)(Abbas and Wink,2009; Modernelli et al.,2015; Niu et al.,2013; Wagner et al.,2015; Xiong et al.,2018). tshuaj yej dub tuaj yeem ua rau muaj kev noj qab haus huv thiab kev ua neej nyob ntawm cov txiv hmab txiv ntoo yoov (Site al., 2011; Wagner li al., 2015), worms (Fei li al., 2017; Xiong li al., 2018), thiab nas (Imran li al. ., 2018; Niu etal.2013). thiab lub ntsiab kev cai mechanism yog kev tiv thaiv zoo ntawm oxidative kev nyuaj siab los ntawm EGCG ntawm kev hloov kho ntawm ROSsignals.cov txiaj ntsig cistanche,Txawm li cas los xij, txawm tias TFs, raws li tus yam ntxwv "golden" cov tebchaw hauv cov tshuaj yej dub, ua lub luag haujlwm hauv kev tswj kev laus thiab kev ua neej nyob tsis paub ntau. Txoj kev tshawb no muab pov thawj qhia tau hais tias TFs kev cuam tshuam yog siv tau rau kev cuam tshuam rau kev noj zaub mov, thiab kev noj qab haus huv mus sij hawm ntev ntawm TFs los ntawm kev noj zaub mov ua rau muaj kev cuam tshuam rau cov txiv neej thiab poj niam yoov. Cov kev tshawb pom no muab cov lus qhia rau kev tshawb fawb ntxiv txog kev ua haujlwm ntawm TFs hauv cov tsiaj txhu ntev.

Theaflavins txhim kho lub hnub nyoog pib microbiota dysbiosis thiab txhawb txoj hnyuv homeostasis Progressive imbalance ntawm proliferative homeostasis thiab regenerative muaj peev xwm yog ib qho cim ntawm kev laus thiab muaj hnub nyoog pib kab mob (Clark et al., 2015; Heintz thiab Mair, 2014; Maynard, 2010 and Weinkov. et al. 2012). Kev mob ntev yog cuam tshuam nrog kev poob ntawm homeostasis thiab nce kev mob qog noj ntshav hauv cov kab mob laus (Bartke li al.,2019; Kapahi li al., 2017; Xiao li al.,2020). Qhov no yog qhov tshwj xeeb tshaj yog nyob rau hauv cov kab mob epithelium xws li plab hnyuv epithelium (Guo li al., 2014; Maynard thiab Weinkove, 2018). Nrog rau tus qauv rau kev txhim kho cov hnub nyoog ntsig txog dysplasia hauv txoj hnyuv laus, peb tau pom tias kev txhim kho kev sib raug zoo ntawm tus tswv-commensal hauv kev laus barrier epithelia tuaj yeem txhawb kev noj qab haus huv thiab kev ua neej. Peb tau siv ntau txoj hauv kev uas tsis muaj kev cuam tshuam los soj ntsuam cov hnyuv ua haujlwm thiab microbiota dynamics thaum laus. Ua ntej, qhov kev ntsuam xyuas Smurf tau qhia txog lub luag haujlwm zoo ntawm TFs hauv kev tswj hwm cov kab mob epithelial barrier tsis ua haujlwm thaum muaj hnub nyoog. Tom qab ntawd, qhov kev ntsuam xyuas kev tiv thaiv kab mob siv ntau yam yoov hloov pauv tau qhia tias TFs ncua sijhawm dhau ntawm ISCs thiab kev hloov pauv ntawm cov hlwb sib txawv. Thaum kawg, kev sib txuas ntawm microbiome tau pom tias TFs txo qis lub hnub nyoog pib nthuav dav ntawm txoj hnyuv microflora. Cov txiaj ntsig no qhia tau hais tias TFs ua lub luag haujlwm tseem ceeb hauv kev ncua kev laus ntawm tus tswv tsev los ntawm kev tswj lub plab homeostasis.

Theaflavins alleviate ntau lub cev tiv thaiv kab mob hauv plab los ntawm kev tswj tsis zoo imd signaling

Nws tau raug pom zoo tias qhov sib npaug ntawm lub plab tiv thaiv kab mob thiab microbiota yuav tsum tau tswj xyuas kom lub tsev nyob mus ntev ntawm cov kab mob epithelia (Guo li al., 2014; Maynard thiab Weinkove, 2018). Cov kev tshawb fawb yav dhau los kuj tau qhia tias qhov mob hnyav heev hauv lub plab yog txuam nrog kev muaj hnub nyoog ntsig txog kev ua haujlwm tsis zoo hauv cov kab mob epithelia thiab yog ib qho cim ntawm kev laus (Clark et al., 2015; Guo et al., 2014; Ji et al., 2019) . Yog li, kev tswj hwm cov neeg noj qab haus huv los ntawm kev khaws cia hauv lub cev tiv thaiv kab mob hauv lub cev hauv cov epithelia yog ib txoj hauv kev zoo los txhawb kev noj qab haus huv thiab kev noj qab haus huv (Bonfini li al.,2016; Broderick, 2016; Clark et al., 2015). uas TFs ncua kev laus, peb tau ua RNA-seq tsom xam siv cov hnyuv dissected nrog lossis tsis muaj TFs kho. Peb cov ntaub ntawv qhia tau hais tias cov pab pawg ntawm cov noob tau nthuav tawm hauv cov hnyuv ntawm cov txiv hmab txiv ntoo tswj yoov piv nrog cov tsiaj kho TFs tau ua kom muaj txiaj ntsig nrog cov noob cais raws li "kev tiv thaiv kab mob" cov noob. Ntxiv bioinformatic tsom xam thiab tau-PCR kev soj ntsuam tau qhia tias qhov kev qhia ntawm Imd-txog AMP noob tau cuam tshuam loj heev los ntawm kev noj zaub mov ntxiv ntawm TFs. Cov txiaj ntsig tau los ntawm CD-1 kev sim nas siv DSS los ua kom tsis muaj zog tiv thaiv kab mob hauv cov hnyuv ntxiv tau lees paub qhov kev xav tias TFs ua haujlwm raws li kev tiv thaiv kab mob hauv plab.

Theaflavins inhibit cov cim los ntawm kev txiav txim siab ntawm amyloid sib dhos ntawm Imd

Yuav ua li cas TFs tsis zoo tswj lub Imd signaling txoj kev? Raws li cov txiaj ntsig ntawm kev txheeb xyuas cov ntaub ntawv pov thawj, peb pom tias TFs yog siv tau rau kev hloov pauv ntawm cov ntsiab lus tseem ceeb hauv Imd signaling pathway. Ntawm qhov txaus siab, peb tau pom qhov pom tseeb ntawm kev sib raug zoo ntawm TFs nrog purified Imd protein los ntawm SPR kev soj ntsuam, uas qhia txog qhov tshwm sim uas TFs tuaj yeem ua lub luag haujlwm hauv kev tswj hwm Imd protein tus cwj pwm lossis nws cov txheej txheem kev hloov pauv tom qab. Cov kev tshawb fawb yav dhau los tau qhia txog lub luag haujlwm tseem ceeb ntawm ubiquitination hauv Imd teeb liab hloov pauv (Kleino thiab Silverman, 2014; Myllymaki li al., 2014), thiab kev tiv thaiv ntawm Imd ubiquitination tau ua rau muaj kev cuam tshuam loj ntawm Imd signaling (Myllymaki li al., 2014; Thevenon et al., 2009). Txawm li cas los xij, peb qhov kev ntsuam xyuas hauv vitro thiab hauv vivoubiquitination tau pom tias TFs tsis tshua muaj kev cuam tshuam rau cov qauv ubiquitination ntawm Imd. Ib txoj kev tshawb fawb tsis ntev los no tau qhia tias kev tsim cov amyloid yuav tsum tau ua kom lub Drosophila Imd txoj hauv kev thaum pom cov kab mob peptidoglycans (Kleino li al., 2017). Qhov zoo siab, peb qhov kev ntsuam xyuas SDD-AGE thiab ThTbinding tau qhia tias kev sib koom ua ke ntawm Imd raug tiv thaiv los ntawm TFs. Kev tiv thaiv ntawm Imd aggregation los ntawm knocking down los yog kev hloov ntawm Imd nyob rau hauv lub plab hlwb thaj tsam reversed cov txiaj ntsig zoo ntawm TFs ntawm lifespan extension. Txhawm rau kom paub meej ntxiv txog peb qhov kev tshawb pom, nws yuav yog qhov txaus siab los tshuaj xyuas seb TFs puas tsis ua haujlwm ntxiv rau kev ua neej nyob rau hauv cov txiv hmab txiv ntoo fliers thaum lub pov tseg cryptic RHIM (cRHIM, amino acids los ntawm 118th mus rau 121h) motif ntawm Imd yog tshwj xeeb knocked tawm puas, Raws li qhov kev tshawb pom tias kev ua txhaum cai hauv Imd yog qhov tseem ceeb rau nws qhov sib sau ua ke thiab tsim amyloid (Kleino li al., 2017). Txawm li cas los xij, peb txoj kev tshawb fawb qhia txog lub luag haujlwm tseem ceeb ntawm TFs hauv kev tswj cov hnyuv homeostasis thiab cov txheej txheem kev laus, tej zaum los ntawm kev hloov pauv Imd coalescence. Cov kev tshawb fawb yav tom ntej yuav tsum tsom mus rau kev tshawb nrhiav cov seem lossis thaj chaw ntawm Imd raug khi los ntawm TFs thiab yuav ua li cas qhov kev khi no ua rau kev tiv thaiv Imd condensation. Tsis tas li ntawd, peb qhov kev tshawb pom hauv cov tsiaj txhu tau qhia tias TFs tuaj yeem tiv thaiv txoj hnyuv tiv thaiv kab mob los ntawm exogenous lom tshuaj lom DSS, qhia txog kev tswj hwm lub luag haujlwm ntawm TFs hauv kev tswj cov hnyuv homeostasis. Nws yog ib qho tsim nyog los tshawb xyuas seb TFs ua lub luag haujlwm hauv kev tswj hwm cov protein sib dhos hauv cov ntaub so ntswg mammalian.

Kab lus no yog muab rho tawm los ntawm zhongwenxie@ahau.edu.cn