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Magranule eBan-Lan-Gen anoderedza chirwere chedextran chinokonzerwa nesodium sulfate-induced relapsing colitis mumakonzo nekuchinja ma microbiota emudumbu uye kudzoreredza kugadzirwa kweSCFA Derived-GLP-1 mudumbu.
Jiao Peng,1-3,*Li Xi,4,*Zheng Lin,3,5 Duan Lifang,1 Gao Zhengxian,2,5 Diehu,1 Li Jie,6 Li Xiaofeng,6 Shen Xiangchun,5 Xiao Haitao21Peking University Shenzhen Hospital Dhipatimendi reMishonga, Shenzhen, People's Republic of China; 2Shenzhen University Health Science Center School of Pharmacy School of Pharmacy, Shenzhen, People's Republic of China; 3Guizhou Medical University Engineering Technology Research Center of Ethnic Medicine and Traditional Chinese Medicine Development and Application Ministry of Education, Guizhou Provincial Key Laboratory of Pharmacy, Guizhou Medical University, Guiyang, People's Republic of China; 4 Dhipatimendi reGastroenterology, Peking University Shenzhen Hospital, Shenzhen, People's Republic of China; 5 Chikoro cheMishonga, Guizhou Medical University, State Key Laboratory of Medicinal Plant Function and Application, Guiyang; 6 Dhipatimendi reLaboratory Medicine, Peking University Shenzhen Hospital, Shenzhen, China [email protected] Shen Xiangchun, Chikoro cheMishonga, Guizhou Medical University, Guizhou, People's Republic of China, 550004, Email [email protected] Chinangwa: Kurapa kwakavakirwa paGLP-1 inzira itsva yekurapa chirwere chemudumbu chinoputika. Magranule eBan-Lan-Gen (BLG) imhando inozivikanwa yeTCM inorwisa utachiona inoratidza kugona kwekudzivirira kuzvimba mukurapa zvirwere zvakasiyana-siyana zvinoputika. Zvisinei, mhedzisiro yayo yekudzivirira kuzvimba pacolitis uye mashandiro ayo haasati ajeka. NZIRA: Kugadzira dextran sodium sulfate (DSS)-induced chronic relapsing colitis mumakonzo. Zviratidzo zvechirwere, zviratidzo zvekukuvara kwehistological, uye mazinga eproinflammatory cytokine zvakaitwa kuti zviongorore mhedzisiro yekudzivirira yeBLG. Mhedzisiro yeBLG pa gut microbiota negut yakaratidzwa nemazinga eGLP-1 muropa uye colonic Gcg, GPR41, uye GRP43 expression, gut microbiota composition, fecal Mazinga eSCFA, uye kuburitswa kweGLP-1 kubva kumasero ekutanga eepithelial embeva colonic kugadzirwa kweGLP-1 inobva kuSCFA. Mhedzisiro: Kurapwa kweBLG kwakaderedza zvakanyanya kurasikirwa nehuremu hwemuviri, DAI, kupfupika kwecolon, kukuvara kwetishu yekoloni, uye mazinga eTNF-α, IL-1β, uye IL-6 mumatishu ekoloni. Pamusoro pezvo, kurapwa kweBLG kunogona kudzoreredza zvakanyanya mazinga eGcg, GPR41 uye GRP43 uye serum GLP-1 mumatishu ekoloni, uye nekuwedzera mabhakitiriya anogadzira SCFA akadai seAkkermansia nePrevotellaceae_UCG-001, uye kuderedza kuwanda kwemabhakitiriya akadai seEubacterium_xylanophilum_group, Ruminococcaceae_UCG-014, Intestinimonas neOscillibacter. Pamusoro pezvo, kurapwa kweBLG kunogona kuwedzera zvakanyanya huwandu hweSCFA mutsvina yemakonzo ekoloni. Panguva imwe chete, kuyedza kwe in vitro kwakaratidzawo kuti tsvina yembeva dzakarapwa neBLG inogona kukurudzira zvikuru masero madiki eMurine colonic epithelial cells anogadzira GLP-1. Mhedziso: Izvi zvakawanikwa zvinoratidza kuti BLG ine simba rekurwisa colitis. BLG ine mukana wekugadzirwa semushonga, kunyanya muchikamu nekuchinja ma microbiota emudumbu uye kudzoreredza kugadzirwa kweGLP-1 inobva mudumbu. Mishonga inovimbisa yechirwere checolitis chisingaperi. Mazwi akakosha: colitis, Ban-Lan-Gen granules, gut microbiota, short-chain fatty acids, GLP-1
Ulcerative colitis (UC) chirwere chinogara kwenguva refu chekuzvimba kwemuura nemudumbu chinoratidzwa nemanyoka anodzokorora, kurwadziwa mudumbu, kudzikira uremu, uye tsvina ine ropa mucopurulent.1 Munguva pfupi yapfuura, kuwanda kweUC kwave kuri kuwedzera munyika dzaimbove dzisina chirwere ichi, kusanganisira China, nekuwedzera kukurumbira kwemararamiro ekuMadokero.2 Kuwedzera uku kunokonzera matambudziko makuru kuhutano hwevanhu uye kune zvakunokanganisa kugona kwevarwere kushanda uye hupenyu hwakanaka. Zvinonyanya kukosha kuti pathogenesis yeUC haisati yanyatsojeka, asi zvinogamuchirwa kuti majini, zvinhu zvakatipoteredza, gut microbiota, uye immune system zvese zvinobatsira mukukura kweUC.3 Kunyangwe pari zvino, hapana mushonga weUC, uye chinangwa chekurapa ndechekudzora zviratidzo zvekiriniki, kukurudzira nekuchengetedza kupora, kukurudzira kupora kwemucosal, uye kuderedza kudzokazve. Kurapa kwekare kunosanganisira aminosalicylates, corticosteroids, immunosuppressants, uye biologics. Zvisinei, mishonga iyi haigone kuwana mhedzisiro inodiwa nekuda kwemigumisiro yayo yakasiyana-siyana.4 Munguva pfupi yapfuura, zvidzidzo zvakawanda zvakaratidza kuti mishonga yechivanhu yeChinese (TCM) yakaratidza kugona kukuru mukubatsira kuderedza UC ine chepfu shoma, zvichiratidza kuti kugadzirwa kwemishonga mitsva yeTCM inzira yekurapa ine tariro yeUC.5-7​​
Banlangen Granules (BLG) mushonga wechivanhu wekuChina unogadzirwa kubva mumvura inotorwa kubva mumudzi weBanlangen.8 Kuwedzera pakushanda kwayo kwekurwisa mavhairasi, BLG inoratidza kugona kwekudzivirira kuzvimba mukurapa zvirwere zvakasiyana-siyana zvinozvimba.9,10 Pamusoro pezvo, maglucosinolates (R,S-goitrin, progoitrin, epiprorubin uye glucoside akatorwa uye akaonekwa kubva muzvimedu zvemvura zveRadix isatidis) uye nucleosides (hypoxanthine, adenosine, uridine uye guanosine) uye indigo alkaloids dzakadai se indigo uye indirubin.11,12 Zvidzidzo zvekare zvakaratidza zvakanaka kuti makemikari adenosine, uridine uye indirubin anoratidza simba rekudzivirira colitis mumhando dzakasiyana dzemhuka dzecolitis.13-17 Zvisinei, hapana zvidzidzo zvakavakirwa pauchapupu zvakaitwa kuti tiongorore kushanda kweBLG mucolitis. Muchidzidzo chino, takaongorora kudzivirira kweBLG pa dextran sodium sulfate (DSS)-induced chronic relapsing colitis mu C57BL/6 mbeva uye vakawana kuti kupiwa BLG nemuromo kwakaderedza zvakanyanya DSS-induced chronic relapsing colon mumakonzo. Kuzvimba, nzira dzayo dzekutonga dzakabatana nekuchinja kwe microbiota yemudumbu uye kudzoreredzwa kwekugadzirwa kweglucagon-like peptide-1 (GLP-1) inobva mudumbu.
Magranule eBLG (asina shuga, Z11020357 yakatenderwa neNMPA; Beijing Tongrentang Technology Development Co., Ltd., Beijing, China; nhamba yebatch: 20110966) akatengwa kubva kumishonga. DSS (Molecular Weight: 36,000–50,000 Daltons) yakatengwa kubva kuMP Biologicals (Santa Ana, USA). Sulfasalazine (SASP) (≥ 98% purity), hematoxylin uye eosin zvakatengwa kubva kuSigma-Aldrich (St. Louis, MO, USA). Mouse TNF-α, IL-1β uye IL-6 luminex Elisa assay kits dzakatengwa kubva kuR&D systems (Minneapolis, MN, USA). Acetic acid, propionic acid, uye butyric acid zvakatengwa kubva kuAladdin Industries (Shanghai, China). 2-Ethylbutyric acid yakatengwa kubva kuMerck KGaA (Darmstadt, Germany).
Mbeva dzechirume dzeC57BL/6 dzine mavhiki matanhatu kusvika masere (huremu hwemuviri 18-22 g) dzakatengwa kubva kuBeijing Wetahe Laboratory Animal Technology Co., Ltd. (Beijing, China) uye dzakachengetwa munzvimbo inodziya ye22 ± 2 °C nechiedza chenguva yerima kwemaawa gumi nemaviri. Mbeva dzakapihwa chikafu chembeva chakajairwa nekuwana mvura yekunwa mahara kwevhiki imwe chete kuti dzijairane nenzvimbo itsva. Mbeva dzakazokamurwa zvisina tsarukano kuita mapoka mana: boka rekudzora, boka remuenzaniso weDSS, boka rakarapwa neSASP (200 mg/kg, remuromo) uye boka rakarapwa neBLG (1 g/kg, remuromo). Sezvakaratidzwa muMufananidzo 1A, zvichienderana nekudzidza kwedu kwapfuura, chirwere chechirwere chepfungwa chinodzokororazve chakakonzerwa nemakonzo nematanho matatu e1.8% DSS kwemazuva mashanu, zvichiteverwa nemvura yakasanganiswa kwemazuva manomwe, zvichienderana nekudzidza kwedu kwapfuura.18 Mbeva dziri mumapoka akarapwa neSASP neBLG dzakarapwa neSASP neBLG, zvichiteerana, zuva rega rega kutanga kubva pazuva re0. Sekureva kwekuyedza kwekutanga, Chiyero cheBLG chakaiswa pa1 g/kg. Panguva iyi, chiyero cheSASP chakaiswa pa200 mg/Kg zvichienderana nezvinyorwa.4 Mapoka evanodzora neDSS model akagamuchira huwandu hwakafanana hwemvura mukati mekuedza uku.
Mufananidzo 1 BLG inovandudza chirwere checolitis chisingaperi chinokonzerwa neDSS mumakonzo. (A) Kugadzira uye kurapwa kwechirwere checolitis chisingaperi chinodzokororwa, (B) kuchinja kwehuremu hwemuviri, (C) chibodzwa chechirwere (DAI), (D) kureba kwekoloni, (E) mufananidzo unomiririra koloni, (F) H&E staining Colon (magnification, ×100) uye (G) chibodzwa chehistological. Data rinoratidzwa seavhareji ± SEM (n = 6).##p < 0.01 kana ###p < 0.001 vs boka rekutonga (Con); *p < 0.05 kana **p < 0.01 kana ***p < 0.001 vs boka reDSS.
Kurema kwemuviri, kugara zvakanaka kwetsvina, uye kubuda ropa mudumbu zvainyorwa zuva nezuva. Disease activity index (DAI) yakaongororwa nekubatanidza huwandu hwehuremu hwemuviri, kugara zvakanaka kwetsvina, uye kubuda ropa mudumbu sezvakatsanangurwa kare.19 Pakupera kwekuyedza, mbeva dzese dzakaurayiwa uye ropa, tsvina neura zvakaunganidzwa kuti zviongororwezve.
Matishu emudumbu akagadziriswa formalin ndokuiswa muparaffin. Zvikamu zve5-micron zvakagadzirwa ndokuzoiswa ruvara rwehematoxylin-eosin (H&E), ndokuzopofumadzwa ndokuiswa ganda sezvakatsanangurwa kare.19
RNA yese ye colon tissue yakabviswa ne Trizol reagent (Invitrogen, Carlsbad, CA), ichiteverwa ne cDNA extraction ne reverse transcriptase (TaKaRa, Kusatsu, Shiga, Japan). Quantitative PCR yakaitwa uchishandisa real-time PCR system ne SYBR Green Master (Roche, Basel, Switzerland). Target gene transcripts dzakagadziriswa kuita β-actin uye data rakaongororwa uchishandisa nzira ye 2-ΔΔCT. Ma gene primer sequences anoratidzwa muTafura 1.
Kupatsanurwa kwemasero emudumbu embeva dzekutanga uye kuchengetwa kwemasero kwakaitwa sezvakatsanangurwa kare.20 Muchidimbu, mapapu embeva dzine mavhiki matanhatu kusvika masere ekuberekwa akatanga kubviswa mushure mekubayiwa nekubviswa kwemuromo wechibereko, ndokuzovhurwa kwenguva refu, ndokurapwa neHanks Balanced Salt Solution (HBSS, isina calcium nemagnesium) ndokuchekwa kuita zvidimbu zvidiki zve 0.5-1 mm. Mushure mezvo, matishu akagayiwa ne 0.4 mg/mL collagenase XI (Sigma, Poole, UK) mu DMEM medium yakasununguka uye akaiswa centrifuge pa 300 xg kwemaminitsi mashanu patembiricha yemukamuri. Dzorerazve pellet muDMEM medium (yakawedzerwa ne 10% fetal bovine serum, 100 Units/mL penicillin, uye 100 µg/mL streptomycin) pa 37 °C uye upfuure ne nylon mesh (hukuru hwepore ~250 µm). Zvimwe zvikamu zve colonic epithelial cells zvakaiswa mundiro dzegirazi uye zvakaiswa ne acetic acid, propionic acid, asidhi yebutyric, uye tsvina yembeva kwemaawa maviri pa37°C, 5% CO2.
Matishu emudumbu akaongororwa nePBS, uye huwandu hwema cytokines IL-6, TNF-α uye IL-1β mumatishu emudumbu hwakaonekwa uchishandisa luminex ELISA assay kits (R&D systems, Minneapolis, MN, USA). Saizvozvowo, huwandu hweGLP-1 mu serum uye culture medium ye primary murine colonic epithelial cells hwakaongororwa neELISA kit (Bioswamp, Wuhan, China) zvichienderana nemirairo yemugadziri.
DNA yese kubva mutsvina yakabviswa uchishandisa kit yekubvisa DNA (Tiangen, China). Hunhu nehuwandu hweDNA zvakayerwa pachiyero che260 nm/280 nm uye 260 nm/230 nm, zvichiteerana. Mushure mezvo, uchishandisa DNA yega yega yakabviswa se template, ma primer chaiwo 338F (ACTCCTACGGGAGGCAGCAG) uye 806R (GGACTACHVGGGTWTCTAAT) akashandiswa kuwedzera nzvimbo dzeV3-V4 dze16S rRNA gene munzvimbo dzakasiyana. Zvigadzirwa zvePCR zvakacheneswa uchishandisa QIAquick Gel Extraction Kit (QIAGEN, Germany), yakayerwa ne real-time PCR, uye yakarongwa uchishandisa IlluminaMiseq PE300 sequencing platform (Illumina Inc., CA, USA). Kune bioinformatics analysis, data processing yakaitwa zvichitevera protocols dzakambotaurwa.21,22 Muchidimbu, shandisa Cutadapt (V1.9.1) kusefa mafaira e express asina kubikwa. MaOTU akaunganidzwa. uchishandisa UPARSE (vhezheni 7.0.1001) ine kupatsanurwa kwakafanana kwe97%, uye UCHIME yakashandiswa kubvisa ma "chimeric sequences". Kuongorora nekupatsanura nharaunda kwakaitwa uchishandisa RDP classifier (http://rdp.cme.msu.edu/) zvichibva pane SILVA ribosomal RNA gene database.
Mazinga emafuta ane ma "short-chain fatty acids" (acetic acid, propionic acid, uye butyric acid) akayerwa sezvakatsanangurwa kare naTao et al., pamwe nekumwe kuchinja.23 Muchidimbu, 100 mg yetsvina yakatanga yaiswa mu 0.4 mL yemvura yakabviswa maion, ichiteverwa ne 0.1 mL ye 50% sulfuric acid uye 0.5 mL ye 2-ethylbutyric acid (internal standard), ndokuzoiswa homogenized uye kupiswa pa 4°C. Kutenderera kwemhepo pa12,000 rpm kwemaminitsi gumi nemashanu paC. Supernatant yakabviswa ne0.5 mL ye ether ndokupinzwa muGC kuti iongororwe. Pakuongorora kwegas chromatography (GC), sampuro dzakaongororwa uchishandisa GC-2010 Plus gas chromatograph (Shimadzu, Inc.) ine flame ionization detector (FID). Kupatsanurwa kwakaitwa uchishandisa ZKAT-624 column, 30 m × 0.53 mm × 0.3 μm (Lanzhou Zhongke Antai Analytical Technology Co., Ltd., China). Data rakatorwa uchishandisa GC solution software (Shimadzu, Inc.). Chikamu chekupatsanurwa chaive 10:1, gasi raitakura raive nitrogen, uye mwero wekuyerera waive 6 mL/min. Vhoriyamu yejekiseni yaive 1 μL. Tembiricha yejekiseni neye detector yaive 300°C. Tembiricha yeovheni yakabatwa pa140°C kwemaminitsi gumi nematatu nehafu, ndokuzowedzerwa kusvika pa250°C pa tembiricha yacho yakamira kwemaminitsi mashanu; 120°C/min.
Data rinounzwa seavhareji ± standard error yeavhareji (SEM). Kukosha kwedata kwakaongororwa neAnova yenzira imwe chete ichiteverwa nebvunzo yaDuncan ye multiple range. GraphPad Prism 5.0 software (GraphPad Software Inc., San Diego, CA, USA) yakashandiswa pakuverenga kwese uye p < 0.05 yakaonekwa seyakakosha muhuwandu hwedata.
Zvinozivikanwa kuti UC chirwere chisingaperi checolitis chinodzokororwa chine marwadzo emudumbu, manyoka uye kubuda ropa. Saka, chirwere chisingaperi checolitis chinokonzereswa neDSS mumakonzo chakagadzwa kuti chiongorore kushanda kweBLG mukurwisa colitis (Mufananidzo 1A). Zvichienzaniswa neboka rekutonga, makonzo ari muboka remuenzaniso weDSS aive akadzikira huremu hwemuviri zvakanyanya uye DAI yakakwira, uye shanduko idzi dzakadzoserwa zvakanyanya mushure memazuva makumi maviri nemana ekurapwa kweBLG (Mufananidzo 1B naC). Kupfupisa colon chiratidzo chakakosha cheUC.Sezvinoratidzwa muMifananidzo 1D naE, hurefu hwemakonzo akagamuchira DSS hwakapfupikiswa zvakanyanya, asi hwakarerutswa nekurapwa kweBLG. Nekuda kweizvozvo, ongororo yehistopathological yakaitwa kuti iongorore kuzvimba kwekoloni. Mifananidzo yakatsvukiswa neH&E uye mamakisi epathological yakaratidza kuti kupihwa kweDSS kwakakanganisa zvakanyanya magadzirirwo ekoloni uye kwakakonzera kuparadzwa kwecrypt, nepo kurapwa kweBLG kwakaderedza zvakanyanya kuparadzwa kwecrypt uye mamakisi epathological (Mufananidzo 1F naG). Zvinonyanya kukosha, mhedzisiro yekudzivirira yeBLG pa1 g/Kg yaive yakafanana neye SASP pa 200 mg/Kg. Pamwe chete, zvakawanikwa izvi zvinoratidza kuti BLG inoshanda mukuderedza kuoma kwechirwere cheDSS-induced chronic relapsing colitis mumakonzo.
TNF-α, IL-1β uye IL-6 zviratidzo zvakakosha zvekuzvimba kwekoloni.Sezvinoratidzwa muMufananidzo 2A, DSS yakakonzera kuwedzera kukuru mukutaura kwemajini eTNF-α, IL-1β uye IL-6 mukoloni zvichienzaniswa neboka rekutonga. Kutungamirirwa kweBLG kunogona kuchinja zvakanyanya shanduko idzi dzinoitwa neDSS. Tevere, takashandisa ELISA kuti tione huwandu hwema cytokines anoputika eTNF-α, IL-1β, uye IL-6 mukoloni. Zvakabuda zvakaratidzawo kuti huwandu hweTNF-α, IL-1β, uye IL-6 mukoloni hwakawedzera zvakanyanya mumakonzo akarapwa neDSS, nepo kurapwa kweBLG kwakaderedza kuwedzera uku (Mufananidzo 2B).
Mufananidzo 2 BLG inodzivirira kuratidzwa kwemajini uye kugadzirwa kwema cytokines eproinflammatory TNF-α, IL-1β uye IL-6 mukoloni yemakonzo akarapwa neDSS.(A) Kuratidzwa kwemajini eColonic eTNF-α, IL-1β uye IL-6; (B) mazinga emapuroteni eColonic eTNF-α, IL-1β uye IL-6. Data rinoratidzwa seavhareji ± SEM (n = 4–6).#p < 0.05 kana ##p < 0.01 kana ###p < 0.001 vs boka rekutonga (Con); *p < 0.05 kana **p < 0.01 vs boka reDSS.
Dysbiosis yemudumbu yakakosha mukukura kweUC.24 Kuti vaongorore kana BLG ichichinja ma microbiota emudumbu emakonzo akarapwa neDSS, 16S rRNA sequencing yakaitwa kuti iongorore nharaunda yemabhakitiriya emukati memudumbu. Dhayagiramu yeVenn inoratidza kuti mapoka matatu aya ane maOTU 385. Panguva imwe chete, boka rega rega raive nemaOTU akasiyana (Mufananidzo 3A). Uyezve, Chao1 index neShannon index zvinoratidzwa muMufananidzo 3B naC zvakaratidza kuti kusiyana kwenharaunda yema microbiota emudumbu kwakadzikira mumakonzo akarapwa neBLG, sezvo Shannon index yakaderera zvakanyanya muboka rakarapwa neBLG. Kuongororwa kwePrincipal component (PCA) ne principal coordinate analysis (PCoA) zvakashandiswa kuona maitiro ekuunganidza pakati pemapoka matatu uye zvakaratidza kuti chimiro chenharaunda yemakonzo akarapwa neDSS chakaparadzaniswa zvakajeka mushure mekurapwa neBLG (Mufananidzo 3D naE). Data iri rinoratidza kuti kurapwa neBLG kwakakanganisa zvakanyanya chimiro chenharaunda yemakonzo ane DSS-induced colitis.
Mufananidzo 3 BLG inochinja kusiyana kwe microbiota yemudumbu mumakonzo ane DSS-induced colitis.(A) Venn diagram yeOTU, (B) Chao1 index, (C) Shannon's richness index, (D) Principal Component Analysis (PCA) score plot yeOTU, (E) OTU Principal Coordinate Analysis (PCoA) score Mufananidzo. Data rinoratidzwa seavhareji ± SEM (n = 6).**p < 0.01 vs DSS group.
Kuti tiongorore shanduko dzakati wandei muhupfu hwemakonzo, takaongorora kuumbwa kwehupfu hwemakonzo pamatanho ese etaxonomic.Sezvinoratidzwa muMufananidzo 4A, phyla huru mumapoka ese yaive Firmicutes neBacteroidetes, ichiteverwa neVerrucomicrobia. Kuwanda kweFirmicutes neFirmicutes/Bacteroidetes kwakawedzera zvakanyanya munharaunda dzemakonzo anorapwa neDSS zvichienzaniswa nembeva dzinodzora, uye shanduko idzi dzakadzoserwa zvakanyanya mushure mekurapwa neBLG. Kunyanya, kurapwa neBLG kwakawedzera zvakanyanya kuwanda kweVerrucobacterium mutsvina yemakonzo ane DSS-induced colitis. Padanho remhuri, nharaunda dzemakonzo anouraya makonzo dzaigarwa neLachnospiriaceae, Muribaculaceae, Akkermansiaceae, Ruminococcaceae nePrevotellaceae (Mufananidzo 4B). Zvichienzaniswa neboka reDSS, kupera kweBLG kwakawedzera kuwanda kweAkkermansiaceae, asi kwakaderedza kuwanda kweLachnospiraceae neRuminococcaceae. Zvikuru, parudzi rwegenus. Padanho repamusoro, utachiona hwematumbu hwakanga hwakabatwa neLachnospira_NK4A136_group, Akkermansia nePrevotellaceae_UCG-001 (Mufananidzo 4C). Izvi zvakaratidzawo kuti kurapwa kweBLG kwakagadzirisa kusawirirana kwehutachiona hwematumbu mukupindura dambudziko reDSS, iro rinoratidzwa nekudzikira kweEubacterium_xylanophilum_group, Ruminococcaceae_UCG-014, Intestinimonas neOscillibacter, uye kuwedzera kweAkkermansia nePrevotellaceae_UCG-001.
Mufananidzo 4 BLG inochinja kuwanda kwe microbiota mudumbu mumbeva dzinonzi DSS-induced colitis.(A) Kuwanda kwe microbiota mudumbu padanho re phylum; (B) Kuwanda kwe microbiota mudumbu padanho remhuri; (C) Kuwanda kwe microbiota mudumbu padanho re genus. Data rinoratidzwa seavhareji ± SEM (n = 6).#p < 0.05 kana ###p < 0.001 vs control (Con) group; *p < 0.05 kana **p < 0.01 kana ***p < 0.001 vs DSS group.
Tichifunga kuti ma "short-chain fatty acids" (SCFAs) ndiwo ma "metabolites" makuru eAkkermansia nePrevotellaceae_UCG-001, nepo acetate, propionate nebutyrate zviri maSCFA akawanda mu intestinal lumen, 25-27 tichiri muchidzidzo chedu. Sezvakaratidzwa muMufananidzo 5, huwandu hwe "fecal acetate, propionate," uye "butyrate" hwakaderera zvakanyanya muboka rakarapwa neDSS, nepo kurapwa kweBLG kwaigona kudzvinyirira kudzikira uku.
Mufananidzo 5. BLG inowedzera huwandu hweSCFAs mutsvina yemakonzo ane DSS-induced colitis.(A) Acetic acid mutsvina; (B) propionic acid mutsvina; (C) butyric acid mutsvina. Data rinoratidzwa seavhareji ± SEM (n = 6).#p < 0.05 kana ##p < 0.01 vs boka rekutonga (Con); *p < 0.05 kana **p < 0.01 vs boka reDSS.
Takaverenga zvakare Pearson correlation coefficient pakati pe genus-level differential SCFA uye fecal microbiota.Sezvakaratidzwa muMufananidzo 6, Akkermansia yakabatana zvakanaka nekugadzirwa kwepropionic acid (Pearson = 0.4866) uye butyric acid (Pearson = 0.6192). Kusiyana neizvi, Enteromonas neOscillobacter zvese zvaive zvakabatana zvisina kunaka nekugadzirwa kweacetate, nePearson coefficients ye0.4709 ne0.5104, zvichiteerana.Saizvozvowo, Ruminococcaceae_UCG-014 yaive yakabatana zvisina kunaka nekugadzirwa kwepropionic acid (Pearson = 0.4508) uye butyric acid (Pearson = 0.5842), zvichiteerana.
Mufananidzo 6 Kuongororwa kwePearson pakati pemaSCFA akasiyana uye hutachiona hwekoloni.(A) Enteromonas ine acetic acid; (B) Concussion bacillus ine acetic acid; (C) Akkermansia vs propionic acid; (D) Ruminococcus_UCG-014 ine propionic acid; (E) Akkermansia ine butyric acid; (F) ) Ruminococcus _UCG-014 ine butyric acid.
Glucagon-like peptide-1 (GLP-1) chigadzirwa cheproglucagon (Gcg) mushure mekushandurwa kwemasero chine hunhu hwekudzivirira kuzvimba.28 Sezvakaratidzwa muMufananidzo 7, DSS yakakonzera kudzikira kukuru mukutaura kweGcg mRNA. Kurapa kweColon neBLG kunogona kuderedza zvakanyanya kudzikiswa kweGcg inokonzerwa neDSS zvichienzaniswa neboka rekutonga (Mufananidzo 7A). Panguva imwe chete, huwandu hweGLP-1 muropa hwakadzikira zvakanyanya muboka rakarapwa neDSS, uye kurapwa kweBLG kunogona kudzivirira kudzikira uku (Mufananidzo 7B). Sezvo ma "short-chain fatty acids" achigona kukurudzira kuburitswa kweGLP-1 kuburikidza neG-protein-coupled receptor 43 (GRP43) uye G-protein-coupled receptor 41 (GRP41) activation, takaongororawo GPR41 neGRP43 mukoloni yemakonzo ecolitis uye takaona kuti kuratidzwa kweGRP43 neGPR41 mukoloni kwakadzikira zvakanyanya mushure mekusangana neDSS, uye kurapwa kweBLG kunogona kubatsira kuderedza uku (Mufananidzo 7C naD).
Mufananidzo 7 BLG inowedzera mazinga eGLP-1 muropa uye kuratidzwa kweGcg, GPR41 uye GRP43 mRNA mumbeva dzakarapwa neDSS.(A) Kuratidzwa kweGcg mRNA mudumbu remudumbu; (B) nhanho yeGLP-1 mudumbu remudumbu; (C) Kuratidzwa kweGPR41 mRNA mudumbu remudumbu; (D) Kuratidzwa kweGPR43 mRNA mudumbu remudumbu. Data rinoratidzwa seavhareji ± SEM (n = 5–6).#p < 0.05 kana ##p < 0.01 vs boka rekutonga (Con); *p < 0.05 vs boka reDSS.
Sezvo kurapwa kweBLG kuchigona kuwedzera mazinga eGLP-1 muropa, kuratidzwa kweGcg mRNA yekoloni, uye mazinga eSCFA yetsvina mumakonzo akarapwa neDSS, takaongorora zvakare acetate, propionate, uye butyrate pamwe chete ne control (F-Con), DSS colitis (F-Con) -DSS) uye BLG-treated colitis (F-BLG) makonzo pakuburitswa kweGLP-1 kubva kumasero eprimary murine colonic epithelial.Sezvinoratidzwa muMufananidzo 8A, masero eprimary mouse colonic epithelial akarapwa ne2 mM acetic acid, propionic acid, uye butyric acid, zvichiteerana, akakurudzira zvikuru kuburitswa kweGLP-1, zvichienderana nezvidzidzo zvekare.29,30 Saizvozvowo, ese F-Con, F-DSS, uye F-BLG (yakaenzana ne0.25 g yetsvina) akakurudzira zvikuru kuburitswa kweGLP-1 kubva kumasero eprimary murine colonic epithelial. Zvinonyanya kukosha kuti huwandu hweGLP-1 hwakaburitswa nemasero eprimary mouse colonic epithelial akarapwa neF-DSS hwakanga hwakaderera kupfuura ihwohwo. yemasero epithelial epithelial embeva eF-Con neF-BLG akarapwa. (Mufananidzo 8B). Ruzivo urwu runoratidza kuti kurapwa kweBLG kwakadzoreredza zvakanyanya kugadzirwa kweGLP-1 inobva kuSCFA mudumbu.
Mufananidzo 8 SCFA inobva kuBLG inokurudzira kuburitswa kweGLP-1 kubva kumasero epithelial epithelial epithelial epithelial epithelial epithelial.(A) Acetic acid, propionic acid, uye butyric acid zvakakurudzira kuburitswa kweGLP-1 kubva kumasero epithelial ... Maawa maviri pa37°C, 5% CO2, zvichiteerana. Huwandu hweGLP-1 hwakaburitswa kubva kumasero epithelial epithelial epithelial ekutanga emurine hwakaonekwa neELISA. Data rinoratidzwa seavhareji ± SEM (n = 3).#p < 0.05 kana ##p < 0.01 vs. blank kana F-Con; *p < 0.05 vs. F-DSS.
Zvidimbu: Ace, acetic acid; Pro, propionic acid; zvisinei, butyric acid; F-Con, tsvina inobva mumbeva dzinodzora; F-DSS, tsvina inobva mumbeva dzinokonzera colitis; F-BLG, kubva muura hwakarapwa neBLG.
Yakanyorwa neWorld Health Organization sechirwere chisingadi kurapwa, UC iri kuva njodzi pasi rose; zvisinei, nzira dzinoshanda dzekufanotaura, kudzivirira, uye kurapa chirwere ichi dzichiri shoma. Nokudaro, pane chikonzero chikuru chekutsvaga nekugadzira nzira itsva dzakachengeteka uye dzinoshanda dzekurapa UC. Mishonga yechivanhu yechiChinese isarudzo inovimbisa nekuti mishonga yechivanhu yakawanda yechiChinese yakaratidzwa kuti inoshanda mukurapa UC muvanhu vechiChinese kwemazana emakore, uye zvese zvinhu zvechisikigo uye zvinhu zvechisikigo zvisingakuvadzi vanhu nemhuka.31,32 Chidzidzo ichi chaive nechinangwa chekutsvaga mushonga wechivanhu wechiChinese wakachengeteka uye unoshanda wekurapa UC uye kuongorora mashandiro ayo.BLG imhando inozivikanwa yemishonga yechiChinese inoshandiswa kurapa furuwenza.8,33 Basa murabhoritari yedu nedzimwe rakaratidza kuti indigo, mushonga wechivanhu wechiChinese wakagadzirwa kubva muzvinhu zvakafanana neBLG, unoratidza kushanda kwakakosha mukurapa UC muvanhu nemhuka.4,34 Zvisinei, mhedzisiro yeBLG yekurwisa colitis nemigumisiro yayo Maitiro acho haana kujeka. Muchidzidzo chedu chazvino, mhedzisiro yedu inoratidza kuti BLG inoderedza kuzvimba kwekoloni kunokonzerwa neDSS, uko kwakabatana nekuchinja kwemudumbu. microbiota uye kudzoreredzwa kwekugadzirwa kweGLP-1 kunobva mudumbu.
Zvinozivikanwa kuti UC inoratidzwa nenguva dzekudzokera shure dzine zviratidzo zvekurapa, zvakaita sekuderera kwehuremu, manyoka, kubuda ropa mudumbu, uye kukuvara kukuru kwemucosal yemudumbu.35 Saka, chirwere checolitis chinogara chichidzoka chakapihwa nekupa 1.8% DSS katatu kwemazuva mashanu, zvichiteverwa nemazuva manomwe ekunwa mvura.Sezvinoratidzwa muMufananidzo 1B, kudzikira kwehuremu uye mamakisi eDAI zvakaratidza kutanga kwechirwere checolitis chinogara chichidzoka. Mbeva dziri muboka rakarapwa neBLG dzakaratidza kupora kubva pazuva rechisere, izvo zvaive zvakasiyana zvikuru nezuva rechi24. Shanduko dzakafanana dzakaonekwawo muDAI score, zvichiratidza kuvandudzika mukuvandudzwa kwechirwere checolitis.Panyaya yekukuvara kwemudumbu uye mamiriro ekuzvimba, kureba kwemudumbu, kukuvara kwemudumbu, uye kugadzirwa kwemajini uye kugadzirwa kwema cytokines eproinflammatory TNF-α, IL-1β, uye IL-6 mudumbu remudumbu zvakavandudzikawo zvikuru mushure mekurapwa neBLG.Pamwe chete, mhedzisiro iyi inoratidza zvakajeka kuti BLG inoshanda mukurapa chirwere checolitis chinogara chichidzoka mudumbu mumbeva.
BLG inoita sei kuti iite mhedzisiro yayo yemishonga? Zvidzidzo zvakawanda zvakapfuura zvakaratidza kuti gut microbiota ine basa guru mukukura kweUC, uye mishonga inotorwa kubva ku microbiome uye microbiome yakabuda senzira inokwezva yekurapa UC. Muchidzidzo chino, takaratidza kuti kurapwa kweBLG kwakakonzera shanduko huru mukuumbwa kwe gut microbiota, zvichiratidza kuti kudzivirira kweBLG kubva kuDSS-induced colitis kwakabatana nekuchinja kwe gut microbiota. Izvi zvinoenderana nepfungwa yekuti kugadzirisa homeostasis ye gut microbiota inzira yakakosha yekunzwisisa kushanda kweTCM preparations.36,37 Zvinonyanya kukosha, Akkermansia ibhakitiriya reGram-negative uye rine anaerobic yakasimba rinogara mu mucus layer ye gut, iro rinoparadza mucins, rinogadzira propionic acid, rinosimudzira goblet cell differentiation, uye rinochengetedza mucosa. basa rekudzivirira hutachiona.26 Ruzivo rwakawanda rwekiriniki nerwemhuka runoratidza kuti Akkermansia ine hukama hwakanyanya nehutachiona hwakanaka,38 uye kupiwa kweAkkermansia spp nemuromo. zvinogona kuvandudza kuzvimba kwemucosal zvakanyanya.39 Data redu razvino rinoratidza kuti kuwanda kweAkkermansia kwakawedzera zvakanyanya mushure mekurapwa kweBLG. Pamusoro pezvo, Prevotellaceae_UCG-001 ibhakitiriya rinogadzira SCFA.27 Ongororo dzakawanda dzakaratidza kuti Prevotellaceae_UCG-001 yakawanikwa muhuwandu hushoma mutsvina yemhuka dzine colitis.40,41 Data redu razvino rinoratidzawo kuti kurapwa kweBLG kunogona kuwedzera zvakanyanya kuwanda kwePrevotellaceae_UCG-001 muura hwembeva dzakarapwa neDSS. Kusiyana neizvi, Oscillibacter ibhakitiriya rinogara richirwisa mesophilic, risingapindi mweya.42 yakashuma kuti kuwanda kweOscillibacter kwakawedzera zvakanyanya mumbeva dzeUC uye kwakabatana zvakanaka nemazinga eIL-6 neIL-1β uye mamakisi ezvirwere.43,44 Zvinoshamisa kuti, kurapwa kweBLG kwakaderedza zvakanyanya kuwanda kweOscillibacter mutsvina yembeva dzakarapwa neDSS. Zvinoshamisa kuti, mabhakitiriya aya akachinjwa neBLG ndiwo anonyanya kugadzira SCFA. mabhakitiriya. Zvidzidzo zvakawanda zvekare zvakaratidza kuti maSCFA anogona kubatsira pakuzvimba kwemudumbu uye kudzivirira epithelial integrity yemudumbu.45,46 Ruzivo rwedu rwazvino rwakaonawo kuti kuwanda kweSCFA acetate, propionate, uye butyrate mutsvina yakarapwa neDSS kwakawedzera zvakanyanya mumbeva dzakarapwa neBLG. Zvikatorwa pamwe chete, zvakawanikwa izvi zvinoratidza zvakajeka kuti kurapwa neBLG kunogona kuwedzera mabhakitiriya anogadzira SCFA anokonzerwa neDSS mumakonzo ane chirwere checolitis chinogara chichidzoka.
GLP-1 imushonga unonyanya kugadzirwa muileum nemuura uye unoita basa rakakosha mukunonoka kubuda kwemudumbu uye kuderedza shuga yeropa mushure mekudya.47 Humbowo hunoratidza kuti dipeptidyl peptidase (DPP)-4, GLP-1 receptor agonist, uye GLP-1 nanomedicine zvinogona kuderedza kuzvimba kwemudumbu mumakonzo.48-51 Sezvakashumwa muzvidzidzo zvekare, kuwanda kweSCFA kwakabatana nemazinga eplasma GLP-1 muvanhu nemakonzo.52 Data redu razvino rinoratidza kuti mushure mekurapwa kweBLG, mazinga eGLP-1 muropa uye kuratidzwa kweGcg mRNA kwakawedzera zvakanyanya.Saizvozvowo, kuburitswa kweGLP-1 kwakawedzera zvakanyanya mumatunhu emudumbu mushure mekukurudzirwa netsvina kubva kumakonzo akarapwa neBLG zvichienzaniswa nekukurudzirwa netsvina kubva kumakonzo akarapwa neDSS. MaSCFA anokanganisa sei kuburitswa kweGLP-1?Gwen Tolhurst et al. yakashuma kuti SCFA inogona kukurudzira kuburitswa kweGLP-1 kuburikidza neGRP43 neGPR41.29 Data redu razvino rinoratidzawo kuti kurapwa kweBLG kunowedzera zvakanyanya kuratidzwa kwemRNA kweGRP43 neGPR41 muura hwembeva dzakarapwa neDSS. Data iri rinoratidza kuti kurapwa kweBLG kunogona kudzorera kugadzirwa kweGLP-1 kwakasimudzirwa neSCFA nekushandisa GRP43 neGPR41.
BLG mushonga weOTC unotengeswa kwenguva refu muChina. Chipimo chepamusoro cheBLG chinobvumirwa mumakonzo ekuKunming i80g/Kg, uye hapana chepfu yakanyanya yakaonekwa.53 Parizvino, chiyero chinokurudzirwa cheBLG (isina shuga) muvanhu i9-15 g/zuva (katatu pazuva). Chidzidzo chedu chakaratidza kuti BLG pa1g/Kg yakavandudza chirwere cheDSS-induced chronic relapsing colitis mumakonzo. Chipimo ichi chakafanana nechiyero cheBLG chinoshandiswa mukiriniki. Chidzidzo chedu chakaonawo kuti mashandiro ayo anokonzerwa, zvishoma nezvishoma, nekuchinja mugut microbiota, kunyanya mabhakitiriya anogadzira SCFA, akadai seAkkermansia nePrevotellaceae_UCG-001, kudzoreredza kugadzirwa kweGLP-1 inobva mugut. Izvi zvakawanikwa zvinoratidza kuti BLG inofanirwa kutariswa zvakanyanya semushonga wekurapa chirwere checolitis. Zvisinei, nzira chaiyo yainogadzirisa nayo gut microbiota ichiri kusimbiswa nemakonzo asina microbiota uye kuisirwa mabhakitiriya etsvina.
Ace, acetic acid; asi, butyric acid; BLG, pandan; DSS, dextran sodium sulfate; DAI, disease activity index; DPP, dipeptidyl peptidase; FID, flame ionization detector; F-Con, control Fecal extracts yemakonzo; F-DSS, fecal extracts yemakonzo eDSS colitis; F-BLG, fecal extracts yemakonzo eBLG-treated colitis; GLP-1, glucagon-like peptide-1; Gcg, glucagon; gas chromatography, gas chromatography; GRP43, G protein-coupled receptor 43; GRP41, G protein-coupled receptor 41; H&E, hematoxylin-eosin; HBSS, Hanks' Balanced Salt Solution; OTC, OTC; PCA, principal component analysis; PCoA, principal coordinate analysis; Pro, propionic acid; SASP, sulfasalazine; SCFA, short-chain fatty acids; Mishonga yechiChinese, mishonga yechivanhu yechiChinese; UC, chirwere chemudumbu chinonzi ulcerative colitis.
Maitiro ese ekuyedza akatenderwa neAnimal Ethics Committee yePeking University Shenzhen-Hong Kong University of Science and Technology Medical Center (Shenzhen, China) zvichienderana neInstitutional Guidelines and Animal Regulations (ethics number A2020157).
Vanyori vese vakapa mipiro yakakosha mukufunga nekugadzira, kutora data, kana kuongorora data nekududzira; vakatora chikamu mukunyora chinyorwa kana kugadzirisa zvakadzama zviri mukati mepfungwa dzakakosha; vakabvuma kuendesa gwaro racho kujenari iripo; pakupedzisira vakabvumidza shanduro yacho kuti ibudiswe; Vane mutoro wezvese zvebasa.
Basa iri rakatsigirwa neNational Natural Science Foundation of China (81560676 na81660479), chirongwa chemhando yepamusoro cheShenzhen University (86000000210), Shenzhen Science and Technology Innovation Committee Fund (JCYJ20210324093810026), uye Guangdong Provincial Medical Science and Technology Research Fund (A2020157 naA2020272), Guizhou Medical University Pharmacy Guizhou Province Funded by Key Laboratory (YWZJ2020-01) uye Peking University Shenzhen Hospital (JCYJ2018009).
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