Tshooj 1: Lub raum Tubular Peroxisomes yog Dispensable rau lub raum ua haujlwm

Yog xav paub ntxiv, hu rautina.xiang@wecistanche.com

Peroxisomes yog tshwj xeeb cellular organelles koom nrog ntau yam txheej txheem metabolic. Hauv tib neeg, kev hloov pauv ua rau poob ntawm peroxisomes ua rau ntau lub cev tsis ua haujlwm (Zellweger's spectrum disorders, ZSD), suav nroglub raum tsis zoo. Txawm li cas los xij, lub luag haujlwm ntawm lub cev (patho) ntawm peroxisomes hauv lub cevraumtseem tsis paub. Peb tau hais txog lub luag haujlwm ntawm peroxisomes hauvlub raum ua haujlwmnyob rau hauv cov nas uas muaj xwm txheej ablation ntawm peroxisomal biogenesis nyob rau hauv lub raum tubule (cKO nas). Kev soj ntsuam kev ua haujlwm tsis tau qhia txog lub raum phenotype hauv cKO nas. Txawm li cas los xij, cov txiv neej cKO nas muaj lub cev qis thiab lub raum hnyav, thiab cov txiv neej laus cKO nas pom qhov txo qis hauv lub raum hnyav thiab lub raum qhov hnyav / lub cev hnyav piv. Kev tsom xam stereological pom tias muaj qhov nce ntawm mitochondria ntom ntom hauv cov tubules ze ze ntawm cKO nas. Integrated transcriptome thiab metabolome analyses qhia qhov kev rov ua haujlwm ntawm ntau txoj hauv kev metabolic, suav nrog cov metabolism ntawm glutathione thiab biosynthesis / biotransformation ntawm ntau chav kawm loj ntawm lipids. Txawm hais tias qhov kev ntsuam xyuas no pom tau tias them nyiajoxidative kev nyuaj siab, kev sib tw nrog kev noj zaub mov tsis zoo tsis ua rau lub raum tsis zoo hauv cKO nas. Peb pom tau hais tias lub raum tubular peroxisomes yog dispensable rau lub raum ua haujlwm. Peb cov ntaub ntawv tseem qhia tau hais tias lub raum tsis zoo hauv cov neeg mob ZSD yog lub hauv paus ntawm extrarenal.

Nyem qhov no kom paub txog cistanche tubulosa extract cov txiaj ntsig

Taw qhia

Peroxisomes yog ib leeg-membrane-bound organelles uas tau pom thawj zaug hauv lub raum nas los ntawm Rhodin hauv xyoo 1954(1). Cov kev kwv yees tam sim no qhia tias peroxisomes muaj ntau dua 50 enzymes koom nrog ntau lub zog ntawm tes, suav nrog -oxidation ntawm cov kab sib txuas ntev fatty acids (VLCFAs), ntev-chain fatty acids (LCFAs), thiab cov saw ntev dicarboxylic acid; a-thiab-oxidation ntawm branched-chain fatty acids; oxidation ntawm prostaglandins thiab leukotrienes; metabolism ntawm amino acids; biosynthesis ntawm ether phospholipids (xws li plasmalogens) thiab kua tsib acids; redox homeostasis; glyoxylate detoxification; thiab ferroptosis. Peroxisomes muaj nyob txhua qhov chaw nyob rau hauv yuav luag tag nrho cov tsiaj cov tsiaj, nrog ntau tshaj plaws nyob rau hauv lub raum proximal tubule cells thiab hepatocytes. Hauv cov hlwb no, peroxisomes nyob li ntawm 3 feem pua ​​​​ntawm cov xov tooj ntawm tes, tab sis lawv cov naj npawb, qhov loj, thiab cov duab sib txawv heev thaum lub sij hawm embryonic thiab postembryonic txoj kev loj hlob (2); lawv tus lej tuaj yeem loj hlob zoo hauv ntau yam kev ntxhov siab (3). Txawm hais tias ntau cov enzymes peroxisomal tau ua tau zoo, cov kev tshawb fawb hais txog lawv cov kev qhia thiab kev ua haujlwm hauv lub raum tsis tshua muaj. Tsis tas li ntawd, tag nrho kev ua haujlwm tseem ceeb ntawm peroxisomes hauv lub raum tseem tsis paub meej.

Cov pov thawj rau lub luag haujlwm ntawm peroxisomes hauv lub raum (patho) physiology tau tshwm sim los ntawm tib neeg kev tshawb fawb noob caj noob ces. Ob hom kev hloov pauv uas ua rau tib neeg peroxisomal teeb meem tau raug txheeb xyuas: (i) kev hloov pauv hauv lub npe hu ua peroxin (PEX) cov noob koom nrog hauv peroxisome biogenesis thiab (ii) kev hloov pauv hauv cov enzymes tshwj xeeb peroxisomal. Thawj hom kev hloov pauv ua rau muaj kev ua haujlwm tsis zoo ntawm peroxisomal ua rau cerebrohepatorenal Zellweger's spectrum disorders (ZSD) (4). Cov menyuam mos uas muaj mob hnyav ntawm ZSD feem ntau tuag thaum thawj xyoo ntawm lub neej los ntawm ntau lub cev tsis ua haujlwm. Txawm hais tias muaj cov kab mob hauv lub raum thiab / lossis lub raum oxalate pob zeb hauv cov neeg mob ZSD tau raug sau tseg zoo xyoo (5, 6), cov txheej txheem molecular ua rau cov kev puas tsuaj no tseem tsis tau paub txog. Lwm qhov ua rau lub raum txawv txav hauv ZSD tsis tau tshawb xyuas vim tias tus kab mob no tau tswj hwm los ntawm cov teeb meem neurological. Qhov cuam tshuam ntawm cov ntaub ntawv nruab nrab lossis me me ntawm ZSD ntawm lub raum kev ua haujlwm tsis tau raug tshuaj xyuas. Cov pov thawj txuas ib leeg peroxisomal enzyme deficiency rau lub raum pathophysiology tseem txwv.

Kev hloov pauv hauv daim siab peroxisomal alanine: glyoxylate aminotransferase encoded los ntawm AGXT noob ua rau thawj hyperoxaluria hom I, ib tus kab mob uas tshwm sim los ntawm kev tso nyiaj ntawm calcium oxalate crystals hauv lub raum (saib xyuas hauv ref. 7). Tsis ntev los no, ib qho kev hloov pauv ntawm autosomal uas ua rau lub raum Fanconi syn-drome, los yog kev ua haujlwm tsis zoo ntawm cov tubules ze ze, tau txheeb xyuas hauv peroxisomal enzyme enoyl-CoA hydratase thiab 3-hydroxy acyl CoA dehydrogenase (EHHADH)(8) . Qhov kev hloov pauv no ua rau muaj kev tsis sib haum xeeb ntawm EHHADH rau mitochondria thiab kev ua haujlwm tsis zoo ntawm mitochondrial. Kev tshawb nrhiav caj ces ua rau cov kab mob sib kis tau pom tias peroxisomal enzyme hydroxy acids oxidase 2 (HAO2) tuaj yeem ua lub luag haujlwm hauv kev tswj ntshav siab (9,10). Txawm li cas los xij, seb qhov phenotype no yog vim qhov hloov pauv HAO2 ua haujlwm hauv lub raum lossis lwm cov ntaub so ntswg tseem tsis paub. Feem ntau, cov ntaub ntawv los ntawm cov qauv tsiaj nrog rau lub raum tshwj xeeb tsis xws luag ntawm peroxisome biogenesis lossis lub raum tshwj xeeb inactivation ntawm ib leeg peroxisomal enzyme tseem ploj lawm. Ntawm no, peb tau hais txog lub luag haujlwm ntawm peroxisomes hauv lub raum ua haujlwm hauv cov txiv neej thiab poj niam nas nrog cov xwm txheej ablation ntawm peroxisomal biogenesis hauv lub raum tubule.

Cov txiaj ntsig

Generation thiab cov yam ntxwv tseem ceeb ntawm tus me nyuam mos txiv neej thiab poj niam nas nrog cov xwm txheej ablation ntawm peroxisomal biogenesis hauv lub raum tubule. Peroxisome biogenesis nyob rau hauv lub raum tubule tau cuam tshuam los ntawm kev tsis ua haujlwm ntawm Pex5 encoding cytosolic peroxisome targeting signal 1 (PTS1) receptor, uas yog qhov tseem ceeb rau kev xa khoom ntawm PTS1 motif-muaj cov proteins rau hauv peroxisomes (11,12). Kev tshem tawm ntawm Pex5 hauv lub raum tubule tau ua tiav siv doxycycline-inducible (DOX-inducible) system(PexSanlo/Pax8-rtTA/LC1 nas, ref.13). Vim tias qhov ua haujlwm tseem ceeb ntawm peroxisomes tuaj yeem nyob ntawm hnub nyoog (14), tus yam ntxwv tseem ceeb ntawm Pex5 tsis txaus hauv lub raum tau ua tiav ob qho tib si hauv cov menyuam mos thiab cov nas laus. Hauv kev sim nrog cov nas me me, kev txiav tawm ntawm floxed Pex5 allele tau ua tiav los ntawm kev tswj hwm ntawm DOX (2 mg / mL hauv dej haus) rau cov poj niam cev xeeb tub ntawm E14 thiab tswj kom txog thaum P7. Me nyuam nas raug txi ntawm P28. Cov nas uas muaj Pex5oilo genotype tau siv los tswj. Raws li tau pom hauv Daim Duab Ntxiv 1A (cov khoom siv ntxiv muaj nyob hauv online nrog rau tsab xov xwm no; https://doi.org/10.1172/jci.insight.155836DS1), DOX kev kho mob tau ua tiav qhov kev txiav tawm ze ntawm floxed Pex5 allele hauv Pexswn/Pax{{ 29}}GTA/LC1 txiv neej thiab poj niam menyuam mos nas. Kev soj ntsuam ntawm lub raum tsis tau qhia txog qhov txawv txav ntawm lub raum lossis lub raum pob zeb hauv cov nas me me uas tsis muaj Pex.5 hauv lub raum tubule (tsis pom). Albuminuria tsis nyob hauv qhov chaw kuaj zis ntawm Pex5- tsis muaj cov nas me me ntawm ob qho tib sipoj niam txiv neej(Cov duab ntxiv 1B). Lub cev hnyav (BW) thiab lub raum hnyav (KW) tab sis tsis yog KW / BW piv tau qis dua hauv txiv neej Pex5-tsis muaj cov nas me me piv nrog cov littermate tswj (Cov duab ntxiv 1C).

Generation thiab cov yam ntxwv tseem ceeb ntawm cov neeg laus txiv neej thiab poj niam nas nrog cov xwm txheej ablation ntawm peroxisomal biogenesis hauv lub raum tubule. Kev tshem tawm Pex5 nyob rau hauv lub raum tubule ntawm cov neeg laus nas yog induced los ntawm 2-lub lim tiam kev kho mob nrog DOX ntawm 8-lub lim tiam-laus Pex5xio/Pax8-ntTA/LC1 txiv neej thiab poj niam nas, nyob rau hauv no hais txog. raws li cKOm thiab cKOf nas, ntsig txog (saib Cov Txheej Txheem). Nyob rau tib lub sijhawm, tib txoj kev kho DOX tau muab rau lawv cov littermate tswj (Pex5a / lx nas, tom qab ntawd hu ua Ctrlm thiab Ctrlf nas, feem). Txhua qhov kev sim ntawm cov nas laus tau ua 4 lub lis piam tom qab qhov kawg ntawm DOX kho. Raws li pom nyob rau hauv daim duab 1A, Pex5 mRNA qhia tau txo qis hauv ob lub raum ntawm cKOm thiab cKOf nas. Txawm li cas los xij, hauv cKOf nas, qee qhov seem ntawm qhov ntev Pex5 mRNA kuj tau kuaj pom, qhia tias tsis tiav kev txiav tawm ntawm floxed allele. Ib yam li ntawd, PEX5 protein zoo li tsis tuaj yeem nyob hauv ob lub raum ntawm cKOm nas tab sis tseem tuaj yeem pom hauv cKOf nas, txawm tias nyob rau theem qis. Qhov sib txawv no tau pom thaum lub raum rho tawm los ntawm cKOm thiab cKOf nas tau thauj khoom ntawm tib lub SDS-PAGE gel thiab immunoblotted ua ke (Daim duab 1B). Ob leeg Kim thiab cKOf nas tau siv tau, tsis pom muaj qhov txawv txav, thiab muaj BW ib txwm piv rau cov nas tswj (Table 1). Qhov sib piv ntawm KW thiab KW / BW tau qis dua hauv cKOm nas piv nrog Ctrl nas tab sis tsis nyob rau hauv cKOf nas piv rau Ctrlf nas (Figures 1, C, thiab D, feem). Kev tshuaj xyuas ntawm 24-teev cov zis thiab cov ntshav plasma tsis tau qhia txog qhov tshwm sim ntawm genotype hauv cov nas ntawm ob leeg poj niam txiv neej, tshwj tsis yog cov plasma potassium qis hauv cKOm nas piv nrog Ctrl nas (Table 1). Albuminuria tsis nyob hauv cov zis ntawm cKOm thiab cKOf nas (Cov duab ntxiv 2A). Tsis muaj kev hloov pauv tag nrho, calcium oxalate deposits, lossis lipid tsub zuj zuj hauv lub raum ntawm cKOm nas (Cov duab ntxiv 2, BD, feem).

Electron microscopy kev soj ntsuam ntawm proximal tubule cells hauv cov neeg laus cKO nas. Electron microscopy kev soj ntsuam ntawm lub raum cortex tau nthuav tawm ntau cov peroxisomes nyob rau hauv cov tubule ze ze ntawm Ctrlm thiab Ctrlf nas (Daim duab 2, A, thiab B, feem). Peroxisomes tsis tshua muaj nyob rau hauv cov tubules ze ze ntawm cKOm thiab cKOf nas (Figure2, Cand D, feem). Cov cuab yeej stereological tau ua haujlwm rau kev soj ntsuam ntau ntawm cov cellular

organelles thiab cellular dimensions nyob rau hauv proximal tubules ntawm Ctrlm thiab cKOm nas. Ob leeg peroxisomes'volume density (number / μm² cytoplasm) thiab feem pua ​​​​ntawm cytoplasm nyob los ntawm peroxisomes nyob rau hauv proximal tubule cells tau poob qis hauv cKOm nas (Daim duab 3, A thiab B, raws li), Hloov pauv, mitochondrial ntim zoo li qhov ntom ntom, Feem pua ​​​​ntawm cytoplasm nyob los ntawm mitochondria nyob rau hauv cov tubule ze ze, tau nce hauv cKOm nas piv nrog Ctrl nas (Daim duab 3, C thiab D, raws li). Kuj-los ntim ntim thiab feem pua ​​​​ntawm cytoplasm nyob los ntawm lysosomes nyob rau hauv cov tubules ze ze tsis txawv ntawm Kim thiab Ctrlm nas (Daim duab 3, E thiab F, raws li). Raws li pom nyob rau hauv daim duab 3G, proximal tubule hlwb los ntawm cKOm nas tended txo cell dav (P= 0.083).

Integrated transcriptomic thiab metabolomic analyses qhia qhov kev rov ua dua tshiab hauv metabolic, antioxidant, thiab lipid synthesis pathways hauv ob lub raum ntawm cKO nas. Kev sib sib zog nqus transcriptome-sequencing thiab metabolome tsom xam tau ua los txheeb xyuas cov kev hloov pauv ntawm cov molecular hauv ob lub raum ntawm cKO nas (GSE179202). Kev sib piv ntawm transcriptomes qhia 1350 cov ntawv teev lus sib txawv hauv ob lub raum ntawm Ctrlm thiab cKOm nas (Daim duab 4A thiab Ntxiv Table 2, FDR< 5%)and="" 121="" transcripts="" differentially="" expressed="" in="" kidneys="" of="" ctrlf="" and="" ckof="" mice="" (figure="" 4b="" and="" supplemental="" table="" 3,="">< 5%).="" 61="" differentially="" expressed="" transcripts="" were="" present="" in="" both="" sexes(figure="" 4c="" and="" supplemental="" figure="" 3).="" enrichment="" analysis="" of="" 100="" genes="" encoding="" proteins="" related="" to="" peroxisomal="" function(kyoto="" encyclopedia="" of="" genes="" and="" genomes="" [kegg]="" pathway="" hsa04146)performed="" on="" transcriptomes="" of="" ctrlm="" and="" ckom="" mice="" revealed="" 52="" transcripts="" either="" up-or="" downregulated="" in="" kidneys="" of="" ckom="" mice="" (figure="" 4d="" and="" supplemental="" figure="" 4).="" gene="" set="" enrichment="" analysis(gsea)="" performed="" on="" the="" kegg="" pathway="" database="" (release="" on="" december="" 11,="" 2020)showed="" downregulation="" of="" pathways="" related="" to="" the="" metabolism="" of="" pyruvate,="" glyoxylate="" and="" dicarboxylate,="" amino="" acids="" (arginine,="" proline,="" cysteine,="" methionine,="" tryptophan,="" alanine,="" and="" histidine),="" or="" glutathione(figure="" 4e="" and="" supplemental="" figure="" 5)="" and="" upregulation="" of="" pathways="" related="" to="" fatty="" acid="" synthesis="" and="" degradation,="" ppar="" signaling,="" and="" abc="" transporters="" (figure="" 4f="" and="" supplemental="" figure="" 6).="" no="" enrichment="" was="" found="" in="" pathways="" linked="" to="" inflammation="" or="" fibrosis.="" targeted="" analysis="" of="" several="" groups="" of="" functionally="" related="" genes="" that="" are="" not="" represented="" in="" kegg="" pathways="" revealed="" substantial="" changes="" in="" the="" expression="" of="" genes="" encoding="" enzymes="" involved="" in="" the="" peroxisomal="" metabolism="" of="" fatty="" acids="" (acsl1/3/4/6,="" acsvl1,="" acot3/4,="" acnat1,="" acox3,="" abcd3,="" ehhadh,="" and="" acaa1b),="" plasmalogen="" biosynthesis="" (far1="" and="" agps),="" bile="" acid="" synthesis(amacr="" and="" hsd17b4),="" and="" reactive="" oxygen="" species="" (ros)="" detoxification(cat="" and="" sod1)="" (supplemental="" figure="" 3).="" alterations="" were="" also="" noted="" in="" the="" expression="" of="" a="" large="" number="" of="" genes="" critical="" for="" membrane="" transport="" processes="" in="" the="" proximal="" tubule,="" thick="" ascending="" limb="" (tal),="" and="" distal="" nephron(supplemental="" figure="" 7="" and="" supplemental="" table="" 4).="" for="" instance,="" in="" the="" proximal="" tubule,="" we="" found="" a="" reduction="" in="" the="" expression="" of="" an="" aquaporin-1="" water="" channel(agp1);="" megalin="" (lrp2);="" phosphate="" transporter="" napi-2a="" (slc34al);="" urate="" transporters="" uratl="" (slc22a12),="" npt1/4="" (sic17al/3),="" and="" oat1="" (slc22a6);="" and="" numerous="" other="" organic="" anion="" and="" amino="" acid="" transporters.="" in="" the="" tal="" and="" distal="" nephron,="" there="" was="" a="" substantial="" increase="" in="" the="" expression="" of="" genes="" encoding="" proteins="" involved="" in="" sodium="" reabsorption:="" nkcc2="" (slc12a1),="" clc-kb="" (clcnkb),="" βenac="" (scnn1b)=""><0.05), and="" ncc="" (slc12a3)(fdr="">

<0.05. (d)="" enrichment="" analysis="" of="" a="" homemade="" gene="" set="" (based="" on="" the="" kegg="" pathway="" mmu04146)="" targeting="" 100="" transcripts="" related="" to="" peroxisomal="" functions,="" in="" ckom="" versus="" ctrlm="" mice.="" (e="" and="" f)="" scatter="" plot="" of="" the="" top="" 25="" most="" downregulated="" (e)="" or="" upregulated="" (f)="" metabolic="" pathways="" in="" ckom="" versus="" ctrlm="" mice,="" based="" on="" an="" untargeted="" gsea="" using="" a="" database="" of="" 543="" kegg="" metabolic="" pathways.="" pathways="" are="" sorted="" by="" their="" absolute="" normalized="" enrichment="" score.="" a="" significant="" pathway="" regulation="" can="" be="" considered="" when="" adjusted="" p="" value="" referred="" to="" as="" "q="" value"="" is=""><0.2" alt="Figure 4. Transcriptional reprogramming in the kidneys of cKO mice. (A and B) Volcano plot representing the relative transcriptional expression of all renal transcripts in cKOm versus Ctrlm (A) or cKOf versus Ctrlf (B). Transcripts depicted in blue are significantly downregulated while transcripts depicted in red are significantly upregulated. (C) Venn diagrams showing the number of transcripts significantly downregulated or upregulated in cKOm (in blue) or cKOf (in pink) mice versus Ctrl mice of the same sex. A significant transcript regulation is considered when the adjusted P value referred to as " fdr"="" is=""><0.05. (d)="" enrichment="" analysis="" of="" a="" homemade="" gene="" set="" (based="" on="" the="" kegg="" pathway="" mmu04146)="" targeting="" 100="" transcripts="" related="" to="" peroxisomal="" functions,="" in="" ckom="" versus="" ctrlm="" mice.="" (e="" and="" f)="" scatter="" plot="" of="" the="" top="" 25="" most="" downregulated="" (e)="" or="" upregulated="" (f)="" metabolic="" pathways="" in="" ckom="" versus="" ctrlm="" mice,="" based="" on="" an="" untargeted="" gsea="" using="" a="" database="" of="" 543="" kegg="" metabolic="" pathways.="" pathways="" are="" sorted="" by="" their="" absolute="" normalized="" enrichment="" score.="" a="" significant="" pathway="" regulation="" can="" be="" considered="" when="" adjusted="" p="" value="" referred="" to="" as="" "q="" value"="" is=""><0.2" width="480" height="320" border="0" vspace="0" style="width: 480px; height: 320px;">

Los ntawm tag nrho ntawm 852 kuaj pom cov metabolites, 207 tau pom qhov sib txawv ntau hauv ob lub raum ntawm Ctrlm thiab cKOm nas, thiab 118 pom qhov sib txawv ntawm ob lub raum ntawm Ctrlf thiab cKOf nas (Table 5, FDR).<5%). seventy-nine="" metabolites="" demonstrating="" differential="" abundance="" were="" common="" in="" both="" comparisons(figure="" 5a="" and="" supplemental="" table="" 6).among="" metabolites="" showing="" the="" most="" significant="" differences="" were="" plasmalogens="" and="" sphingomyelins(decreased="" abundance)and="" glutathione-related="" metabolites="" and="" dicarboxylic="" acids="" (increased="" abundance)="" in="" kidneys="" of="" both="" ckom="" and="" ckof="" mice="" (figure="" 5b="" and="" supplemental="" figure="" 8,="" respectively).="" global="" analysis="" of="" metabolome="" confirmed="" that="" plasmalogens="" and="" sphingomyelins="" constituted="" a="" majority="" of="" metabolites="" showing="" a="" decreased="" abundance="" in="" kidneys="" of="" both="" ckom="" and="" ckof="" mice="" (figure="" 5c).lcfa="" dicarboxylates,="" very="" long-chain="" fatty="" acyl-carnitines,="" phosphatidylcholines,="" and="" phosphatidylethanolamines="" were="" present="" among="" metabolites="" with="" increased="" abundance,="" in="" addition="" to="" glutathione-related="" metabolites="" and="" dicarboxylic="" acids(figure="">

<0.05. metabolites="" are="" identified="" by="" their="" biochemical="" name="" and="" sorted="" by="" related="" metabolisms="" and="" subclasses="" of="" metabolites.="" for="" each="" metabolite,="" the="" individual="" expression="" of="" 6="" ctrl="" and="" 6="" cko="" mice="" normalized="" between="" 0="" and="" 1="" and="" the="" log2="" -transformed="" mean="" fold="" change="" of="" expression="" (log2fc)="" in="" cko="" versus="" ctrl="" mice="" are="" given,="" for="" both="" sexes.="" for="" calculation="" of="" the="" mean="" fc="" of="" expression,="" missing="" values="" (depicted="" in="" gray)="" have="" been="" replaced="" by="" the="" minimum="" value="" of="" both="" genotypes="" from="" the="" same="" sex."="" alt="Figure 5. Remodeling of the renal metabolome in cKO mice. (A) Venn diagrams representing the number of detected renal metabolites showing a significantly decreased or increased abundance in cKOm (in blue) or cKOf (in pink) mice versus Ctrl mice of the same sex. (B) Volcano plot representing the relative abundance of all detected metabolites in kidneys of cKOm versus Ctrlm. Metabolites depicted with blue dots are significantly less abundant, and transcripts depicted with red dots are significantly more abundant in kidneys of cKOm mice as compared with Ctrlm mice. The names of some representative metabolites are depicted using colors shared for related metabolites. (C and D) Heatmaps of metabolites showing a significantly decreased (C) or increased (D) abundance in cKO mice of both sexes as compared with Ctrl mice. A significant difference of abundance is considered when adjusted P value from a 2-way ANOVA referred to as " fdr"="" is=""><0.05. metabolites="" are="" identified="" by="" their="" biochemical="" name="" and="" sorted="" by="" related="" metabolisms="" and="" subclasses="" of="" metabolites.="" for="" each="" metabolite,="" the="" individual="" expression="" of="" 6="" ctrl="" and="" 6="" cko="" mice="" normalized="" between="" 0="" and="" 1="" and="" the="" log2="" -transformed="" mean="" fold="" change="" of="" expression="" (log2fc)="" in="" cko="" versus="" ctrl="" mice="" are="" given,="" for="" both="" sexes.="" for="" calculation="" of="" the="" mean="" fc="" of="" expression,="" missing="" values="" (depicted="" in="" gray)="" have="" been="" replaced="" by="" the="" minimum="" value="" of="" both="" genotypes="" from="" the="" same="" sex."="" width="480" height="320" border="0" vspace="0" style="width: 480px; height: 320px;">

Kev tsom xam txoj hauv kev sib koom ua ke (MetaboAnalyst 5.0) ntawm cov ntawv sau tseg thiab cov metabolites nthuav tawm ntau ntxiv lossis txo qis hauv ob lub raum ntawm cKOm nas piv nrog Ctrl nas tau txheeb xyuas 22 downregulated thiab 7upregulated pathways (Daim duab 6, A thiab B, feem; FDR<0.1; supplemental="" table="" 7).="" nitrogen="" metabolism,="" pyruvate="" metabolism,="" glycolysis,="" and="" gluconeogenesis="" were="" identified="" among="" the="" downregulated="" pathways="" while="" fatty="" acid="" degradation="" was="" identified="" among="" the="" upregulated="" pathways="" (figure="" 6,="" a="" and="" b,="" respectively).="" glutathione="" metabolism="" and="" retinol="" metabolism="" were="" present="" among="" both="" up-and="" downregulated="" pathways="" (figure="" 6,="" a="" and="" b).="" detailed="" analysis="" of="" transcripts="" and="" metabolites="" related="" to="" glutathione="" metabolism="" identified="" 16="" transcripts="" and="" 3="" metabolites="" with="" decreased="" abundance="" in="" kidneys="" of="" ckom="" mice="" (figure="" 6,="" c="" and="" d,="" respectively),="" along="" with="" 6="" transcripts="" and="" 8="" metabolites="" exhibiting="" increased="" abundance(figure="" 6,="" e="" and="" f,="" respectively).="" the="" oxidized="" form="" of="" glutathione(gssg)was="" present="" in="" ckom="" but="" not="" in="" ctrlm="" mice,="" thereby="" suggesting="" oxidative="" stress="" in="" ckom="" mice(figure="">

<0.1. the="" size="" of="" each="" dot="" depends="" on="" the="" percentage="" of="" all="" transcripts="" and="" metabolites="" ("compounds")="" of="" the="" pathway="" that="" are="" significantly="" affected="" in="" ckom="" mice.="" (c="" and="" d)="" relative="" expression="" of="" glutathione-related="" transcripts="" (c)="" and="" metabolites="" (d)="" significantly="" less="" abundant="" (fdr="" <="" 0.05)="" in="" ckom="" mice="" as="" compared="" with="" ctrlm="" mice.="" (e="" and="" f)="" relative="" expression="" of="" glutathione-related="" transcripts="" (e)="" and="" metabolites="" (f)="" significantly="" more="" abundant="" (fdr="" <="" 0.05)="" in="" ckom="" mice="" as="" compared="" with="" ctrlm="" mice.="" individual="" values="" from="" 6="" ckom="" and="" 6="" ctrlm="" mice="" are="" depicted="" after="" transformation="" from="" raw="" individual="" data:="" values="" of="" metabolite="" abundance="" have="" been="" divided="" by="" the="" median="" value="" of="" both="" genotypes,="" while="" values="" of="" transcript="" expression="" from="" both="" genotypes="" have="" been="" normalized="" between="" 0="" and="" 1.="" box="" and="" whiskers="" represent="" the="" mean="" and="" the="" sem,="" respectively.="" nd,="" not="" detected."="" alt="Figure 6. Joint analysis of transcriptional and metabolic changes in cKOm mice. (A and B) Scatter plot of significantly downregulated (A) or upregulated (B) metabolic pathways, based on a joint pathway analysis of both regulated transcripts and modulated metabolites in kidneys of cKOm versus Ctrlm mice. Pathways are sorted by their absolute impact, and a significant pathway regulation is considered when adjusted P value referred to as " fdr"="" is=""><0.1. the="" size="" of="" each="" dot="" depends="" on="" the="" percentage="" of="" all="" transcripts="" and="" metabolites="" ("compounds")="" of="" the="" pathway="" that="" are="" significantly="" affected="" in="" ckom="" mice.="" (c="" and="" d)="" relative="" expression="" of="" glutathione-related="" transcripts="" (c)="" and="" metabolites="" (d)="" significantly="" less="" abundant="" (fdr="" <="" 0.05)="" in="" ckom="" mice="" as="" compared="" with="" ctrlm="" mice.="" (e="" and="" f)="" relative="" expression="" of="" glutathione-related="" transcripts="" (e)="" and="" metabolites="" (f)="" significantly="" more="" abundant="" (fdr="" <="" 0.05)="" in="" ckom="" mice="" as="" compared="" with="" ctrlm="" mice.="" individual="" values="" from="" 6="" ckom="" and="" 6="" ctrlm="" mice="" are="" depicted="" after="" transformation="" from="" raw="" individual="" data:="" values="" of="" metabolite="" abundance="" have="" been="" divided="" by="" the="" median="" value="" of="" both="" genotypes,="" while="" values="" of="" transcript="" expression="" from="" both="" genotypes="" have="" been="" normalized="" between="" 0="" and="" 1.="" box="" and="" whiskers="" represent="" the="" mean="" and="" the="" sem,="" respectively.="" nd,="" not="" detected."="" width="480" height="320" border="0" vspace="0" style="width: 480px; height: 320px;">

Kev sib tw pub mis ntau tsis ua rau muaj kev ntxhov siab oxidative ntxiv hauv ckKOmice. Kev hloov pauv hauv glutathione-txog txoj hauv kev, depletion ntawm plasmalogens, thiab txo qis ntawm catalase pom zoo oxidative kev nyuaj siab thiab / los yog rov kho cov kab mob antioxidant sib txawv hauv ob lub raum ntawm cKOm nas. Txhawm rau kuaj cov kev xav no, peb tau twv cKOm nas nrog kev noj zaub mov muaj roj ntau (HFD) rau 4 lub lis piam. Qhov kev sib tw no tsis ua rau albuminuria tab sis nce zis ntim thiab tso zis ntawm calcium thiab urate. Tsis muaj qhov sib txawv ntawm qhov ntsuas ntshav plasma, suav nrog calcemia thiab uricemia (Cov Lus Qhia Ntxiv 8). Tsis muaj tag nrho cov kev hloov pauv morphological lossis lipid tsub zuj zuj hauv ob lub raum ntawm HFD-kev sib tw Ctrlm thiab cKOm nas (Daim duab 7A). Tag nrho thiab nonenzymatic antioxidant muaj peev xwm raws li kev soj ntsuam los ntawm Trolox assay (Daim duab 7B) nrog rau cov ntaub so ntswg theem ntawm malondialdehyde, ib qho cim ntawm polyunsaturated fatty acid peroxidation (Daim duab 7C), tsis txawv ntawm kev kho mob thiab genotypes. Qhov ntau ntawm lipid peroxidation khoom 4-hydroxynonenal (4-HNE) tau nce hauv Ctrlm nas kho nrog HFD piv nrog Ctrl nas ntawm kev tswj kev noj zaub mov, tab sis tsis muaj qhov sib txawv tau pom hauv cKOm nas kho nrog 2 kev noj haus (Daim duab 7D).