Front Psychiatry. 2018; 9: 81.
Idasindikizidwa pa intaneti 2018 Mar 12. do: 10.3389 / fpsyt.2018.00081
PMCID: PMC5858605
PMID: 29593587
Minho Lee,1,† Hyeyoung Cho,2,3,† Seung Hyun Jung,2,4 Seon-Hee Yim,2 Sung-Min Cho,2,3 Ji-Won Chun,5 Soo-Hyun Paik,5 Yae Eun Park,6,7 Dong Huey Cheon,7,8 Ji Eun Lee,7 Jung-Seok Choi,9 Dai-Jin Kim,5,* ndi Yeun-Jun Chung1,2,3,*
Kudalirika
Background
Kugwiritsa ntchito kwambiri intaneti komanso masewera a pa intaneti ndi vuto la matenda amisala lomwe limatchedwa kuti masewera a pa intaneti (IGD). Mafayilo osinthika a microRNA (miRNA) adanenedwa m'magazi ndi muubongo wamatenda a odwala omwe ali ndi vuto linalake lamaganizidwe amisala ndipo adanenedwa ngati biomarkers. Komabe, sipanapezeke malipoti onena za mbiri ya miRNA mu IGD.
Njira
Kuti tidziwe miRNA yolumikizidwa ndi IGD, tidasanthula ma profiles a miRNA of 51 samples (25 IGD and 26 control) pogwiritsa ntchito TaqMan Low Density miRNA Array. Pofuna kutsimikizika, tinachita kuchuluka kwa PCR olembedwa ngati ma 36 odziimira (20 IGD ndi 16 yolamulira).
Results
Mwakufufuza komanso kutsimikizira pawokha, tazindikira miRNAs zitatu (hsa-miR-200c-3p, hsa-miR-26b-5p, hsa-miR-652-3p) zomwe zidalembedwa mopepuka mu gulu la IGD. Anthu omwe ali ndi kusintha kwa miRNA konse anali ndi chiopsezo chachikulu cha IGD kuposa omwe alibe [osasinthika [OR) 22, 95% CI 2.29-211.11], ndipo ma ORs adakulitsa kumwa modalira ndi kuchuluka kwa maRNA. Mitundu yomwe yakonzedweratu ya maRNA atatuwa idalumikizidwa ndi njira zamatumbo. Tidasanthula kuchuluka kwa mapuloteni amitundu itatu yakumunsi yomwe yakulowera chakumadzulo ndipo tatsimikizira kuti mawu a GABRB2 ndi DPYSL2 anali okwera kwambiri pagulu la IGD.
Kutsiliza
Tawona kuti mawu a hsa-miR-200c-3p, hsa-miR-26b-5p, ndi hsa-miR-652-3p adalembedwa modabwitsa m'magulu a IGD. Zotsatira zathu zidzakhala zothandiza kumvetsetsa pathophysiology ya IGD.
Introduction
Kugwiritsa ntchito kwambiri intaneti komanso masewera opezeka pa intaneti sikuti kumangochitika kumayiko omwe ali ndi zida zambiri zopezera intaneti, koma vuto la matenda amisala lomwe limatchedwa kuti intaneti ya masewera (IGD) (1-3). Malinga ndi malipoti a miliri, kuchuluka kwa IGD mu achinyamata kumasiyana m'maiko onse, kuyambira 0.8 mpaka 26.7% (4). Makamaka, kafukufuku akuwonetsa kuchuluka kwa ziwonetsero zapamwamba kuposa 10% mwa achinyamata akumayiko ambiri aku Asia monga South Korea, China, Taiwan, Hong Kong, ndi Singapore (4). IGD imalumikizidwa ndi kusokonezeka mu chizindikiritso, maubwenzi a psycho-moyo, komanso moyo watsiku ndi tsiku; mwachitsanzo, kutsika kwamaphunziro kapena ntchito (4-7). IGD tsopano yaphatikizidwa mu Gawo III (Conditions for More Study) ya kukonzanso kwachisanu kwa Diagnostic and Statistical Manual of Mental Disorder (DSM-V) (8). Komabe, ngakhale ndizakufunika kwachikondwerero, ndizochepa zomwe zimadziwika pokhudzana ndi maselo a maselo a IGD.
Kafukufuku waposachedwa kwaposachedwa akuwonetsa ngati mtundu wa IGD (9, 10). Vink et al. anafufuza za kusiyana komwe kumagwiritsa ntchito Internet mokakamiza ndi mapasa a achinyamata a 5,247 monozygotic ndi dizygotic ku Netherlands Twin Register ndipo adanenanso kuti kusiyana kwa 48% kufotokozedwa ndi majini (9). Li et al. adawona mapawiri awiri a 825 amapasa achichepere aku China ndipo adati zomwe zimachokera ku genetic zidafotokoza 58-66% ya kusiyana (10). Momwemo, ma polymorphisms amtundu omwe akukhudzidwa ndi neurotransication, ccinition, ndi chidwi monga dopamine receptor D2 gene (DRD2), genate wa catecholamine-O-methyltransferase (COMT), genotonin transporter gene (5HTTLPR), ndi cholinergic receptor nicotinic alpha 4 gene (CHRNA4) adanenedwa kuti amagwirizana kwambiri ndi chizolowezi cha intaneti (11-13). Posachedwa, Kim et al. adayang'ana mitundu ya mitundu yopitilira 100 yogwirizana ndi kupanga, kuchitapo, ndi kagayidwe ka ma neurotransmitters mwa kuwunika kwotsatira kwa m'badwo wina ndipo adanenanso kuti rs2229910 ya NTRK3 gene limalumikizidwa ndi IGD (14).
Kuphatikiza pa majini, zimadziwikanso kuti neurobehavioral phenotypes imayang'aniridwa ndi ma RNA osakhazikitsa kuphatikizapo ma MicroRNA (miRNAs) (15, 16). miRNA ndi ma cell ochepa a RNA osakhala ndi ma cell a RNA (pafupifupi 20-23 nucleotides m'litali), omwe amaletsa mawu osonyeza mtundu wa protein-coding mwa ma mRNA amisala ndikuchita mbali yofunika kwambiri pamatenda a pathophysiological a matenda osiyanasiyana (17). Maumboni awonetsa kuti ma MRNA ndi ochulukirapo mkati mwa dongosolo lamanjenje la anthu (CNS) ndipo amachitapo kanthu bwino paziwonetsero zama genes awo omwe akhudzidwa ndi chitukuko ndi kusasitsa kwa dongosolo la CNS (15). Inde, kafukufuku waposachedwa waonetsa kuti mbiri ya miRNA imasinthidwa mu minofu ya ubongo ya odwala omwe ali ndi matenda amisala, kuwonetsa kuti mafotokozedwe awo akhoza kukhala opanga ma biomarkers pazovuta zamisala15, 16, 18). Mwachitsanzo, kudzera mu kusanthula kwa postmortem, Lopez et al. adati mawu a miR-1202, omwe amawongolera mafotokozedwe a metabotropic glutamate receptor-4 gene ndikuwonetseratu kuyankha kwa antidepressant, adagwiritsidwa ntchito kale mu preortal cortex tishu a odwala akulu okhumudwa (19). Ponena za kuwunika kwa biomarker, njirayi ili ndi malire podziwika chifukwa kuchita biopsy ya mins ya CNS pakuwunika sikungatheke. Popeza miRNAs imatha kupezeka m'magazi (plasma kapena seramu), maRNA oyendayenda ali ndi mwayi wotsimikizika monga osagwiritsa ntchito ma biomarkers omwe ali ndi vuto la neuropsychiatric. Komabe, mpaka pano, sipanapezeke kafukufuku wokhudza kuyendetsa ma profayilo a miRNA ku IGD. Kumvetsetsa bwino ma profiles a miRNA ozungulira kungathandize kumveketsa bwino kwamangidwe a IGD ndikuwongolera kutanthauzira kwachipatala.
Phunziroli, tidafuna kuzindikira zolembedwa za miRNA zogwirizana ndi IGD powona mosiyanasiyana maRNA a plasma miRNA pakati pa magulu a IGD ndi magulu owongolera komanso kuwunika zomwe zingawathandize.
Zida ndi njira
Nkhani Zophunzira
Tidayesa achinyamata a 3,166 (zaka za 12-18 zaka) pogwiritsa ntchito bao la DSM-V IGD. Mwa iwo, 251 (amuna a 168 ndi akazi a 83) adapezeka kuti ali ndi IGD molingana ndi njira ya DSM-V (8). Anthu angapo a 91 (ma 49 IGDs ndi 42 zowongolera) adapereka chidziwitso phunziroli. Mwa iwo, anthu anayi adasiyidwa malinga ndi njira zopatula. Pomaliza, anthu a 87 (45 IGD maphunziro ndi 42 owongolera athanzi) adalembetsa nawo phunziroli. Pakati pawo, omwe anali nawo pagululi la 51 (odwala 25 IGD ndi ma 26 zowongolera) adalembedwa ngati zomwe zapezeka kuchokera ku 2014 mpaka 2016. Otsatira ena a 36 (odwala a 20 IGD ndi zowongolera za 16) adalembedwa ngati chitsimikiziro chodziyimira payokha chochokera ku 2016. Onse omwe anali nawo anali anthu aku Korea, omwe adalembetsa kuchokera ku Seoul St. Mary's Hospital (Seoul, South Korea) ndi Seoul National University Boramae Hospital (Seoul, South Korea). Onse omwe adachita nawo kafukufuku adawunikiridwa ndi dokotala wamisala yochokera ku Korea Kiddie Ndondomeko ya Ogwirizana ndi Matenda ndi Schizophrenia (K-SADS-PL) (20). Ophunzira onse adamaliza gawo la block Design ndi Vocabulary la Korea-Wechsler Intelligence Scale for Children, 4th edition (K-WISC-IV) (21). Kusasinthika kuyesedwa ndi Barratt Impulsiveness Scale (BIS) (22). Miyezo ya Behavioral Inhibition System (BInS) ndi Behavioral Activation System (BAS) anayeza kuti ayesere kukula kwa umunthu (23). Njira zochotsedwera zinaphatikizapo zovuta zam'mbuyomu kapena zamakono zamankhwala (mwachitsanzo, matenda a shuga), matenda amitsempha (mwachitsanzo, zovuta za kugunda, kuvulala pamutu), matenda amisala (mwachitsanzo, kusokonezeka kwakukulu, nkhawa), kubweza m'maganizo, kapena kuzunza zilizonse. , fodya, cannabis, mowa). Zomwe zimachitika phunziroli mwachidule Table1.1. Kafukufukuyu adavomerezedwa ndi Institutional Review Board of the Catholic University Medical College of Korea (MC16SISI0120). Onse omwe atenga nawo mbali ndi makolo awo adapereka chidziwitso cholembedwa.
Gulu 1
Makhalidwe ambiri pazophunziridwa.
| anapeza | Kuvomereza | Kuphatikiza | |||||||
|---|---|---|---|---|---|---|---|---|---|
| Control | IGD | Pchizindikiro | Control | IGD | Pchizindikiro | Control | IGD | Pchizindikiro | |
| N | 26 | 25 | 16 | 20 | 42 | 45 | |||
| Zaka (zaka) | |||||||||
| Pakati (min-max) | 13 (12 - 17) | 13 (12 - 15) | 0.759 | 15 (13 - 18) | 14.5 (12 - 18) | 0.628 | 14 (12 - 18) | 14 (12 - 18) | 0.509 |
| Ma ola osewera pa intaneti (h) | |||||||||
| Pakati (min-max) | 5.25 (2 - 17) | 18 (6 - 46) | 1.27E − 6a | 5.5 (2 - 23) | 8 (1 - 112) | 0.374 | 5.5 (2 - 23) | 14 (1 - 112) | 1.63E − 5a |
| Chuma chanyumba mwezi (KRW miliyoni) | |||||||||
| Pakati (min-max) | 5 (1 - 9) | 3 (1 - 9) | 0.588 | 4 (4 - 4) | 2 (2 - 2) | 1.000 | 5 (1 - 9) | 3 (1 - 9) | 0.460 |
| Maphunziro (zaka) | |||||||||
| Pakati (min-max) | 8 (7 - 9) | 8 (7 - 9) | 0.584 | 12 (12 - 12) | 6 (6 - 13) | 0.305 | 8 (7 - 12) | 8 (6 - 13) | 0.269 |
| K-WISC: kapangidwe ka block | |||||||||
| Pakati (min-max) | 10.5 (4 - 17) | 10 (4 - 16) | 0.544 | 10 (3 - 16) | 12.5 (4 - 15) | 0.125 | 10 (3 - 17) | 11 (4 - 16) | 0.598 |
| K-WISC: mawu | |||||||||
| Pakati (min-max) | 9 (5 - 17) | 7 (5 - 13) | 0.174 | 9.5 (8 - 15) | 11.5 (5 - 15) | 0.595 | 9 (5 - 17) | 9 (5 - 15) | 0.527 |
| KS | |||||||||
| Pakati (min-max) | 24 (17 - 36) | 37 (22 - 51) | 3.81E − 6a | 29 (17 - 34) | 59 (22 - 108) | 1.2E − 5a | 25 (17 - 36) | 40 (22 - 108) | 2.05E − 10a |
| bis | |||||||||
| Pakati (min-max) | 63 (35 - 75) | 67.5 (45 - 81) | 0.080 | 61 (45 - 79) | 63 (32 - 82) | 0.835 | 62 (35 - 79) | 65 (32 - 82) | 0.240 |
| anatsalira | |||||||||
| Pakati (min-max) | 31 (15 - 40) | 31 (13 - 51) | 0.558 | 36.5 (22 - 48) | 34 (27 - 52) | 1.000 | 32 (15 - 48) | 34 (13 - 52) | 0.637 |
| MABATA | |||||||||
| Pakati (min-max) | 18 (10 - 26) | 17.5 (13 - 27) | 0.642 | 18.5 (12 - 25) | 20 (13 - 21) | 0.138 | 18 (10 - 26) | 19 (13 - 27) | 0.302 |
IGD, odwala pa intaneti omwe ali ndi vuto la masewera; KS, Kulembetsa kwa Mtanda Waku Internet wa Korea; BIS, Barratt Impulsivity Scale; BAS, Njira Yogwirizira Yogwira Ntchito; BInS, Behavioral Inhibition System; KRW, Won waku Korea.
aP <0.05 (Mayeso a Mann-Whitney – Wilcoxon).
TaqMan Low Density miRNA Array (TLDA) Kafukufuku
Magazi oyipitsa adasonkhanitsidwa kuchokera kwa aliyense wogwira nawo ntchito ndikuusamutsira ku labotale mkati mwa 4 h kuti muchepetse magazi a m'magazi. Choyerekezedwacho chinali chapakati pa 3,000 rpm pa 10 mphindi kutentha kwa chipinda. Kenako, supernatant (plasma wosanjikiza) amatengedwa popanda kuipitsa maselo a m'magazi. MaRNA oyendayenda adachotsedwa pogwiritsa ntchito TaqMan miRNA ABC Chiyeretso Kit (Human Panel A; Thermo Fisher Science Science, Waltham, MA, USA) malinga ndi malangizo a wopanga. Mwachidule, 50 µL ya zitsanzo za plasma ndi 100 µL ya buC ya buger idasakanizidwa. Pambuyo poyeserera ndi mikanda yamagnetic yotsutsana ndi miRNA, mikanda yozungulira yozungulira yozungulira imatulutsidwa kuchokera ku mikanda ndi 100 µL ya buution yopangira. Mu gawo lazopeza, ma 381 miRNA adayesedwa kuchokera ku zitsanzo za ma 51 plasma (ma 25 IGDs ndi ma 26 oyang'anira) pogwiritsa ntchito TaqMan miRNA ABC Kuyeretsa Kit (Human Panel A; Thermo Fisher Science Science, Waltham, MA, USA) malinga ndi malangizo a wopanga. Megaplex reverse transcript ndi pre-amplation reaction zimachitika pofuna kuwonjezera kuchuluka kwa cDNA pakuwunikira kwa miRNA pogwiritsa ntchito MegaplexPreAmp Primers Human Pool A ndi TaqManPreAmp Master Mix (Thermo Fisher Science). Pulogalamu ya TLDA A v2.0 (Thermo Fisher Science Scient) idayendetsedwa pa ViiA7 yeniyeni PCR dongosolo (Thermo Fisher Science Science) kuti iwunikenso kufotokozera kwa maRNA. Zambiri zosawerengeka zidakonzedwa pogwiritsa ntchito ExpressionSuite Software v1.0.4 (Thermo Fisher Science Science) kuti zitsimikizire Ct pazomwe zili ndi maRNA.
Kusanthula Kwathunthu kwa TLDA
Tidayesa kaye poyambira kuzungulira (Ct mtengo) wa miRNA iliyonse. miRNAs yokhala ndi Ct mtengo> 35 amawerengedwa ngati osawoneka ndipo sanaphatikizidwe pakuwunika komwe kudachitika. Mitengo yonse ya Ct idasinthidwa kukhala mtengo wa Ct wa miR-374b (valueCt mtengo), imodzi mwama miNA omwe amadziwika bwino kwambiri omwe amafalikira m'madzi a m'magazi (24). Chiwerengero chosinthika cha log2 (ΔΔCt value) cha mawerengedwa chimawerengeredwa pogwiritsa ntchito mitundu yoyenera yolemba ngati calibrator phukusi la HTqPCR ku Bioconductor (25). Quantification wachibale (RQ) ya chandamale chilichonse cha miRNA amatanthauzidwa kuti 2-DACC. Pakuyesa kopitilira muyeso wa kusiyana pakati pa magulu awiri, tinagwiritsa ntchito kusanthula kosasintha (SVA) kuti tipeze ma heterogeneities monga zotsatira za batchi pazoyeserera pogwiritsa ntchito sva phukusi ku Bioconductor (26). maRNA ndi a P-phindu <0.05 idawonedwa kuti ndi yosiyana kwambiri pakati pa magulu awiri.
Kuwongolera Kwakutsogolo kwa Gene
Pakuwunika kofufuza kwamtundu, tinagwiritsa ntchito ToppFun ku ToppGene Suite (27) kuti liphunzitse bwino Gene Ontology (GO) (28) njira, njira, ndi mawu. Monga momwe mungagwiritsire ntchito njirayi, tinagwiritsa ntchito 1,230 yolosera za mtundu wa miRNA. Kusanthula kwa Pwayway kunagwiritsidwa ntchito kuti apeze njira zazikulu zamitundu yomwe yakonzedwa molingana ndi KEGG, BioCarta, Reactome, GeneMAPP, ndi MSigDBin njira zapamwamba za ToppGene. Kufunika kwa mawu olemeretsa ogwira ntchito adatsimikizika potengera momwe Bonferroni-adasinthira P-chachidziwikire.
Kukhazikika Kotsatsa kwa PCR (qRT-PCR) Kuvomerezeka ndi Kuchita Zinthu
Kuti mutsimikizire ma 10 miRNA omwe adafotokozedwa mosiyanasiyana mu gawo lazopeza, qRT-PCR idachitika pogwiritsa ntchito TaqMan MicroRNA Assay (miR-15b-5p, #000390; miR-26b-5p, #000407; miR-29b # 3; miR-000413b-125p, #5; miR-000449c-200p, #3; miR-002300-337p, #5; miR-002156-411p, #5; miR-001610-miR -UMNUMX; -423p, #5; and miR-002340-483p, #5) ndi ViiA002338 system (Life Technologies) molingana ndi protocol ya wopanga. Ma nanogram khumi a RNA yathunthu adasinthidwa kukhala woyamba-strand cDNA ndi primers enieni a miRNA ogwiritsa ntchito TaqMan MicroRNA Reverse Transcript Kit (#652, Life Technologies), yotsatiridwa ndi PCR yeniyeni yokhala ndi TaqMan Probes. RQ ya miRNA iliyonse imatanthauzidwa kuti 3−ΔCt, komwe ΔCt ndi kusiyana pamizere yolowera yachitsanzo, yofanana ndi yolimbana ndi miRNA yotsika (miR-374b-5p, #001319). Zotsatira zonse za PCR zidachitika m'mitundu itatu, ndipo makhalidwe awo a Ct adachitidwa. Tidawerengera log2 fold-change ratio (ΔΔCt) ya miRNA iliyonse chimodzimodzi ndi kuwunika kokhazikika. Mayeso omwe sanali a parametric Mann-Whitney-Wilcoxon adayesedwa kuti ayesere kusiyana kwa mawu a maRNA m'magulu awiri okhala ndi cholowa PChiwonetsero cha 0.05.
Western Blot Analysis
Gawo lililonse la seramu lidatha mapuloteni apamwamba a 14 okwera kwambiri (albumin, immunoglobulin G, immunoglobulin A, serotransferrin, haptoglobin, alpha-1 macroglobulin, alpha-2ote Alugulin Aphin Apholopotein, alpha-1ote acid. , apolipoprotein A-II, C3, ndi transthyretin) pogwiritsa ntchito mzati wa MARS-14 (4.6 × 50 mm, Agilent Technology, Santa Clara, CA, USA) kusanachitike kumadzulo kwa blot. Gawo losasunthika lomwe linapezedwa kuchokera kumzati wa MARS-14 lidalimbikitsidwa pogwiritsa ntchito fayilo ya Amicon Ultracel-3 centrifugal (3 kDa cutoff), kenako ndende ya protein idatsimikizika pogwiritsa ntchito njira ya bicinchoninic acid. Ndalama zomwezo (kuchokera ku 10 mpaka 30 µg) zowongolera ndi ma IGD seramu zitsanzo zidasiyanitsidwa pa 4-20% Mini-PROTEAN TGX precast gel (Bio-Rad, CA, USA) ndikusamutsira ku membala wa polyvinylidene difluoride. Pambuyo pake, nembanemba idatsekedwa mu TBS-T (190 mM NaCl, 25 mM Tris-HCl, pH 7.5, ndi 0.05% Tween 20) yokhala ndi mkaka wopanda mkaka wama 5% wopanda mafuta pamafiriji a 30. Zomwe zimapangidwa panthawiyo zidapangidwa ndi ma antibodies oyamba motsutsana ndi DPYSL2 (1: 500, Novus Biologicals, Littleton, CO, USA), GABRB2 (1: 1000, Abcam, Cambridge, MA, USA), ndi CNR1 (1: 100: 4: 1: 500: 15: 1: 500: 5: 4: 1: 1,000: 1: 1,000: 1: 1: XNUMX: XNUMX: XNUMX: XNUMX: XNUMX: XNUMX , Inc., Santa Cruz, CA, USA), DUSPXNUMX (XNUMX: XNUMX, MybioSource, San Diego, CA, USA), ndi PIXNUMX (XNUMX: XNUMX, MybioSource, San Diego, CA, USA) ku TBS-T yokhala ndi XNUMX % mkaka wowuma wopanda mafuta ku XNUMX ° C usiku wonse, kenako ndi ma antibodies oyenerera omwe ali anti-mouse (XNUMX: XNUMX, Santa Cruz Biotechnology) kapena anti-akalulu a mbuzi (XNUMX: XNUMX, Cell Signaling, Beverly, MA, USA ) cholumikizidwa ndi horseradish peroxidase pamoto kutentha kwa XNUMX h. Kuzindikira kwa chizindikiro kunachitika pogwiritsa ntchito chemiluminescence ndi ECL reagent (GE healthcare, Piscataway, NJ, USA). Tinafotokozera zotsatira zakumadzulo pogwiritsa ntchito pulogalamu yaulele ya TotalLab XNUMXD (non-linear Dynamics, Newcastle upon Tyne, UK). Kenako, kuchuluka kwa ma densitometry kuwerengera kudawerengera gawo la densitometry mulingo uliwonse monga wafotokozedwera kwina (29). Monga njira yolamulira masanjidwe, seramu yoyesedwa yochokera ku 46 IGD ndi zitsanzo zowongolera zinagwiritsidwa ntchito pakuyesera kulikonse. Kufunikira kwa Statist kunatsimikiziridwa pogwiritsa ntchito mayeso osakhala a parametric Mann - Whitney-Wilcoxon PChiwonetsero cha 0.05.
Results
Makhalidwe a Nkhani Zophunzira
Zowerengera za anthu komanso zamankhwala pazophunzirazi zikuwonetsedwa mu Gome Table1.1. Tikayerekezera IGD ndikuwongolera magulu malinga ndi Korea Internet Addiction Proneness Scale (K-Scale) monga tafotokozera kwina (20, 30), gulu la IGD linawonetsa mtengo wapakatikati wa K-Scale kuposa gulu lolamulira (37 vs. 24, P = 3.81 × 10-6) (Tebulo (Table1) .1). Nthawi yama Median sabata yomwe imagwiritsidwa ntchito pamasewera pa intaneti mu gulu la IGD inali yayitali kwambiri kuposa nthawi yowongolera (18 vs. 5.25 h, P = 1.27 × 10-6). Pomwe panalibe kusiyana kwakukulu pakati pa magulu awiri azaka, ndalama zapanyumba pamwezi, nthawi yayitali yophunzira, mapangidwe a block, ndi zotsatira zowoneka bwino za K-WISC, BIS, BInS, ndi BAS.
Kusiyanitsa Kwina miRNA Pakati pa IGD ndi Maulamuliro
Kuti tidziwe miRNA yolumikizidwa ndi IGD, tidatengera njira ziwiri (zopezera komanso zotsimikizika pawokha). Mapangidwe apangidwe ndi malingaliro onse akuwonetsedwa mu Chithunzi S1 mu Zowonjezera Zowonjezera. Mu gawo lazopeza, tidasanthula ma profiles a miRNA of 51 sampuli (ma 25 IGDs ndi ma 26 zowongolera) pogwiritsa ntchito mtundu wa miRNA wokhala ndi ma 384 miRNA. Ziwonetsero zama 10 miRNA zinapezeka kuti ndizosiyana kwambiri pakati pa IGD ndi magulu owongolera (Gome (Table2) .2). Magawo omasulira a 10 miRNA awa akuwonetsedwa pa Chithunzi Chithunzi1.1. Mwa iwo, awiri (hsa-miR-423-5p ndi hsa-miR-483-5p) adalembetsa ndipo asanu ndi atatu (hsa-miR-15b-5p, hsa-miR-26b-5p, hsa-miR-29p-XN hsa-miR-3b-125p, hsa-miR-5c-200p, hsa-miR-3c-337p, hsa-miR-5-411p, gulu la hsa-miR-5-652p) lidasungidwa m'gulu la IGD.
Gulu 2
Kusintha kofotokozedwa mosiyanasiyana ma microRNA (miRNAs) ndi kusintha kwa khola.
| miRNA | anapeza | Kuvomereza | Kuphatikiza | |||
|---|---|---|---|---|---|---|
| Pchizindikiro | Kusintha kwamitundu | Pchizindikiro | Kusintha kwamitundu | Pchizindikiro | Kusintha kwamitundu | |
| hsa-miR-15b-5p | 0.033 | 0.829 | 0.694 | 1.119 | 0.381 | 0.947 |
| hsa-miR-26b-5pa | 0.008 | 0.871 | 0.049 | 0.841 | 0.013 | 0.857 |
| hsa-miR-29b-3p | 0.005 | 0.400 | 0.560 | 1.187 | 0.089 | 0.647 |
| hsa-miR-125b-5p | 0.021 | 0.582 | 0.290 | 0.950 | 0.069 | 0.723 |
| hsa-miR-200c-3pa | 0.011 | 0.336 | 0.003 | 0.542 | 2.93 × 10-5 | 0.415 |
| hsa-miR-337c-5p | 0.009 | 0.385 | 0.582 | 0.872 | 0.020 | 0.553 |
| hsa-miR-411-5p | 0.004 | 0.322 | 0.336 | 1.282 | 0.158 | 0.595 |
| hsa-miR-423-5p | 0.026 | 1.387 | 0.189 | 0.955 | 0.518 | 1.175 |
| hsa-miR-483-5p | 0.018 | 1.861 | 0.765 | 1.413 | 0.211 | 1.647 |
| hsa-miR-652-3pa | 0.019 | 0.715 | 0.049 | 0.877 | 0.011 | 0.782 |
amiRNAs zimasinthika kwambiri pakupezeka komanso kutsimikizika kumayikidwa mosasintha.
QRT-PCR Chitsimikiziro cha Amaunika maRNA
Kuti tiwonetsetse ma 10 ofuna kusintha mamina a 20, tidayeseza qRT-PCR ndimayimidwe odziyimira pawokha (16 IGDs and 1 control) (Table S200 in Supplementary Material). Zitatu mwa miRNAs izi (hsa-miR-3c-26p, hsa-miR-5b-652p, ndi hsa-miR-3-XNUMXp) zidalembedwa mozama mu gulu la IGD loyikira (Table2) .2). MaRNA ena atatu (hsa-miR-337c-5p, hsa-miR-125b, ndi hsa-miR-423-5p) adalembedwanso m'gulu la IGD koma osachita kwambiri. Kusala miRNAs zinayi (hsa-miR-15b-5p, hsa-miR-29b-3p, hsa-miR-411-5p, ndi hsa-miR-423-5p) zidafotokozedwa motsutsana muyeso wokhazikitsidwa. Tidaphatikiza zopezedwa komanso zotsimikizika (zowerengeka za 45 IGD maphunziro ndi zowongolera za 42), miRNA itatu yotsimikizika inali yofunika kwambiri (Tebulo (Table2) .2). Zambiri, malo a chromosomal, machitidwe okhwima, ndi kuchuluka kwa mawu mu CNS mwa miRNA atatuwa akupezeka mu Table S2 ku Supplementary Material.
Zotsatira za Synergistic Zomwe Zimasinthidwa Pomwe Pano pa Ma MiRNA Atatu pa IGD Ngozi
Kuti tiwone momwe zotsatira za maRNA atatuwo tawonera, magawo atatu a magawo anayi (omwe anali ndi 0, 1, 2, kapena 3 miRNA kusintha). kusintha kwa miRNA kumatanthauziridwa ndi mtengo wa RQ monga tafotokozera mu Gawo "Zida ndi njira. ”Chifukwa onse atatu okhala ndi maRNA adalembedwa mgulu la IGD, miRNA yomwe mtengo wake wa RQ womwe unali m'munsi mwake udasinthidwa ngati wosinthika. Zambiri mwatsatanetsatane wa mtengo wamtundu uliwonse wa RQ pazopangira maRNA atatuwo zimapezeka mu Table S3 mu Supplementary Material. Pa gulu lililonse, zosamvetsetseka zimawerengeredwa ngati chiwerengero cha zowongolera kukhala zofanana ndi za IGD, ndiye kuti OR iliyonse imawerengeredwa pogawa zovuta za gulu lililonse mosagwirizana ndi gulu lililonse popanda kusintha kwa miRNA. Anthu omwe ali ndi kusintha kwa miRNA atatu adawonetsa kuwopsa kwa 22 nthawi zambiri kuposa omwe alibe kusintha kwa miRNA (OR 22, 95% CI 2.29-211.11). ORs adawonetsa kuwonjezeka ndi kuchuluka kwa miRNA yosinthika kuchokera ku 0 kupita ku 3 (r2 = 0.996) (Chithunzi (Figure22).
GO ndi Kusanthula kwa Pathway kwa Target Genes ya maRNA
Kuti mumvetsetse momwe zigawo zitatu za maRNA zimalembedwera kwambiri m'gululi la IGD, mitundu yawo yolosera idanenedweratu kuti adzagwiritsa ntchito miRWalk 2.0 database (31). Mitundu yonse ya 1,230 idanenedweratu kuti ikhale pansi pamipanda ya ma algorithms anayi (miRWalk, miRanda, RNA22, ndi Targetscan) pogwiritsa ntchito nkhokwe ya miRWalk (32-34) (Table S4 mu Zowonjezera Zowonjezera). Gene anakhazikitsa kuwunikira kolimbikitsa kugwiritsa ntchito ToppFun ku ToppGene Suite kuwonetsa kuti mitundu yomwe akufuna kutsata ya maRNA amenewa idalumikizidwa kwambiri ndi njira zachitukuko monga "chiwongolero cha Axon" ndi mawu a Go monga "neurogeneis" (Table S5 in Supplementary Material).
Kulingalira kwa Chiwonetsero cha Chiyembekezo Cholosera
Mwa mitundu yotsika yomwe ili pamunsi mwa mitundu itatu ya maRNA, 140 idanenedweratu nthawi imodzi kwa miRNA ziwiri kapena zingapo (Table S4 in Supplementary Material). Kuti tiwone ngati kuchuluka kwa mapuloteni am'magawo am'munsi otsika ndizosiyana pakati pa IGD ndi magulu owongolera, tinasankha mitundu ya 2 (DUSP4 ndi PI15), zomwe zimanenedweratu kuti zigawo zotsika za mitundu yonse ya 3 miRNA ndi mitundu yowonjezera ya 3 (GABRB2, DPYSL2ndipo CNR1) kuchokera kwa omwe ananenedweratu za 2 miRNAs ndikufufuza zakumadzulo ndi zitsanzo za plasma zochokera ku 28 IGDs ndi 28 zowongolera zomwe zikupezeka poyesa. Tidafanizira zomwe zigwiriziro zisanu zomwe zikupezeka pakati pa IGD ndikuwongolera magawo poyesa kulimba kwa gulu ndi malo monga amafotokozera kwina (29). Pakati pawo, magulu omasulira a DPYSL2 (28 IGDs and 28 control, P = 0.0037) ndi GABBR2 (27 IGDs ndi 28 zowongolera, P = 0.0052) anali okwera kwambiri pagulu la IGD (Chithunzi (Figure3) .3). Komabe, sitinathe kuwona zosiyana za CNR1 (P = 0.0853), DUSP4 (P = 0.5443), ndi PI15 (P = 0.6346).
Kukambirana
Adanenedwa kuti miRNA ikuphatikizidwa ndi chitukuko cha neuronal (35, 36), ndi mawonekedwe osiyanitsa amatsenga a ubongo miRNA amawonedwa mu matenda amisala monga schizophrenia (37). Chifukwa chake, zikuwonekeratu kuti kufalitsa ma profayer a miRNA kungakhale kothandiza kwa ma biomarkers a IGD. MaRNA oyendayenda afotokozedwa ngati ma biomarkers amisala yama neuropsychiatric yamavuto osiyanasiyana (38-40); Komabe, ma molekyulu omwe adayambitsa chitukuko cha IGD sanadziwikebe kwambiri ngakhale akufunika m'chipatala komanso chikhalidwe. Makamaka, sipanapezeke maphunziro pa ma IGNA ogwirizana ndi IGD. Zolinga za kafukufukuyu zinali ziwiri. Choyamba, tinayesa kupeza ma plazma miRNA ophatikizidwa ndi IGD. Chachiwiri, tidawunikira tanthauzo la kubadwa kwa miRNA ofuna kudziwa kuwonetsa mapuloteni ndi GO amitundu yomwe ikutsikira. Kupitilira kuwunika kwa miRNA ma profiles ndi kutsimikizika kwa kutsika kwa ofuna, tapeza kuti mawu atatu a maRNA (hsa-miR-200c-3p, hsa-miR-26b-5p, ndi hsa-miR-652-3p) anali otsika kwambiri mwa odwala a IGD kuposa owongolera. Ngakhale mawonekedwe a anthu ena asanu ndi awiri a MRNA sanatchulidwe pamawuwo, zitha kukhala zabodza chifukwa cha zitsanzo zazing'ono pakuphunziraku. Pazidziwitso zathu, iyi ndi yoyamba lipoti lothekera kuti mafayilo amwazi a MRNA atha kukhala othandiza pa biomarkers a IGD. Kuphatikiza zolemba zitatu za miRNA zitha kukhala chida chovuta kwambiri chodziwitsa anthu omwe ali pachiwopsezo cha IGD.
Ma miRNA omwe adadziwika mu phunziroli akuti ali ndi vuto la mitsempha yosiyanasiyana. Chithunzi cha hsa-miR-200c m'magazi akuti chakhala chikucheperachepera pamavuto angapo amisala monga psyizophrenia (41) ndi malo akulu okhumudwitsa (42). miR-200c idanenedwa kuti ikuwonetsedwa kwambiri pazigawo za synaptic kuposa kuchuluka kwa forebrain (43) komanso kuphatikizidwa ndi kufa kwa neuronal cell (44). Kutengera ndi malipoti am'mbuyomuwa, miR-200c imagwira nawo ntchito mu neurodevelopment ndipo imatha kulumikizidwa ndi vuto la neuropsychiatric ngati mawu ake ali operewera. Kafukufuku angapo adalimbikitsa kuyanjana pakati pa miR-652 ndi chiopsezo cha zovuta za neuropsychiatric. Zofanana ndi njira yathu, kudziwa ma biomarker a schizophrenia, Lai et al. adachita kusanthula kwa TLDA ndi odwala a schizophrenia komanso kuwongolera nthawi zonse, ndipo adapeza kuti miRNA zisanu ndi ziwiri kuphatikiza hsa-miR-652 zidafotokozedwa mosiyanasiyana mwa odwala a schizophrenia (45). Pakufufuza kwotsatira, adapanga njira yolosera pogwiritsa ntchito miRNA expression data ndipo adasiyanitsa bwino schizophrenia pakuwongolera koyenera (46). Mawu osintha a hsa-miR-652 adawonedwanso mu zidakwa (47). Hsa-miR-26b adapezeka kuti adamulowetsa pakusiyanitsa kwa ma cell a neuronal (48). Perkins et al. adanenanso kuti hsa-miR-26b adayang'aniridwa mu preortalal cortex ya odwala a schizophrenia (49).
Ngakhale palibe umboni wachindunji wothandizirana pakati pa mawu omwe ali ndi ziwonetserozi za ma maRN ndi ma pathophysiology a IGD, titha kudziwa kuti kusowa kwa miRNA uku kungaphatikizidwe ndi pathophysiology ya IGD kutengera malipoti angapo am'mbuyomu amtundu wapansi womwe tidaneneratu . Mitundu ina yotsika ya mitundu itatu ya maRNA monga GABRB2, CNR1, NRXN1ndipo DPYSL2 akuti akuphatikizidwa ndi vuto la neuropsychiatric. Gamma-aminobutyric acid (GABA) ndi chachikulu choletsa ma neurotransmitter mu CNS. Kuwonongeka kwa GABA receptor, kumakhudzidwa ndi zovuta za neuropsychiatric kuphatikizapo chizolowezi, nkhawa, komanso kukhumudwa (50), zomwe ndizophatikizanso zazikulu za IGD (8). Ma polymorphisms amtundu wa GNA receptor genes akuti akuphatikizidwa ndi mankhwala osokoneza bongo ndi schizophrenia (51, 52). Dihydropyrimidinase-monga 2 (DPYSL2) ndi membala wa banja la protein yotsutsa ya collapsin, yemwe amathandizira pamsonkhano wa microtubule, siginecha ya synaptic, ndikuwongolera kukula kwa axonal. Chifukwa chake, molekyulu'yi akuti ndiwothetsera mavuto a matenda amisala (53, 54). Polymorphism mu DPYSL2 jini amadziwikanso kuti amayanjana ndi vuto la kumwa mowa (55). Malipoti apakale ndi deta yathu ikuwonetsa kuti kuwonjezeka kwa GABRB2 ndi DPYSL2, kutsikira kwa zotsika za miRNAs, zomwe zikuwoneka kuti zili ndi tanthauzo pa pathogenesis yamavuto am'mimba kuphatikizira IGD. Cannabinoid receptor mtundu 1 (CNR1) ndi presynaptic heteroreceptor yomwe imasintha kumasulidwa kwa neurotransmitter komanso zosokoneza mu signn cannabinoid zimagwirizanitsidwa ndimatenda osiyanasiyana a neuropsychiatric (56). Ma polymorphism amtundu wa CNR1 jini amadziwika kuti amalumikizidwa ndi kudalira mankhwala ku Caucasians (57). Mu mtundu wa rat, activation ya ventral hippocampus CNR1 imasokoneza chikhalidwe chazomwe zimachitika komanso kuzindikira (58). Kusintha kwa ma genetic mu banja la NRXN amadziwika kuti akukumana ndi mavuto osiyanasiyana amitsempha yama bongo kuphatikizira kukopeka (59).
Kuti tipeze tanthauzo la kubadwa kwa anthu atatu omwe ali ndi maRNA mwanjira yowongoka, tidasanthula kuchuluka kwa mapuloteni amtundu wawo wotsikira. Chifukwa cha kupezeka pang'ono kwa zitsanzo za plasma, za ovomerezeka a 140 (zonenedweratu monga kutsika kwa 2 kapena miRNAs), tidasanthula zigawo za 5 (GABRB2, DPYSL2, CNR1, DUSP4, ndi PI15) kuchokera kumadzulo kwa blotNXXBB ndipo DPYSL2 anali wamkulu kwambiri pagulu la IGD. Malipoti apakale ndi deta yathu ikuwonetsa kuti kuwonjezeka kwa GABRB2 ndi DPYSL2, zigawo zotsikira za maRNA omwe adalembedwa, atha kukhala ndi tanthauzo ku pathogenesis yamavuto a neuropsychiatric kuphatikiza IGD. Zotsatira za kusanthula kwa njira ya GO ndi njira za njira za chitukuko cha neural zimathandiziranso kutanthauzira kwa ma neurobiological omwe amalembera miRNA. Chosangalatsa china chinali zotsatira za kusinthana kwofanana kwa maRNA. Anthu omwe ali ndi vuto la allRNUMX miRNAs adawonetsa chiopsezo cha 2 nthawi zochulukirapo kuposa omwe alibe zolembedwa, ndipo ma OR amakula modalira. Ngakhale CI pazosinthidwa zitatuzi zinali zazikulu chifukwa cha kukula kwachitsanzo, mawonekedwe abwino omveka (r2 = 0.996) imathandizira kulumikizana kwa ma miRNA atatu.
Ngakhale tinapeza zolemba za IGD zogwirizana ndi miRNA komanso anthu omwe ali ndi kusintha kwa miRNA yonse anali ndi chiopsezo cha 22 nthawi zambiri kuposa omwe sanasinthidwe ndi miRNA, pali zoperewera zingapo mu kafukufukuyu. Choyamba, kukula kochepa kakulidwe kumachulukitsa mwayi wosowa zina zofunikira za maRNA. Chachiwiri, popeza deta yathu sinali yokwanira kufotokoza ngati mapulogalamu a plasma miRNA ali ndi chifukwa kapena sachita chilichonse, sitingathe kutsimikizira maumbidwe a zolembedwazi zosagwiritsidwa ntchito pachipatala. Powonjezera miRNA ndikuwunika kwawo komwe kumatsika ndikugwiritsa ntchito ubongo waumunthu kuchokera ku banki ya ubongo kungapereke yankho lolunjika. Kusanthula kwa minyewa yaubongo ndi mtundu wa nyama yamagetsi kungakhalenso kothandiza. Chachitatu, chifukwa cha kupezeka pang'ono kwa zitsanzo za plasma, tidangoyang'ana mamolekyulu asanu ozama. Kuwunikira zigawo zina zotsika kwambiri ndi zitsanzo zazikulu kumathandizira kuti mumve mozama mamangidwe a IGD.
Mwachidule, kudzera pa genome-wide screen of miRNA expression profiles and itself solidified, tapeza ma IGNA atatu omwe ali ndi-IGR (hsa-miR-200c-3p, hsa-miR-26b-5p, hsa-miR-652-3p). Mitundu yawo yambiri yotsikira imanenedwa kuti ili ndi vuto la mitsempha yosiyanasiyana, ndipo kuyesa kotsimikizika kwa kusintha kosinthika kwa mitundu yotsikira iyi kumathandizira kutanthauza kwa maRNA omwe atchulidwa phunziroli. Tidapeza kuti anthu omwe ali ndi vuto la zigawo zitatu zonsezi ali pachiwopsezo chachikulu cha IGD. Pamodzi ndi zodziwika bwino za ngozi kapena zachilengedwe komanso njira zodziwira, zomwe tikupeza zingathandizire kulowerera koyambirira kuti athandize anthu omwe ali pachiwopsezo chachikulu cha IGD.
Chikhalidwe
Kafukufukuyu adavomerezedwa ndi Institutional Review Board of the Catholic University Medical College of Korea (MC16SISI0120). Onse omwe atenga nawo mbali ndi makolo awo adapereka chidziwitso cholembedwa.
Zopereka za Wolemba
ML ndi HC adathandizanso chimodzimodzi patsamba ili. ML, D-JK, ndi Y-JC adapanga kafukufukuyu. SJ, S-MC, YP, DC, ndi JL adachita zoyesa ndikuwonetsa deta. J-WC, S-HP, J-SC, ndi D-JK adatenga zitsanzo zamagazi ndi zambiri zamankhwala. ML, HC, S-HY, ndi Y-JC adasanthula deta. ML, HC, S-HY, ndi Y-JC adalongosola zolemba pamanja. Y-JC amayang'anira ntchitoyi.
Kutsutsana kwa Chidwi
Olembawo akunena kuti kufufuza kunkachitika popanda mgwirizano uliwonse wa zamalonda kapena zachuma zomwe zingatengedwe kuti zingatheke kukangana.
Mawu a M'munsi
Ngongole. Ntchitoyi idathandizidwa ndi thandizo lochokera ku Brain Research Program kudzera ku National Research Foundation of Korea (NRF), yothandizidwa ndi Ministry of Science and ICT and future Planning (NRF-2015M3C7A1064778).
Zowonjezera Zowonjezera
Zowonjezera Zowonjezera pa nkhaniyi zitha kupezeka pa intaneti pa http://www.frontiersin.org/articles/10.3389/fpsyt.2018.00081/full#supplementary-material.
Zothandizira
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