Investigation on the changes of intestinal flora in children with cerebral palsy using high-throughput sequencing

doi:10.11852/zgetbjzz2019-1418

Abstract

Abstract: Objective To explore the changes of intestinal flora in children with cerebral palsy by high-throughput sequencing. Methods Thirty-four children with cerebral palsy diagnosed in Longgang District Social Welfare Center and eating normally were selected as the cerebral palsy group,and 14 children with cerebral palsy accompanied by epilepsy and eating normally were selected as the cerebral palsy accompanied by epilepsy group,meanwhile 22 healthy children were selected as the healthy group.Total DNA was extracted and amplified from fecal specimens of all subjects.The 16SrRNA gene was sequenced with high throughput and the sequencing results were analyzed biologically. Results 1) The intestinal flora structure of healthy group was significantly different from that of cerebral palsy group and cerebral palsy accompanied by epilepsy group (R²=0.410,P=0.001;R²=0.450,P=0.001),but the intestinal flora structure of cerebral palsy group was similar to that of cerebral palsy accompanied by epilepsy group (R²=0.025,P=0.272).2) The abundance of Prevotella in children with cerebral palsy and cerebral palsy increased significantly (32.68% in cerebral palsy group and 34.34% in cerebral palsy accompanied by epilepsy group). Conclusions Prevotella abundance increases significantly in children with cerebral palsy and cerebral palsy accompanied by epilepsy who ate normally.Prevotella may be a protective bacteria of children with cerebral palsy who ate normally.

Key words: high-throughput sequencing, cerebral palsy, intestinal flora

摘要： 目的利用高通量测序方法,探讨脑瘫儿童肠道菌群的变化。方法选择龙岗区社会福利中心诊断为脑瘫且正常进食的34例患儿作为脑瘫组;同时选取脑瘫伴癫痫且正常进食的20例患儿作为脑瘫伴癫痫组;另外选取同龄健康儿童22例作为健康组。留取所有受试者的粪便标本,提取总DNA并扩增,采用对16SrRNA基因进行高通量测序,对测序结果进行生物学分析。结果 1)健康组与脑瘫组、脑瘫伴癫痫组儿童相比肠道菌群结构差异有统计学意义(R²=0.410、P=0.001;R²=0.450、P=0.001),但脑瘫组与脑瘫伴癫痫组儿童相比肠道菌群结构较为相似,差异无统计学意义(R²=0.025、P=0.272)。2)脑瘫组和脑瘫伴癫痫组儿童肠道菌群中Prevotella丰度均明显增加(脑瘫组均值为32.68%,脑瘫伴癫痫组均值为34.34%)。结论正常进食的脑瘫组、脑瘫伴癫痫组儿童中Prevotella丰度明显增加,Prevotella也许是脑瘫儿童的“保护菌属”。

关键词: 高通量测序, 脑瘫, 肠道菌群

CLC Number:

R742.3

ZHOU Bo-lan, HUANG Cong-fu, PENG Yuan-ping, LI Yin-hu, LI Xiu-yun, WANG Pei-qin, WU Gen-feng, JIAO Guang. Investigation on the changes of intestinal flora in children with cerebral palsy using high-throughput sequencing[J]. journal1, 2020, 28(10): 1077-1082.

周波兰, 黄从付, 彭远平, 李寅虎, 李秀云, 汪培勤, 伍根峰, 焦光. 高通量测序探讨脑瘫儿童肠道菌群的变化[J]. 中国儿童保健杂志, 2020, 28(10): 1077-1082.

References

[1] 中国康复医学会儿童康复专业委员会,中国残疾人康复协会小儿脑性瘫痪康复专业委员会,《中国脑性瘫痪康复指南》编委会.中国脑性瘫痪康复指南(2015):第一部分[J].中国康复医学杂志,2015,30(7):747-753.
[2] 王姣姣,张瑞光,董晓荣.肠道菌群与大脑功能相互联系的研究进展[J].中华神经医学杂志,2018,17(12):1281-1286.
[3] 黄从付,彭远平,李寅虎,等.脑瘫伴癫痫儿童肠道菌群变化[J].中国微生态学杂志,2019,31(4):402-406.
[4] Huang CF,Li YH,Li DF,et al.Distinct gut microbiota composition and functional category in children with cerebral palsy and epilepsy[J].Front Pediatr,2019,7:394.
[5] Wang Y,Kasper LH.The role of microbiome in central nervous system disorders[J].Brain Behav Immun,2014,38:1-12.
[6] Tlaskalova-Hogenova H,Stepankova R,Kozakova H,et al.The role of gut microbiota (commensal bacteria) and the mucosal barrier in the pathogenesis of inflammatory and autoimmune diseases and cancer:contribution of germ-free and gnotobiotic animal models of human diseases[J].Cell Mol Immunol,2011,8(2):110-120.
[7] Galland L.The gut microbiome and the brain[J].J Med Food,2014,17(12):1261-1272.
[8] De Filippis F,Pellegrini N,Laghi L,et al.Unusual sub-genus associations of faecal prevotella and bacteroides with specific dietary patterns[J].Microbiome,2016,4(1):57.
[9] De Filippo C,Cavalieri D,Di Paola M,et al.Impact of diet in shaping gut microbiota revealed by a comparative study in children from Europe and rural Africa[J].Proc Natl Acad Sci USA,2010,107:14691-14696.
[10] Arumugam M,Raes J,Pelletier E,et al.Enterotypes of the human gut microbiome[J].Nature,2011,473:174-180.
[11] Kang DW,Park JG,Ilhan ZE,et al.Reduced incidence of Prevotella and other fermenters in intestinal microflora of autistic children[J].PLoS One,2013,8(7):e68322.
[12] Huws SA,Kim EJ,Lee MRF,et al.As yet uncultured bacteria phylogenetically classified as Prevotella,Lachnospiraceaeincertaesedis and unclassified Bacteroidales,Clostridiales and Ruminococcaceae may play apredominant role in ruminalbiohydrogenation[J].Environ Microbiol,2011,13:1500-1512.
[13] Dyall SC.Interplay Between n-3 and n-6 long-chain polyunsaturated fatty acids and the endocannabinoid system in brain protection and repair[J].Lipids,2017,52(11):885-900.
[14] Tremlett H,Bauer KC,Appel-Cresswell S,et al.The gut microbiome in human neurological disease[J].Ann Neurol,2017,81(3):369-382.
[15] Tao J H,Duan J A,Jiang S,et al.Simultaneous determination of six short-chain fatty acids in colonic contents of colitis mice after oral administration of polysaccharides from chrysanthemum morifolium Ramat by gas chrom-atography with flame ionization detector[J].J Chromatogr B Analyt Technol Biomed Life Sci,2016,1029-1030:88-94.
[16] Barcelo A,Claustre J,Moro F,et al.Mucin secretion is modulated by luminal factors in the isolated vascularly perfused rat colon[J].Gut,2000,46(2):218-224.
[17] Goverse G,Molenaar R,Macia L,et al.Diet-derived short chain fatty acids stimulate intestinal epithelial cells to induce mucosal tolerogenic dendritic cells[J].J Immunol,2017,198(5):2172-2181.
[18] Rooks MG,Garrett WS.Gut microbiota,metabolites and host immunity[J].Nat Rev Immunol,2016,16(6):341-352.
[19] Nastasi C,Candela M,Bonefeld C M,et al.The effect of short-chain fatty acids on human monocyte-derived dendritic cells[J].Sci Rep,2015,5:16148.
[20] Jiang SY,Wei CC,Shang TT,et al.High glucose induces inflammatory cytokine through protein kinase C-induced toll-like receptor 2 pathway in gingival fibroblasts [J].Biochem Biophys Res Commun,2012,427(3):666-670.
[21] Vieira A T,Macia L,Galvao I,et al.A role for gut microbiota and the metabolite-sensing receptor GPR43 in a murine model of gout[J].Arthritis Rheumatol,2015,67(6):1646-1656.
[22] Wu GD,Chen J,Hoffmann C,et al.Linking long-term dietary patterns with gut microbial enterotypes[J].Science,2011,334(6052):105-108.
[23] Kovatcheva-datchary P,Nilsson A,Akrami R,et al.Dietary fiber-induced improvement in glucose metabolism is associated with increased abundance of prevotella[J].Cell Metab,2015,22(6):971-982.
[24] Mengxiang Z,Wei M,Juan Z,et al.Analysis of gut microbiota profiles and microbe-disease associations in children with autism spectrum disorders in China[J].Sci Rep,2018,8:13981.
[25] Parthasarathy G,Chen J,Chen X,et al.Relationship between microbiota of the colonic mucosa vs feces and symptoms,colonic transit,and methane production in female patients with chronic constipation[J].Gastroenterology,2016,150(2):367-379.
[26] Khan TJ,Ahmed YM,Zamzami MA,et al.Atorvastation treatment modulates the gut microbiota of the hyperchol-esterolemic patients[J].Omics,2018,22(2):154-163.
[27] Gomez-arango LF,Barrett HL,Mcintyre HD,et al.Connections between the gut microbiome and metabolic hormones in early pregnancy in overweight and obese women[J].Diabetes,2016,65(8):2214-2223.
[28] Kverka M,et al.Oral administration of Parabacteroides distasonis antigens attenuates experimental murine colitis through modulation of immunity and microbiota composition[J].Clin Exp Immunol,2011,163(2):250-259.
[29] Arpaia N,et al.Metabolites produced by commensal bacteria promote peripheral regulatory T-cell generation[J].Nature,2013,504(7480):451-455.
[30] Sears CL.In celebration of Sydney M.Finegold,M.D.:bacteroides fragilis in the colon:the good & the bad[J].Anaerobe,2012,18(2):192-196.
[31] Rehman A,Rausch P,Wang J,et al.Geographical patterns of the standing and active human gut microbiome in health and IBD[J].Gut,2016,65(2):238-248.
[32] Li YN,Huang F,Cheng HJ,et al.Intestine-derived Clostridium leptum induces murine tolerogenic dendritic cells and regulatory T cells in vitro[J].Hum Immunol,2014,75(12):1232-1238.
[33] Clavel T,Gomes-neto JC,Lagkouvardos I,et a1.Deciphering interactions between the gut microbiota and the immunesystem via microbial cultivation and minimal microbiomes[J].Immunol Rev,2017,279(1):8-22.
[34] Zhang J,Guo Z,Xue Z,et al.A phylo-functional core of gut microbiota in healthy young Chinese cohorts across lifestyles[J].ISME J,2015,9:1979-1990.
[35] Weiguang C,Fanlong L,Zongxin L,et al.Human intestinal lumen and mucosa-associated microbiota in patients with colorectal cancer[J].PLoS One,2012,7(6):e39743.
[36] Quitadamo P,Thapar N,Staiano A,et al.Gastrointestinal and nutritional problems in neurologically impaired children [J].Eur J Paediatr Neurol,2016,20(6):810-815.