Molecular Diagnosis Of Pediatric Patients With Citrin Deficiency In China:SLC25A13 Mutation Spectrum And The Geographic Distribution

Objective: Citrin deficiency（CD）is a Mendelian disease due to biallelic mutations of SLC25A13 gene.Neonatal intrahepatic cholestasis caused by citrin deficiency（NICCD）is the major pediatric CD phenotype currently,and its definite diagnosis relies on SLC25A13 genetic analysis.China is a vast country with a huge population,but the SLC25A13 genotypic features of CD patients in our country remains far from being well clarified.As a result,many CD patients were misdiagnosed or not diagnosed yet.This study aimed to investigate the SLC25A13 mutation spectrum and their geographic distribution,and to provide reliable evidences for the establishment of relevant molecular diagnostic targets in different geographic areas of China.Subjects and methods: The research subjects included 304 patients suspected to have NICCD in our Department of Pediatrics from the beginning of March,2013 to the end of March,2017,as well as their parents.And,other 119 CD patients diagnosed by our group from July,2005 to the end of February,2013,and their parents,were also enrolled in this study.Besides the conventional DNA analyses,c DNA cloning and Western blot analysis of citrin protein were carried out to definitely diagnose pediatric CD patients.Meanwhile,the SLC25A13 mutation spectrum of all the diagnosed CD patients was explored.Moreover,the geographic distribution of the SLC25A13 mutations and genotypes,as well as the allelic heterogeneity,were compared among the north,border and south regions relevant to the boundary of the Yangtze River by means of Chi-square test.Results:（1）This study diagnosed 204 new CD patients in mainland China and identified 7novel deleterious SLC25A13 mutations,i.e.c.493C>T（p.Q165X）,c.755-1G>C（p.252fs269X）,c.845_c.848+1del G（p.D283 fs X285）,c.933_c.933+1ins GCAG（p.A312 fs X317）,c.1381G>T（p.E461X）,c.1706₁707del TA（p.S331 fs X363）and [c.329-154_c.468+2352del2646;c.468+2392_c.468+2393ins23]（p.E110fs127X）.（2）Among the 323 CD patients diagnosed by our group thus far,44 SLC25A13 mutations/variations were detected,including 14 missense mutations,7 deletion,12 nonsense,4 splice-site,3 insertion,1 duplication,1 pathogenic SNP1 aberrant transcript and 1 complex mutation.（3）The 8 mutations c.475C>T（p.Q159X）,c.775C>T（p.Q259X）,c.851₈54del4,c.1078C>T（p.R360X）,IVS11+1G>A,c.1364G>T （p.R455L）,c.1399C>T（p.R467X）and IVS16ins3 kb demonstrated significantly different geographic distribution.（4）Among the total 58 identified genotypes,c.851₈54del4/c.851₈54del4,c.851₈54del4/IVS16ins3 kb,c.851₈54del4/c.1399C>T（p.R467X）and IVS16ins3kb/IVS11+1G>A presented different geographic distribution.（5）The northern population had a higher level of SLC25A13 allelic heterogeneity than those in the south.Conclusions: Via conventional molecular analysis,cDNA cloning and Western blot analysis of AGC2,204 new CD patients in mainland China were diagnosed and 7 novel deleterious SLC25A13 mutations identified,enriching the SLC25A13 mutation spectrum.The SLC25A13 mutation spectrum in the hitherto largest CD cohort of 323 cases and their different geographic distribution formed a substantial contribution to the in-depth understanding of the genotypic feature of CD patients in China,providing reliable evidences for NICCD definite diagnosis and for the development of molecular diagnostic strategies in different Chinese areas.