A New Mutation Of SLC19A2 In A Zhuang Ethnic Family With Thiamine-responsive Megaloblastic Anemia

Background and purposeThe thiamine-responsive megaloblastic anemia(TRMA)syndrome is a rare autosomal recessive genetic disease.The main clinical manifestations are: large cell anemia,hyperglycemia,and gradually increasing sensory nerves.Deaf.In addition,there are other uncommon features,including stroke-like seizures,thrombocytopenia,ataxia,arrhythmia,retinal degeneration,and short stature.Complex and diverse clinical features often lead to misdiagnosis and mistreatment of TRMA.Genetic analysis revealed that the only pathogenic gene of TRMA is the SLC19A2 gene,which is located on 1q23.3,and encodes a polypeptide chain thiamine transporter-1(THTR-1)consisting of 497 amino acids.At present,at least 45 mutations in the SLC19A2 gene are found in different races to cause TRMA,most of which are missense mutations and nonsense mutations,followed by insertion and deletion mutations.Thiamine is an essential vitamin in life.However,the human body lacksthe ability to synthesize thiamine and must be taken from exogenous foods.The thiamine phosphate obtained in food is converted into free thiamine by intestinal phosphatase and absorbed by intestinal epithelial cells.The thiamine that enters the cell is converted to thiamine pyrophosphate(TPP)by thiamine pyrophosphorokinase and becomes a key enzyme in the process of glucose metabolism(pyruvate dehydrogenase,?-ketoglutarate dehydrogenase,transketolase)is involved in cellular energy metabolism.We performed a gene mutation analysis on a TRMA family of Zhuang nationality,determined the cause of the disease from the genetic level,and further used high performance liquid chromatography to detect the proband's thiamine supplementation and thiamine supplementation(30 mg/d)for 3 months.After the relative concentration of whole blood TPP,provide an important basis for disease diagnosis and treatment.MethodWe collect clinical data on a family of Zhuang people with TRMA.Based on the clinical data,mutation analysis was performed on all exons and their flanking sequences of the SLC19A2 gene.Peripheral blood lymphocytes from probands and healthy controls were isolated and RNA was extracted.The expression of SLC19A2 gene was detected by RT-PCR.Computer analysis predicts the transmembrane region,conservation,and three-dimensional structure of THTR-1.The relative concentrations of whole blood TPP in healthy controls,probands before thiamine supplementation,and thiamine(30 mg/d)supplementation were measured by high performance liquid chromatography.ResultThe mutations in the exons and their flanking sequences of the SLC19A2 gene were analyzed in this family member.The homozygous insertion mutationof c.1409 insT was detected in exon 6 of the SLC19A2 gene in the proband.The parents and sisters were carriers of the heterozygous mutation.RT-PCR results showed that there was no difference in the mRNA expression level of SLC19A2 between the proband and healthy control group(P<0.05).The transmembrane region of human THTR-1 amino acid sequence was predicted and the results showed that amino acids 471-497 were located in the cell membrane and in the cells;the conservation of the amino acid sequences(471-497)of the THTR-1part of 10 species was compared.Some of the amino acids are still highly conserved;THTR-1 three-dimensional structure predictions show that mutations result in a conformational change of the protein.HPLC results showed that the relative concentration of whole blood TPP in the healthy control group was6648.00±680.82 mV*s,and the relative concentration of TPP in the whole blood before the proband supplemental thiamine was 2403 mV*s,compared with the healthy control group.The difference was statistically significant(P = 0.000);the relative concentration of whole blood TPP after 3 months of thiamine supplementation(30 mg/d)in the proband was 6185 mV*s,which was not statistically significant compared with the healthy control group.(P = 0.203).Conclusion1.A new mutation(c.1409insT)was found in the SLC19A2 gene of the thiamine-responsive megaloblastic anemia syndrome family in Zhuang by genetic testing.2.The relative concentration of thiamine pyrophosphate in the proband's whole blood was significantly reduced,probably due to dysfunction of the thiamine transporter-1.