Reference
Complete the bibliographic reference for the article according to AJE format.

Shaw GM, et al. Genetic variation of infant reduced folate carrier (A80G) and risk of orofacial and conotruncal heart defects. Am J Epidemiol 2003 Oct;158: 747-52.

Category of HuGE information
Specify the types of information (from the list below) available in the article:

1. Prevalence of gene variant
2. Gene-disease association
3. Gene-environment interaction
4. Gene-gene interaction
5. Genetic test evaluation/monitoring

1. Prevalence of gene variant
2. Gene-disease association
3. Gene-environment interaction

Study hypotheses or purpose
The authors study hypotheses or main purpose for conducting the study.

Infants homozygous for the RFC1 (SLC19A1) G80/G80 genotype would be at increased risk of orofacial clefts or conotruncal heart defects because of an impaired ability to transport folates to the cytoplasm of a critical cellular population. Further, that elevation in maternal serum folate levels resulting from periconceptional supplementation of folic acid might improve the activity of the poorly functioning variant form of the transport protein. In evaluation of the gene-environment interaction, risk was predicted to be higher among infants homozygous for RFC1 (SLC19A1) G80/G80 whose mothers did not use periconceptional vitamin supplements containing folic acid, compared with infants homozygous for RFC1 (SLC19A1) A80/A80 whose mothers used folic acid supplements in the periconceptional period.

Gene(s)
Identification of the following:

1. Gene name
2. Chromosome location
3. Gene product/function
4. Alleles
5. OMIM #

1. Gene name: SLC19A1- solute carrier family 19 (folate transporter), member 1 (alias: RFC1)
2. Chromosome location: 21q22.3
3. Gene product/function: Transporter for the intake of folate. Uptake of folate in human placental choriocarcinoma cells . occurs by a novel mechanism called potocytosis which functionally couples three components, namely the folate receptor, the folate transporter, and a v-type h+-pump.
4. Alleles: A80G single nucleotide polymorphism
5. OMIM #: 600424

Environmental factor(s)
Identification of the major environmental factors studied (infectious, chemical, physical, nutritional, and behavioral)

Maternal periconceptional intake of vitamins containing folic acid maternal use of cereal

Health outcome(s)
Identification of the major health outcome(s) studied

1. Cleft lip with or without cleft palate (CLP)
2. Cleft palate (CP)
3. Conotruncal heart defects

1. Case-control
2. Cohort 
3. Cross-sectional
4. Descriptive or case series
5. Clinical trial
6. Population screening

1. Case-control

1. Disease case definition
2. Exclusion criteria
3. Gender
4. Race/ethnicity
5. Age
6. Time period
7. Geographic location
8. Number of participants

1. Disease case definition: liveborn Infants with conotruncal heart defects, isolated cleft palate, or cleft lip with or without cleft palate
2. Exclusion criteria: Infants with orofacial anomalies with other major anomalies and infants diagnosed with single-gene disorders, trisomies, or Turner’s syndrome (45,X)
3. Gender: male and female
4. Race/ethnicity: White Hispanic, White non-Hispanic, and other
5. Age: up to 1 year after birth
6. Time period: deliveries if infants or fetal deaths between January 1987- December 1988 for conotruncal defects and deliveries if infants or fetal deaths between January 1987- December 1989 for conotruncal defects
7. Geographic location: most California counties
8. Number of participants: 207 (87% of total eligible) conotruncal heart defect cases and 489 (85% of total eligible) of orofacial defect cases.

1. Control selection criteria
2. Matching variables
3. Exclusion criteria
4. Gender
5. Race/ethnicity
6. Age
7. Time period
8. Geographic location
9. Number of participants

1. Control selection criteria: infants with no major congenital anomalies identified before the first birthday
2. Matching variables: geographic area
3. Exclusion criteria: major congenital anomalies identified before the first birthday
4. Gender: male and female
5. Race/ethnicity: same as cases
6. Age: same as cases
7. Time period: same as cases
8. Geographic location: same as cases
9. Number of participants: 734 controls (76 %of total eligible).

1. Environmental factor
2. Exposure assessment
3. Exposure definition
4. Number of participants with exposure data (% of total eligible)

1. Environmental factor: maternal use of vitamin supplement containing folic acid
2. Exposure assessment: telephone interview elicited
3. Exposure definition: “Use” defined as starting vitamin use anytime during the 1 month before conception through the end of the second month after conception and “nonuse” defined as starting vitamin use in the third month after conception
4. Number of participants with exposure data:
   Conotruncal heart defect cases and controls; 398
   Orofacial defect cases and controls; 782

1. Gene
2. DNA source
3. Methodology
4. Number of participants genotyped (% of total eligible) 

1. Gene: SLC19A1
2. DNA source: dried newborn-screening blood specimens on filter paper
3. Methodology: Extract DNA using Puregene DNA Extraction Kit (Gentra Systems, Minneapolis, Minnesota) and a 140 bp fragment in exon 2 amplified using PCR, then restriction fragment length polymorphism analysis (New England Biolabs, Beverly, Massachusetts) created by HhaI (GCG/C) restriction site at position 80.
4. Number of participants genotyped: Cases: One hundred sixty three of the infants with conotruncal heart defects (79% of total eligible). Four hundred twenty eight infants with orofacial clefts (305 with CLP and 123 with CP, 88% of total eligible).
5. Controls: 364 were randomly sampled and genotyped (239 for control infants born in 1987-1988) of 652 for whom DNA was available.

1. Prevalence of gene variant
2. Gene-disease association
3. Gene-environment interaction
4. Gene-gene interaction
5. Genetic test evaluation/monitoring

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An increased risk of conotruncal defects but not isolated cleft defects among infants homozygous (G80/G80) or heterozygous for RFC1 (SLC19A1) A80G polymorphism was found. Some evidence suggestive of an interaction between the infant G80/G80 genotype and maternal supplemental vitamin use was observed, but only for conotruncal defects. The authors suggest that the elevated risk for only conotruncal defects may reflect a true association with one potentially folate-responsive defect, but not with another. Alternatively, such differences may reflect a chance association of RFC1 (SLC19A1) and conotruncal defects. Elevated risks for conotruncal heart defects associated with the G allele could also be interpreted as a deficit of conotruncal cases among infants with the A80/A80 (referent) genotype, suggesting that the A80/A80 genotype is associated with a reduced risk of conotruncal defects. These findings require replication, given that this appears to be the first epidemiologic study investigating the contribution of RFC1 (SLC19A1) to the risk of conotruncal and orofacial cleft defects. There is also no explanation for why heterozygotes were observed to be at higher risk than homozygotes for conotruncal heart defects, although one could postulate selection against homozygotes (such as fetal loss).

As the author’s state, the study was limited to infant genotype. Maternal genotype may provide additional information and explanation for results. The article presents results of gene-nutrient interaction but odds ratios do not compare presence vs. absence of nutrient, e.g., as measured by plasma folate levels.