Diagnosis Of Adult-type Hypolactasia Research Articles

Adult-type hypolactasia in children: A genetic perspective

Adult-type hypolactasia is an autosomal recessive condition resulting from the physiological decline of the lactase enzyme activity in intestinal cells which occurs in a significant proportion of the global population. Mechanisms proposed to explain the occurrence of adult-type hypolactasia include: (a) decreased production of lactase, (b) synthesis of an inactive high molecular weight lactase, (c) defective post-translational modifications, and (d) susceptibility of lactase to luminal proteases during weaning due to change in levels of sialylated and fucosylated enzymes in the small intestine. Recently, the C/T-13910 polymorphism on chromosome 2q21 in North-European populations has been found to be associated with lactase activity and its genetic typing is advocated as a first-stage screening test for adult hypolactasia. Available biochemical methods for diagnosis of adult-type hypolactasia consists of the lactose tolerance test (LTT), which measures the increase in blood glucose after an oral lactose load, and the hydrogen breath test (HBT), that is based on the determination of exhaled hydrogen produced by the bacterial flora in the colon after an oral lactose load. These methods require significant procedural time both for the patient and the processing clinical laboratory. The present review describes genetic regulation of lactase expression and various single nucleotide polymorphisms (SNPs) associated with adult-type hypolactasia and lactase persistence in different human populations.

Accuracy of lactase gene C/T-13910 polymorphism and hydrogen breath test in a gastroenterology outpatient clinic: a retrospective study

Adult type hypolactasia is the most prevalent carbohydrate malabsorption. To assess the distribution of lactase gene C/T-13910 polymorphism and the accuracy and concordance of a genetic test and H(2) breath test in the diagnosis of adult type hypolactasia. 496 patients with symptoms of lactose intolerance were enrolled in a retrospective study who underwent genetic test using TaqMan polymerase chain reaction and H(2) breath test. The prevalence of C/T-13910 genotypes was: CC 48.1%, TC: 40.5%, and TT: 11.4%. When the genetic test was taken as reference, the sensitivity of the breath test was 84.3%, with a specificity of 95.7%, a positive predictive value of 96.7% and negative predictive value of 80.4%. Conversely, the accuracy of genetic test was: sensitivity 96.6%, specificity 80.4%, positive predictive value 84.3% and negative predictive value 95.7%. The concordance value between the two tests (kappa index) was 0.78. The results were discordant in 11.1% of the cases. In symptomatic patients, the lactase non-persistence genotype CC occurred in almost half of the patients. Both the genetic and the breath tests are sufficiently accurate, with good predictive value and they can be used to set up the diagnosis. Discordant results should be carefully interpreted.

Effect of C/T −13910 cis-acting regulatory variant on expression and activity of lactase in Indian children and its implication for early genetic screening of adult-type hypolactasia

Background Absorption of milk sugar (lactose) is regulated by the activity of lactase enzyme in gut wall. Intestinal lactase activity declines during childhood in majority of human populations leading to adult-type hypolactasia (primary lactose malabsorption), limiting the use of fresh milk due to lactose intolerance. Aim of this study was to correlate lactase expression and activity with C/T −13910 variant in Indian children, determine the age of onset of down-regulation of lactase activity and assess the applicability of the C/T −13910 variant as a diagnostic marker for identifying children genetically inclined to develop adult-type hypolactasia. Methods Intestinal biopsies were obtained from 176 children aged 1–16 years undergoing routine endoscopy for various abdominal complaints. The biopsies were assayed for lactase, sucrase and maltase activities and genotyped for C/T 13910 variant using PCR-RFLP analysis. The functional effect of the C/T − 13910 variant on expression of lactase mRNA and protein in these children was examined using reverse-transcription PCR and western blotting. Results Among the 176 children investigated in our study, 56.8% (100/176) carried the C/C − 13910 genotype, which has been associated with the onset of adult-type hypolactasia, while 40.9% (72/176) carried the C/T − 13910 genotype and 2.3% (4/176) the T/T − 13910 genotype. There was a significant correlation between lactase activity and C/T − 13910 variant ( P < 0.001). The mean level of lactase activity among children with C/C − 13910 genotype was 15.9 U/g protein and with C/T and T/T − 13910 genotypes was 30.9 U/g protein. The age of onset of down-regulation of lactase activity in children with C/C − 13910 genotype was between 3 and 5 years and keeping 10 U/g protein lactase activity as cut off, adult-type hypolactasia was evident in all the individuals > 8 years of age for this genotype. C/C − 13910 genotype was associated with low expression of lactase mRNA and protein compared with C/T genotype. Considering lactase activity of 10 U/g protein as gold standard, predictive value of genetic test based on C/T − 13910 variant for adult-type hypolactasia was 100% in children > 8 years of age. Conclusion C/T − 13910 cis-acting regulatory variant located ≈ 14 kb upstream of lactase gene (LCT) completely correlates with lactase phenotype in Indian children. The genetic testing for the C/T − 13910 variant may be helpful in the diagnosis of adult-type hypolactasia in Indian children.

Evaluation of a novel reverse-hybridization StripAssay for typing DNA variants useful in diagnosis of adult-type hypolactasia

Background Adult-type hypolactasia is a genetically determined inability to digest lactose after weaning. Two single-nucleotide polymorphisms ( C-13910T, G-22018A) located upstream of the lactase gene ( LCT) within the gene MCM6 are associated with the lactase persistence/non-persistence trait in patients of European descent. Therefore, the genotyping of these SNPs has been established as a diagnostic tool for adult-type hypolactasia. We have recently shown that several novel allelic variants located in close proximity to the C-13910T SNP interfere with the diagnostic accuracy of real-time PCR-based genotyping methods. Methods We describe here the validation of a comprehensive reverse-hybridization teststrip-based assay for the detection of common and novel LCT SNPs ( C-13907G, C-13910T, T-13913C, G-13914A, T-13915G, and G-22018A). This assay is based on multiplex DNA amplification and ready-to-use membrane teststrips containing variant-specific oligonucleotide probes immobilized as an array of parallel lines. Results We evaluated the novel reverse-hybridization StripAssay on 125 DNA samples in comparison to LightCycler analysis and sequencing. The outcome of StripAssay genotyping was found to be completely concordant with that obtained by sequencing. Conclusions The StripAssay represents an accurate and robust screening tool to identify multiple LCT/MCM6 variants in a rapid manner. It overcomes diagnostic pitfalls that were reported and allows the simultaneous genotyping of closely spaced LCT variant sites in a single-step diagnostic approach .

Genetic testing improves the diagnosis of adult type hypolactasia in the Mediterranean population of Sardinia

Recently, the C/T-13910 polymorphism on chromosome 2q21 in North-European populations has been found completely associated with lactase activity and its genetic typing proposed as first-stage screening test for adult hypolactasia. However, the C/T-13910 variant in some sub-Saharan African groups is not a predictor of lactase persistence. In this work, we wanted to verify if in the Mediterranean island of Sardinia, located in Southern Europe, the C/T-13910 polymorphism may be useful or not for the diagnosis of adult type hypolactasia. Validation study of a genetic testing for adult type hypolactasia in Sardinians. Brotzu Hospital and Microcitemico Hospital, Cagliari, Italy. The sample consisted in 84 Sardinian individuals (63 women and 21 men; range 20-73 years) selected from a group of 832 patients. Genetic testing was compared to an improved test obtained by a combination of different breath hydrogen tests and clinical assessment. We found that all 49 individuals with lactose malabsorption, demonstrated by a combination of different breath hydrogen tests and clinical assessment, carried the C/C-13910 genotype associated with lactase non-persistence, 23 individuals with lactose normal absorption carried the C/T-13910 genotype associated with lactase persistence and only one person with the above phenotype showed a discordant C/C-13910 genotype. The genetic testing showed very high sensitivity, specificity, positive and negative predictive values of 100, 95.8, 98 and 100%, respectively. Sardinians, unlike some ethnic groups in sub-Saharan Africa, show the same genetic association of hypolactasia with the C/T-13910 variant as other North-European populations. The genetic testing for the C/T-13910 variant may contribute to improving the diagnosis of adult type hypolactasia.

P0066 PP GENETIC TESTING OF ADULT-TYPE HYPOLACTASIA

Introduction: Adult-type hypolactasia affects most of world’s human population and limits the use of fresh milk due to lactose intolerance. The diagnosis of adult-type hypolactasia has been difficult to establish due to unsatisfactory diagnostic methods. We have identified a DNA variant residing 13910 bp upstream of the lactase-phlorizin hydrolase (LPH) gene at chromosome 2q21–22 that shows complete association with lactase non-persistence/persistence trait (Enattah 2002, Kuokkanen 2003). The genotype C/C-13910 is associated with adult-type hypolactasia and genotypes C/T-13910 and T/T-13910 with lactase persistence. We have assessed the applicability of the C/T-13910 variant as a diagnostic test at the time of downregulation of the lactase activity in children of African, Finnish and other Caucasian origins. Methods: Intestinal biopsies were obtained from 333 children and adolescents aged from 0.1 to 20 years undergoing upper gastrointestinal endoscopy because of unspecified abdominal complaints. The biopsies were assayed for lactase, sucrase and maltase activity and genotyped for the C/T-13910 variant using PCR-minisequencing (Messer 1966, Syvänen 1994). Results: The prevalence of the genotype C/C-13910 defining adult-type hypolactasia was 14.5% among Finnish (n=256), 95.4% among African (n=65) and 75.0% among the other Caucasian children (n=12). The majority of Finnish children (92%) with C/C-13910 genotype had high lactase activity (>10 U/g protein) before the age of five years. A gradual decline took place between 5–12 years of age. The decline of the lactase activity in African children occurred somewhat earlier than in Finnish children as 30% of the children already had low levels (<10 U/g protein) at the age of five years. In Finnish children with C/C-13910 genotype the probability of low lactase activity (<10 U/g protein) was 8% before five years of age, 63% between 6–11 years of age and 100% after 12 years of age. The C/C-13910 genotype was associated with low lactase activity (<10 U/g protein) in all children >12 years of age in all ethnic groups studied giving a sensitivity of 94% and specificity of 100% for this genetic test defining adult-type hypolactasia. Conclusion: Genetic testing of adult-type hypolactasia is a clinical aid in diagnosis of abdominal complaints in populations like those of Northern Europeans where the prevalence of adult-type hypolactasia is relatively high and consumption of dairy products common. This test is easily performed with a semiautomated analysis from a drop of peripheral blood without fasting making this test suitable not just for diagnostic purposes at individual level but also as a screening method for population studies. Reference(S):

Morphological method for the diagnosis of human adult type hypolactasia.

The primary adult type hypolactasia is the most common form of genetically determined disaccharidase deficiency. This study examined a large and homogeneous population of the south of Italy: surgical biopsy specimens of proximal jejunum from 178 adult subjects have been assayed for disaccharidase activities; the expression of lactase protein and lactase activity has also been investigated on tissue sections by immunomorphological and enzymohistochemical techniques. Histograms of lactase to sucrase ratio were found to provide a useful distribution of the lactase activity; a lactase to sucrase ratio of 0.17 was found to show discrimination between tissues with persistence of high lactase activity and tissues with adult type hypolactasia. In all 28 subjects with persistent high lactase activity, a uniform distribution of lactase protein and lactase activity in all villus enterocytes was detected, whereas in all 150 subjects with adult type hypolactasia a variable number of villus enterocytes failed to express the lactase. Moreover in hypolactasic samples the lactase activity on tissue sections was constantly detected later than in samples with persistent high lactase activity. The absolute correlation between the immunohistochemical and enzymohistochemical features and the assessment of lactase activity in intestinal homogenates suggests that the morphological criteria are an alternative method for the diagnosis of adult type hypolactasia in human biopsy specimens from proximal small jejunum.