Juan Carlos Feoli-Fonseca

Absence of Association of Thrombophilia Polymorphisms with Intrauterine Growth Restriction

BACKGROUND Previous data have demonstrated associations between thrombophilia polymorphisms in pregnant women and an increased risk of intrauterine growth restriction in their offspring, but this finding remains uncertain. METHODS We performed a hospital-based case-control study and a family-based study including 493 newborns with intrauterine growth restriction (defined by birth weight below the 10th percentile for gestational age and sex according to Canadian norms) and 472 controls (with birth weight at or above the 10th percentile). We determined the presence or absence in newborns and their parents of the following polymorphisms: methylenetetrahydrofolate reductase (MTHFR) C677T and A1298C, factor V Leiden G1691A, and prothrombin G20210A. Mothers were interviewed to obtain information on other risk factors for intrauterine growth restriction. RESULTS The risk of intrauterine growth restriction was not increased among mothers carrying a polymorphism associated with thrombophilia. In the case-control study, the odds ratios associated with two copies of the variant, after adjustment for newborn genotype and other risk factors, were 1.55 for MTHFR C677T (95 percent confidence interval, 0.83 to 2.90) and 0.49 for MTHFR A1298C (95 percent confidence interval, 0.25 to 0.93); heterozygotes for factor V Leiden had an odds ratio of 1.18 (95 percent confidence interval, 0.54 to 2.55), and heterozygotes for prothrombin G20210A had an odds ratio of 0.92 (95 percent confidence interval, 0.36 to 2.35). These polymorphisms in the newborn were not associated with an increased risk. Newborns who were homozygous for the MTHFR C677T variant had a decreased risk of intrauterine growth restriction (odds ratio after adjustment for mothers genotype and other confounders, 0.52 [95 percent confidence interval, 0.29 to 0.94]). The results of the family-based study supported those of the case-control study. CONCLUSIONS Our findings do not indicate that there are associations between maternal or newborn polymorphisms associated with thrombophilia and an increased risk of intrauterine growth restriction.

Familial lipoprotein lipase deficiency in infancy: Clinical, biochemical, and molecular study

OBJECTIVES To describe the characteristics of lipoprotein lipase (LPL)-deficient patients seen in infancy and to evaluate the safety and efficacy of severe fat restriction. METHODS Children <1 year old presenting with chylomicronemia between 1972 and 1995 were identified, and their clinical courses were reviewed retrospectively. RESULTS LPL deficiency was demonstrated in 16 infants who presented with irritability (n = 7), lower intestinal bleeding (n = 2), pallor, anemia, or splenomegaly (n = 5), and a family history or fortuitous discovery (n = 2). All plasma samples were lactescent at presentation. Chylomicronemia responded rapidly to dietary fat restriction, and it was possible to maintain satisfactory metabolic control for a prolonged period of time. Only 1 adolescent girl had an episode of pancreatitis associated with the use of oral contraceptives. No persistent adverse effects on growth were seen. We obtained abnormal values for serum iron, alkaline phosphatase, and total calcium. CONCLUSIONS The presentation of LPL deficiency is heterogeneous during infancy. Close dietary monitoring is required to avoid nutritional deficiencies. Estrogen therapy should be avoided in LPL-deficient patients.

Human papillomavirus (HPV) study of 691 pathological specimens from Quebec by PCR-direct sequencing approach.

Human papillomaviruses (HPV) are etiological agents of cervical cancer. In order to address clinical demand for HPV detection and sequence typing, mostly in pre‐cancerous cervical lesions, we applied our two‐tier PCR‐direct sequencing (PCR‐DS) approach based on the use of both MY09/MY11 and GP5 + /GP6 + sets of primers. We tested 691 pathological specimens, all of which were biopsies, 75% of which were diagnosed histologically as cervical intraepithelial neoplasia (CIN) grades I–III. In total, 484 samples (70%) tested HPV‐positive, yielding 531 HPV sequences from 47 HPV types, including two novel types. Four most frequently found HPV types accounted for 52.9% of all isolates: HPV6, 16, 11, and 31 (21.5%, 20.0%, 7.0%, and 4.5%, respectively). Some interesting results are the following: all currently known high‐risk HPV (14 types) and low‐risk HPV (6 types) were detected; HPV18 was not the 1st or 2nd but rather the 4th–5th most frequent high‐risk HPV type; the highest detection rate for HPV (86%) among samples suspected to be HPV‐infected was found in the youngest age group (0–10 years old), including 70% (44/63) “genital” HPV types; HPV types of undetermined cervical cancer risk represented 19% and of the total HPV isolates but were strongly increased in co‐infections (36.5% of all isolates). To our knowledge, this is the largest sequencing‐based study of HPV. The HPV types of unknown cancer risk, representing the majority of the known HPV types, 27 of the 47 types detected in this study, are not likely to play a major role in cervical cancer because their prevalence in CIN‐I, II, and III declines from 16% to 8% to 2.5%. The two‐tier PCR‐DS method provides greater sensitivity than cycle sequencing using only one pair of primers. It could be used in a simple laboratory setting for quick and reliable typing of known and novel HPV from clinical specimens with fine sequence precision. It could also be applied to anti‐cancer vaccine development. J. Med. Virol. 63:284–292, 2001.

A two-tier polymerase chain reaction direct sequencing method for detecting and typing human papillomaviruses in pathological specimens.

An in-house polymerase chain reaction direct sequencing (PCR-DS) approach for HPV detection and typing was developed, taking advantage of two widely used pairs of human papillomavirus (HPV)-specific PCR primers, MY09/MYII and GP5/GP6, and 33P-labeled dideoxynucleotides. In this study, 105 pathological specimens were examined: 89|X% were diagnosed as cervical intraepithelial neoplasia (CIN) grade I-III, 76. 2|X% were HPV-positive by PCR-DS. The PCR using GP5/GP6 (first tier) and MY09/MY11 primers (second tier for the GP5/GP6-negative samples) detected additional 15|X%-25|X% HPV-positive samples compared with each pair used separately. Direct sequencing was then used to type the HPV. A readout of a sequence as short as 34 nucleotides within a specific region in the LI gene is sufficient to type known or novel sequences. Because of its high sensitivity and cost-effectiveness, the two-tier PCR-DS was adopted by the authors as the current method of choice for HPV diagnosis with ultimate sequence precision

Small babies receive the cardiovascular protective apolipoprotein ε2 allele less frequently than expected

A newborn whose weight for gestational age and sex is less than expected, based on population standards, is considered as having intrauterine growth restriction (IUGR); a cut off at less than the 10th centile is often used to define IUGR. Causes of IUGR remain unclear although a number of fetal and maternal risk factors have been identified.1,2 Increased early morbidity and mortality, as well as, possibly, less than optimal neuropsychological development, have been reported as consequences of IUGR.2,3 In addition, small size at birth has been associated with health problems in adulthood such as coronary heart disease and dyslipidaemia.4,5 The association between restricted fetal growth and adult chronic diseases (often referred to as the Barker hypothesis) is now considered robust and possibly causal.6 Apolipoprotein E (apoE) is one of the key regulators of plasma lipid levels as it affects hepatic binding, uptake, and catabolism of several classes of lipoproteins.7 The apolipoprotein E gene (APOE) codes for the apoE protein; in animal models, underexpression of the APOE gene and lack of the apoE protein result in increased susceptibility to atherosclerosis,8,9 whereas gene overexpression displays anti-inflammatory, antiproliferative, and atheroprotective properties.10 ApoE has also emerged as a central factor in various biological processes such as immunoregulation, control of cell growth and differentiation,11 and brain development.12 The three common allelic variants at the APOE locus (e2, e3, e4) code for three major apoE protein isoforms (E2, E3, E4). These isoforms differ from one another only by single amino acid substitutions, yet these changes exhibit functional consequences at both the cellular and molecular levels.13,14 In previous studies, children who carry the e4 allele and those who carry the e2 allele have been shown to have, respectively, higher and lower total …

Identification of two novel LDL receptor gene defects in French‐Canadian pediatric population: Mutational analysis and biochemical studies

Familial hypercholesterolemia (FH) is at least twofold more prevalent in French Canadians from Québec than in most Western populations. Although our recent data confirmed this high frequency of heterozygous FH in our pediatric population with hypercholesterolemia, none of the five established molecular defects for the French‐Canadian population was detected in 29% of the unrelated French‐Canadian children characterized by a persistent increase in LDL (low density lipoprotein receptor) cholesterol and a positive parental history of hyperlipidemia (Assouline et al., 1995). To probe for new mutations, six of these molecularly undiagnosed children were investigated as index patients. By using single‐strand conformation polymorphism analysis and DNA sequencing, two novel mutations were identified in two of these subjects: (1) 7‐base pair (bp) duplication following nucleotide 681 (according to the cDNA sequence) in exon 4 (681ins7), which causes a frameshift, the introduction of a stop at codon 208, and premature chain termination, and (2) A to G change in exon 8 substituting a tyrosine for a cysteine at amino acid 354 (Y354C). A third subject carried the recently reported exon 10 mutation (Y468X), whereas the remaining three patients demonstrated various known polymorphisms with no effect on gene product. Rapid molecular assays were developed to detect the two new mutations as well as the Y468X mutation. Screening of our cohort showed heterozygosity in 1/88, in 2/88, and in 2/88 of patients for the 681ins7, the Y354C, and the Y468X mutations, respectively. Hum. Mutat. 9:555–562, 1997.

Determinants of Lipid Level Variability in French-Canadian Children With Familial Hypercholesterolemia

The wide variability in the biochemical expression of familial hypercholesterolemia (FH) is only partly explained by mutational heterogeneity in the low density lipoprotein receptor (LDLR) gene. In the current study, we measured this biochemical variability in a group of children heterozygous for the >15-kb LDLR gene deletion (n=67) and examined the contribution of apolipoprotein (apo) E and B allelic variations to this phenotypic variability. Variances of total cholesterol (TC), LDL-C, and apoB concentrations and of the ratio of TC to high density lipoprotein cholesterol (HDL-C) were increased in FH subjects compared with controls. However, after taking the means into account, the coefficients of variation showed that the variability of LDL-C and apoB concentrations was smaller for FH than for controls and that the variability of TC and of the ratio TC to HDL-C was similar between both groups. The ϵ2/3 genotype was associated with lower mean TC, LDL-C, and apoB concentrations in FH. The magnitude of this effect was smaller in controls than in FH. Indeed, the percentages of total variance of TC, LDL-C, and apoB attributable to the apoE locus were 19.9%, 18.1%, and 11.8%, respectively, in FH cases and 5.9%, 7.4%, and 6.0%, respectively, in controls. We did not detect any effect of the apoB insertion/deletion polymorphism on lipid traits in FH children. However, in controls, we observed a strong interaction between apoE and apoB genotypes on apoB concentrations and on TC to HDL-C ratios. Our study reemphasizes the important role of apoE in lipid metabolism and illustrates that the effects of allelic variations on lipid traits are context dependent.

Direct Human Papillomavirus (HPV) Sequencing Method Yields a Novel HPV in a Human Immunodeficiency Virus-Positive Quebec Woman and Distinguishes a New HPV Clade

Papillomaviruses of supergroup A exhibit genital tropism and are best known as etiologic agents for benign and malignant cervical lesions in women. A polymerase chain reaction direct sequencing approach with P-33-labeled dideoxynucleotides was used to detect and type human papillomaviruses (HPVs) in cervical biopsies. A novel sequence was found in condylomatous specimens from a human immunodeficiency virus-positive French Canadian woman. The viral gene L1 was sequenced completely, yielding a novel HPV type of supergroup A, named JC9710. This is related to a previously described HPV type from New Mexico, CP8061, and to Colobus monkey papillomavirus 1. Sequence similarity searches and phylogenetic analyses with different software packages clustered the three viral types as a new clade, for which the next available number, A15, was proposed.

Human papillomavirus PCR direct sequencing study of cervical precancerous lesions in Quebec children.

Editor,—Similarly to adult pathology, human papillomavirus (HPV) infection is the most common sexually transmitted disease in adolescent girls, whose prevalence is 16% according to one US study.1 However, little or no HPV sequencing data from paediatric specimens are available. We used our two tier polymerase chain reaction (PCR) direct sequencing (PCR-DS) approach2 to study cervical biopsies from 44 adolescent Quebec girls (14–17 years old). They originated from various social and ethnic groups, as well as geographically distinct areas of Quebec. Written informed consent about the use of the specimens was obtained from the ethics committee of this institution. All biopsies were analysed for histological changes and presence of HPV specific DNA. Most of them (n = 36) were diagnosed as cervical intraepithelial neoplasia (CIN), seven as inflammatory changes, and one as …