Affected Sib-pair Method Research Articles

Affected sib-pair analysis of the GLUT1 glucose transporter gene locus in non-insulin-dependent diabetes mellitus (NIDDM): evidence for no linkage.

Despite the strong evidence for a major role played by genetic factors in the aetiology of non-insulin-dependent diabetes mellitus (NIDDM), the genes involved are still unknown. Association studies of candidate genes for the inheritance of NIDDM have so far yielded inconclusive results. Some evidence exists for an association between NIDDM and the glucose transporter gene GLUT1, involved in basal glucose transport, although this has not been confirmed. In the present study we have tested the hypothesis of linkage between NIDDM and the GLUT1 gene, using affected sib-pairs. With this method the concordance observed for a given gene marker is compared with that expected under the assumption of no linkage between that marker and the disease. Fifty-four pedigrees (22 Italians and 32 British), for a total of 82 sib-pairs were studied by the affected sib-pair method proposed by Weeks and Lange, using two restriction fragment length polymorphisms (RFLPs) at the GLUT1 locus, the MspI RFLP, at an estimated 0.171 recombination frequency from the GLUT1 gene, and the XbaI RFLP, located within the GLUT1 gene and previously shown to be associated with the disease. Results showed that the MspI marker and NIDDM segregate independently; for the XbaI RFLP, linkage could be shown only if the results were weighted by the allele frequency [f(p) = 1/p], and only in the Italian and the combined (Italian and British) sib-pair groups. Multilocus analysis with both markers was also negative. We conclude that the GLUT1 gene is very unlikely to play a major role in the aetiology of NIDDM, although an accessory role cannot be excluded, and studies of the gene sequence should help to clarify this question.

Linkage analysis in nuclear families. 1: Optimality criteria for affected sib-pair tests.

The affected sib-pair method can be applied to investigate linkage between a marker locus and a disease. Several statistics have been proposed to test if the observed pattern of marker alleles shared identically by descent (ibd) is compatible with the null hypothesis of no linkage. Here, we consider different optimality criteria for sib-pair linkage tests. While for recessive inherited diseases the mean test is found to be uniformly (in theta) most powerful, it can also be shown that, irrespective of the mode of inheritance, the mean test is the locally optimal test.

Genetic approaches to common diseases.

Combined molecular and epidemiological studies are advancing our understanding of the genetic basis of multifactorial diseases. Several of the results obtained during the past year highlight methodological issues associated with these approaches. For example, the affected sib-pair method has been applied successfully to detect linkage between the angiotensinogen gene and susceptibility to hypertension, and a large multi-centre epidemiological study has demonstrated association of a polymorphism of the angiotensin-converting enzyme gene with increased risk of myocardial infarction. The study of Mendelian forms of multifactorial diseases has also led to many new results. These include the characterization of mutations in the glucokinase gene in maturity onset diabetes of the young, localization to chromosome 2 of a gene involved in familial colon cancer, and localization to chromosome 19 of a gene responsible for hemiplegic migraine. New insights have been provided into the genetics of multifactorial disorders such as diabetes and hypertension through the study of animal models. Localization of susceptibility loci in such models has recently led to the identification of new candidate genes that may be implicated in disease.

Linkage analysis of Alzheimer's disease with methods using relative pairs

Four relative-pair methods for detecting genetic linkage were applied to familial Alzheimer's disease data. Results obtained using an extended Haseman-Elston test and a weighted rank pairwise correlation test, which both use information from all relative pairs, were consistent with previously published likelihood results and appear to be more powerful than affected sib pair methods.

Homozygous parent affected sib pair method for detecting disease predisposing variants: application to insulin dependent diabetes mellitus.

For complex genetic diseases involving incomplete penetrance, genetic heterogeneity, and multiple disease genes, it is often difficult to determine the molecular variant(s) responsible for the disease pathogenesis. Linkage and association studies may help identify genetic regions and molecular variants suspected of being directly responsible for disease predisposition or protection, but, especially for complex diseases, they are less useful for determining when a predisposing molecular variant has been identified. In this paper, we expand upon the simple concept that if a genetic factor predisposing to disease has been fully identified, then a parent homozygous for this factor should transmit either of his/her copies at random to any affected children. Closely linked markers are used to determine identity by descent values in affected sib pairs from a parent homozygous for a putative disease predisposing factor. The expected deviation of haplotype sharing from 50%, when not all haplotypes carrying this factor are in fact equally predisposing, has been algebraically determined for a single locus general disease model. Equations to determine expected sharing for multiple disease alleles or multiple disease locus models have been formulated. The recessive case is in practice limiting and therefore can be used to estimate the maximum proportion of putative susceptibility haplotypes which are in fact predisposing to disease when the mode of inheritance of a disease is unknown. This method has been applied to 27 DR3/DR3 parents and 50 DR4/DR4 parents who have at least 2 children affected with insulin dependent diabetes mellitus (IDDM). The transmission of both DR3 and DR4 haplotypes is statistically different from 50% (P < 0.05 and P < 0.001, respectively). An upper estimate for the proportion of DR3 haplotypes associated with a high IDDM susceptibility is 49%, and for DR4 haplotypes 38%. Our results show that the joint presence of non-Asp at DQ beta position 57 and Arg at DQ alpha position 52, which has been proposed as a strong IDDM predisposing factor, is insufficient to explain the HLA component of IDDM predisposition.

Relationship of HLA to Schizophrenia not supported in multiplex families

The role of the human histocompatibility complex (HLA) in the pathogenesis of schizophrenia has been suggested in previous reports. We conducted a genetic study in 33 new families. Our linkage analysis, which used the affected sib-pair method, did not provide evidence for nonrandom assortment. Moreover, the results of an association study using the “haplotype relative risk” method failed to confirm the positive association between HLA A9 and schizophrenia. Taken together, our data did not support any relationship of HLA type to schizophrenia.

The genetics of IgG4 deficiency: role of the immunoglobulin heavy chain constant region and HLA loci.

IgG4 deficiency is very common (1/400 in the Italian population) and provides a good model for analyzing the genetic factors involved in Ig subclass deficiencies. We have previously reported an association between some immunoglobulin heavy chain constant region (IGHC) polymorphisms and the IgG4 deficiency. The associated polymorphisms spanned the region between the GP and the G4 genes. A larger sample composed of 50 healthy blood donors with IgG4 deficiency (less than 0.001 g/l IgG4), not carrying homozygous gene deletions, together with 82 first-degree relatives is now examined. The results confirmed the association of the deficiency with IGHC polymorphisms, and detected a new association with the HLA-D locus with a strong additive effect between the two systems. However, despite these associations and a highly significant risk for IgG4 deficiency within families, close linkage with either IGHC or HLA loci was not apparent by the affected sib pair method. These findings suggest that several concomitant, possibly cooperating, genetic factors may be involved in IgG4 deficiency.

Sib pair linkage analysis of renin gene haplotypes in human essential hypertension.

Although essential arterial hypertension is believed to have a strong genetic predisposition, the gene(s) responsible are unknown. The mechanisms underlying the regulation of blood pressure and experimental studies place the renin gene among the main candidate genes that need to be tested in humans. We tested the hypothesis of a linkage between the renin gene and essential hypertension using the affected sib pair method. Siblings (133 subjects, 52.1 +/- 10.9 years) from 57 families were selected for sustained hypertension (160.7 +/- 22.9/99.5 +/- 12.8 mmHg with 80% of patients under antihypertensive treatment), of early onset (40.7 +/- 12.0 years), in the absence of obesity, diabetes mellitus, and secondary hypertension. Eight renin haplotypes were generated from three diallelic renin restriction fragment length polymorphisms (RFLPs) (TaqI, HinfI, HindIII) located throughout the renin gene. The allelic concordance between the sib pairs was analyzed by identity by state relationships for 98 sib pairs (41 for 41 couples, 39 for 13 trios, 18 for 3 quartets). Allelic frequencies in the 57 hypertensive probands were similar to those observed among 102 hypertensive subjects studied previously. Six of eight possible haplotypes were observed, the informativity of the marker corresponded to 70% of heterozygosity. Allelic concordance for all sib pairs according to sibship size was not significantly different from that expected under the hypothesis of no linkage (t = 0.52, P = 0.15) reflecting only a small excess of renin alleles shared by the hypertensive sibs (1.44 +/- 0.6 vs 1.36 +/- 0.6). Likewise the linkage hypothesis was unsupported by weighted estimates to correct for possible bias due to large sibship size. Thus, the sib pair analysis suggests that the renin gene does not have a frequent role in the pathogenesis of essential hypertension; further more powerful linkage studies or other approaches will be needed to detect contributions at the renin locus to the heritability of essential hypertension.

Sampling strategies for linkage studies

Broadly there are three alternative sampling strategies for linkage studies of complex disorders such as schizophrenia. These are to select either affected sib pairs (and their parents) or small-to-medium-size pedigrees, or large pedigrees. The advantages and disadvantages of each are discussed. Our studies show that, even in the presence of heterogeneity, linkage can be found in clinically realistic sample sizes of nuclear families or medium-sized pedigrees. However, relying on affected sib pair methods in less satisfactory.

Affected kin‐pair IBD methods: Genetic models

Cases of interest using affected sib-pair methods to distinguish between recessive and additive (dominant) modes of inheritance of a disease-predisposing gene involve goodness-of-fit tests with a small expected number in the "share-zero parental haplotypes" category, as well as an unknown parameter, the frequency of the disease-predisposing allele. Our simulations demonstrate that the real significance level of the chi-square test using the three-haplotype-sharing IBD values (share 2, 1, and 0 parental haplotypes) is close to the assumed (.05) level in these cases, so that the haplotype-sharing classes do not have to be lumped, which would leave no degrees of freedom for a statistical test. The validity of the chi-square approximation in cases of small expected frequencies has previously been described, but the situations that have been considered do not cover the very small values in the share-zero category that are often expected in the affected sib-pair analysis, nor do they involve estimation of an unknown parameter. Although including IBD values from affected kin pairs other than sibs can be a very powerful tool in demonstrating linkage of a marker and disease, these pairs do not add power, in fact they reduce the power, of the chi-square tests of goodness-of-fit of modes of inheritance.

The Application of Molecular Genetics to the Study of Familial Arterial Hypertension

The hereditary nature of familial hypertension has been clearly established by a number of clinical studies. Most of the present work has been concentrated on the correlation between various phenotypic traits and the level of blood pressure. The development of molecular genetics allow now to establish a link between high blood pressure and specific phenotypes. As analyzed in this paper, several strategies can be used for the genetic study of arterial hypertension: linkage studies in informative families, population association studies, analysis of subjects with contrasted predisposition to high blood pressure, affected sib-pair method. The identification of the loci implicated in blood pressure regulation and which contribute to the development of arterial hypertension can then be performed in this clinical material by two main approaches. One is based on the study of candidate genes, genes whose products are known to participate in blood pressure regulation, such as those of the renin-angiotensin system which are examplified. The other involves testing a series of markers distributed randomly throughout the genome in order to establish a link between increased blood pressure and a particular region of the genome.

Co‐segregation of HLA and rheumatoid arthritis in multicase families

Inheritance of parental HLA haplotypes was examined in the offspring of 95 multicase rheumatoid arthritis (RA) families. Overall, in these families there was no evidence of preferential transmission of one parental haplotype, although this might have been expected given the loading of these families with RA cases. However, there was a difference in inheritance when the affected and non-affected offspring were compared. A co-segregation analysis showed that the inheritance of parental HLA haplotypes was different between the affected and the unaffected offspring. Unlike a previous report, no difference was demonstrated in this study between the offspring of affected and non-affected parents. Similarly, the affected offspring of HLA DR4 heterozygote parents were more likely to inherit HLA DR4 than the non-affected offspring. It is concluded first that linkage studies of RA using the affected sib-pair method are not invalidated, which would have been the case in the presence of preferential transmission of HLA to all offspring. Secondly, HLA and specifically HLA-DR4 does co-segregate with RA, and, finally, parental RA status, independent of DR4, has little influence in explaining the genetic susceptibility to RA.

Utility of the affected sib pair method to detect linkage of a sex-linked dominant disease with a sex-linked recessive trait.

Use of affected sib-pairs for linkage determination of a sex-linked dominant disease with a recessive X-chromosome marker will require extremely large sample sizes, particularly when the marker is marginally polymorphic. In addition, this method should not be used without caution on data collected for a traditional LOD score analysis, without considering the parental mating types involved in the informative families. Nevertheless, ascertainment adjustment and correction for dependence of multiple pairs of affected sib-pairs in families suggests that this method provides linkage inference concordant with that of the traditional LOD score analysis.

Genetics of IDDM: Evidence for complex inheritance with HLA

Analysis of the Fifth Genetic Analysis Workshop (GAW5) insulin-dependent diabetes mellitus (IDDM) data leads to the following conclusions: 1) With a maximum-likelihood affected sib pair method, there is strong evidence for linkage with HLA and no evidence for linkage with INS, Gm, or Km. 2) Susceptibility as defined by HLA genotypes is very complex. Each DR allele has a unique susceptibility, and DR3 and DR4 haplotype associations for DR 3/4 genotypes are different from those for 3/X and 4/X. 3) Risk is substantially higher in sibships with an affected father compared to those with an affected mother. This excess cannot be attributed to transmission distortion at HLA.

The impact of varying diagnostic thresholds on affected sib pair linkage analysis.

For Mendelian disorders, it is usually simple to classify individuals as either affected or unaffected. By contrast, for "complex" phenotypes, the diagnostic boundary of the disorder is often uncertain. This paper explores the following question: to most efficiently detect linkage in such a complex phenotype by the affected sib pair method, where should the diagnostic threshold be drawn? The model assumes that the disorder is due to a generalized two-allele single major locus (SML) where liability in each genotype is normally distributed. Evidence for linkage between the marker and disease loci is highly dependent on the location of the threshold. The relationship between the placement of the threshold and population linkage information (PLI) is Gaussian-like. At high thresholds, linkage efficiency (LE) (or the amount of linkage information per affected sib pair) is high but PLI is low because the number of affected sib pairs is very small. At low thresholds, the number of affected sib pairs is high, but PLI is low because LE is very low. The model is applied to published SML parameters for schizophrenia, and maximal PLI is achieved at thresholds broader than those for schizophrenia alone.

Lack of association and linkage between HLA and familial polyposis coli.

We investigated possible association of and linkage between HLA and familial polyposis coli (FPC). In 182 individuals from 66 pedigrees of FPC and 108 individuals from a normal population, HLA-A, -B, and -C antigens were determined. When the frequencies of HLA antigens in 66 unrelated patients and in normal controls were compared, no association of FPC with HLA was observed. For the linkage analysis, HLA haplotypes of 17 affected sib pairs were investigated by the affected sib pair method. The number of pairs which shared two, one, and no haplotypes identical by descent was not significantly different from the number expected with random occurrence (P greater than 0.95). Finally, seven families were analyzed using Morton's sequential test. A maximum lod score of -0.056 at a recombination fraction of 0.4, and a lod of -3.089 at a recombination fraction of 0.05 were obtained. Therefore, there is neither an association of nor linkage between FPC and HLA.

Relationship of HLA to schizophrenia in 10 nuclear families

We tested the hypothesis of linkage of HLA (human leukocyte antigens) and schizophrenia in 10 nuclear families that were ascertained on the basis of at least two siblings affected with chronic schizophrenia (SCH), as defined by Research Diagnostic Criteria, with or without superimposed affective disorder. All available parents and siblings were interviewed and typed for HLA antigens at the A,B,C, and Dr loci. Lod scores for linkage between HLA and a putative SCH-locus were calculated using the program LIPED, assuming several possible models for the mode of transmission of the SCH locus. Linkage was also tested with DSM-III schizotypal and paranoid personality disorders considered as part of the illness phenotype. The lod score analysis indicated that for most models of inheritance of schizophrenia, close linkage to HLA could be excluded. The results did not change if individuals with “spectrum” disorders were considered affected. We also tested for linkage using the affected sib-pair method which does not require any assumptions about the mode of transmission of the illness. The results of this analysis also indicated that linkage to HLA was very unlikely. If our results are considered together with other published studies, a susceptibility locus for schizophrenia or “schizophrenia spectrum” is not likely to be in the HLA region of chromosome 6.

HLA antigens in schizophrenia: a family study.

HLA haplotypes A and B were studied in eight families with two or more schizophrenic members. All index cases and all but two sick relatives satisfied Research Diagnostic Criteria for the diagnosis of schizophrenia; 45 subjects were studied. Data were analysed by sequential tests for the detection of linkage and by the affected sib-pair method. The results do not support a linkage between HLA and schizophrenia.

861 THE MOST FREQUENT HUMAN AUTOSOMAL RECESSIVE DISEASE

Nonclassical steroid 21-hydroxylase deficiency (nc21OHD) is an autosomal recessive disease which results in a phenotypically variable syndrome of postnatal hyperandrogenism. Its prevalence is unknown because basal serum 17-OHP levels are not sufficiently elevated for detection with the microfilter paper technique used in screening for classical 21-OHD. We therefore ascertained the frequency of the nc21OHD gene using ethnic group-specific HLA-B-nc21OHD associations in conjunction with ACTH testing in obligate heterozygote parents. Confirmation of this approach was obtained by the affected sib pair method of Thomson and Bodmer. The gene frequency for nc21OHD was highest in Ashkenazi Jews (19.1%) and was also high in Hispanics (13.6%), Yugoslavs (12.5%), and Italians (5.8%). In other Caucasians studied it was 0.1%. Disease frequencies were 1/27 for Ashkenazi Jews, 1/53 for Hispanics, 1/63 for Yugoslavs, 1/333 for Italians, and 1/1000 for other Caucasians. Carriers of the HLA-B14 marker had a 32-fold increased risk of nc21OHD compared to controls. Linkage disequilibrium betweeen HLA-B14 and nc21OHD was significant in Ashkenazi Jewish, Hispanic, and Italian patients, but not in Yugoslavs or other Caucasians.Conclusion: It appears that nc21OHD is the most frequent autosomal recessive genetic disorder in man.

Affected sib pair method for searching major gene in multifactorial disease.

The affected sib pair method to detect the effect of HLA-linked disease genes has been explained from a practical viewpoint. It is based on the assumption that HLA haplotypes as marker are used to trace the inheritance pattern of closely linked disease susceptibility genes. Looking at families in which two or more siblings had disease in question, the nonrandom zygotic assortment of HLA haplotypes would be a demonstration of disease gene.