Does 77C-->G in PTPRC modify autoimmune disorders linked to the major histocompatibility locus?

A 77G allele of the gene encoding CD45, also known as the protein tyrosine phosphatase receptor-type C gene (PTPRC), has been associated with multiple sclerosis (MS). Here we determine allele frequencies in large numbers of MS patients, primary immunodeficiencies linked to major histocompatibility complex (MHC) locus and over 1,000 controls to assess whether aberrant splicing of PTPRC caused by the 77C-->G polymorphism results in increased susceptibility to these diseases. Our results show no difference in the frequency of the 77G allele in patients and controls and thus do not support a causative role for the polymorphism in the development of disorders with a strong autoimmune component in etiology.

Epstein-Barr virus genome studies in Burkitt's and non-Burkitt's lymphomas in Uganda.

Burkitt's lymphoma (BL) has been widely investigated and has attracted attention because of the possible etiologic role of the Epstein-Barr virus (EBV). To further determine the role of EBV in the causation of this tumor, we measured EBV-specific nuclear antigen (EBNA) and EBV DNA using immunofluorescence and nucleic acid hybridization techniques, respectively. Of 34 BL biopsies, 27 tissues (79%) were EBNA-positive, whereas none of the 25 non-BL biopsy tissues were EBNA-positive. Of 15 BL tumors tested, 14 (93%) were EBV DNA-positive with a mean of 39 (range, 8-86) EBV genome equivalents per cell. Each of the 15 non-BL biopsy specimens subjected to nucleic acid hybridization had less than two virus genome equivalents per cell, although all had serologic evidence of past EBV infection. The findings further supported the possible etiologic role of EBV in African BL and negated the passenger hypothesis. The EBV genome could, therefore, be used as a separating marker between African BL and non-BL lymphomas.

Documentation of Epstein-Barr virus infection in immunodeficient patients with life-threatening lymphoproliferative diseases by Epstein-Barr virus complementary RNA/DNA and viral DNA/DNA hybridization.

Tissues from patients thought to have Epstein-Barr virus (EBV)-induced lymphoproliferative diseases were probed for EBV genomes using 2 independent hybridization techniques. Tissues from six patients with the X-linked lymphoproliferative syndrome, all five renal allograft recipients with immunoblastic sarcoma, and eight patients with diverse types of immunodeficiency and lymphoproliferative diseases such as fatal infectious mononucleosis or malignant lymphoma associated with antecedent immunodeficiency contained significant numbers of EBV genome equivalents per cell. The use of 2 hybridization probes is recommended to confirm the presence of EBV genomes. The finding of significant numbers of EBV genomes in tissues from patients with immunodeficiency suggests that EBV is the etiological agent of the associated lymphoproliferative diseases.

Documentation of Epstein-Barr virus infection in immunodeficient patients with life-threatening lymphoproliferative diseases by clinical, virological, and immunopathological studies.

Multiple methods, pedigree analysis, clinical evaluation, and Epstein-Barr virus (EBV)-specific serology, EBV DNA hybridization of tissues to probe for viral genome, staining of touch imprints for EBV nuclear-associated antigen, establishment of spontaneous infected B-cell lines from peripheral blood or tissues, examination of peripheral blood smears, and hematopathology studies, were used to study seven patients with the X-linked lymphoproliferative syndrome and seven additional patients with life-threatening EBV-associated diseases. These studies demonstrated EBV in the tissues of all 14 patients and immunodeficient antibody responses to EBV were documented. This virus can produce various life-threatening lymphoproliferative diseases in a variety of immunodeficient patients.

Segregation of a totally skewed pattern of X chromosome inactivation in four familial cases of Rett syndrome without MECP2 mutation: implications for the disease.

BACKGROUND: Rett syndrome is a neurodevelopmental disorder affecting only girls; 99.5% of Rett syndrome cases are sporadic, although several familial cases have been reported. Mutations in the MECP2 gene were identified in approximately 70-80% of sporadic Rett syndrome cases. METHODS: We have screened the MECP2 gene coding region for mutations in five familial cases of Rett syndrome and studied the patterns of X chromosome inactivation (XCI) in each girl. RESULTS: We found a mutation in MECP2 in only one family. In the four families without mutation in MECP2, we found that (1) all mothers exhibit a totally skewed pattern of XCI; (2) six out of eight affected girls also have a totally skewed pattern of XCI; and (3) it is the paternally inherited X chromosome which is active in the patients with a skewed pattern of XCI. Given that the skewing of XCI is inherited in our families, we genotyped the whole X chromosome using 32 polymorphic markers and we show that a locus potentially responsible for the skewed XCI in these families could be located on the short arm of the X chromosome. CONCLUSION: These data led us to propose a model for familial Rett syndrome transmission in which two traits are inherited, an X linked locus abnormally escaping X chromosome inactivation and the presence of a skewed XCI in carrier women.

Identification of four novel polymorphisms in the calcitonin/alpha-CGRP (CALCA) gene and an investigation of their possible associations with Parkinson disease, schizophrenia, and manic depression.

We identified novel polymorphisms in the calcitonin/CGRPalpha (CALCA) gene by direct sequencing of genomic DNA and subsequent genotyping by RFLP (restriction fragment length polymorphism) detection and investigated association with neurological or psychiatric disease. Four novel polymorphic alleles were found: two (g.979G>A and g.4218T>C) represented single nucleotide polymorphisms (SNPs), one consisted of two coupled SNPs in close vicinity to each other (g.1210T>C and g.1214C>G), and one was an intronic 16-bp microdeletion (2919-2934del16). One of the SNPs (g.4218T>C) causes a non-synonymous amino acid change (Leu66Pro) in the third exon, an exon common to both procalcitonin and pro-alpha-CGRP. In a subsequent association study, frequencies of the identified polymorphisms in Parkinson and schizophrenia patients were compared with frequencies in the normal population. No statistically significant association was found in our material. The 16-bp microdeletion polymorphism was present in a family with multiple cases of unipolar or bipolar depressive disorder. Using this polymorphism as marker, cosegregation with the phenotype was observed in the majority of individuals.

Larger CAG expansions in skeletal muscle compared with lymphocytes in Kennedy disease but not in Huntington disease.

The size of CAG repeats was compared in lymphocytes and skeletal muscle from nine patients with Huntington disease (HD) and two patients with Kennedy disease (KD). In HD, the number of CAG repeats did not differ between lymphocytes and skeletal muscle. In the two KD patients, however, the CAG expansion was larger in muscle than in lymphocytes. The difference in trinucleotide expansion between lymphocytes and muscle cells is not a universal phenomenon in trinucleotide repeat disorders, but seems to occur in disorders primarily affecting the neuromuscular system.

C4342T-mutation in the SCN4A gene on chromosome 17q in a Swedish family with paramyotonia congenita (Eulenburg)--correlations with clinical, neurophysiological and muscle biopsy data.

Genetic analysis of the adult muscle sodium channel alpha-subunit, SCN4A gene on chromosome 17q, was performed by means of PCR technique in a Swedish family with paramyotonia congenita (Eulenburg) (PMC). The mutation was found in four family members and consisted of a C to T transition affecting the fourth domain of the sodium channel protein. This mutation has earlier been described in other families with paramyotonia congenita. All family members carrying the mutation had cold-induced paradoxical myotonia, myotonic bursts on EMG, and a type IIB atrophy on muscle biopsy. Three of them had slight CK elevation and two had episodes of paralysis. On the basis of clinical findings in this family, persistent proximal muscle weakness, myopathic EMG abnormalities, a type IIB atrophy on muscle biopsy and no symptoms but other signs of muscle affection, were earlier suggested as clinical features of PMC. However, genetic analysis revealed that family members with these symptoms and findings did not have the mutation, indicating that these features are not due to PMC.

Welander distal myopathy is not linked to other defined distal myopathy gene loci.

Welander distal myopathy is an autosomal dominant disorder with late onset that affects extensor muscles of the hands and the feet. The disorder is considered as the most prevalent of the distal myopathies and is almost only seen in Sweden and in some parts of Finland. On clinical, morphological and genetical grounds the disorder is clearly separated from other distal myopathies. We have performed linkage analysis with the MLINK program in a total of six families with microsatellite markers dispersed throughout the genome and report exclusion for the localisation of the gene of 64% of the human genome. These studies have clearly separated Welander distal myopathy from previously mapped forms of distal myopathy such as the Miyoshi myopathy by excluding linkage to chromosome 2. The region on 14q that has been suggested to house the gene of the distal myopathy described by Laing et al. (Am J Hum Genet 1995;56:422-7), has as well been excluded by several markers.

Welander hereditary distal myopathy, a molecular genetic comparison to hereditary myopathies with inclusion bodies.

Welander distal myopathy (WDM) is an autosomal dominant disorder with late onset predominantly affecting distal extensor muscles of the hands and the feet. The disorder is considered as the most common of the distal myopathies but is almost only seen in Sweden and some parts of Finland. The finding of rimmed vacuoles in muscle biopsies from patients with moderate and severe symptoms constitutes one similarity with hereditary inclusion body myopathy (HIBM) sparing the quadriceps as described by Argov and Yarom [Argov Z, Yarom R. J Neurol Sci 1984;64:33-43]. The question has been raised whether some of the different forms of distal myopathy might be allelic. In previous reports the gene defects for HIBM and autosomal recessive hereditary distal myopathy with rimmed vacuoles (DMRV) have been mapped to chromosome 9pl-q1. The Finnish tibial muscular dystrophy (TMD) that displays similar histopathological findings has recently been linked to chromosome 2q. We have investigated the regions of interest with dispersed microsatellite markers in four well-described pedigrees, and this study now excludes the regions on chromosome 9pl-q1 and 2q from linkage to WDM both by haplotype analysis and linkage analysis with the MLINK program. WDM, showing morphological similarities with HIBM, is clearly separated from the disorders mapped to chromosomes 9 and 2 on clinical and genetical grounds.

Welander distal myopathy--an overview.

Welander distal myopathy has an autosomal dominant inheritance and a late onset. The onset of symptoms is in the hands and gradually distal muscles of the lower extremities are involved. The most-affected muscles are the long extensors of the hands and feet. CK-values are normal or slightly elevated. There is never any cardiac involvement in Welander distal myopathy. Neurophysiological findings are of both myopathic and neuropathic character. Histopathological findings in muscle biopsies are mainly of myopathic type and include rimmed vacuoles which correspond to autophagic vacuoles on the ultrastructural level. Tubulo-filamentous inclusions with a diameter of 16-21 nm, i.e. of the same type as found in patients with Inclusion Body Myositis, are found in the sarcoplasm and in myofibre nuclei. A neurogenic component in Welander distal myopathy has been suggested, on the grounds of a sensory dysfunction, neuropathic findings on neurophysiology and muscle biopsy and a decrease of A-delta nerve fibres on sural nerve biopsy. Genetic analysis has excluded linkage to other defined distal myopathies and hereditary Inclusion Body Myopathy loci.

Variation of CTG-repeat number of the DMPK gene in muscle tissue.

Myotonic dystrophy (DM) is associated with an unstable expansion of CTG repeats located in the 3' untranslated region of a protein kinase-encoding gene (DMPK) on chromosome 19 (19q13.3). The CTG repeat number varies between 5 and 37 in lymphocytes of normal individuals, whereas DM patients may have expansions from 50 to several thousand copies. Although the CTG expansions related to myotonic dystrophy (DM) are usually larger in muscle compared to peripheral blood, the variation in repeat number in non-dystrophic muscle is not known. In order to investigate if there is a variation, the CTG-repeat number was determined in percutaneous muscle biopsies obtained from 86 individuals without any muscle disorder or with a neuromuscular disorder without any clinical or histopathological signs of DM. The number of CTG repeats varied between 5 and 28, this being within the normal range reported for peripheral blood. A major sharp peak at n = 5 (27%) and a broader peak at n = 8-17 (56%) with peak values at n = 12 and 14 (11 and 14%, respectively) were observed. Alleles with 19 or more repeats amounted to 17% with a small peak at n = 20 and 21 (6 and 4%, respectively). It is concluded that the normal variation of CTG-repeat number in skeletal muscle is within the range found in peripheral blood, although there is a slight shift in the overall frequency distribution towards alleles with CTG repeat numbers in the higher range.

A second locus for autosomal dominant myopathy with proximal muscle weakness and early respiratory muscle involvement: a likely chromosomal locus on 2q21.

We recently mapped a locus for a new variant of autosomal dominant myopathy (Swedish families) with proximal muscle weakness, early respiratory muscle involvement, and unique muscle biopsy findings to chromosomal region 2q24-31. In this study, a French family with a similar clinical phenotype and pathology (muscle biopsy) was investigated to see whether the disease gene associated with the myopathy is mapped to the same region as the one in the Swedish families; however, chromosomal region 2q24-q31 was completely excluded. In order to localise the disease gene for the French family, a genome-wide scan was performed using polymorphic microsatellite markers. A maximum two-point lod score of 2.11 (the highest lod score that can be achieved in this family) was obtained for the markers in the region between D2S1272 and D2S1260, spanning 4 cM. This result suggests that the gene responsible for the French form is likely to be located on chromosome 2q21.

Lack of evidence of permanent engraftment after in utero fetal stem cell transplantation in congenital hemoglobinopathies.

The use of fetal hematopoietic stem cells for in utero transplantation to create permanent hematochimerism represents a new concept in fetal therapy. In one fetus with alpha-thalassemia, one with sickle cell anemia, and one with beta-thalassemia, we have transplanted fetal liver cells obtained from legal abortions in gestational weeks 6-11. The fetus with alpha-thalassemia was transplanted twice during pregnancy, in the 15th (20.4 x 10(8) cells/kg) and in the 31st weeks of gestation (1.2 x 10(8) cells/kg), and is now two years of age. One fetus with sickle cell anemia received its transplant in the 13th week of gestation (16.7 x 10(8) cells/kg), and is now one year old. The fetus with beta-thalassemia was transplanted in 18th week (8.6 x 10(8) cells/kg), and is now three months old. Engraftment was evaluated by chromosomal analysis (sex chromosomes), red cell phenotyping, HLA class I and II typing, and PCR (polymerase chain reaction) for Y chromosome-specific sequences and DNA polymorphisms in cord and peripheral blood. The children with alpha- and beta-thalassemia underwent bone marrow aspirations at 3 and 7 months of age, respectively. In neither of these cases were we able to detect convincing evidence of stem cell engraftment. Thus, the administration of fetal stem cells to fetal recipients after the 12th week of gestation did not result in permanent hematochimerism. It remains to be determined whether the engraftment process can be promoted by earlier transplantations and/or higher cell doses.

Segregation analysis of the X-chromosome in a family with Rett syndrome in two generations.

We report on the first family in which Rett syndrome (RTS) appeared in two consecutive generations. The index case is a 12-year-old girl (classical RTS); her maternal aunt, age 44 years, has mild RTS. Clinically, the family illustrates the wide phenotypic variability between cases, particularly in severity of neurological manifestations. We have analyzed the short arm of the X-chromosome of the family with gene technology. This did not uncover any genetic marker for diagnosis, but it did suggest how the syndrome might have segregated in the family. A cytogenetic analysis gave no information about chromosome abnormalities.

Reactions to predictive testing in Huntington disease: case reports of coping with a new genetic status.

A predictive testing program for Huntington disease has been available in Stockholm, Sweden since October 1990. Psychosocial assessments were performed throughout the testing program to evaluate the impact of the risk situation itself and the effect of predictive testing, and to identify those individuals who were most vulnerable to severe stress and anxiety reactions. All subjects underwent neurological, neuropsychological, and psychiatric examinations. Individuals undergoing predictive testing were assessed twice by a genetic counsellor before receiving their results, and at 10 days (gene carriers only) and then 2, 6, 12, and 24 months after receiving the results. The process of coping with the test results and the psychological adjustment to knowledge about new genetic status have been shown to vary considerably. In this report, we describe the results obtained from two gene carriers and two noncarriers. The four persons chosen represent different ways of coping with the outcome of the test and of integrating knowledge about their genetic status into everyday life. These cases illustrate common themes and recurrent problems often surfacing during the counselling and testing process. The longitudinal evaluations provide information about the impact, adaptation, and long-term effects of living with a new genetic status.

Aicardi syndrome: early neuroradiological manifestations and results of DNA studies in one patient.

A patient with Aicardi syndrome is presented. We report results of DNA analysis from the patient and her parents with probes mapped to Xp21.3-22.3 in an attempt to localize a deletion in this region. No signs of a microdeletion could be detected, using 5 different DNA markers. Further, it is suggested that a specific combination of cerebral abnormalities may be characteristic of the syndrome and that antenatal ultrasonographic diagnosis may be feasible.

Prader-Willi syndrome in a brother and sister without cytogenetic or detectable molecular genetic abnormality at chromosome 15q11q13.

We report on a 12-year-old boy and his 7-year-old sister with the Prader-Willi syndrome. They both had severe initial hypotonia with feeding problems and later developed an increasing appetite. Both sibs have almond-shaped eyes, triangular mouth, hypogonadism, retarded growth, and mental retardation. An older brother suffered from severe hypotonia and died at 7 days of age. The children have normal chromosomes by high-resolution technique and have inherited the same chromosomes 15 short arm polymorphisms from their parents. The family was informative for one of four DNA markers specific for the 15q11q13 region. No deletion was found using this marker. The parents were healthy and unrelated. Autosomal recessive inheritance or a paternally inherited submicroscopic deletion are possible explanations for the sib occurrence in this family.

NURR1 mutations in cases of schizophrenia and manic-depressive disorder.

Transgenic mice lacking the nuclear orphan transcription factor Nur-related receptor 1 (Nurr1) fail to develop mesencephalic dopamine neurons. There is a highly homologous NURR1 gene in humans (formerly known as NOT) which therefore constitutes a good candidate gene for neurologic and psychiatric disorders with an involvement of the dopamine neuron system, such as Parkinson's disease, schizophrenia, and manic-depression. By direct sequencing of genomic DNA, we found two different missense mutations in the third exon of NURR1 in two schizophrenic patients and another missense mutation in the same exon in an individual with manic-depressive disorder. All three mutations caused a similar reduction of in vitro transcriptional activity of NURR1 dimers of about 30-40%. Neither of these amino acid changes, nor any sequence changes whatsoever, were found in patients with Parkinson's disease or control DNA material of normal populations. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:808-813, 2000.