Enhanced frequency of a PTPRC (CD45) exon A mutation (77C-->G) in systemic sclerosis.

A point mutation in exon A (C to G transversion at position 77) of human PTPRC (CD45) has recently been associated with the development of multiple sclerosis (MS) for at least a subgroup of patients. In the present report, we studied the frequency of the 77C-->G transversion in two other autoimmune diseases namely systemic sclerosis (SSc) and systemic lupus erythematosus (SLE). The mutation was found with significantly enhanced frequency in patients suffering from SSc suggesting that PTPRC could play a role as susceptibility gene not only in MS but also in other autoimmune diseases. Further understanding of the mode of interaction of mutant PTPRC with other susceptibility genes may uncover mechanisms common in various autoimmune disorders.

Residual chloride secretion in intestinal tissue of deltaF508 homozygous twins and siblings with cystic fibrosis. The European CF Twin and Sibling Study Consortium.

BACKGROUND & AIMS: Cholinergic stimulation of chloride secretion is impaired in the intestines of patients with cystic fibrosis (CF). However, intestinal chloride secretion has been observed in patients with mild CF mutations. The aim of this study was to investigate residual Cl(-) secretion in the intestine of DeltaF508 homozygous CF patients, and examine the contribution of cystic fibrosis transmembrane conductance regulator (CFTR) and alternative Cl(-) conductances. Twins and siblings with identical CFTR genotypes were investigated to determine the impact of factors other than CFTR on chloride secretion. METHODS: Chloride secretion in rectal tissue was investigated by applying Ca(2+) and adenosine 3',5'-cyclic monophosphate (cAMP)-linked agonists before and after the inhibition of alternative Cl(-) conductances with 4,4'-diisothiocyanostilbene-2, 2'-disulfonic acid (DIDS). RESULTS: cAMP-mediated Cl(-) secretion was observed in 73% of patients, and 20% showed DIDS-sensitive Ca(2+)-activated Cl(-) secretion. This DIDS-sensitive alternative chloride conductance was seen only in CF patients who also responded to cAMP agonists. Chloride secretion was more concordant within monozygous twins than within dizygous pairs. CONCLUSIONS: These results suggest the presence of CFTR-mediated Cl(-) secretion in a subgroup of patients, implying that a portion of deltaF508 CFTR can be processed in vivo and function as a chloride channel in the apical membrane of intestinal cells. Moreover, a considerable number of deltaF508 homozygous patients express chloride conductances other than CFTR in their intestinal epithelia.

Genetic variation at 9 autosomal microsatellite loci in Asian and Pacific populations.

Genetic variation at 9 autosomal microsatellite loci (CFS1R, TH01, PLA2A, F13A1, CYP19, LPL, D20S481, D20S473, and D20S604) has been characterized in 16 Asian and Oceanic populations, mostly from mainland and insular Southeast Asia. The neighbor-joining tree and the principal coordinates analysis of the genetic relationships of these populations show a clear separation of Papua New Guinea Highlanders and, to a lesser extent, Malayan aborigines (Orang Asli or Semai) from the rest of the populations. Although the number of markers used in this study appears to be inadequate for clarifying the patterns of genetic relationships among the studied populations, in the principal coordinates analysis a geographic trend is observed in the mainland and insular Southeast Asian populations. Furthermore, in an attempt to contrast the extent of variation between autosomal and Y-chromosome-specific microsatellite loci and to reveal potential differences in the patterns of male and female migrations, we have also compared genetic variation at these 9 autosomal loci with variation observed at 5 Y-chromosome-specific microsatellites in a common set of 14 Asian populations.

Analysis of five Y-specific microsatellite loci in Asian and Pacific populations.

We have analyzed five Y-specific microsatellite loci (DYS388, DYS390, DYS391, DYS394, DYS395) in 17 Asian and Pacific populations representing a broad geographical area and different linguistic families, with an emphasis on populations from mainland and insular Southeast Asia. Analysis of gene diversity indicates that several of the studied populations have experienced substantial genetic isolation, and a reduction in male effective sizes (viz. the Northeast Indian populations Nishi, Adi and the Taiwanese aboriginals). The average values of the F(ST) and ((ST) statistics indicate a high degree of genetic differentiation among these populations at the five Y-specific markers (F(ST) =0.21 and ((ST) = 0.33, based on individual loci; F(ST) = 0.09 and ((ST) = 0.36, based on haplotypes), which conform to the expectation of a fourfold smaller effective size of the Y-linked loci compared with the autosomal loci. Dendrogram and principal coordinates analysis, with few exceptions, show a major separation between mainland and insular populations. Among the mainland populations, the Tibeto-Burman speakers from Northeast India cluster in a well-defined group, supported by high bootstrap values. The Southern Chinese, Northern Thai, So, and Cambodian also are integral to this cluster. The other major cluster is rather heterogeneous and includes, among others, the Austronesian-speaking populations. The Samoans of the Pacific, with a distinctive pattern of allelic distributions, stand as an outlier in the tree and PC representations. Although trends of genetic affinities among ethnically and geographically related populations are evident from the Y-specific microsatellite data, microsatellites are not optimal for deciphering complex migratory patterns of human populations, which could possibly be clarified by using additional and more stable genetic markers.

Development of microchimerism in pediatric patients after living-related liver transplantation.

Microchimerism has been suggested to play an important role in the long-term acceptance of allogeneic organ grafts by transplant patients and for the maintenance of a state of donor-specific low responsiveness. In order to elucidate the kinetics of the development of chimerism we have performed a follow-up analysis in 10 pediatric patients with living-related liver transplantation (LRLTx). Blood samples obtained during the first 6 months and at 18 months post-transplant and skin biopsies taken at one month were analysed for the presence of donor cells by PCR using donor-specific HLA-DRB1 primer pairs or primers for a Y chromosome-specific sequence. Furthermore 13 long-term patients more than 2 yr after LRLTx were studied at two different time points. In the follow-up studies donor cells could be demonstrated in the blood of all patients immediately after transplantation. After a gradual decline all patients became chimerism-negative for several weeks or months. At 6 months, however, in five of eight patients tested and at 18 months in six of nine patients donor cells had reappeared. This biphasic pattern in the development of chimerism is proposed to reflect the occurrence of different donor-derived cell populations in the recipient. The population giving rise to the first wave of chimerism probably represents matured cells with a limited lifespan which are released from the graft into the circulation of the recipient during the first weeks after transplantation. The population of cells occurring with the second wave of chimerism is likely to have been generated by donor-derived cells with stem cell potential located either in the graft or in the hematopoetic organs of the recipient after emigration from the graft. This model may be able to explain fluctuations in the incidence and degree of microchimerism described in other patient populations during the first year post-transplant. Of the 13 long-term patients, chimerism could be demonstrated in 11. In seven patients it was detected in both blood and skin, in three patients the results obtained for blood and skin were discordant. In one patient only blood was analysed. It is not clear whether the negative results really reflected the absence of chimerism or whether the number of donor cells was below the level of detectability.

Distribution and evolution of CTG repeats at the myotonin protein kinase gene in human populations.

We have analyzed the CTG repeat length and the neighboring Alu insertion/deletion (+/-) polymorphism in DNA samples from 16 ethnically and geographically diverse human populations to understand the evolutionary dynamics of the myotonic dystrophy-associated CTG repeat. Our results show that the CTG repeat length is variable in human populations. Although the (CTG)5 repeat is the most common allele in the majority of populations, this allele is absent among Costa Ricans and New Guinea highlanders. We have detected a (CTG)4 repeat allele, the smallest CTG known allele, in an American Samoan individual. (CTG) > or = 19 alleles are the most frequent in Europeans followed by the populations of Asian origin and are absent or rare in Africans. To understand the evolution of CTG repeats, we have used haplotype data from the CTG repeat and Alu(+/-) locus. Our results are consistent with previous studies, which show that among individuals of Caucasian and Japanese origin, the association of the Alu(+) allele with CTG repeats of 5 and > or = 19 is complete, whereas the Alu(-) allele is associated with (CTG)11-16 repeats. However, these associations are not exclusive in non-Caucasian populations. Most significantly, we have detected the (CTG)5 repeat allele on an Alu(-) background in several populations including Native Africans. As no (CTG)5 repeat allele on an Alu(-) background was observed thus far, it was proposed that the Alu(-) allele arose on a (CTG)11-13 background. Our data now suggest that the most parsimonious evolutionary model is (1) (CTG)5-Alu(+) is the ancestral haplotype; (2) (CTG)5-Alu(-) arose from a (CTG)5-Alu(+) chromosome later in evolution; and (3) expansion of CTG alleles occurred from (CTG)5 alleles on both Alu(+) and Alu(-) backgrounds.

Development, stability, and clinical correlations of allogeneic microchimerism after solid organ transplantation.

To assess the development, stability, and clinical relevance of donor-type microchimerism, skin and blood were analyzed in heart (n = 53) and liver (n = 18) transplant recipients by nested polymerase chain reaction. Microchimerism was detectable in 40 (75%) and 13 (72%) patients after heart and liver transplantation, respectively. In heart transplantation, chimerism-positive patients showed a lower frequency of acute rejection as compared with negative patients, although this was only of borderline statistical significance. Repeated intraindividual analyses demonstrated variable patterns of microchimerism over time, but changes did not correlate to the clinical state. In liver transplantation, chimeric state showed no clear correlation with the patients' immunological situation. Our results demonstrate that peripheral microchimerism frequently develops after different types of organ transplantation and represents a dynamic process but without diagnostic value to predict the immunological risk for individual patients.

A point mutation in the human CD45 gene associated with defective splicing of exon A.

CD45 is a receptor-type protein tyrosine phosphatase involved in the regulation of lymphocyte activation. Different CD45 isoforms are generated by alternative splicing of three variable exons (A, B and C). The pattern of CD45 splicing depends upon cell type and state of activation. CD45RA isoforms (containing exon A-encoded sequences) can usually be found on a subset of resting T cells, but not on activated T cells. We have recently described a variant pattern of CD45RA expression which is characterized by continuous expression of CD45RA molecules on activated and memory T cells. Here, we demonstrate that this phenotype is associated with heterozygosity for a point mutation at nucleotide position 77 of exon A, leading to a C-->G transition. This mutation does not change the protein sequence of the CD45RA isoform. We conclude that position 77 is part of a motif necessary for splicing of exon A, which supports the hypothesis that sequences within exons have significant effects on alternative splicing. The mutation of this motif might prevent binding of a transacting splice factor. In the heterozygous state, this mutation is not associated with impaired T cell reactivity. Functional consequences of the homozygous state remain to be elucidated.

Recurrence of hepatitis B virus cirrhosis and hepatocellular carcinoma: an indication for retransplantation?

Hepatitis B virus reinfection as well as recurrence of hepatocellular carcinoma are frequent complications in liver allograft recipients. This is the report of a patient who had two liver transplantations with a 6-year interval, both for the same indication-postnecrotic cirrhosis and liver cancer. Following first and second transplantation, hepatitis B reinfection occurred at 12 and 6 months, and allograft cirrhosis developed after 20 and 10 months, respectively. Intrahepatic recurrence of hepatocellular carcinoma was found after 69 months. Two years following retransplantation, the patient is tumor-free, and has normal graft function. In selected patients with hepatitis B-related liver disease, and hepatocellular carcinoma liver replacement is indicated when combined with effective immunoprophylaxis, and other adjuvant therapy. Late recurrence of both diseases is unusual, and retransplantation may come into question.

Population genetics of dinucleotide (dC-dA)n.(dG-dT)n polymorphisms in world populations.

We have characterized eight dinucleotide (dC-dA)n.(dG-dT)n repeat loci located on human chromosome 13q in eight human populations and in a sample of chimpanzees. Even though there is substantial variation in allele frequencies at each locus, at a given locus the most frequent alleles are shared by all human populations. The level of heterozygosity is reduced in isolated or small populations, such as the Pehuenche Indians of Chile, the Dogrib of Canada, and the New Guinea highlanders. On the other hand, larger average heterozygosities are observed in large and cosmopolitan populations, such as the Sokoto population from Nigeria and German Caucasians. Conformity with Hardy-Weinberg equilibrium is generally observed at these loci, unless (a) a population is isolated or small or (b) the repeat motif of the locus is not perfect (e.g., D13S197). Multilocus genotype probabilities at these microsatellite loci do not show departure from the independence rule, unless the loci are closely linked. The allele size distributions at these (CA)n loci do not follow a strict single-step stepwise-mutation model. However, this features does not compromise the ability to detect population affinities, when these loci are used simultaneously. The microsatellite loci examined here are present and, with the exception of the locus D13S197, are polymorphic in the chimpanzees, showing an overlapping distribution of allele sizes with those observed in human populations.

Patterns of donor-type microchimerism after heart transplantation.

Allogeneic microchimerism of donor-type has been demonstrated in stable patients in the long-term after organ transplantation. We have analysed microchimerism in skin and blood of 47 heart-transplanted patients after transplantation with polymerase-chain-reaction amplification specific for donor HLA-DRB1. Microchimerism was detectable in 50% of the patients in the first 6 months, in 100% between 6 months and 2 years, and in 58% in the third postoperative year or later. The state of chimerism was not related to acute or chronic rejections. Patterns of microchimerism after heart transplantation may be dynamic, but any association with clinical and immunological variables remains to be elucidated.

Selected di- and tetranucleotide microsatellites from chromosomes 7, 12, 14, and Y in various Eurasian populations.

Microsatellite polymorphisms of nine Eurasian populations (> 1200 chromosomes) were analyzed for the following loci: i) intronic (gt)n stretches of three T cell receptor (TCR) B loci on chromosome 7 (TCRBV6S1, TCRBV6S3, TCRBV6S7); ii) an intergenic (gt)n repeat in the region between the TCRDV3 and TCRAJ61 elements on chromosome 14; iii) two tetranucleotide simple repeats (D12S66, D12S67), not linked to known genes on chromosome 12; iv) a Y-chromosomal (gata)n polymorphism (DYS19). In general, allele frequencies and heterozygosity rates were similar, but specific alleles were missing in one or more populations. Distinct DYS19 alleles predominated in particular cohorts. Different allele frequencies were observed for the TCR loci in European and Asian populations. Tetranucleotide polymorphisms were distributed normally, whereas TCR alleles displayed bimodal frequency profiles. For TCRBV6S1 and TCRBV6S7, this profile reflects a diallelic protein polymorphism that correlates exactly with the length of the intronic repeats.