Non-motor symptoms and quality of life in dopa-responsive dystonia patients.

BACKGROUND: In patients with GTP-cyclohydrolase deficient dopa-responsive dystonia (DRD) the occurrence of associated non-motor symptoms (NMS) is to be expected. Earlier studies report conflicting results with regard to the nature and severity of NMS. The aim of our study was to investigate the prevalence of psychiatric disorders, sleep problems, fatigue and health-related quality of life (HR-QoL) in a Dutch DRD cohort. METHODS: Clinical characteristics, motor symptoms, type and severity of psychiatric co-morbidity, sleep problems, fatigue and HR-QoL were assessed in DRD patients with a confirmed GCH1 mutation and matched controls. RESULTS: Twenty-eight patients were included (18 adults and 10 children), from 10 families. Dystonia symptoms were well-controlled in all patients. According to the DSM IV patients significantly more often met the criteria for a lifetime psychiatric disorder than controls (61% vs. 29%, p < 0.05). In particular the frequencies of generalized anxiety and agoraphobia were higher in patients (both 29% vs. 4%, p < 0.05). Patients scored significantly higher on daytime sleepiness than controls (ESS, 11.2 vs 5.7, p < 0.05). Adult patients had significantly lower scores on the mental component of the HR-QoL (47 vs. 54, p < 0.05) than controls mainly associated with (worse) quality of sleep. CONCLUSION: NMS were highly prevalent in our cohort of DRD patients, despite adequate treatment of motor symptoms. Our findings support the accumulating evidence of an important non-motor phenotype in DRD, with possible involvement of serotonergic mechanisms. This highlights the need to address NMS and the underlying neurobiology in patients with DRD.

Differentiating among children with PDD-NOS, ADHD, and those with a combined diagnosis on the basis of WISC-III profiles.

Pervasive Developmental Disorder-Not Otherwise Specified (PDD-NOS) and Attention Deficit/Hyperactivity Disorder (ADHD) have partly overlapping symptoms. It can also be debated whether a third diagnostic category exists: children with a combined diagnosis. In this study an attempt was made to distinguish among the three groups on the basis of intelligence (WISC-III) profiles. It was found that the PDD-NOS group had higher verbal and performance IQ's, as well as higher WISC-III index scores than the ADHD group. Subtests Block Design and Mazes discriminated best. It was concluded that based on intelligence scores, only PDD-NOS and ADHD emerged as distinct categories, whereas the combined diagnosis did not. Future research on the distinctiveness of these diagnostic groups, however, should include variables other than IQ.

Hypervariable region diversity of hepatitis C virus and humoral response: comparison between patients with or without cirrhosis.

To investigate the potential clinical utility of antibody response to HVR1 of HCV, the genomic and amino acid diversity of HVR1 was compared between two groups of four chronic HCV carriers with or without liver cirrhosis. Peptides corresponding to the deduced COOH- and NH2-terminal amino acid sequences of HVR1 were synthesised to assess the reactivity of patient sera to autologous and homologous HVR1 epitopes by enzyme-linked immunosorbent assay. HCV chronic carriers had significantly more frequent cross-reactivity with homologous C- than N-terminal HVR1 peptides. Twelve cirrhotic and eleven noncirrhotic patients had a similar frequency of cross-reactivity with either C- or N-terminal HVR1 peptides. However, noncirrhotic patients had a significantly higher level of C-terminal HVR1 antibody cross-reactivity than cirrhotic patients. In HCV chronic carriers, the magnitude of the immune response to but not the frequency of cross-reactivity with C-terminus HVR1 peptides differ between patients with and without liver cirrhosis.

Molecular genotyping of HIV-1 in 61 patients with AIDS from Lomé, Togo.

To study the distribution of HIV types and genotypes, in Lomé, Togo, a random population of patients who met the clinical criteria of the Bangui definition of AIDS and were positive with two independent screening assays for antibodies to HIV-1 group M, HIV-2, and HIV-1 group O was selected. HIV RNA from serum samples was reverse-transcribed and amplified with degenerate primers annealing to conserved regions of the HIV-1, HIV-2, and HIV-O gag gene. Amplicons were directly sequenced using an automated sequencer. A 262-271-bp (strain-dependent) fragment of the gag gene from each patient was phylogenetically analyzed and compared to the corresponding gag sequences of published HIV-1 sequences of known African genotypes. Genotype A was found in 48 of 60 patient amplicons (80%), subdivided into two clusters. Ten patients (16.7%) were HIV-1 genotype G; one was genotype D and one genotype H. HIV-1 genotype B was not found. Amplicons from two patients contained sequence ambiguities, requiring cloning and sequencing of the gag insert. One patient (T52) was apparently infected with HIV-1 genotypes A and G; whereas HIV-1 from patient T139 was of genotype A, with 2/10 clones having a three-codon insertion at nucleotide position 1142 of the gag gene. HIV-1 genotype A is dominant in Togo; genotype G is frequent and genotype B has not been found.

A pro-B-cell stage characterized by germline Ig transcription without surrogate light chain expression.

B-cell commitment is characterized by the expression of specific membrane proteins and the rearrangement and expression of immunoglobulin (Ig) heavy (H) and light (L) chain genes. At early stages of B-cell development, unrearranged Ig loci are transcribed, which correlates with these regions becoming accessible for Ig gene rearrangement. Some germline transcripts can be translated into protein and potentially play a role in cell signaling during B-cell development. In this report an early stage in human B-cell development is characterized using Epstein-Barr virus (EBV)-transformed cell lines from patients with a severe combined immunodeficiency (SCID). These lines were shown to produce germline constant (C) gene transcripts from the IGH and IGK loci. We demonstrate here that these cells are committed to the B-cell lineage as substantiated by expression of CD79a and CD79b. No surrogate light chain (SLC) gene transcription was detected, indicative of a very early differentiation stage. From these cell lines two types of germline IgV gene transcripts were isolated: a transcript containing the IGKV4-1 gene and a germline IGHV-1 transcript nearly identical to IGHV1/OR15-1 (HC15-1, DP-1), an orphon VH gene on chromosome 15. Germline VH transcripts originating from the VH locus on chromosome 14 could not be detected. It is of interest that, apart from Ig V and C genes (non-functional), V genes that reside outside the Ig locus are a target for the transcription factors that are postulated to initiate Ig gene rearrangement early in B-cell development.

Immunoglobulin heavy chain germ-line JH-C mu transcription in human precursor B lymphocytes initiates in a unique region upstream of DQ52.

From human precursor B cells which had both immunoglobulin (Ig) heavy (H) chain loci in germ-line configuration, various IgH chain germ-line transcripts were isolated and sequenced. These transcripts were shown to contain sequences derived from the JH region, the IgH chain enhancer element or the Ig switch region. A number of isolated cDNA clones contained sequences at their 5' end that were derived from a single exon located just upstream of DQ52, designated the mu o' element. Sequence analysis of a 920-bp genomic DNA segment, containing the mu o' exon and its 5' flanking region, revealed the presence of various conserved motifs for DNA-binding proteins, such as E2A, Ets, NF-kappa B and AP-2, which have previously been found in the IgH and L chain enhancers. We propose that the activity of the mu o' element, resulting in germ-line transcription of the DQ52-JH gene segment, is required to generate full accessibility for the V(D)J recombinase.

Diversity of immunoglobulin kappa light chain gene rearrangements and evidence for somatic mutation in V kappa IV family gene segments in X-linked agammaglobulinemia.

X-linked agammaglobulinemia (XLA) is a humoral immunodeficiency disease in man, characterized by an arrest in B lymphocyte differentiation at the precursor B cell stage. The structure of expressed immunoglobulin (Ig) kappa light (L) chain rearrangements of nine B lymphoblastoid cell lines from one XLA patient was investigated by amplification of cDNA by the polymerase chain reaction using 5' V kappa family-specific primers and a 3' kappa constant region primer. Members of all four V kappa gene families were found to be utilized in Ig kappa L chain rearrangements at frequencies that were consistent with random V kappa family usage. There was no preference for usage of any particular kappa joining segment. Additional diversity was generated by deletions and random nucleotide insertions at the site of juxtaposition. Particular V kappa members seemed to be overrepresented in the sample. The observed homology of the V kappa I, V kappa II and V kappa III region sequences, both to each other and to known germ-line V kappa sequence indicated the absence of somatic mutations in the majority of these expressed Ig genes. In contrast of the single-member V kappa IV family four different sequences were found to be expressed. That these sequences were mutated derivatives of a germ-line V kappa IV element was substantiated both by sequence analysis and oligonucleotide hybridization. This finding shows that the mutation process can occur in early stages of B cell development i.e. before H chain class switch has occurred. The presence of these mutations is probably independent of clonal expansion since XLA patients are unable to respond to antigen. We conclude that the differentiation arrest in XLA does not preclude early onset of somatic mutation events in V kappa gene segments.

Immunoglobulin kappa light chain germ-line transcripts in human precursor B lymphocytes.

B lymphoblastoid cell lines (BLCL), established from bone marrow and peripheral blood mononuclear cells from two severe combined immunodeficiency (SCID) patients, manifested a complete absence of genomic rearrangements of the immunoglobulin (Ig) heavy (H) and light (L) chain loci. The BLCL contained germ-line transcripts of the Ig kappa region locus of approximately 1.2 kilobase (kb). By cDNA cloning and sequence analysis the transcripts were shown to consist of a C kappa segment, a J kappa 1 gene segment, 160 base pairs (bp) of J kappa 1 5' intervening sequence, containing the heptamer/nonamer recombination recognition sequences and at the 5' end a 523-bp segment designated human kappa zero, The first 206 bp of this 5' segment were homologous to the reported murine kappa zero region. Genomic restriction mapping and DNA sequence analysis demonstrated that the human kappa zero segment is located approximately 4 kb upstream of J kappa 1. The kappa zero segment contains a putative promoter region with an OCT2 binding site, and has a splice donor site to accomplish splicing to an acceptor site 160 bp upstream of J kappa 1. Expression of the kappa zero gene segment was found in BLCL derived from normal fetal bone marrow, in which both Ig kappa loci were in the germ-line configuration. These findings indicate that the described transcripts are not only present in SCID, but also in normal developing pre-B lymphocytes. The expression of germ-line Ig kappa L chain transcripts may be associated with the locus becoming accessible to gene rearrangement.

Restricted utilization of germ-line VH3 genes and short diverse third complementarity-determining regions (CDR3) in human fetal B lymphocyte immunoglobulin heavy chain rearrangements.

Twenty-one independent immunoglobulin heavy chain VH3DJH rearrangements were cloned and sequenced from livers of human fetuses at 7, 13 and 18 weeks of gestation. The VH elements expressed were not somatically mutated. Eight out of the estimated 30 VH3 elements were utilized with a preference for five of them. One of these VH3 sequences, designated FL13-28, represented a thus-far unknown VH3 gene segment. From the six functional JH elements the JH3 and JH4 segments were utilized preferentially and from the estimated 30 D segments the DQ52 element and the Dxp family were found to rearrange frequently. D elements were utilized both in normal and inverted orientation, as single copies or in D to D fusions. Addition of N nucleotides, removal of nucleotides from the coding sequences and utilization of DIR elements (D genes with irregular recombination signals) further expanded the third complementarity-determining region (CDR3) diversity. One fourth of the fetal CDR3 regions lacked N regions. Due to utilization of DQ52, the relative absence of N regions and extensive exonuclease activity operating on the D elements, the fetal CDR3 regions were significantly shorter than those found in adult B lymphocytes.

X-linked agammaglobulinemia.

X-linked agammaglobulinemia (XLA) patients manifest a very low production of immunoglobulins (Ig) of all classes and plasma cells are virtually absent. The XLA gene plays a crucial role in the transition of pre-B cells to later B cell stages, as hardly any slg-positive B lymphocytes can be detected. In the bone marrow almost normal numbers of pre-B lymphocytes are present. These cytoplasmatic C mu+ pre-B lymphocytes appear to express truncated M heavy chain molecules lacking the variable region segment. The T lymphocyte compartment is intact: the numbers of mature T cell receptor (TcR) alpha beta expressing T lymphocyte populations and their proliferative responses to antigens are normal. That the B cells are primary and exclusively affected was proven by X-chromosome inactivation studies. There is no evidence that the XLA gene is directly involved in the Ig gene rearrangements since B lymphoblastoid cell lines (BLCLs) established from peripheral blood of XLA patients were found to produce IgM molecules composed of complete Ig heavy and light chains and were shown to contain normal VHDJH recombinations. The data do not exclude the involvement of the XLA gene in a B cell specific process that makes the Ig loci accessible for recombination. Investigations on the degree of diversity of immunoglobulins generated by XLA patients exposed no limitations in the VH family usage. Sequence analysis of expressed VH3 and VH4 rearrangements however revealed that some genetic elements of the Ig locus might be over-represented and that a high portion of rearrangements was generated by unconventional mechanisms. By restriction length polymorphism (RFLP) and pulsed field gel electrophoreses analyses the XLA gene was mapped to an 8- to 12-Mb DNA fragment located in the Xq22 region. The known location of the XLA gene on the X-chromosome with closely linked RFLP markers and the availability of X-chromosome inactivation assays provides methods for carrier detection and prenatal diagnosis.

Diversity of immunoglobulin heavy chain gene segment rearrangement in B lymphoblastoid cell lines from X-linked agammaglobulinemia patients.

X-linked agammaglobulinemia (XLA) is characterized by an arrest in early B lymphocyte differentiation. Precursor B cells are present in the bone marrow (BM), whereas peripheral blood B cell numbers are severely decreased. A series of Epstein-Barr virus (EBV)-transformed B lymphoblastoid cell lines (BLCL) was established from peripheral blood of three XLA patients belonging to one pedigree. These BLCL manifested productive VHDJH rearrangements and a random utilization of the VH families. The CDR3 regions of the rearrangements varied in length from 12 to 47 nucleotides and included N regions in all cases. The results supported the conclusion that the few B lymphocytes in peripheral blood of XLA patients exhibit all mechanisms that generate immunoglobulin (Ig) heavy (H) chain diversity. However, no evidence for somatic mutation was found. Within the VH3 family 50% of the expressed VH gene segments belonged to a single subgroup and within the VH4 family a preferential utilization of one VH4 gene element was observed. The utilization of H chain joining (HH) elements was biased to JH4 and JH6 and a high percentage of the CDR3 regions was found to be generated by unconventional mechanisms, such as multiple D usage and the fusion of D elements to D segments with irregular recombination recognition signals. These unique features of the recombined and expressed VHDJH regions in XLA may explain the inability of XLA patients to respond to a variety of antigens. Alternatively, they could be secondary to a B lymphocyte maturation defect in XLA.

[Food hygiene and the prevalence of diarrhea and vomiting in independent elderly]. / Voedselhygiëne en het vóórkomen van diarree en braken bij zelfstandig wonende ouderen.

Food hygiene and the prevalence of diarrhea and vomiting were investigated in a population of self-supporting elderly aged 65 to 80 years. Two hundred and fourteen of them were interviewed at home. During this interview the temperature of the refrigerator and the cellar was measured. The prevalence of diarrhea in the 3 months preceding the interview was 20%, the prevalence of vomiting was 13%. By means of the interview at the respondents' homes and the temperature measurement, food hygiene was evaluated. Seventy percent of the respondents appeared to act in a hazardous way on one or more of the selected items. After excluding those who might have been vomiting or suffering from diarrhea due to certain chronic conditions, a statistically significant association between food hygiene evaluation and diarrhea in the preceding 3 months was found. The 3-month prevalence of the 90 persons with hazardous practices was 22%; in the group of 35 persons without such practices the prevalence was 6%. It is hard to compare these results with those of other studies because of different definitions and different methods. The impression that this age-category is a high risk group for food hygiene cannot be confirmed nor ruled out.

DNA sequences at the ends of the genome of bacteriophage Mu essential for transposition.

We have determined the minimal DNA sequences at the ends of the genome of bacteriophage Mu that are required for its transposition. A mini-Mu was constructed on a multicopy plasmid that enabled the manipulation of the DNA sequences at its ends without affecting the genes essential for transposition. The genes A and B, which were cloned outside the ends of the mini-Mu on the same plasmid, were both needed for optimal transposition. In our experimental system the predominant end products of the transposition are cointegrates both in the presence and in the absence of B. Two regions ending approximately 25 and 160 bp from the left end and one ending approximately 50 bp from the right end appear to be essential for optimal transposition. Overlapping with these regions, a 22-base-pair sequence was recognized with the consensus Y-G-T-T-C-A-Y-T-N-N-A-A-R-Y-R-C-G-A-A-A-A, where Y and R represent any pyrimidine and purine, respectively. At the left end these sequences occur as direct repeats; at the right end this sequence is inverted with respect to those at the left end.