Variability in upper motor neurone-type dysarthria: an examination of five cases with dysarthria following cerebrovascular accident

The degree of diversity in the nature and extent of the physiological deficits which occur in subjects with dysarthria with similar neurological damage is demonstrated through the individual assessment profiles of five subjects with dysarthria following upper motor neurone (UMN) damage. The perceptual profiles of each subject were compiled using perceptual ratings of deviant speech parameters, intelligibility ratings from the Assessment of Intelligibility of Dysarthric Speech (ASSIDS), and perceptual judgements of subsystem function determined from the Frenchay Dysarthria Assessment (FDA). For each individual, the perceptual profile of their speech impairments was compared and contrasted with the objective results of spirometric and kinematic assessments of respiratory function aerodynamic and electroglottographic evaluations of laryngeal function, pressure and strain gauge evaluations of articulatory function, and nasal accelerometric assessments of nasality. The outcomes of the individual perceptual and physiological profiles are discussed with respect to the presence of differential subsystem impairments both within each subject and between subjects with similar underlying pathophysiological deficits. The importance of interpreting the instrumental findings with respect to the interdependency of each of the motor speech subsystems, the limitations of perceptual assessments, and the advantages of utilising both perceptual and physiological analyses in the process of identifying treatment goals is discussed.     

Anti-idiotype monoclonal antibodies specific for the MOPC167 anti-phosphocholine transgene-encoded antibody

Four rat x mouse hybridomas secreting monoclonal anti-idiotypic (anti-Id) antibodies (MAb) specific for the transgene-encoded antibody of the 207-4 transgenic mouse line, which carries the VH1/V kappa 24 gene segments of the IgA, phosphocholine-(PC) specific MOPC167 myeloma, were developed from a fusion of Ag8-X63.653 mouse cells with spleen cells from a rat immunized with MOPC167 and HPCM27 anti-PC antibodies. The anti-Id MAb were shown by ELISA to be specific for PC-binding proteins of VH1/V kappa 24 H and L chains of various isotypes. They did not bind VH1/V kappa 22, VH1/V kappa 8, or VH1/V kappa 1 PC-binding proteins or other IgA or IgM myeloma proteins. Analysis by flow cytometry demonstrated that these MAb bind to the transgene-encoded membrane immunoglobulin (sIgM) as expressed on > 95% of the B220 positive 207-4 spleen cells. All four MAb were able to inhibit the binding of MOPC167 to PC conjugated to bovine serum albumin. Differences in fine specificity of binding were demonstrated by differential staining of spleen cells of the 216-7 mu kappa delta Mem MOPC167 transgenic mice. In these mice endogenous H chains associate with the transgene encoded L chain to form MOPC167 crossreactive idiotopes. Two of the MAb, 28-4-3 and 28-6-20, stained significant numbers of cells, while MAb 28-5-15 did not bind to 216-7 cells. Three of the MAb, 28-5-15, 28-6-20, and 28-4-3, when conjugated to Sepharose beads, were able to induce DNA synthesis in cultures of 207-4 transgenic spleen cells. None of the MAb were able to induce an antibody response in vivo. These MAb should prove useful in staining PC-transgenic B cells for flow cytometry studies and in defining early cellular events in the activation of idiotype positive B cells by anti-Id antibodies.     

Neurotoxin binding and allosteric modulation at receptor sites 2 and 5 on purified and reconstituted rat brain sodium channels

Purified and reconstituted sodium channels have previously been shown to be functional in voltage-dependent ion conductance and in high affinity binding of tetrodotoxin and saxitoxin at neurotoxin receptor site 1 and alpha-scorpion toxins at receptor site 3, but high affinity binding of neurotoxins at receptor sites 2, 4, and 5 has not been demonstrated. The pyrethroid insecticide RU39568 enhances the specific binding of [3H]batrachotoxinin A 20-alpha-benzoate (BTX-B) to neurotoxin receptor site 2 on purified and reconstituted sodium channels up to 500-fold, reducing the Kd to 1.5 nM. Brevetoxins and alpha-scorpion toxins cause further allosteric enhancement of BTX-B binding. The pyrethroids deltamethrin and bifenthrin and the nonpyrethroid insecticide 2,2-bis(p-chlorophenyl)trichloroethane can partially substitute for RU39568 in enhancing BTX-B binding, but other pyrethroids are inactive. The brevetoxin PbTx-1 binds specifically to neurotoxin receptor site 5 on purified and reconstituted sodium channels with a Kd value of approximately 30 nM. Brevetoxin binding is enhanced up to 2-fold by the combination of batrachotoxin and RU39568. The allosteric enhancement of BTX-B binding by RU39568 is voltage dependent, decreasing progressively with depolarization to 0 mV. In contrast, PbTx-1 binding is not voltage dependent and PbTx-1 reduces the voltage dependence of the effect of RU39568. The results demonstrate restoration of high affinity binding and allosteric interactions of ligands at neurotoxin receptor sites 2 and 5 on purified and reconstituted sodium channels and provide an experimental approach to covalent labeling and identification of the peptide components of those receptor sites.     

Complex CD44 splicing combinations in synovial fibroblasts from arthritic joints

CD44 is a broadly expressed cell surface glycoprotein which is the major cell surface receptor for the glycosaminoglycan, hyaluronan. In humans, alternative splicing of up to 9 variant exons (v2-v10) into CD44 mRNA, together with post-translational modification via glycosylation and chondroitin sulfate attachment has the potential of generating a large number of CD44 isoforms. Insertion of these various exons has the potential to change the functional capacities of the molecule and has implications in disease. We have analyzed CD44 splice variant expression in cultured VCAM-1-positive synovial fibroblasts isolated from patients with osteo- or rheumatoid arthritis and from normal synovium. Rheumatoid and osteoarthritic tissue express CD44 splice variants at the cell surface level. At the mRNA level exons v3, v6, v7, v8, v9 and v10 were detected in different splicing combinations. Rheumatoid tissue showed high expression, osteoarthritic tissues showed great variation. In contrast, non-inflamed tissue showed no splicing events. Our results indicate that the nature of CD44 splice variant expression may be linked to the inflammatory state of the synovial joint.     

Evidence of androgen receptor expression in lichen sclerosus: an immunohistochemical study

OBJECTIVE: While topical androgen administration is widely used in the treatment of lichen sclerosus of the vulva, localization and level of expression of androgen receptor (AR) have not been described previously. METHODS: Thirty-nine paraffin-embedded punch biopsies of patients with lichen sclerosus of the vulva were examined. Androgen receptor, estrogen receptor (ER), and progesterone receptor (PR) expression in lichen sclerosus and in normal vulvar skin were investigated by immunohistochemistry. RESULTS: Five tissue specimens (12.8%) of lichen sclerosus showed nuclear staining with anti-AR in the parabasal cell layers of the epidermis. Median age of patients with positive nuclear staining for AR versus women without AR expression was 71 (range, 63-78) and 66.5 (range, 38-91) years, respectively. Estrogen receptor expression was present in only one patient. Nuclear staining reaction for PR expression was absent in all cases. Four of the five AR-positive women reported no complaints and therefore received no topical testosterone therapy. CONCLUSION: Our results suggest a lack of complaints in AR-positive lichen sclerosus patients. Our findings could justify a larger study comparing symptoms of patients with and without AR expression.     

A phenotypic host range alteration determines RD114 virus restriction in feline embryonic cells

We have characterized the restriction mechanism for RD114 virus replication in embryonic feline cells (FeF). By comparing growth properties of the virus in FeF cells with its behavior in a fetal feline glial cell line (G355) permissive for RD114, we showed that both cell lines were readily infectible by virus grown in permissive cells and that no significant differences in viral integration or viral RNA expression could be detected. However, analysis of viral protein expression revealed differences in viral env gene processing in the two cell types. Envelope precursor pR85 was produced, but the expected processed gp70 product was detectable only in permissive (G355) cells. An envelope product of 85 kDa was packaged into virions produced by FeF cells, while virions produced by G355 cells contained the expected RD114 gp70. While the gp85 env-containing virions were infectious for permissive G355 cells, they were unable to infect FeF cells. The block to infection by the gp85-containing particles in FeF cells could be abrogated by treatment with the glycosylation inhibitor tunicamycin. Our results indicate that restriction of RD114 virus involves a novel mechanism dependent on two factors: altered glycosylation of the envelope to a gp85 form and an altered RD114 receptor in FeF cells.     

RAG1 mediates signal sequence recognition and recruitment of RAG2 in V(D)J recombination

Recent studies have demonstrated that DNA cleavage during V(D)J recombination is mediated by the RAG1 and RAG2 proteins. These proteins must therefore bind to the recombination signals, but the specific binding interaction has been difficult to study in vitro. Here, we use an in vivo one-hybrid DNA binding assay to demonstrate that RAG1, in the absence of RAG2, can mediate signal recognition via the nonamer, with the heptamer acting to enhance its binding. A region of RAG1 with sequence similarity to bacterial invertases is essential for DNA binding. Localization of RAG2 to the signal is dependent upon the presence of RAG1 and is substantially more efficient with a 12 bp spacer signal than with a 23 bp spacer signal.     

The molecular characterization of transport vesicles

Secretion, endocytosis and transport to the lytic compartment are fundamental, highly coordinated features of the eukaryotic cell. These intracellular transport processes are facilitated by vesicles, many of which are small (100 nm or less in diameter) and 'coated' on their cytoplasmic surface. Research into the structure of the coat proteins and how they interact with the components of the vesicle membrane to ensure the selective packaging of the cargo molecules and their correct targeting, has been quite extensive in mammalian and yeast cell biology. By contrast, our knowledge of the corresponding types of transport vesicles in plant cells is limited. Nevertheless, the available data indicate that a considerable homology between plant and non-plant coat polypeptides exists, and it is also suggestive of a certain similarity in the mechanisms underlying targeting in all eukaryotes. In this article we shall concentrate on three major types of transport vesicles: clathrin-coated vesicles, COP-coated vesicles, and 'dense' vesicles, the latter of which are responsible for the transport of vacuolar storage proteins in maturing legume cotyledons. For each we will summarize the current literature on animal and yeast cells, and then present the relevant data derived from work on plant cells. In addition, we briefly review the evidence in support of the 'SNARE' hypothesis, which explains how vesicles find and fuse with their target membrane.     

The C825T polymorphism in the human G-protein beta3 subunit gene is not associated with diabetic nephropathy in Type I diabetes mellitus

In Type I (insulin-dependent) diabetes mellitus a genetic predisposition exists to nephropathy and is related to parental hypertension. Enhanced G-protein activation, a cellular phenotype observed in cultured cells from patients with essential hypertension, was recently documented in Type I diabetic subjects with nephropathy. This enhanced G-protein activation has been associated with a genetic variant in the G-protein beta3 subunit, GNB3. A C-->T polymorphism at position 825 in exon 10 is associated with G-protein activation, the T allele associated with enhanced activity. Furthermore the T allele was observed more frequently in a group with essential hypertension. In this report we have analysed the role of the C825T polymorphism in the predisposition to diabetic nephropathy in Type I diabetes. We have investigated the frequency of this polymorphism in a large case-control study and found no association of the T allele with diabetic nephropathy. Specifically carriage of the T allele as CT or TT was observed in 49% of 200 Type I diabetic control subjects with normoalbuminuria (diabetes duration 24 years) compared with 53% of 216 Type I diabetic subjects with nephropathy (overt proteinuria or end-stage renal failure). Within this group we have also examined the inheritance of C825T alleles in a family study and found no evidence for excess transmission of the T allele to Type I diabetic offspring with nephropathy (T allele transmitted to 51% of nephropathy offspring, C allele transmitted to 49% of nephropathy offspring, p = 0.79). In none of the Type I diabetic datasets examined was there any effect of genotype on variation in systolic or diastolic blood pressure. In conclusion we can find no evidence for the C825T polymorphism of the beta3 G-protein subunit as a major gene in the susceptibility to diabetic nephropathy in Type I diabetes.