Peripersonal space boundaries around the lower limbs.

Neurophysiological investigations in non-human primates have shown that bi- and tri-modal fronto-parietal neurons exist that respond to touch on the body and visual (and/or auditory) stimuli near the body. The receptive fields of these neurons extend into space around the body, producing a zone wherein multisensory information is readily integrated. This space around the body, known as peripersonal space (PPS), has also been investigated behaviourally in humans. Some studies have focused on how far into depth the spatial boundaries of PPS extend. Most of these investigations have focused on the upper body (e.g., hands, face, trunk), while little is known about the size of PPS for the lower body (i.e. legs and feet). Thus, the aim of the current study was to delineate a PPS boundary around the lower limbs in healthy participants using a multisensory interaction task. Participants made speeded responses to the presence of vibrations applied to the toes while a task-irrelevant visual stimulus approached towards (Experiment 1) or receded from (Experiment 2) the feet. Participants responded significantly faster to tactile stimuli when the visual stimulus was within approximately 73 cm from the feet, but only when it approached (and not receded from) the legs. This is the first study, to our knowledge, to outline the size of PPS for the lower limbs. These findings could provide insight into the mechanisms underlying multisensory integration in the lower limbs, and add to the current body of knowledge on PPS representations.

Immunomodulatory therapy of eosinophil-associated gastrointestinal diseases.

Eosinophil-associated gastrointestinal disorders (EGIDs), including eosinophilic esophagitis (EE) and eosinophilic gastroenteritis (EG), are a spectrum of increasingly recognized inflammatory diseases characterized by gastrointestinal symptoms and eosinophilic infiltration of the gastrointestinal tract. Significant morbidity is associated with the development of esophageal strictures in some patients. Immune-mediated reactions to food allergens appear to drive the inflammation in a subset of patients, especially those with solitary EE, but dietary interventions remain difficult in EE and are less effective in EG. Despite the increasing incidence of these disorders and their increased recognition by physicians, there are currently no medications that either United States or European Union regulatory agencies have specifically approved for use in EGIDs. This lack of safe and effective therapies for EGIDs is a major obstacle in the care of these patients and underscores the need for new therapeutic approaches. This review briefly discusses the currently available 'off label' drug treatments for EGIDs, most notably topical and systemic corticosteroids. Pathogenesis studies of EGIDs suggest possible therapeutic targets, and conversely, clinical trials of mechanistically-targeted therapeutics give insight into disease pathogenesis. Thus, EGID pathogenesis is discussed as an introduction to mechanistically-targeted immunotherapeutics. The two biologic categories that have been used in EGIDs, anti-IgE (omalizumab) and anti-IL-5 (SCH55700/reslizumab and mepolizumab), are discussed. Because there are similarities in the pathogenesis of EGIDs with asthma and atopic dermatitis, biologic therapeutics currently in early trials for asthma management are also briefly discussed as potential therapeutic agents for EGIDs. Given the deficiencies of current therapeutics and the rapidly advancing knowledge of the pathogenesis of these disorders, EGIDs are an ideal model for translating recent advances in understanding immunopathogenesis into mechanistically-based therapeutics. Further understanding of the early events in pathogenesis is also needed to develop preventive and disease-modifying treatments.

ERBIN deficiency links STAT3 and TGF-ß pathway defects with atopy in humans.

Nonimmunological connective tissue phenotypes in humans are common among some congenital and acquired allergic diseases. Several of these congenital disorders have been associated with either increased TGF-ß activity or impaired STAT3 activation, suggesting that these pathways might intersect and that their disruption may contribute to atopy. In this study, we show that STAT3 negatively regulates TGF-ß signaling via ERBB2-interacting protein (ERBIN), a SMAD anchor for receptor activation and SMAD2/3 binding protein. Individuals with dominant-negative STAT3 mutations (STAT3mut ) or a loss-of-function mutation in ERBB2IP (ERBB2IPmut ) have evidence of deregulated TGF-ß signaling with increased regulatory T cells and total FOXP3 expression. These naturally occurring mutations, recapitulated in vitro, impair STAT3-ERBIN-SMAD2/3 complex formation and fail to constrain nuclear pSMAD2/3 in response to TGF-ß. In turn, cell-intrinsic deregulation of TGF-ß signaling is associated with increased functional IL-4Rα expression on naive lymphocytes and can induce expression and activation of the IL-4/IL-4Rα/GATA3 axis in vitro. These findings link increased TGF-ß pathway activation in ERBB2IPmut and STAT3mut patient lymphocytes with increased T helper type 2 cytokine expression and elevated IgE.

Levels of p53 protein increase with maturation in human hematopoietic cells.

Transfection of the wild-type p53 gene into malignant cell lines usually results in an inhibition of proliferation. However, the physiological function of the endogenous p53 gene product has been difficult to ascertain. In order to examine whether p53 is involved in the regulation of proliferation and/or differentiation of hematopoietic tissue, we modified a recently developed flow cytometric assay to assess p53 protein expression in normal human hematopoietic cells, primary leukemias, and selected leukemia cell lines. In normal human bone marrow, p53 protein was not detected in the proliferative, progenitor cell populations identified by the cell surface antigens CD34 (progenitor cells of multiple lineages) or glycophorin (erythroid precursors). In contrast, low but detectable levels of p53 protein were observed in the nonproliferative, mature lymphoid, granulocytic, and monocytic cell populations. Similarly, p53 levels increased and DNA synthesis decreased during 12-O-tetradecanoylphorbol-13-acetate-induced differentiation of ML-1 myeloblastic leukemia cells. Both of these results suggest that endogenous, wild-type p53 protein may play a role in hematopoietic cell maturation, possibly by contributing to the inhibition of proliferation that occurs during terminal differentiation. Leukemia cells deviated from this pattern of expression: (a) in contrast to the normal, proliferative bone marrow progenitor cells, a significant percentage of patient leukemia samples expressed detectable levels of p53 protein; and (b) leukemia cell lines exhibited lineage-specific abnormalities in p53 expression, with overexpression in lymphoid cell lines and lack of expression in myeloid cell lines.

Flow cytometric detection of rare normal human marrow cells with immunophenotypes characteristic of acute lymphoblastic leukemia cells.

Rare subpopulations of normal marrow B lymphoid cells expressing immunophenotypes typically found in B-lineage acute lymphoblastic leukaemias (ALL) were sought by multiparameter flow cytometry. First, CD34+ marrow leukocytes were isolated by immune adherence using immunomagnetic microspheres, and analyzed for coexpression of the following pairs of membrane antigens: CD34 CD22; CD34 CD20; and CD10 CD22. Terminal deoxynucleotidyl transferase expression was not assessed. All three antigen combinations were found on small percentages of the CD34-enriched cell population. Second, unseparated normal low density marrow leukocytes were examined by 'gating' on cells with the right-angle light scatter of lymphoid cells, plus either CD34+ or CD10+ immunofluorescence. This independent approach confirmed that rare subsets of normal cells coexpress 'immature' and 'mature' differentiation antigens. In addition, remission marrow cells were examined from two children who had completed therapy for ALL two and four months earlier. Both specimens had a more than threefold increase in CD34+ cells over normal marrow, and cells coexpressing immature and mature cell surface antigens were easily detected. These findings demonstrate that immunophenotypes characteristic of B-lineage ALL, previously labeled 'asynchronous' with respect to the developmental sequence of the majority of normal B lymphoid cells, exist at low frequency in normal human bone marrow.

Biogenesis of the bundle-forming pilus of enteropathogenic Escherichia coli: reconstitution of fimbriae in recombinant E. coli and role of DsbA in pilin stability--a review.

Enteropathogenic Escherichia coli (EPEC) adhere to tissue culture cells in a distinct pattern known as localized adherence (LA). We have defined two loci necessary for LA. A plasmid-encoded gene cluster encodes bundlin, the major structural subunit of a type-IV fimbria called the bundle-forming pilus (BFP), a prepilin peptidase necessary for processing of pre-bundlin to its mature form, and twelve other proteins. Under the control of an exogenous promoter, these 14 genes are sufficient for the biogenesis of BFP in a heterologous E. coli host. The chromosomal gene dsbA, which encodes a periplasmic disulfide-bond oxidoreductase, is also required for LA. In the absence of DsbA protein, bundlin is made but rapidly degraded. Pre-bundlin is also rapidly degraded in the absence of DsbA, suggesting that the prepilin is a transcytoplasmic protein simultaneously accessible to enzymes on both sides of the inner membrane. These studies offer a fresh perspective on the biogenesis of type-IV pili.

The use of mirtazapine in a patient with chronic pain.

Antidepressant drugs that act on serotonin and noradrenergic systems may be analgesic. The newer antidepressant mirtazapine (Remeron) has activity on noradrenergic and serotonergic transmission and is approved for the treatment of a Major Depressive Disorder. This paper describes a case that suggests that mirtazapine may also be useful in the treatment of chronic pain.

Trichotillomania resulting in a trichobezoar: a case report.

This paper presents a case of a 19-year-old, 10 1/2-week-pregnant woman with trichotillomania that resulted in a trichobezoar. The case illustrates typical presentation, patient behavior, symptomatology, and physical findings of patients with trichobezoars. The hypothesized methods for trichobezoar formation, complications, and treatment are discussed. The diagnostic criteria, epidemiology, and treatment of trichotillomania are also discussed.

Data collection efforts in New England.

Amid growing demands for statistical compilations about home care, the eight state home care associations of the National Association for Home Care's Region I ventured into the arena of data collection. What have they learned?

Nuclear oncoprotein expression as a function of lineage, differentiation stage, and proliferative status of normal human hematopoietic cells.

Relative levels of the nuclear oncoproteins c-myb, c-myc, and c-fos were determined in selected subpopulations of normal human bone marrow (BM) cells using a flow cytometric assay which simultaneously detects a cell-surface antigen (as a marker of lineage and stage of maturation) and levels of an intracellular protein. At least two monoclonal antibodies directed against each oncoprotein and specific peptide inhibition controls were used for these determinations. Hematopoietic progenitor cells (CD34+) express the highest levels of c-myb and c-myc, whereas c-fos levels in CD34+ progenitor cells are similar to c-fos levels in mature monocytes and granulocytes. Granulocytes are the only hematopoietic cells examined which do not express detectable levels of c-myb and c-myc. The levels of these oncoproteins in these normal, unstimulated BM cell populations were more closely linked to lineage and maturation stage than to the proliferative status of the given population, as determined by either DNA staining or expression of the cell-cycle specific nuclear protein, Ki67. This flow cytometric assay helps in interpreting the significance of oncoprotein levels in leukemia cells by allowing direct comparisons of a leukemia with the phenotypically similar "normal counterpart control" cell population in normal BM.

Effects of bfp mutations on biogenesis of functional enteropathogenic Escherichia coli type IV pili.

Enteropathogenic Escherichia coli expresses a type IV fimbria known as the bundle-forming pilus (BFP) that is required for autoaggregation and localized adherence (LA) to host cells. A cluster of 14 genes is sufficient to reconstitute BFP biogenesis in a laboratory strain of E. coli. We have undertaken a systematic mutagenesis of the individual genes to determine the effect of each mutation on BFP biogenesis and LA. Here we report the construction and analysis of nonpolar mutations in six genes of the bfp cluster, bfpG, bfpB, bfpC, bfpD, bfpP, and bfpH, as well as the further analysis of a previously described bfpA mutant strain that is unable to express bundlin, the pilin protein. We found that mutations in bfpB, which encodes an outer membrane protein; bfpD, which encodes a putative nucleotide-binding protein; and bfpG and bfpC, which do not have sequence homologues in other type IV pilus systems, do not affect prebundlin expression or processing but block both BFP biogenesis and LA. The mutation in bfpP, the prepilin peptidase gene, does not affect prebundlin expression but blocks signal sequence cleavage of prebundlin, BFP biogenesis, and LA. The mutation in bfpH, which is predicted to encode a lytic transglycosylase, has no effect on prebundlin expression, prebundlin processing, BFP biogenesis, or LA. For each mutant for which altered phenotypes were detected, complementation with a plasmid containing the corresponding wild-type allele restored the wild-type phenotypes. We also found that association of prebundlin or bundlin with sucrose density flotation gradient fractions containing both inner and outer membrane proteins does not require any accessory proteins. These studies indicate that many bfp gene products are required for biogenesis of functional type IV pili but that mutations in the individual genes do not lead to the identification of new phases of pilus assembly.

Role of BfpF, a member of the PilT family of putative nucleotide-binding proteins, in type IV pilus biogenesis and in interactions between enteropathogenic Escherichia coli and host cells.

Adherence of enteropathogenic Escherichia coli (EPEC) to epithelial cells is dependent on a type IV fimbria, termed the bundle-forming pilus (BFP). A cluster of 14 genes is required for expression of BFP. The eighth gene in the cluster, bfpF, encodes a putative nucleotide-binding protein which resembles the PilT protein of Pseudomonas aeruginosa. It has been proposed that PilT is required for the retraction of the P. aeruginosa pilus, which results in twitching motility. To test the role of BfpF in BFP function and EPEC pathogenesis, two different mutations were constructed in the bfpF gene, one in the cloned gene cluster in a laboratory E. coli strain and one in wild-type EPEC. Neither mutation affected prepilin synthesis, leader sequence processing, or pilus biogenesis. However, both mutations resulted in increased localized adherence. In addition, the EPEC bfpF mutant displayed increased aggregation. The EPEC bfpF mutant was not deficient in attaching and effacing activity or invasion capacity. These results suggest that BfpF decreases aggregation and adherence by EPEC but that subsequent steps in EPEC pathogenesis do not require this protein.

The Wiskott-Aldrich syndrome.

The Wiskott-Aldrich Syndrome (WAS) is an inherited immunodeficiency caused by a variety of mutations in the gene encoding the WAS protein (WASp). WASp is expressed in hematopoetic cells and facilitates the reorganization of the actin cytoskeleton in response to many important cell stimuli. Extensive study of WAS and more recently WASp has given great insight into the relevance of this molecule and related molecules to both basic cell biology and human immune defenses.

A third secreted protein that is encoded by the enteropathogenic Escherichia coli pathogenicity island is required for transduction of signals and for attaching and effacing activities in host cells.

Enteropathogenic Escherichia coli strains are able to signal host cells, cause dramatic cytoskeletal rearrangements, and adhere intimately to the cell surface in a process known as the attaching and effacing effect. A pathogenicity island of 35 kb known as the locus of enterocyte effacement (LEE) is necessary and sufficient for this effect. The LEE encodes an outer membrane adhesin called intimin, a type III secretion apparatus, and the EspA and EspB secreted proteins. The DNA sequence of the region between espA and espB revealed a new gene, espD. The product of espD was demonstrated by using a T7 expression system. We constructed a nonpolar mutation in espD and found that the mutant is incapable of the signal transduction events that lead to activation of the putative intimin receptor in host cells and that the mutant fails to induce the attaching and effacing effect. These phenotypes were restored to the mutant by complementation with a plasmid containing the cloned espD locus. We demonstrated by immunoblotting and microsequencing that the EspD protein is secreted via the type III apparatus. Thus, we describe a novel locus encoding a secreted protein that is required for attaching and effacing activity.

A cluster of fourteen genes from enteropathogenic Escherichia coli is sufficient for the biogenesis of a type IV pilus.

Enteropathogenic Escherichia coli (EPEC) adhere to epithelial cells in microcolonies, a pattern termed localized adherence (LA). LA is dependent upon the presence of 50-70 MDa plasmids, termed EPEC adherence factor (EAF) plasmids. Expression of an EAF plasmid-encoded type IV fimbria, the bundle-forming pilus (BFP), is associated with the LA phenotype. TnphoA insertions in bfpA, the gene encoding the major structural subunit of the BFP, abolish LA. While bfpA::TnphoA mutants cannot be complemented for LA by plasmids carrying the bfpA gene alone in trans, this work shows that they can be complemented by plasmids carrying the bfpA gene, as well as approximately 10 kb of downstream sequence, suggesting that such mutations have polar effects on downstream genes. The identification and characterization of a cluster of 13 genes immediately downstream of bfpA are described. The introduction into a laboratory Escherichia coli strain of a plasmid containing these 14 bfp gene cluster genes, along with pJPN14, a plasmid containing another fragment derived from the EAF plasmid, confers LA ability and BFP biogenesis. However, when a mutation is introduced into the last gene of the bfp cluster, neither LA nor BFP biogenesis is conferred. This work also provides evidence to show that the fragment cloned in pJPN14 encodes a factor(s) which results in increased levels of the pilin protein. Finally, it is shown that expression of the 14 genes in the bfp cluster from an IPTG-inducible promoter, in the absence of pJPN14, is sufficient to reconstitute BFP biogenesis in a laboratory E. coli strain, but is insufficient for LA. This is the first report demonstrating the reconstitution of a type IV pilus in a laboratory E. coli strain with a defined set of genes. The BFP system should prove to be a useful model for studying the molecular mechanisms of type IV pilus biogenesis.

Stress analyses of glenoid components in total shoulder arthroplasty.

Finite element analysis was used to characterize the local stresses at the bone-implant interface of 2 different types of glenoid components presently used in unconstrained total shoulder arthroplasty. A series of 2-dimensional finite-element meshes was developed to model the glenoid in 2 mutually perpendicular planes with and without implanted components. One of the implants modeled was a cemented all-polyethylene component, and the second was an uncemented metal-backed component. A variety of parameters were studied including the resultant loading direction (concentric versus eccentric), keel geometry, subchondral bone integrity, and cement mantle size. Results of the analyses show that the cemented all-polyethylene design demonstrated an overall stress pattern that was closer to that of the intact glenoid. When the effects of concentric and eccentric loading conditions were compared, the overall stress magnitudes in the subchondral bone were found to be much lower with the uncemented metal-backed component than with its cemented all-polyethylene counterpart. This finding suggests that some degree of stress shielding may be associated with the metal-backed component. In addition, under both the concentric and eccentric loading conditions, extremely high stress regions were found within the polyethylene near the polyethylene-metal interface of the uncemented metal-backed component.

Attaching and effacing of host cells by enteropathogenic Escherichia coli in the absence of detectable tyrosine kinase mediated signal transduction.

An unusual mutant of enteropathogenic E. coli (EPEC), deficient in its ability to invade host cells, was evaluated. The gene interrupted by the transposon in this mutant was located within a region of the EPEC chromosome devoted to secretion of proteins required for signal transduction. The mutant did not secrete detectable levels of the EspB protein, previously shown to be required for attaching and effacing, and did not induce detectable tyrosine phosphorylation of a 90 kDa host cell protein, previously associated with attaching and effacing and invasion. No quantitative or qualitative defect in the ability of the mutant to induce attaching and effacing effects was observed. Moreover, attaching and effacing by wild-type EPEC was unaffected by high doses of the tyrosine kinase inhibitor genistein. These results indicate that attaching and effacing activity can occur in the absence of detectable EspB secretion and tyrosine kinase mediated signal transduction.