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Biomedical subjects

J M Johnson

Publications and source records attributed to J M Johnson.

At least 37 records · Page 2Linked to original sources
PubMed2001

Influence of isometric exercise on blood flow and sweating in glabrous and nonglabrous human skin.

The distribution of the reflex effects of isometric exercise on cutaneous vasomotor and sudomotor function is not clear. We examined the effects of isometric exercise by different muscle masses on skin blood flow (SkBF) and sweat rate (SR) in nonglabrous skin and in glabrous skin. The latter contains arteriovenous anastomoses (AVAs), which cause large fluctuations in SkBF. SkBF was measured by laser-Doppler flowmetry (LDF) and reported as cutaneous vascular conductance (CVC; LDF/mean arterial pressure). SR was measured by capacitance hygrometry. LDF and SR were measured at the sole, palm, forearm, and ventral leg during separate bouts of isometric handgrip (IHG) and isometric leg extension (ILE). CVC and its standard deviation decreased significantly during IHG and ILE in the palm and sole (P < 0.05) but not in the forearm or leg (P > 0.05). Only palmar SR increased significantly during IHG and ILE (P < 0.05). We conclude that the major reflex influences of isometric exercise on the skin include AVAs and palmar sweat glands and that this is true for both arm and leg exercise.

Adult↗
PubMed2001

Institutional learning curve for sentinel node biopsy at a community teaching hospital.

Sentinel lymph node biopsy (SLB) is gaining popularity as an alternative to axillary lymph node dissection for breast cancer staging. Although publications have described the inherent learning curve few have analyzed actual performance in community hospitals. This study analyzes the institutional learning curve for SLB in a community teaching hospital without a formal sentinel node credentialing policy. We conducted an analysis of the initial 96 SLBs performed by 15 general surgeons over a 34-month period. The main outcomes were rate of identification of sentinel node and accuracy of SLB. Overall SLB was successful in identifying one or more sentinel nodes (mean = 2.2) in 73 per cent of attempted cases. There were marked differences in performance of individual surgeons; identification rates varied from 25 to 100 per cent. Only one surgeon performed more than 15 procedures during the study period. Nineteen of 21 cases with positive nodes were correctly characterized (sensitivity = 90.5%; 95% confidence interval = 76-100%; false negative rate = 9.5%). Our institutional learning curve was longer than high-volume individual experiences published in the literature, with a lower rate of sentinel node identification. SLB appears to be sensitive for detecting malignancy, but the small number of patients with positive nodes in our series limits our conclusions. The marked variability in individual surgeon performances and the slow rate of overall improvement in our institution suggest a need for a formalized policy for SLB training.

Breast Neoplasms↗
PubMed2001

Telepathology in India.

Telepathology is the most recent addition to the pathologist's diagnostic tools. It is the acquisition of macroscopic and microscopic images for electronic transmission for diagnosis, consultation and/or education. With the addition of the personal computer at the pathologist's desktop, the stage has been set for one of the greatest advantages the Internet has to offer. Telepathology in India is in infancy, and we at PathoIndia (www.Pathoindia.com) have started a series of publication images from interesting cases in the form of weekly quiz. After cases are published, hundreds of pathologists from around the world are invited by e-mail to send in their diagnosis and comments. The responses to this quiz suggest that telepathology is catching on in the pathology community. Another intention of this series is to identify and select qualified international and Indian pathologists who would be willing to help colleagues from India requesting second opinions online.

Humans↗
PubMed2000

Mechanism of capsid assembly for an icosahedral plant virus.

Capsids of spherical viruses share a common architecture: an icosahedral arrangement of identical proteins. We suggest that there may be a limited number of common assembly mechanisms for such viruses. Previous assembly mechanisms were proposed on the basis of virion structure but were not rigorously tested. Here we apply a rigorous analysis of assembly to cowpea chlorotic mottle virus (CCMV), a typical, small, positive-strand RNA virus. The atomic resolution structure of CCMV revealed an interleaving of subunits around the quasi-sixfold vertices, which suggested that capsid assembly was initiated by a hexamer of dimers (Speir et al., 1995, Structure 3, 63-78). However, we find that the capsid protein readily forms pentamers of dimers in solution, based on polymerization kinetics observed by light scattering. Capsid assembly is nucleated by a pentamer, determined from analysis of the extent of assembly by size-exclusion chromatography. Subsequent assembly likely proceeds by the cooperative addition of dimers, leading to the T = 3 icosahedral capsid. At high protein concentrations, the concentration-dependent nucleation reaction causes an overabundance of five-dimer nuclei that can be identified by classical light scattering. In turn these associate to form incomplete capsids and pseudo-T = 2 capsids, assembled by oligomerization of 12 pentamers of dimers. The experimentally derived assembly mechanisms of T = 3 and pseudo-T = 2 CCMV capsids are directly relevant to interpreting the structure and assembly of other T = 3 viruses such as Norwalk virus and pseudo-T = 2 viruses such as the vp3 core of blue tongue virus.

Bromovirus↗
PubMed2000

Modulation of the IL-10/IL-12 cytokine circuit by interferon-beta inhibits the development of epitope spreading and disease progression in murine autoimmune encephalomyelitis.

IFN-beta has been shown to be effective in the treatment of multiple sclerosis (MS). However, the primary mechanism by which IFN-beta mediates its therapeutic effect remains unclear. Recent studies indicate that under defined conditions, IFN-beta may downregulate DC expression of IL-12. We and others have shown that IFN-beta may also downregulate IL-10. In light of the recently proposed paradigm that an IL-10/IL-12 immunoregulatory circuit controls susceptibility to autoimmune disease, we examined the effect of IFN-beta on the development and behavior of the autoreactive T cell repertoire during experimental autoimmune encephalomyelitis (EAE), an animal model sharing many features with MS. SWXJ mice were immunized with the immunodominant p139-151 determinant of myelin proteolipid protein (PLP), and at onset of EAE were treated every other day with IFN-beta. After eight weeks of treatment, we assessed autoreactivity and observed no significant IFN-beta effect on splenocyte proliferation or splenocyte production of IFN-gamma, IL-2, IL-4, or IL-5 in response to the priming determinant used to initiate disease. However, in IFN-beta treated mice, the cytokine profile in response to the priming immunogen was significantly skewed toward an increased production of IL-10 and a concurrent decreased production of IL-12. Moreover, the in vivo modulation of the IL-10/IL-12 immunoregulatory circuit in response to the priming immunogen was accompanied by an aborted development of epitope spreading. Our results indicate that IFN-beta induces a reciprocal modulation of the IL-10/IL-12 cytokine circuit in vivo. This skewed autoreactivity establishes an inflammatory microenvironment that effectively prevents endogenous self-priming thereby inhibiting the progression of disease associated with epitope spreading.

Adjuvants, Immunologic↗
PubMed2000

Tax oncoprotein trans-represses endogenous B-myb promoter activity in human T cells.

The B-myb gene was identified on the basis of its homology with the protooncogene c-myb, homolog of the avian myeloblastosis virus (AMV) and avian leukemia virus (E26) transforming genes. Several studies using antisense constructs or antisense oligonucleotides as well as overexpression experiments suggest that B-Myb plays an important role in the transition from G(1) to S phase of the cell cycle and that B-Myb expression is cell cycle regulated. We have previously demonstrated that the human T cell lymphotropic virus type 1 (HTLV1) trans-activator Tax is able to repress transcription from c-myb promoter reporter constructs as well as from the endogenous c-myb promoter in human T cells and that this effect is mediated through inhibition of the c-Myb trans-activating functions. Here we report that both HTLV-1 as well as HTLV-2 Tax proteins inhibit c-Myb trans-activation in mouse embryo fibroblasts (MEFs). In addition to c-Myb, B-Myb expression is also markedly downregulated in HTLV-1-transformed cells at both RNA and protein levels. Furthermore, by using a Jurkat T cell line stably transfected with a tax gene driven by a cadmium-inducible promoter (JPX9), we were able to demonstrate that Tax directly represses the endogenous B-myb promoter in T cells. Because c-Myb and B-Myb have been involved in cell cycle progression, our results suggest that Tax, by repressing both c-Myb and B-Myb endogenous promoters, may bypass their requirement for cell cycle progression in HTLV-1-transformed T cells.

Animals↗
PubMed2000

The MHC class I heavy chain is a common target of the small proteins encoded by the 3' end of HTLV type 1 and HTLV type 2.

Human T cell leukemia/lymphotropic virus types 1 and 2 are two immunologically and phylogenetically related retroviruses that differ in their pathogenicity in vivo. The overall genetic structure of HTLV-1 and -2 is similar. Each contains a unique region at the 3' end of the genome, designated the pX region. p12(I) is a membrane-associated protein encoded by the open reading frame I (ORF I) region of HTLV-1, which lies within the pX region. A corresponding protein, p10(I) is encoded by the ORF I region of HTLV-2 and an additional protein, p11(V), is encoded by ORF V, which overlaps the HTLV-2 ORF I region. As with HTLV-1, the small proteins encoded by the pX region of HTLV-2 appear to be dispensable for viral replication and cellular transformation in vitro. However, the small open reading frames of both viruses are important for viral replication in vivo, which suggests they may play an important role during the viral life cycle. This study was undertaken to investigate and compare the cellular targets of the p10(I), p11(V), and p12(I) putative proteins.

3' Untranslated Regions↗
PubMed2000

The HTLV-I orfI protein is recognized by serum antibodies from naturally infected humans and experimentally infected rabbits.

The mechanism of T-cell transformation by human T-cell lymphotropic virus type I (HTLV-I), though not completely understood, appears to involve the interactions of several viral and cellular proteins. One of these viral proteins, p12(I), encoded by HTLV-I orfI, is a weak oncogene that binds the 16-kDa subunit of the vacuolar ATPase and interacts with the immature beta and gamma(c) chains of the IL-2 receptor. We have expressed the singly spliced orfI cDNA in the baculovirus system and used the recombinant protein as a tool to assess the presence of antibodies in naturally or experimentally infected hosts. In addition, rabbit antisera were raised against various p12(I) synthetic peptides and used to identify three antigenic regions within p12(I), one between the two putative transmembrane regions of p12(I) and two at the carboxy-terminus of the protein. More importantly, sera from a naturally infected human (1 of 32) and experimentally infected rabbits (9 of 20) recognized the rp12(I), demonstrating orfI expression and immunogenicity in vivo. Taken together these data provide the first evidence of orfI expression during HTLV-I infections.

Amino Acid Sequence↗
PubMed2000

Predicting ligand-binding function in families of bacterial receptors.

The three-dimensional fold of a new protein sequence can often be inferred directly from sequence homology to a protein of known structure. The function of a new protein sequence is more difficult to predict, however, since homologues can have different molecular and cellular functions. To develop and automate computational methods for determining molecular function, we have analyzed ligand-binding specificity in two related families of binding proteins. One of these families includes Escherichia coli lactose repressor and ribose-binding protein, and the other includes E. coli sulfate- and phosphate-binding proteins. These proteins have similar folds but varying specificity, binding many different small molecules, including mono- and disaccharides, purines, oxyanions, ferric iron, and polyamines. Starting from template structural alignments, alignments of over 90 sequences per family were generated by iterative database searches with hidden Markov models. Phylogenetic trees were made of full-length sequences and of subsets of residues lining the binding cleft, to determine whether subbranches of the trees correlate with ligand-binding preference. Automated analyses of residues in the binding pocket were also used to predict ligand-binding function for many uncharacterized database sequences and to identify specific side chain-ligand contacts in proteins without solved structures. Our results demonstrate the utility of anchoring functional annotation within a protein family context.

Amino Acid Sequence↗
PubMed2000

Transactivation and growth suppression by the gut-enriched Krüppel-like factor (Krüppel-like factor 4) are dependent on acidic amino acid residues and protein-protein interaction.

Gut-enriched Krüppel-like factor (GKLF or KLF4) is a pleiotropic (activating and repressive) transcription factor. This study characterizes the mechanisms of transactivation by GKLF. Using a GAL4 fusion assay, the activating domain of murine GKLF was localized to the 109 amino acid residues in the N-terminus. Site-directed mutagenesis showed that two adjacent clusters of acidic residues within this region are responsible for the activating effect. Transactivation by GKLF involves intermolecular interactions as demonstrated by the ability of wild-type, but not mutated, GKLF to compete with the N-terminal activation domain. In addition, wild-type adenovirus E1A, but not a mutated E1A that failed to bind p300/CBP, inhibited transactivation by the N-terminal 109 amino acids of GKLF, suggesting that p300/CBP are GKLF's interacting partners. A physical interaction between GKLF and CBP was demonstrated by glutathione- S -transferase pull-down and by in vivo co-immuno-precipitation experiments. We also showed that the two acidic amino acid clusters are essential for this interaction, since GKLF with mutations in these residues failed to co-immunoprecipitate with CBP. Importantly, the same mutations abrogated the ability of GKLF to suppress cell growth as determined by a colony suppression assay. These studies therefore provide plausible evidence for a structural and functional correlation between the transactivating and growth-suppressing effects of GKLF.

Amino Acids↗
PubMed2000

A survey of smoking cessation knowledge, training, and practice among U.S. Army general medical officers.

BACKGROUND: Historically, cigarette-smoking rates have been higher among military personnel than among civilians, although recently these rates have decreased. METHODS: In March 1997, a questionnaire assessing (1) training received on smoking cessation, (2) objective knowledge of smoking-cessation techniques, (3) frequency of practice habits, and (4) personal tobacco use among physicians, was successfully mailed to 232 of the total population of 279 Army general medical officers (GMOs). RESULTS: One-hundred-fifty (65%) GMOs returned questionnaires. Of these, 3.3% reported personal cigarette smoking, and 7.3% regularly used smokeless tobacco. During internship, few (13%) GMOs received smoking-cessation training. Primary care programs provided training more frequently than did surgery internship programs. The mean score on the objective knowledge portion was 72%. GMOs had a variable practice pattern in their use of smoking-cessation techniques (percent answering "usually" or "always"): helping patients set quit dates (35%), offering to prescribe the nicotine patch (59%), referring patients to a behavior-modification program (86%). Physicians who received training during internship were significantly more likely (p < 0.01) to help their patients set a quit date. Training did not result in a statistically increased frequency of other practice habits. CONCLUSIONS: GMOs received minimal training on smoking cessation during internship. GMOs refer patients to smoking-cessation classes, reflecting the strategy of the Army Health Promotion program. Strategies to increase the frequency that GMOs prescribe nicotine replacement and assist patients in setting a quit date are needed. Military smoking-cessation efforts may provide valuable lessons for the civilian community.

Adult↗
PubMed2000

Female reproductive hormones and thermoregulatory control of skin blood flow.

Changes in progesterone and estrogen levels over the course of the menstrual cycle, with oral contraceptives or with hormone replacement therapy alter thermoregulatory control of skin blood flow and body temperature at rest and during exercise. We review here recent evidence concerning the sites and potential mechanisms of these influences.

Body Temperature Regulation↗
PubMed1999

A theoretical model successfully identifies features of hepatitis B virus capsid assembly.

The capsids of most spherical viruses are icosahedral, an arrangement of multiples of 60 subunits. Though it is a salient point in the life cycle of any virus, the physical chemistry of virus capsid assembly is poorly understood. We have developed general models of capsid assembly that describe the process in terms of a cascade of low order association reactions. The models predict sigmoidal assembly kinetics, where intermediates approach a low steady state concentration for the greater part of the reaction. Features of the overall reaction can be identified on the basis of the concentration dependence of assembly. In simulations, and on the basis of our understanding of the models, we find that nucleus size and the order of subsequent "elongation" reactions are reflected in the concentration dependence of the extent of the reaction and the rate of the fast phase, respectively. The reaction kinetics deduced for our models of virus assembly can be related to the assembly of any "spherical" polymer. Using light scattering and size exclusion chromatography, we observed polymerization of assembly domain dimers of hepatitis B virus (HBV) capsid protein. Empty capsids assemble at a rate that is a function of protein concentration and ionic strength. The kinetics of capsid formation were sigmoidal, where the rate of the fast phase had second-power concentration dependence. The extent of assembly had third-power concentration dependence. Simulations based on the models recapitulated the concentration dependences observed for HBV capsid assembly. These results strongly suggest that in vitro HBV assembly is nucleated by a trimer of dimers and proceeds by the addition of individual dimeric subunits. On the basis of this mechanism, we suggest that HBV capsid assembly could be an important target for antiviral therapeutics.

Capsid↗
PubMed1999

Visinin-like protein (VILIP) is a neuron-specific calcium-dependent double-stranded RNA-binding protein.

Double-stranded RNA-binding proteins function in regulating the stability, translation, and localization of specific mRNAs. In this study, we have demonstrated that the neuron-specific, calcium-binding protein, visinin-like protein (VILIP) contains one double-stranded RNA-binding domain, a protein motif conserved among many double-stranded RNA-binding proteins. We showed that VILIP can specifically bind double-stranded RNA, and this interaction specifically requires the presence of calcium. Mobility shift studies indicated that VILIP binds double-stranded RNA as a single protein-RNA complex with an apparent equilibrium dissociation constant of 9.0 x 10(-6) M. To our knowledge, VILIP is the first double-stranded RNA-binding protein shown to be calcium-dependent. Furthermore, VILIP specifically binds the 3'-untranslated region of the neurotrophin receptor, trkB, an mRNA localized to hippocampal dendrites in an activity-dependent manner. Given that VILIP is also expressed in the hippocampus, these data suggest that VILIP may employ a novel, calcium-dependent mechanism to regulate its binding to important localized mRNAs in the central nervous system.

3' Untranslated Regions↗
PubMed1999

Treatment with BBB022A or rolipram stabilizes the blood-brain barrier in experimental autoimmune encephalomyelitis: an additional mechanism for the therapeutic effect of type IV phosphodiesterase inhibitors.

We examined the treatment effects of two structurally distinct phosphodiesterase type IV (PDE IV) inhibitors, BBB022 and rolipram, in murine and rat models of experimental autoimmune encephalomyelitis (EAE). Based on our data, we propose a mechanism of action which may supplement immunomodulatory effects of PDE IV inhibitors. In particular, PDE inhibitors promote elevation of intracellular cAMP levels, increasing the electrical resistance of endothelial monolayers by stabilizing intercellular junctional complexes. Such an effect on central nervous system (CNS) vascular endothelium has the potential to reduce disease severity in EAE, because both inflammatory cells and humoral factors readily cross a disrupted blood-brain barrier (BBB). In this report, we demonstrate the capacity of BBB022 and rolipram to decrease clinical severity of EAE. further, PDE IV inhibitors significantly reduced BBB permeability in the spinal cords of mice with EAE. These results provide evidence that PDE IV-inhibitors may exert therapeutic effects in EAE by modifying cerebrovascular endothelial permeability, reducing tissue edema as well as entry of inflammatory cells and factors.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1999

Alignment and structure prediction of divergent protein families: periplasmic and outer membrane proteins of bacterial efflux pumps.

Broad-specificity efflux pumps have been implicated in multidrug-resistant strains of Pseudomonas aeruginosa and other Gram-negative bacteria. Most Gram-negative pumps of clinical relevance have three components, an inner membrane transporter, an outer membrane channel protein, and a periplasmic protein, which together coordinate efflux from the cytoplasmic membrane across the outer membrane through an unknown mechanism. The periplasmic efflux proteins (PEPs) and outer membrane efflux proteins (OEPs) are not obviously related to proteins of known structure, and understanding the structure and function of these proteins has been hindered by the difficulty of obtaining reasonable multiple alignments. We present a general strategy for the alignment and structure prediction of protein families with low mutual sequence similarity using the PEP and OEP families as detailed examples. Gibbs sampling, hidden Markov models, and other analysis techniques were used to locate motifs, generate multiple alignments, and assign PEP or OEP function to hypothetical proteins in several species. We also developed an automated procedure which combines multiple alignments with structure prediction algorithms in order to identify conserved structural features in protein families. This process was used to identify a probable alpha-helical hairpin in the PEP family and was applied to the detection of transmembrane beta-strands in OEPs. We also show that all OEPs contain a large tandem duplication, and demonstrate that the OEP family is unlikely to adopt a porin fold, in contrast to previous predictions.

Amino Acid Sequence↗