PubMed Health⌕ Search

Biomedical subjects

R F Howard

Publications and source records attributed to R F Howard.

At least 37 records · Page 2Linked to original sources
PubMed1996

Increased prevalence of systemic sclerosis in a Native American tribe in Oklahoma. Association with an Amerindian HLA haplotype.

OBJECTIVE: To investigate a high prevalence of systemic sclerosis (SSc; scleroderma) in a well-defined population of 21,255 Choctaw Indians residing in 8 southeastern Oklahoma counties who were "users" of Indian Health Services. METHODS: A case-control study of 12 SSc cases and 48 matched non-SSc controls (4 per case) was conducted to investigate potential occupational, residential, and infectious exposures, as well as genetic factors which might predispose to SSc. HLA class II alleles were determined by DNA oligotyping, and class I and III alleles were defined serologically. RESULTS: The prevalence of SSc in full-blooded Choctaws was at least 8/1,704, or 469/100,000 (95% confidence interval [95% CI] 203-930) over the 4-year interval 1990-1994 and was significantly higher than that among non-full-blooded Choctaws (6/19,551, or 31/100,000) (P = 0.00001, odds ratio [OR] = 15.4, 95% CI 4.9-49.8). The overall prevalence of SSc in Oklahoma Choctaws (66/100,000) also was significantly higher than that in other Native Americans in Oklahoma (9.5/100,000) (P = 10(-6), OR = 6.95, 95% CI 3.3-13.7), who showed a prevalence similar to that reported for whites (2.1-25.3/100,000). Among the SSc cases, there was striking homogeneity of disease expression with the majority exhibiting diffuse scleroderma, pulmonary fibrosis, and autoantibodies to topoisomerase I. No environmental exposures were found to be in excess among cases versus controls. The strongest risk factor for SSc in cases (100%) versus controls (54%) was an HLA haplotype bearing the alleles B35, Cw4, DRB1*1602 (DR2), DQA1*0501, and DQB1*0301 (DQ7) (P = 0.002, Pcorr = 0.036, OR = 21, 95% CI 2.9-437). Survey of another group of Choctaws residing in another state revealed no cases of SSc despite a high frequency of the same HLA haplotype. CONCLUSION: Full-blooded Choctaw Native Americans living in southeastern Oklahoma have the highest prevalence of SSc yet found in any population. A major risk factor for disease is a uniquely Amerindian HLA haplotype; however, additional genes and/or an as-yet-unidentified environmental exposure seem likely.

Adult↗
PubMed1995

The secretary pathway of plasmodium falciparum regulates transport of p82/RAP1 to the rhoptries.

The rhoptries of Plasmodium falciparum are formed during a restricted period in the asexual erythrocytic cycle. The steps required for rhoptry biogenesis and the pathway for targeting proteins to the rhoptries have not been elucidated. Using the maturation of the Rhoptry-Associated Protein 1 (RAP-1) gene product to study these steps, it is reported here that a secretory pathway controls transport of protein complexes containing RAP-1 products to the rhoptries. Both brefeldin A (BFA) and low temperature reversibly block the processing of an 86-kDa precursor (Pr86) to the mature 82-kDa RAP-1 product (p82). Furthermore, the points of action of BFA and low temperature appear to overlap since their sequential application reversibly prevents Pr86 processing. Treatment of intact cells with N-ethylmaleimide, which prevents the fusion of transport vesicles with Golgi membranes in other eukaryotic cells, irreversibly blocks processing of Pr86. The role of the secretory pathway in targeting p82 protein complexes to the rhoptries product of RAP-1. These in vitro results also reveal that the RAP-1 product contains a cleavable N-terminal signal peptide and appears to be initially synthesized as an 84-kDa protein. The above data indicate that transport of p82 to the rhoptries is regulated by the secretory pathway and that the RAP-1 primary translation product differs in apparent molecular weight from the in vivo precursor Pr86. Our results suggest that rhoptry biogenesis is controlled in part by the secretory pathway and that the RAP-1 gene product acquires a previously undetected protein modification during its maturation.

Animals↗
PubMed1994

Expansion of selected V delta 1+ gamma delta T cells in systemic sclerosis patients.

We have previously shown an increased percentage of gamma delta T cells expressing the TCR V delta 1 gene segment in the peripheral blood and bronchoalveolar lavage fluid of patients with systemic sclerosis (SSC). To estimate clonality of these V delta 1+ T cells, the diversity of V delta 1 junctional regions (V-D-J) was examined using a reverse transcriptase-PCR to amplify TCR delta-chain transcripts isolated from PBMC, lung, esophagus, stomach, or skin of patients and controls. Limited diversity of V delta 1-J delta junctional regions in SSC patients was demonstrated by comparing the size distribution of PCR-amplified junctional region cDNA from patients with that of controls. Sequence analyses confirmed that V delta 1-J delta junctional regions from the blood of SSc patients had less diversity than those from controls, in that a significantly higher proportion of sequences were repeated in patients (54.4 vs 19.4% in controls). Evidence for selection of the V delta 1+ T cells in the tissues of SSC patients came from the findings that the same V delta 1-J delta junctional sequences persisted in an individual patient over time and that identical junctional sequences were isolated from multiple sites. Analysis of deduced amino acid sequences revealed two clusters of similarities among the junctional regions from patients. These data suggest that expansion of V delta 1+ gamma delta T cells may be Ag driven in SSC patients.

Adult↗
PubMed1994

Depth of epidural space in children.

We have studied the depth of epidural space in 586 children who had lumbar epidural block as part of their anaesthetic management. The mean depth of epidural space in neonates was 1 cm (SD 0.2, range 0.4-1.5 cm). The depth of epidural space in older infants and children correlated significantly with age and weight with regression equations of depth (cm) = 1 + 0.15 x age (years) and depth (cm) = 0.8 + 0.05 x weight (kg) respectively.

Adolescent↗
PubMed1993

Reactivity profile of human anti-82-kilodalton rhoptry protein antibodies generated during natural infection with Plasmodium falciparum.

Immunization of monkeys with the 82-kDa rhoptry protein (p82) of Plasmodium falciparum can protect them against a lethal blood stage challenge, and monoclonal antibodies to p82 inhibit parasite growth in vitro. The role that a p82-specific immune response might play in human immunity to the parasite is not known. To determine to what extent humans produce antibodies to p82 following infection with P. falciparum, sera from individuals believed to be hyperimmune, semi-immune, or never infected with the parasite were examined. Portions of the p82 gene were expressed separately as fusion proteins and used on immunoblots to test for antibodies to the recombinant proteins. All but 1 of the 30 immune sera possessed antibodies to p82, while nonimmune sera produced, at best, only a marginal signal to the fusion proteins. The signal intensity produced with the human immune sera depended on the region of p82 being assayed, with the N-terminal 37% of p82 producing stronger signals than more C-terminal parts of p82. This N-terminal domain contains a tandem octapeptide repeat (consensus KSSSPSXT/V) of the structure (repeat)2-Q-T-S-G-S/L-(repeat)3. It is shown here that the sequence of this repetitive motif is conserved among four parasite isolates at both the nucleotide and amino acid levels; the five-residue repeat interruption peptide QTSGS/L separating the two sets of repeats contains the only amino acid substitution (Ser or Leu) detected in this region to date. Despite their conservation of structure, the repeats do not appear to be the only epitope recognized by the human antibodies, since sera which recognize the N-terminal fusion protein containing the repeats also bind a related protein after truncation and removal of the repeats. These results indicate that the structurally conserved p82 molecule contains multiple B-cell epitopes and is likely to be immunogenic in most individuals during natural infections with P. falciparum. These observations are consistent with the idea that antibodies to p82 generated during parasite infection have a role in the development of immunity to the organism.

Amino Acid Sequence↗
PubMed1993

An 11-year-old girl with a left-ankle injury.

The following case is presented to illustrate the roentgenographic and clinical findings of a condition of interest to the orthopaedic surgeon. The initial history, physical findings, and roentgenographic examinations are found on this page. The final clinical and roentgenographic differential diagnoses are presented on the following pages.

Ankle Injuries↗
PubMed1990

Intravenous immunoglobulin therapy in systemic lupus erythematosus-associated thrombocytopenia.

Seven patients with thrombocytopenia and systemic lupus erythematosus were treated with intravenous (IV) doses of human immunoglobulin to assess clinical response and to examine the mechanism of action of IV immunoglobulin in these patients. Five of 7 patients had a greater than 50% increase in their platelet counts. Four of these patients had a sustained benefit of at least 6 months duration. The initial effectiveness of IV immunoglobulin therapy was not dependent on the reduction of levels of circulating platelet-binding IgG or circulating immune complexes.

Antigen-Antibody Complex↗
PubMed1990

Plasmodium falciparum: hetero-oligomeric complexes of rhoptry polypeptides.

We have previously described several monoclonal antibodies (McAbs) which specifically recognize antigens in the rhoptries of Plasmodium falciparum and which immunoprecipitate polypeptides of 82, 70, 67, 39, and 37 kDa. We now show that only p82, p70, p67, and a 86-kDa precursor (Pr86) of p82 possessed epitopes for these McAbs. These four proteins were not synthesized until schizogony. These results and proteolysis experiments indicated that Pr86, p82, p70, and p67 were the products of the same gene, whereas the dissimilar digestion patterns of p39 and p37 suggested that p39 was encoded by a second gene and p37 by yet another. Complexes of these proteins (termed RI complexes) are maintained by noncovalent interactions since the ionic detergent SDS was sufficient to dissociate them into individual polypeptides. Sucrose gradient centrifugation demonstrated that RI complex formation was not dependent on the presence of antibody and that these complexes had higher sedimentation rates than the 185-kDa P. falciparum merozoite glycoprotein. Covalent crosslinking with the reversible, homobifunctional, primary amine-specific reagent 3,3'-dithiobis(sulfosuccinimidylpropionate) followed by RI McAb immunoprecipitation resulted in purification of intact complexes which were not dissociable by SDS alone. Immunodepletion experiments with a subtype of RI McAb which does not immunoprecipitate p37 suggested that the binding of p39 and p37 to the other RI proteins was mutually exclusive. Therefore, the minimal composition of the RI complexes is one molecule of Pr86, p82, p70, or p67 and one of p39 or p37. The epitopes of Pr86, p82, p70, and p67 for the RI McAbs were sensitive to disulfide bond reduction. Surprisingly, reduction increased their electrophoretic mobilities. This enhanced mobility could not be accounted for by post-translational glycosylation, phosphorylation, or acylation, or by covalent attachment via the sulfhydryl moiety of cysteine residues to additional parasite proteins. We suggest that, due to an asymmetric distribution of amino acids in the Pr86-class molecules, SDS binding results in a lower charge to mass ratio in the native folded polypeptides and a higher charge to mass ratio upon disulfide bond reduction and unfolding of the polypeptides.

Animals↗
PubMed1990

Cross-reactive asparagine-rich determinants shared between several blood-stage antigens of Plasmodium falciparum and the circumsporozoite protein.

From a Plasmodium falciparum cDNA expression library derived from mRNA of the asexual blood stages, we isolated and sequenced five different cDNA clones whose predicted protein products were unusually rich in asparagine (Asn). Two of the clones, R5 and G5, contain tandem imperfectly repeated sequences based on Asn-Asn-Thr (NNT) and Asn-Asn-Met (NNM) respectively. The other three, E4, C5 and R13, as well as G5, contain stretches of polyasparagine varying in length from 2 to 26 residues. Results of DNA blotting experiments with the individual cDNA sequences as probes suggest that each of the five clones corresponds to a different P. falciparum gene. The fragments of P. falciparum proteins expressed by the cDNA clones shared cross-reactive antigenic determinants which were present on multiple P. falciparum proteins. In immunoblotting experiments, owl monkey antibodies selected for binding to the polypeptide expressed by clone E4, C5 or G5 reacted with the expressed proteins from all 5 clones, and with at least 10 proteins from schizont infected erythrocytes. The cross-reactive epitopes could be modeled by two Asn-rich peptide structures: (1) (NNT)8, whose sequence was based on the R5 repeat; and (2) (NPNA)6, whose sequence was based on the Asn-rich repeat of the P. falciparum circumsporozoite protein (CSP). Antibodies that bound to each peptide were selected from sera of immune monkeys that had never been exposed to sporozoites. The selected antibodies bound all 5 expressed proteins in immunoblotting assays and also bound to several proteins from parasitized erythrocytes. Such cross reactivity between the CSP repeating unit and several blood-stage antigens has not been previously reported.

Amino Acid Sequence↗
PubMed1988

Characterization of a high-molecular-weight phosphoprotein synthesized by the human malarial parasite Plasmodium falciparum.

During its intra-erythrocytic cycle, Plasmodium falciparum synthesizes a protein of apparent Mr 250,000-300,000. Its precise size is dependent on the P. falciparum isolate examined. This protein contains phosphate covalently bound to one or more serine residues and hence is termed PP300. Monoclonal antibody, McAb4-1F, binds to PP300 on immunoblots of protein extracts from all parasite isolates tested, both those exhibiting and those lacking the knob phenotype. Using McAb4-1F, the polypeptide was shown to be physically associated with the plasma membrane in a membrane-isolation procedure. However, in an indirect immunofluorescence assay the McAb appeared to bind to antigen associated with the erythrocyte plasma membrane in parasitized cells. However, it reacted only to fixed, not unfixed, parasitized erythrocytes indicating that the epitope is not normally exposed to extracellular antibodies. Clone 29-2 was isolated by a McAb4-1F immunoscreen of a P. falciparum complementary DNA (cDNA) expression library created in pUC8. Rat anti-clone serum which was raised to the purified protein encoded by the lacZ-29-2 fusion in pUC8 reacted with PP300 in immunoblots of parasite antigen. In Southern-blot analyses of parasite DNA digested with EcoRI, HindIII, or EcoRV, the 29-2 DNA insert hybridized to more than one fragment even though the insert lacked internal sites for these enzymes. In addition, hybridization studies were conducted using two oligodeoxy-nucleotides which were constructed based on the sequence of a cDNA clone which encoded part of a similar high-molecular-weight P. falciparum protein [Coppel et al., Mol. Biochem. Parasitol. 20 (1986) 265-277]. Analysis of these results indicates that the two cDNA sequences are parts of the same gene or a family of related genes.

Amino Acid Sequence↗
PubMed1986

Conservation and antigenicity of N-terminal sequences of GP185 from different Plasmodium falciparum isolates.

Complementary DNA (cDNA) clones for GP185, a major antigenically diverse glycoprotein of Plasmodium falciparum, were isolated from a cDNA library of the Honduras I/CDC (Honduras I) isolate, and 1052 bp were sequenced. The expression of cDNA fragments in Escherichia coli using the vector pCQV2 allowed verification of the reading frame. This GP185 cDNA sequence, like the cDNA sequence for a homologous gene of the K1 isolate [Hall et al., Nature 311 (1984) 379-382], codes for a polypeptide which is truncated due to multiple, in-frame stop codons. This polypeptide corresponds to the N-terminal 15% of the proposed coding region of the GP185 gene [Holder et al., Nature 317 (1985) 270-273]. Comparison of the nucleotide sequences for the GP185 gene of Honduras I and five other isolates indicated that there are two areas of conserved DNA sequence, one of 310 bp (beginning 181 bp upstream from the proposed initiation codon) and the other of greater than or equal to 360 bp (located entirely within the coding region), separated by a region encoding isolate-specific tandem amino acid repeats. Rat antiserum was raised to a fusion protein derived from the conserved regions and the intervening repeat region of this Honduras I protein. This antiserum bound GP185 on immunoblots of the homologous Honduras I isolate and the heterologous K1 isolate, which has different tandem repeats. Serum from owl monkeys and humans previously infected with P. falciparum reacted with the fusion protein on immunoblots demonstrating that determinants in the N-terminal 15% of GP185 were immunogenic in infected individuals and suggesting that some of these sites are conserved among isolates.(ABSTRACT TRUNCATED AT 250 WORDS)

Antigens, Protozoan↗
PubMed1985

Two Plasmodium falciparum merozoite surface polypeptides share epitopes with a single Mr 185 000 parasite glycoprotein.

The malarial parasite Plasmodium falciparum synthesizes a major glycoprotein (gp) of Mr 185 000 during its asexual blood cycle. Immunoprecipitation of [35S]methionine- or [3H]glucosamine-labeled schizont antigens indicated that two groups of polypeptides were distinguished with anti-gp 185 mouse monoclonal antibodies: group A was composed of glycosylated molecules of Mr 185 000, 120 000, 90 000, 88 000, 46 000, and 40 000 while group B contained, in addition to gp 185, polypeptides of Mr 152 000, 106 000 and 83 000. The latter polypeptides lacked detectable amounts of radiolabeled saccharide. The smaller Mr polypeptides were specifically immunoprecipitated and not merely coprecipitated with gp 185. Our results suggest that gp 185 contains at least two structurally distinct domains which may be processed independently into either the group A or group B polypeptides. Although gp 185 may not be a merozoite surface protein, representative group A and group B-specific monoclonal antibodies bound to surface antigens of the merozoite as demonstrated by immunolabeling followed by electron microscopy. Therefore, at least one group A antigen and one group B antigen appeared to be on the extracellular surface of the merozoite. The proteins found in immunoprecipitates after both (1) sonication in aqueous medium and ultracentrifugation and (2) solubilization and phase separation of parasite molecules with Triton X-114 suggested that the group A and group B polypeptides and glycoproteins are either soluble or peripheral membrane proteins. Some of these, therefore, may be components of the surface coat of the merozoite.

Animals↗
PubMed1985

Recognition of a Mr 56K glycoprotein on the surface of Plasmodium falciparum merozoites by mouse monoclonal antibodies.

Hybridomas were prepared from mice repeatedly injected with disrupted Plasmodium falciparum (FVO isolate) schizonts and merozoites. Antibodies secreted by two of these hybridomas were shown by immunoelectron microscopy to bind to the surface of merozoites from the FVO isolate. These monoclonal antibodies (McAb) reacted with the FVO and Geneva isolates by an indirect fluorescence antibody test (IFAT) and immunoprecipitated a protein of relative molecular weight (Mr) 56K from both isolates. The 56K protein could be labeled with [35S] methionine and [3H]glucosamine. Glycosidase treatment of the affinity-purified polypeptide proved that the [3H]glucosamine had been incorporated into sugar side chains and that this protein (called gp56) was glycosylated. The anti-gp56 McAb did not react by IFAT or immunoprecipitation with four isolates (Honduras I, Indochina I, Tanzania I, and Kenya) that lack gp56 but contain major glycoproteins of Mr 50K. Antibodies from an Aotus monkey immune to the FVO isolate immunoprecipitated gp56 from both the FVO and Geneva isolates, but did not immunoprecipitate the 50K glycoproteins from the other four isolates. Extraction experiments conducted with the nonionic detergent Triton X-114 indicate that some of the gp56 molecules are hydrophilic and that the others are either hydrophobic or interact with hydrophobic molecules. These results, together with the electron microscopic data, suggest that the hydrophilic gp56 is a component of the extracellular matrix and that the hydrophobic gp56 may be associated with the plasma membrane of the merozoite.

Animals↗