Biomedical subjects

T Humphreys

Publications and source records attributed to T Humphreys.

At least 19 recordsLinked to original sources

PubMed1992

N-(4'-hydroxyphenylacetyl)palytoxin: a palytoxin prodrug that can be activated by a monoclonal antibody-penicillin G amidase conjugate.

Palytoxin (PTX), one of the most toxic nonprotein molecules known, is cytotoxic at picomolar concentrations against a wide variety of cell types. In contrast to most cytotoxins, PTX exerts its activity extracellularly. A method for targeting PTX to tumor cells is described in which a monoclonal antibody-enzyme conjugate activates a PTX prodrug at surfaces of tumor cells. The prodrug, N-(4'-hydroxyphenylacetyl)palytoxin (NHPAP), was prepared by reacting PTX with an active ester of 4-hydroxyphenylacetic acid. NHPAP was 1000 times less toxic than PTX to a panel of carcinoma and lymphoma cell lines. The cytotoxic activity of the combination of penicillin G amidase from Escherichia coli with NHPAP was equal to PTX. Two cell lines that were multidrug resistant showed no enhanced resistance to NHPAP +/- penicillin G amidase. Immunologically specific activation of NHPAP took place when H2981 cells (L6 antigen positive) were treated with the monoclonal antibody conjugate L6-penicillin G amidase followed by NHPAP. This system is distinguished from other prodrug activation schemes, since the released drug exerts its activity extracellularly, has high potency, and may be able to overcome the multidrug resistant phenotype.

Acrylamides↗

PubMed1992

Preformatted charts improve documentation in the emergency department.

STUDY OBJECTIVES: To determine if the use of programmed charts with complaint-specific entry criteria results in improved documentation of patient encounters and better clinical outcome. DESIGN: Prospective study. SETTING: Emergency department of an urban university hospital. TYPE OF PARTICIPANTS: Female patients presenting to the emergency department with gynecologic complaints of abdominal pain, bleeding, or vaginal discharge. INTERVENTIONS: Programmed and blank charts were provided randomly for physicians in the ED. MEASUREMENTS: Chart scores based on documentation criteria for patient history, physical examination, laboratory studies, diagnosis, and discharge instructions and patient outcome scores of 0% to 100% based on the persistence of their complaints at the time of the follow-up interview. MAIN RESULTS: Overall documentation of history, physical examination, and laboratory studies was more complete on programmed charts than on blank charts (81.1% vs 71%, P less than .0001). The patient history portion of the charts was found to benefit the most from the use of programmed charts (74.8% vs 60.1%, P less than .0001). Although programmed charts demonstrated better documentation, there was no statistically significant correlation with patient outcome parameters or with patient satisfaction with the quality of medical care. However, more patients whose physicians used programmed charts were satisfied with their physicians' explanations of their problem (chi 2 = 5.2, P less than .02). CONCLUSION: Programmed charts improve documentation by facilitation of the documentation process and allow more time for patient-physician interaction. Quality of documentation alone, however, is not a reliable indicator of patient outcome or of the quality of care received.

Abdominal Pain↗

PubMed1992

Comparison of restriction fragment length polymorphisms of proto-oncogenes in native Hawaiians and other ethnic groups in Hawaii.

The relative genetic diversity of selected proto-oncogenes in the native Hawaiian gene pool was examined by comparing the restriction fragment length polymorphisms of these genes in a group of 23 individuals with at least part Hawaiian ancestry, and in 20 individuals from other ethnic groups. Twenty-one combinations of the proto-oncogenes, c-fms, c-myc, L-myc, c-Ha-ras, and c-Ki-ras, tested with 1 or more of the restriction enzymes Bam HI, Eco RI, Hind III, Pst I, Pvu I and Kpn I were examined. Sixteen of these did not exhibit RFLPs in Hawaiians or in other ethnic groups. Four of the combinations exhibiting RFLPs in native Hawaiians exhibited similar-sized restriction fragments in the other ethnic groups. Only in the case of c-myc digested with Pst I were 5 individuals of Hawaiian ancestry found to have an RFLP which has not been detected in other ethnic groups. These 5 cases exhibited a 13-kb c-myc fragment in addition to the 5.5-kb fragment found in most Hawaiians and always present in other ethnic groups. The presence in Hawaiians of most RFLPs found in other ethnic groups indicates that the genetic diversity of proto-oncogenes in the gene pool of native Hawaiians is not substantially less than that of other ethnic groups.

Blotting, Southern↗

PubMed1991

Activation of the L1 late H2B histone gene in blastula-stage sea urchin embryos by Antennapedia-class homeoprotein.

The L1 late H2B histone gene of the sea urchin Strongylocentrotus purpuratus is transcriptionally activated in late blastula stage embryos by a mechanism that depends on an enhancer element located 3' of the gene (Zhao et al., 1990). A protein factor, designated H2B abp 1, binds this element at a site that resembles the consensus recognition sequence of Antennapedia-class homeodomain proteins. We demonstrate here that Antennapedia (Antp) and Hbox4 proteins, members of the Antennapedia class of homeoproteins from Drosophila and sea urchin respectively, bind the L1 H2B abp 1 site, and that the Drosophila Antp protein acts through this site to trans-activate the L1 H2B gene, in vivo. In addition, RNA gel blot analysis demonstrated that Hbox4 transcripts accumulate in developing embryos with a time course that closely resembles that of H2B adp 1 DNA binding activity and the activity and the transcription rate of the L1 late H2B gene. Finally, we show that antibody prepared against the sea urchin Hbox4 protein, a member of the Abd-B subclass of the Antennapedia class, specifically inhibits binding of the H2B abp 1 factor to the L1 H2B enhancer, suggesting that H2B abp 1 is encoded by Hbox4 or a closely related gene.

Animals↗

PubMed1990

The transposable element Uhu from Hawaiian Drosophila--member of the widely dispersed class of Tc1-like transposons.

We report the complete nucleotide sequence of the transposable element Uhu from the vicinity of the alcohol dehydrogenase (Adh) gene of Drosophila heteroneura (an endemic Hawaiian Drosophila). The complete element is about 1650 base-pairs (bp) long, has 46-50 base-pair inverse imperfect repeats at it's ends, and contains a large open reading frame potentially encoding a 192 amino acid protein. We demonstrate that Uhu belongs to a class of transposable elements which includes Tc1 from Caenorhabditis elegans, Barney from Caenorhabditis briggsae, and HB1 from Drosophila melanogaster. All of these elements share significant sequence similarity, are approximately 1600 base pairs long, have short inverse terminal repeats (ITRs), contain open reading frames (ORFs) with significant sequence identity, and appear to insert specifically at TA sequences generating target site duplications.

Alcohol Dehydrogenase↗

PubMed1990

Characterization and expression of two sea urchin homeobox gene sequences.

We describe two homeobox sequences, TgHbox5 and TgHbox6, isolated from the Hawaiian sea urchin Tripneustes gratilla using a Drosophila Sex combs reduced probe. Sequence analysis shows that the encoded TgHbox5 homeodomain shares only 30-52% amino acid identity with homeodomains encoded by previously characterized genes, establishing that it is a divergent homeobox that is not in any known class of homeoboxes. TgHbox5 is expressed in the embryo as two major developmentally regulated transcripts. one at 5.0 kilobase (kb) appearing by blastula stage and the other at 2.7 kb appearing at pluteus stage. Multiple transcripts from TgHbox5 are present at a much lower level in adult tissues and are predominantly expressed in small and large intestines. The TgHbox6 homeobox is an Antenna-pedia-class homeobox, which appears not to be expressed during embryogenesis but produces abundant 3.6 and 3.2 kb transcripts in the six adult tissues examined.

Amino Acid Sequence↗

PubMed1989

Progressively restricted expression of a homeo box gene within the aboral ectoderm of developing sea urchin embryos.

A homeo box-containing gene, Hbox1 is expressed in an unusual and highly conserved spatial pattern in embryos of two different species of sea urchin, Tripneustes gratilla and Strongylocentrotus purpuratus. Hybridization in situ shows that this mRNA accumulates initially throughout the aboral ectoderm; however, between blastula and pluteus stages, the region containing Hbox1 mRNA retracts gradually until only a small area around the vertex is labeled in pluteus larvae. Aboral ectoderm appears cytologically uniform and also accumulates uniform levels of other tissue-specific mRNAs. Therefore, the Hbox1 pattern reveals a previously unsuspected heterogeneity of aboral ectoderm cells and a polarity within this tissue. In S. purpuratus, the Hbox1 gene product probably is not involved in initial specification of cell fate, as this message does not achieve a significant fraction of its peak abundance until almost hatching blastula stage, well after the time aboral ectoderm cells have initiated a tissue-specific program of gene expression. RNA blot and RNase protection analyses revealed low levels of Hbox1 mRNA in all adult tissues examined. However, this message was not detectable in mature eggs, suggesting that the Hbox1 gene does not have a maternal function. In addition to highly conserved spatial and temporal patterns of expression, the homeo box genes of these two urchin species also are conserved highly in sequences outside the homeo domain, despite the divergence of these two species (30-45 my). Two notable features of the protein shared with several vertebrate homeo proteins are a short conserved sequence encoded by an exon upstream of that encoding the homeo domain and a large region of high serine and proline content.

Amino Acid Sequence↗

PubMed1988

Stage- and tissue-specific expression of two homeo box genes in sea urchin embryos and adults.

We report the isolation of two different homeo box genes, HB3 and HB4, from the Hawaiian sea urchin Tripneustes gratilla. DNA sequencing revealed a definitive Antennapedia (Antp) class homeo box in each gene. Southern transfer hybridizations showed the genes to be single-copy. A 5.7-kb transcript of the HB3 gene was found in ovary, testis, small intestine and gastrula poly(A)+ RNA. The HB4 gene produces three transcripts. A 3.7-kb and a 4.4-kb transcript are expressed during embryogenesis. A 3.5-kb transcript appears in each of the adult tissues studied. The HB4 gene appears to be the sea urchin cognate of the Drosophila infrabdominal-7 (iab-7) gene, the mouse Hox 1.7 and Hox 3.2 genes and the Xenopus X1Hbox 6 gene. An examination of Antp class homeo box genes in deuterostomes indicates that a chromosomal duplication has taken place in the evolutionary line leading to the vertebrates after the divergence of the echinoderms. Thus, the sea urchin represents the primitive condition.

Amino Acid Sequence↗

PubMed1988

An engrailed class homeo box gene in sea urchins.

The homeo box, a conserved DNA element first recognized in Drosophila development-controlling genes, is present in the genomes of many higher metazoan species and provides a valuable probe for the isolation of regulatory genes from diverse phylogenetic groups. We have employed these probes to isolate and study the homeo-box genes in sea urchins. As in other species, the sea urchin homeo boxes fall into at least two classes defined by nucleotide sequence similarity to the homeo boxes of the Drosophila Antennapedia (Antp) and engrailed (en) genes. In this study, we characterize the only detectable sea urchin en class homeo box. Its nucleotide sequence similarity and lack of an intron indicate that it is more closely related to the two mouse en class homeo boxes than to the two Drosophila en class homeo boxes. These relationships are most parsimoniously explained if the single sea urchin en class homeo-box gene represents the primitive condition and the two mouse and the two Drosophila en class homeo-box genes represent independent duplications which occurred in the evolutionary lines leading to the vertebrates and arthropods, respectively. The most abundant en class gene transcripts detected by gel transfer analysis of RNA extracted from sea urchin tissues were found in Aristotle's lantern. Rare transcripts were present in ovary, testis and coelomocytes.

Amino Acid Sequence↗

PubMed1988

A gene expressed in the endoderm of the sea urchin embryo.

Using a previously cloned, developmentally regulated mRNA sequence expressed predominantly in the endoderm of sea urchin pluteus larvae, we isolated genomic clones and additional cDNA clones to define the gene and the protein it encodes. Nucleic acid sequencing revealed that the gene consists of four exons interrupted by three introns and spans approximately 3600 bp. It encodes a low-molecular-weight protein with polar ends. A stretch of Glu and Asp residues at its carboxyl terminus suggests that it is a nucleic acid-binding protein and a stretch of four Lys residues near the amino terminus suggests a nuclear localization signal.

Animals↗

PubMed1988

Sea urchin morphogenesis and cell-hyalin adhesion are perturbed by a monoclonal antibody specific for hyalin.

We have generated and characterized a monoclonal antibody (McA Tg-HYL) that recognizes sea urchin hyalin as evidenced by immunofluorescence staining of the hyaline layer (HL) and immunoblot staining of the hyalin protein band. On immunoblots of HL extracts only the hyalin protein reacted with McA Tg-HYL. Immunoprecipitates of radioactive proteins from embryos incubated with [35S]methionine yielded radioactive hyalin and 190, 140 and 105 x 10(3) Mr proteins associated with hyalin. McA Tg-HYL was generated against Tripneustes gratilla embryos but reacts with hyalin from the distantly related sea urchin species, Colobocentrotus atratus, Strongylocentrotus purpuratus, Arbacia punctulata, Lytechinus variegatus and Lytechinus pictus. Developing embryos of the above-mentioned six species were treated with McA Tg-HYL and did not gastrulate or form arms. Observations of treated embryos revealed areas of separation of the hyaline layer from the underlying embryonic cells, suggesting that McA Tg-HYL was interfering with binding of the cells to the HL. Using the centrifugation-based adhesion assay of McClay et al. (Proc. natn. Acad. Sci. U.S.A. 78, 4975-4979, 1981), Fab' fragments of McA Tg-HYL were found to inhibit cell-hyalin binding. McA Tg-HYL did not inhibit hyalin gelation in vitro or the reaggregation of dissociated blastula cells. We postulate that McA Tg-HYL recognizes an evolutionarily conserved hyalin domain involved in cell-hyalin binding and required for normal epithelial folding.

Animals↗

PubMed1986

Length and sequence polymorphisms in the ribosomal gene spacer of the Hawaiian sea urchin, T. gratilla.

Blot-hybridization of sea urchin (Tripneustes gratilla) genomic DNA with a cloned rDNA probe revealed individual variation in the length of the rDNA repeat unit and also in the non-transcribed spacer sequences. The number of distinct rDNA repeat subclasses distinguishable within any one sea urchin was limited and usually 2 to 3. However, examination of a number of sea urchins indicated a large number of distinct rDNA repeat types in the population as a whole; all of the rDNA repeat types in nine individuals were different. The presence of limited heterogeneity in the rDNA repeats of single individuals, with may different repeat types in the population as a whole, suggests that rDNA variants can be rapidly and selectively propagated within a chromosomal lineage.

Animals↗

PubMed1986

Stage-specific expression of a homeo box-containing gene in the non-segmented sea urchin embryo.

Hybridization of Drosophila homeo box DNA probes to Southern transfers of genomic DNA from the Hawaiian sea urchin Tripneustes gratilla has revealed that the sea urchin genome contains at least five homeo boxes. Examination of the DNA from several individuals shows that the sequences flanking these homeo boxes exhibit little restriction fragment length polymorphism, indicating they are more highly conserved than the majority of sea urchin DNA. Several clones in a T. gratilla genomic DNA library which hybridized with Drosophila homeo box probes were identified, and one found to be transcribed during embryogenesis was selected for further study. Southern transfer hybridizations showed the cloned gene to be single-copy. DNA sequencing of the sea urchin gene defined a homeo box 70-73% homologous to the Drosophila homeo box probes and an encoded homeo domain 78-88% homologous to those encoded by the probes. Hybridization of DNA probes from the sea urchin homeo box-containing gene to Northern transfers of embryonic RNA demonstrated that the gene produces two transcripts of 6.9 kb and 7.7 kb. Transcripts first accumulate at blastula stage and increase to a maximum level at gastrula stage before decreasing considerably in abundance by pluteus stage.

Amino Acid Sequence↗

PubMed1986

Tumor promoters and diacylglycerol activate the Na+/H+ antiporter of sea urchin eggs.

Various tumor promoters (TPA, lyngbyatoxin and aplysiatoxin) and diacylglycerol induced cytoplasmic alkalinization of sea urchin eggs independently of intracellular Ca2+ release. This response stimulated protein synthesis and was blocked by amiloride or a lack of extracellular Na+, procedures which inhibit the Na+/H+ antiporter. These results suggest that the antiporter which is responsible for cytoplasmic alkalinization in sea urchin eggs is activated directly or indirectly by protein kinase C in a Ca2+-independent manner.

Amiloride↗

PubMed1985

Effects of bepridil on force development and transmembrane electrical activity of adult canine Purkinje strands: comparison with nisoldipine and lidocaine.

Superfusion of a bepridil containing normal Tyrode solution produced a concentration-dependent decrease in force development and shortening of action potential duration in isolated Purkinje strands. At a concentration close to the 50% inhibitory concentration for effects on force development and action potential duration, bepridil blocked 'slow' action potentials in potassium depolarized, isoproterenol-restored Purkinje strands. When the concentration-response relations for decreasing force development and shortening action potential duration were compared to those of nisoldipine and other calcium entry inhibitors, bepridil was the least potent. Bepridil also showed a concentration-dependent effect on the maximum rate of rise of the Purkinje fiber action potential upstroke similar to, though less potent than that of lidocaine. In addition, like lidocaine, bepridil shifted the membrane responsiveness curve of the Purkinje strands to the left. These findings indicate that bepridil produced effects that are consistent with its classification as a calcium entry inhibitor (class IV antiarrhythmic), as well as effects that are 'lidocaine-like' (class I antiarrhythmic).

Animals↗

PubMed1985

Characterization of toposomes from sea urchin blastula cells: a cell organelle mediating cell adhesion and expressing positional information.

Cell adhesion in the sea urchin blastula is mediated by a 22S genus-specific glycoprotein complex consisting initially of six 160-kDa subunits that are processed proteolytically as development proceeds. Noncytolytic removal of the 22S particle from the surface with either 2.5% butanol or trypsin renders dissociated cells reaggregation incompetent, and addition restores reaggregation and development. Polyclonal antibodies against the 22S complex prevent reaggregation in a genus-specific manner while monoclonal antibodies stain cell surface structures in a pattern consistent with a code that specifies the position of a cell in the embryo by a unique combination of subunits in its cell adhesion particles. The existence of similar particles in Drosophila and amphibian embryos suggests that these glycoprotein complexes are a general class of organelles, the toposomes, that in the embryo mediate cell adhesion and express positional information.

Animals↗

PubMed1985

Structural analysis of ribosomal RNA genes from the Hawaiian sea urchin species, Tripneustes gratilla.

Ribosomal gene sequences (rDNA) were isolated from the genomic DNA of the Hawaiian sea urchin Tripneustes gratilla by cloning in plasmid and phage vectors. The rRNA coding regions in four clones were localized by hybridizing Southern transfers of endonuclease-digested DNA with 32P-labeled 26S rRNA and 32P-labeled 18S rRNA. Three of the rDNA clones were isolated from a library of DNA from a single sea urchin and represent the two major types of rDNA repeats present in that individual's genome. Both repeat types appear identical within their rRNA coding regions but are dissimilar in an area of the nontranscribed spacer (NTS) adjacent to the 3' end of the 26S coding region. The cloned rDNA repeats were shorter than their genomic counterparts due to deletions occurring within internally repetitious NTS domains, probably as a result of unequal recombination during phage propagation.

Animals↗

PubMed1985

Transcription of similar sets of rare maternal RNAs and rare nuclear RNAs in sea urchin blastulae and adult coelomocytes.

We studied the sequences transcribed in the rare class of hnRNA and the rare maternal RNA set in blastula embryos and a tissue of adult sea urchins, coelomocytes. About 26% of labelled single-copy DNA formed hybrids which bound to hydroxyapatite after three cycles of hybridization with nuclear RNA from blastulae and coelomocytes. This corresponds to transcription of about 50% of the single-copy genome by both cell populations. To compare the rare hnRNA sequences synthesized by blastulae and coelomocytes directly, labelled single-copy DNA was hybridized with blastula nuclear RNA to high RNA C0t, fractionated into sequences complementary and non-complementary to blastula nuclear RNA by chromotography on hydroxyapatite, and then each fraction was rehybridized with nuclear RNA from blastulae and coelomocytes. About 62% of the labelled DNA complementary to blastula nuclear RNA and about 1.5% of the labelled DNA non-complementary to blastula nuclear RNA hybridized with nuclear RNA from both cell populations. Thus, coelomocytes and blastula embryos transcribe essentially the same single-copy sequences in the rare hnRNA class. A probe for the rare maternal RNA set was isolated by hybridizing single-copy DNA with total egg RNA to high RNA C0t. 65-67% of this probe hybridized with whole-cell RNA from eggs, blastulae, plutei and coelomocytes demonstrating that essentially all rare maternal RNAs are present, and presumably transcribed, in blastulae, plutei and coelomocytes.

Animals↗