Treatment of Refractory Gastrointestinal Bleeding in Patients With Portal Hypertension: A Case Series and Treatment Algorithm.

In patients with portal hypertension, ectopic varices can develop at any site along the gastrointestinal tract outside the classically described gastroesophageal location. Like esophageal variceal hemorrhage, bleeding from ectopic varices can be life-threatening. Diagnosis and treatment of ectopic varices can be challenging; to date, no effective treatment algorithm has been described. A systematic teamwork approach to diagnosing and treatment of ectopic varices is required to successfully manage hemorrhage from ectopic varices.

An evaluation of optimal sampling strategy and adaptive study design.

We have evaluated the utility of optimal sampling strategy coupled with adaptive study design in the determination of individual patient and population pharmacokinetic parameter values. In 9 patients with cystic fibrosis receiving a short (1 minute) infusion of ceftazidime pharmacokinetic parameter values were determined with a nonlinear least-squares estimator analyzing a traditional, geometrically spaced set of 12 postinfusion serum samples drawn over 8 hours. These values were compared with values generated from four sample subsets of the 12 obtained at optimal times and analyzed by nonlinear least-squares estimator, as well as a maximum a posteriori probability Bayesian estimator with prior distributions placed on beta and clearance. The four sampling times were determined according to an adaptive design optimization technique that employs sequential updating of population prior distributions on parameter values. Compared with the 12-point determination, the four optimal points analyzed with the maximum a posteriori probability Bayesian estimator faithfully reproduced both microscopic and hybrid pharmacokinetic parameter values for individual patients and, consequently, also produced accurate measures of population central tendency and dispersion. This has important implications in being able to more efficiently derive target patient population pharmacokinetic information for new drugs. This should also allow generation of better concentration-effect relationships in populations of interest.

A diazepam binding inhibitor (DBI) homolog from the tobacco hornworm, Manduca sexta.

A cDNA clone encoding a protein with 42-51% identity to the mammalian diazepam binding inhibitors (DBIs) has been isolated and sequenced from a midgut cDNA library of the tobacco hornworm, Manduca sexta. The putative M. sexta DBI is 90 residues in length and shares the predicted internal a-helices common to the mammalian DBIs. Sequence alignments indicate that the M. sexta DBI contains three potential proteolytic cleavage site lysines in the same positions as the mammalian DBIs. High DBI mRNA levels were found in larval midgut, larval fat body, and adult ovary. DBI mRNA was also detected in larval prothoracic glands, larval nerve cord, and adult testis. These results suggest that the putative M. sexta DBI is an important gene product with a high degree of identity to the known mammalian DBIs. The M. sexta DBI may therefore be the functional homolog to the mammalian DBIs.

Glutathione S-transferases from larval Manduca sexta midgut: sequence of two cDNAs and enzyme induction.

Two glutathione S-transferase (GST) clones from a larval midgut cDNA library of the tobacco hornworm, Manduca sexta were sequenced. The nucleotide sequence of the first clone, M. sexta GST1, encoded a protein of 217 amino acids with a predicted molecular weight of 24,644 and isoelectric point of 4.8. The M. sexta GST1 was 45.9-48.6% identical to GSTs from Musca domestica and several Drosophila species. The M. sexta GST2 cDNA encoded a protein of 203 amino acids with a predicted molecular weight of 23,596 and isoelectric point of 5.5. The M. sexta GST2 shared 44.8-50.0% sequence identity to a second cluster of insect GSTs from M. domestica, D. melanogaster and Anopheles gambiae. GST1 and GST2 were only 24.1% identical in amino acid sequence. The divergence of these two classes of insect GSTs occurred before the radiation of Diptera and Lepidoptera. Northern analysis of the expression of these GSTs showed increased GST1 mRNA levels in midguts of larvae fed diets containing 2-undecanone, or phenobarbital. Midgut and fat body cytosolic GST activities were induced when larvae were fed diets containing 2-tridecanone, 2-undecanone, or phenobarbital. Partial purification of midgut GSTs by size-exclusion and glutathione affinity chromatography resulted in a series of isoelectric focusing bands, with the major one corresponding to the predicted isoelectric point of the M. sexta GST1. In summary, two midgut GSTs have been identified on the basis of cDNA sequence and one of these, GST1, was inducible by dietary chemicals.

Inducible P450s of the CYP9 family from larval Manduca sexta midgut.

Several related cytochrome P450 cDNAs belonging to the CYP9 family have been cloned from the midgut of larval tobacco hornworms, Manduca sexta. The first P450, CYP9A2, was obtained by RT-PCR using degenerate primers. Northern blot analysis of expression in the midgut using the CYP9A2 probe revealed a significant induction by a variety of chemicals. Diets supplemented with the wild tomato compound 2-undecanone caused a dose-dependent induction which peaked after 48 h. Induction was also observed after addition to the diet of indole-3-carbinol, phenobarbital, 2-tridecanone and xanthotoxin. Neither alpha-pinene, clofibrate nor nicotine were effective inducers. The CYP9A2 probe hybridized to two mRNA species, one of 2. 0 kb and another of 4.2 kb, suggesting cross-hybridization to other P450 mRNAs. Additional P450 clones of the CYP9 family were then obtained and sequenced. Northern hybridization revealed that the 4.2 kb band also hybridized to CYP9A4 whereas the 2.0 kb hybridized to CYP9A5. Despite being 91% identical, CYP9A4 and CYP9A5 were induced differentially by clofibrate and xanthotoxin. Multiple P450 genes from various families are therefore induced in Lepidoptera in response to plant allelochemicals or xenobiotics.

Pilot trial of prophylactic ursodiol to decrease the incidence of veno-occlusive disease of the liver in allogeneic bone marrow transplant patients.

Ursodiol is a hydrophilic, non-hepatotoxic bile salt indicated for the medical treatment of cholesterol gallstones. This pilot study explored the use of prophylactic ursodiol in an attempt to decrease the incidence and severity of veno-occlusive disease (VOD) of the liver following allogeneic bone marrow transplantation (BMT). Between February 1991 and January 1992, 22 consecutive patients undergoing BMT for hematologic malignancies received the BU(4)/CY(2) preparative regimen and CSA/MTX for GVHD prophylaxis. Ursodiol, 600-900 mg daily by mouth was begun at least 1 day prior to beginning the preparative regimen. Results for this pilot group were compared to a control group of 28 consecutive patients transplanted between June 1989 and January 1991 with the same regimen without ursodiol. There were no significant differences in disease or clinical status between the groups pretransplant. However, mean baseline AST levels were significantly higher in the ursodiol group, 28.0 U/l vs 18.1 U/l in the control group (p = 0.001). The median maximum bilirubin observed post-transplant was 2.35 mg/dl (range 0.9-45) in the ursodiol group, and 5.05 mg/dl (range 0.7-29.4) in controls. The incidence of VOD was 2/22 (9.1%) in the ursodiol group and 18/28 (64.3%) in controls (p = 0.0001). Death due to VOD occurred in 1/22 patients (4.5%) in the ursodiol group and in 6/28 (21.4%) controls (p = 0.12). Our data suggest that ursodiol may decrease the incidence of VOD in allogeneic BMT patients.

Carmustine, Ara C, cyclophosphamide and etoposide with autologous bone marrow transplantation in relapsed or refractory lymphoma: a dose-finding study.

The purpose of this study was to define the dose-limiting non-hematologic toxicity of carmustine, Ara C, cyclophosphamide and etoposide (BACE). Between October 1986 and March 1990, 37 patients with relapsed or refractory lymphoma received escalating doses of combination chemotherapy followed by autologous bone marrow transplant (ABMT). Twenty patients with Hodgkin's disease (HD) and 17 patients with intermediate or high grade non-Hodgkin's lymphoma (NHL) initially received conventional-dose therapy with either a 7 week course of modified MACOP-B or a single dose of cyclophosphamide (CY) at 2 g/m2 depending on prior therapy and response. Regardless of response, patients then received escalating doses of BACE, toxicity permitting. Ten patients obtained complete responses (CR) and 12 patients were partial responders (PR), CR+PR (75%) with modified MACOP-B and 7 (64%) patients obtained PR with CY. The maximum-tolerated dose (MTD) for BACE was determined to be carmustine 700 mg/m2, Ara C 1500 mg/m2, CY 150 mg/kg and etoposide 1500 mg/m2. When Ara C was escalated from 1500 mg/m2 to 3000 mg/m2 holding the other drugs at the prior doses, the next two patients died secondary to diffuse alveolar damage. Overall and event-free survivals are identical with 14 of 37 patients (38%) alive with a median follow-up of 61 months (range 38-79 months). Ten patients were treated at the MTD, none of whom died a toxic death and 3 (30%) are alive with a median follow-up of 42 months (range 38-52 months). We defined the MTD and BACE showing pulmonary toxicity to be the dose-limiting non-hematologic toxicity.(ABSTRACT TRUNCATED AT 250 WORDS)

Molecular phylogeny of glutathione-S-transferases.

The glutathione-S-transferase (GST) protein superfamily is currently composed of nearly 100 sequences. This study documents a greater phylogenetic diversity of GSTs than previously realized. Parsimony and distance phylogenetic methods of GST amino acid sequences yielded virtually the same results. There appear to be at least 25 groups (families) of GST-like proteins, as different from one another as are the currently recognized classes. This diversity will require the design of a new nomenclature for this large protein superfamily. There is one well-supported large clade containing the mammalian mu, pi, and alpha classes as well as GSTs from molluscs, helminths, nematodes, and arthropods.

Neuroactive steroids modulate crustacean locomotor activity.

We examined the effects of neuroactive steroids known to modulate gamma-aminobutyric acid(A) (GABA)(A) receptor activity, on locomotor activity in a submerged circular open-field apparatus. Juvenile male lobsters, Homarus americanus, were treated with a single administration of an agonist, 3alpha-hydroxy-5alpha-pregnan-20-one (allopregnanolone, 3alpha,5alpha-TH PROG), an antagonist, pregnenolone sulfate (PREGS), or vehicle alone. 3alpha,5alpha-TH PROG treatment (125 and 250 microg) significantly reduced while PREGS significantly elevated locomotor activity in a dose-dependent manner similar to diazepam. PREGS increased locomotor activity at 30 and 60 microg, while diminishing such activity and altering locomotor patterns at 120 microg. These results suggest that neuroactive steroids may affect crustacean GABA receptors in a fashion similar to the GABA(A) type found in the vertebrates, and that they may be involved in the regulation of locomotor behavior.

Environmental endocrine disruption in decapod crustacean larvae: hormone titers, cytochrome P450, and stress protein responses to heptachlor exposure.

A variety of enzymes and other proteins are produced by organisms in response to xenobiotic exposures. Cytochrome P450s (CYP) are one of the major phase I-type classes of detoxification enzymes found in terrestrial and aquatic organisms ranging from bacteria to vertebrates. One of the primary functions of stress proteins (HSPs) is to aid in the recovery of damaged proteins by chaperoning their refolding. These and other biomarkers of xenobiotic exposure and resulting effects have not been studied in crustacean larvae. This information is of potential importance for environmental management and risk assessment. In this work, we have given Homarus americanus larvae single 24 h exposures to the cyclodiene pesticide heptachlor, a known environmental endocrine disruptor (EDC) on different days of the 1st larval instar. We followed these larvae during the first larval stage for effects on timing of ecdysis to 2nd stage, ecdysteroid molting hormone titers, and alterations in the levels of cytochrome P450 CYP45 and HSP70 proteins. Delays in ecdysis were correlated with alterations in ecdysteroid levels. This result provides clues that this pesticide may function as an environmental endocrine disruptor in crustaceans. CYP45 and HSP70 levels were significantly elevated for several days following heptachlor exposure. The elevation in HSP70 was prolonged depending on the day of pesticide exposure and this was directly related to the increase in mortality. These results demonstrate the utility of these measurements as potential biomarkers in crustacean larval developmental toxicology and EDC effects research.

Competition for space among sessile marine invertebrates: changes in HSP70 expression in two Pacific cnidarians.

The role of stress proteins-either constitutive (HSC) or inducible (HSP)-of the HSP70 family in intra- and interspecific competition for space was examined in two sessile Pacific cnidarians. Anthopleura elegantissima, an intertidal anemone, and Corynactis californica, a subtidal corallimorpharian, express HSP70 in the absence of apparent physical stress. HSP70 protein expression is concentrated in the tentacles of A. elegantissima when the animal is exposed to contact with other benthic organisms. Under the same conditions, however, HSP concentrations are similar in the body and tentacles of C. californica. When two different clones of A. elegantissima interact in the field, the outside polyps (warriors) express more HSP70 than the inside ones (2.4 versus 0.6 ng HSP70/microg Protein). When different C. californica clones interact, HSP70 expression in the outside and inside polyps is similar (1.5 versus 1.8 ng HSP70/microg P) and is fairly constant in the corallimorpharian in the different interspecific encounters. HSP70 expression is related to the different kinds of aggression encountered by both cnidarians. HSP70 expression may be involved in the recovery of tissues damaged by the allelochemical, cytotoxical, or corrosive substances produced by different enemies. C. californica clones appear prepared for war, as evidenced by the high constant expression of HSP70 in the polyps. A. elegantissima exhibits differential HSP70 expression depending on the identity of each neighboring intra- or interspecific sessile competitor. We propose that stress proteins can be used to quantify space competition or aggression among sessile marine invertebrates.

Identification of a new cytochrome P450 family, CYP45, from the lobster, Homarus americanus, and expression following hormone and xenobiotic exposures.

A new cytochrome P450, the first member of the CYP45 family, was identified from the hepatopancreas of the American lobster, Homarus americanus. The lobster CYP45 shares significant sequence homologies to the vertebrate CYP3 and the invertebrate CYP6, CYP9, CYP28, and CYP30 families, perhaps indicating a common ancestor of these P450s. Of seven tissues examined, CYP45 was expressed only in the hepatopancreas, the crustacean equivalent of the vertebrate liver, pancreas, and intestine. Over the course of the lobster molt cycle, CYP45 expression mirrored the hemolymph titer of ecdysteroids, suggesting its potential involvement in molting hormone dynamics. This idea was strengthened further by ecdysteroid treatment of intermolt-stage lobsters during the lowest hemolymph titers and CYP45 expression levels. Significant elevations in hepatopancreas CYP45 mRNA levels were elicited by such injections over a 2- to 4-day interval. Similar experiments were performed by intubating juvenile lobsters with various xenobiotics. Induction of CYP45 expression occurred following phenobarbital and heptachlor administration, but not by beta-naphthoflavone. Hormonal and xenobiotic modulation of lobster CYP45 expression provides a potential pathway for endocrine disruption in lobsters.

Cytochrome P450 enzymes belonging to the CYP4 family from marine invertebrates.

Six new cytochrome P450s assigned to the CYP4 family were identified from marine invertebrates belonging to the arthropod, mollusc, and echinoderm phyla. These are the first reported members of the CYP4 gene family from marine invertebrates, and extends the finding of the CYP4 family of cytochrome P450s to molluscs and echinoderms. Members of each phyla (echinodermata, arthropoda (crustacea), and mollusca) expressed genes belonging to the CYP4C subfamily in their respective digestive tissues. A mollusc, the mussel Mytilus galloprovincialis, expressed a gene belonging to a new CYP4 subfamily, CYP4Y. In Northern blotting experiments with digestive tissues of M. galloprovincialis, the expression of the CYP4Y1 gene was found to be inhibited by increasing concentrations of the hydrocarbon beta-naphthoflavone. Thus, the potential use of marine invertebrate CYP4 genes as biomarkers of xenobiotic exposures may be warranted.