PubMed Health⌕ Search

Biomedical subjects

J L Bennett

Publications and source records attributed to J L Bennett.

At least 19 recordsLinked to original sources
PubMed1992

Characterization of a Xenopus laevis ribonucleoprotein endoribonuclease. Isolation of the RNA component and its expression during development.

In order to facilitate studies of the assembly and transport of the site-specific RNase mitochondrial RNA processing (MRP) ribonucleoprotein, we have characterized it from Xenopus laevis cells. X. laevis RNase MRP displayed a similar spectrum of cleavage activity to that produced by previously isolated mammalian nuclear enzymes. A 277-nucleotide RNA component of the ribonucleoprotein was identified; the gene for the RNA was isolated, sequenced, and found to be 66 and 63% similar to mouse and human RNase MRP RNAs, respectively. Despite the evolutionary distance from its mammalian counterparts, X. laevis RNase MRP RNA contains five regions of homology to the mammalian RNase MRP RNA. Four of these regions correspond to those previously identified as conserved between RNase MRP and RNase P RNAs; the fifth encompasses nucleotides recently discovered to be sufficient for autoantigen binding. The expression and assembly of Xenopus RNase MRP RNA were examined in frog oocytes and developing embryos. RNase MRP RNA was expressed throughout oogenesis; it started to accumulate at stage I and reached a maximum in stage IV. During embryogenesis RNase MRP RNA expression began to elevate at approximately stage 22 and continued to rise through the swimming tadpole stage. When injected into the nucleus of mature oocytes, the X. laevis RNase MRP RNA gene was expressed accurately, and transcripts were packaged into immunoprecipitable particles.

Animals↗
PubMed1992

High-performance liquid chromatographic method for the analysis of Oltipraz in human serum and urine.

A rapid, sensitive procedure for the analysis of Oltipraz in serum and urine using high-performance liquid chromatography was developed. The proposed method illustrates recovery of Oltipraz from biological fluids was greater than 80%. Detection and separation of Oltipraz required as little as 1 ml of serum or urine. Oltipraz was detectable when 2 ng or more of drug was present in 1 ml of serum or urine; the method is highly reproducible when 5 ng/ml or more Oltipraz is present in the biological fluid.

Chromatography, High Pressure Liquid↗
PubMed1992

Clinical pharmacology studies of oltipraz--a potential chemopreventive agent.

Pharmacological studies on Oltipraz [4-methyl-5(pyrazinyl-2)-1-2-dithiole-3-thione)] were conducted with normal healthy subjects using various doses and schedules. Administration of single doses (1, 2 and 3 mg/kg) resulted in detectable drug levels in the serum (mean peak serum concentrations 16, 61 and 205 ng, respectively) and urine. The t1/2 was short (4.4, 4.1 and 5.3 hours respectively) and no steady state was achieved after multiple daily doses for 12 days. Introduction of a loading dose during the first day produced a steady state when 1.5 and 2.0 mg/kg/day were used. Daily administration of Oltipraz sustained the steady state with insignificant variations. Consumption of a high fat diet increased the serum and urine concentrations of Oltipraz (30-60%) compared to the low fat diet. Two subjects experienced flatulence during the administration of the drug. One subject developed numbness and pain in the thumbs with occurrence of small purplish-black spots resembling those observed in subacute endocarditis. These changes disappeared 10 days after discontinuation of the drug. No changes in peripheral blood counts, biochemical profile or thyroid function tests were observed after four weeks of Oltipraz. Further studies with a larger number of healthy subjects are needed for clarification of the safety and biological efficacy of small doses of Oltipraz during chronic administration.

Adult↗
PubMed1992

Schistosoma mansoni: patch-clamp study of a nonselective cation channel in the outer tegumental membrane of females.

An apparent ion channel with a conductance of 295 pS is present in isolated inside-out patches of outer tegumental membrane taken from female Schistosoma mansoni. With positive voltages applied to the intracellular face of the patch, percentage open time for the channel was 0 to 50; with negative voltages applied, percentage open time was greater than 99. Step changes in applied voltage characteristically induced opening-closing activity. However, there was no maintained applied voltage at which there was a high level of sustained opening-closing activity. The 295 pS conductance was by far the most commonly occurring conductance but it appears to result from cooperativity among several channels, the unitary conductance for the channel averaging 95 pS. Alterations in the Na+ or K+ concentration ratios changed the reversal potential for this conductance but alterations in the Cl- concentration did not. From this it is concluded that this channel is selective for Na+ or K+ over Cl- and it appears to be a nonselective cation channel.

Animals↗
PubMed1992

Praziquantel: physiological evidence for its site(s) of action in magnesium-paralysed Schistosoma mansoni.

The mechanism whereby praziquantel produces a contraction and subsequent flaccid paralysis (a loss of sensitivity to subsequent stimuli) of Schistosoma mansoni in a medium containing an elevated Mg2+:Ca2+ ratio was investigated. In RPMI, praziquantel produced a concentration-dependent tonic contraction of the parasite with an EC50 of 200 nM. Magnesium inhibited the contraction in such a manner as to convert the tonic contraction to a phasic one without altering the peak force generated. The Mg(2+)-dependent block was non-competitive with praziquantel but was competitive with extracellular Ca2+, ratios of 7.5:1;Mg2+:Ca2+ being needed to inhibit the tonic contraction and to induce flaccid paralysis. Flaccid paralysis was associated with a reduced ability of the parasite to take up 45Ca2+ from the bath compared to parasites that had not entered into flaccid paralysis and flaccid paralysis was reversible. Recovery from flaccid paralysis was accelerated by treatments that are expected to increase Ca2+ uptake by the parasite. At a concentration of 500 nM, praziquantel produced 2 distinct phasic contractions in intact parasites incubated in an elevated [Mg2+] medium but only 1 phasic contraction in parasites lacking their surface tegumental membranes. In zero Ca2+ I-RPMI, 10 microM praziquantel produced a phasic contraction of intact parasites but did not stimulate contraction of detegumented parasites until Ca2+ was reintroduced into the bath. These results indicate that praziquantel interacts with specific Ca(2+)-permeable sites in the tegumental and sarcoplasmic membranes of the parasite and that under these conditions of elevated Mg2+:Ca2+ ratios, these sites become blocked by Mg2+, leading to flaccid paralysis of the parasite.

Animals↗
PubMed1992

Neurobiology of parasitic flatworms: how much "neuro" in the biology?

The nervous systems in the parasitic Platyhelminthes have generally been considered to be degenerate and of marginal significance, but recent studies have shown these systems to be more significant in the biology of these animals than formerly believed. There are many similarities in the construction and apparent neurochemistry of the nervous systems in the parasitic forms as well as in the free-living Turbellaria. In all forms there appears to be a large neurohormonal component. Though the nervous system appears to be important for many aspects of parasitic flatworm biology, little direct or specific information about the physiology of these systems is yet available.

Animals↗
PubMed1991

Impairment of mitochondrial transcription termination by a point mutation associated with the MELAS subgroup of mitochondrial encephalomyopathies.

Defects in mitochondrial DNA (mtDNA) are associated with several different human diseases, including the mitochondrial encephalomyopathies. The mutations include deletions but also duplications and point mutations. Individuals with MELAS (mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes) carry a common A-to-G substitution in a highly conserved portion of the gene for transfer RNA(Leu(UUR)). Although the MELAS mutation may be comparable to the defect in the tRNA(Lys) gene associated with MERRF (myoclonus epilepsy associated with ragged-red fibres), it is also embedded in the middle of a tridecamer sequence necessary for the formation of the 3' ends of 16S ribosomal RNA in vitro. We found that the MELAS mutation results in severe impairment of 16S rRNA transcription termination, which correlates with a reduced affinity of the partially purified termination protein for the MELAS template. This suggests that the molecular defect in MELAS is the inability to produce the correct type and quantity of rRNA relative to other mitochondrial gene products.

Base Sequence↗
PubMed1991

Oesophageal varices in a region of the Sudan endemic for Schistosoma mansoni.

In a field study of two villages in the Gezira, an area of the Sudan endemic for Schistosoma mansoni, liver ultrasonography was used to detect subjects with Symmers' hepatic periportal fibrosis, some of whom underwent oesophagoscopy to detect oesophageal varices. The prevalence of oesophageal varices in subjects undergoing oesophagoscopy was 54 per cent and 67 per cent respectively, occurring mainly in males aged about 30 years. The varices were usually asymptomatic. Symptomatic varices (with a positive history of haematemesis) occurred in 4 per cent and 3 per cent respectively of subjects with sonographic evidence of liver periportal fibrosis. By detecting oesophageal varices in an asymptomatic phase, hepatic ultrasonography and fibreoptic oesophagoscopy may elucidate the natural history of the varices and their response to periodic anti-schistosomal chemotherapy.

Adult↗
PubMed1991

Purification and characterization of 3-hydroxymethylglutaryl-coenzyme A reductase of Schistosoma mansoni: regulation of parasite enzyme activity differs from mammalian host.

The enzyme 3-hydroxymethylglutaryl-CoA (HMG-CoA) reductase plays a critical role in regulating the production of cholesterol, dolichols, and ubiquinones in mammals. The inhibition of this enzyme in Schistosoma mansoni is accompanied by a cessation of egg production by the female parasite and a reduced ability of the parasite to properly glycoslyate their proteins. Furthermore, we recently demonstrated that mevinolin, if given continuously over a period of 10-14 days, is a potent antischistosomal drug. In this paper, we describe the properties of purified HMG-CoA reductase from S. mansoni. Using affinity chromatography, we were able to obtain a 417-fold purification of the enzyme which had Km values similar to the rat enzyme for HMG-CoA and NADPH. The Ki value for mevinolin, a potent and selective inhibitor of the rat reductase (Ki = 0.6 nM), was significantly higher (Ki = 46 nM) for the schistosome enzyme. SDS-PAGE and HPLC of the purified enzyme resulted in the appearance of a single protein, which had a molecular weight (66,000) in the range reported for the rat enzyme. Parasite reductase activity, unlike that of its host, did not display a circadian rhythm. Furthermore, agents which elevate (cholestyramine) or decrease (cholesterol) mammalian reductase activity had no effect on the parasite enzyme. Our results suggest that the mechanism which regulates production of the parasite's enzyme may differ from its mammalian host.

Acyl Coenzyme A↗
PubMed1991

The schistosomicidal compound Ro 15-5458 causes a reduction in the RNA content of Schistosoma mansoni.

The effect of Ro 15-5458 (10-2-(diethylamino)ethyl-9-acridanone(2-thiazolin- 2-yl)hydrazone) on the steady-state RNA levels of Schistosoma mansoni was studied after dosing the host with 15 mg kg-1 and retrieving parasites. Total RNA content of parasites recovered from the host 12, 72 and 96 h after dosing was reduced by 14, 30 and 41%, respectively. Quantitative filter hybridization of blots of RNA extracted from treated and control parasites with specific probes indicated a decline in actin and superoxide dismutase mRNA as well as rRNA of treated parasites. The decline was observed 12 h after dosing, 48 h before parasites showed drug-induced changes in other vital biological processes. A prominent drug-induced reduction was seen on the 1.9 kb actin mRNA compared to the 1.4 kb. The same dose of the drug did not alter the actin mRNA content of the host liver. Similarly, the administration of the inactive structural analogue Ro 21-6787 (10-2-(diethylamino)ethyl-9-acridanone) was without any effect. We propose that the actions of Ro 15-5458 and/or its products are directed towards the inhibition of the expression of parasite genes.

Acridines↗
PubMed1991

Studies on muscle cells isolated from Schistosoma mansoni: a Ca(2+)-dependent K+ channel.

Muscle cells from adult male Schistosoma mansoni have been isolated using a combination of papain digestions and mechanical dissociation procedures. The muscle fibres isolated in this way lacked nuclei but they did contract and relax in response to high [K+], a response which was blocked in the presence of Co2+. From this we conclude that the isolation procedure yields viable muscle fibres useful for physiological studies. Patch-clamp recordings taken from the isolated fibres show a variety of discrete ionic conductances. In inside-out patches one prominent channel was a Ca(2+)-activated K+ channel with a conductance of 195 pS and a selectivity greater than 10:1 for K+ over Na+, Cs+ or NH4+. Percentage open time was dependent on [Ca2+] at the intracellular face. With [Ca2+] at 1 microM or greater percentage open time was greater than 95%; at 0.1 microM it was less than 2%. No voltage sensitivity could be detected in the voltage range from -50 to -10 mV membrane potential. Ba2+ (10 mM), but neither tetraethylammonium nor 3,4-diaminopyridine blocked the channel from the intracellular face. This Ca(2+)-activated K+ channel in the muscle membrane of this acoelomate animal is similar in most respects to the maxi-K+ channels which have been described in a variety of cells from more highly evolved animals.

Animals↗
PubMed1991

Neurobiology of parasitic platyhelminths: possible solutions to the problems of correlating structure with function.

This paper provides an overview of research on the nervous system of parasitic platyhelminths. We have emphasized studies concerned with the physiological, pharmacological and biochemical nature of the major small molecule neurotransmitters of these parasites. We have attempted to provide a critical review of the work by focusing on important unresolved issues. Finally, we have focused on some recent work in our laboratory, using patch-clamp recording techniques and quantitative fluorescence cytometry, as an example of newer methods that will hopefully resolve some of the unanswered questions concerning the nervous system of these parasites.

Acetylcholine↗
PubMed1991

Nuclear RNase MRP processes RNA at multiple discrete sites: interaction with an upstream G box is required for subsequent downstream cleavages.

RNase MRP is a site-specific endoribonuclease that processes primer RNA from the leading-strand origin of mammalian mitochondrial DNA replication. It is present in active form as isolated from the nucleus, suggesting a bipartite cellular location and function. The relatively high abundance of nucleus-localized RNase MRP has permitted its purification to near homogeneity and, in turn, has led to the identification of protein components of this ribonucleoprotein. Analysis of the mode of RNA cleavage by nuclear RNase MRP revealed the surprising and unprecedented ability of the endonuclease to process RNA at multiple discrete locations. Substrate cleavage is dependent on the presence of a previously described G-rich sequence element adjacent to the primary site of RNA processing. Downstream cleavage occur in a distance- and sequence-specific manner.

Animals↗
PubMed1991

Association of the therapeutic activity of praziquantel with the reversal of Symmers' fibrosis induced by Schistosoma mansoni.

The impact of antischistosomal chemotherapy on the most important complication of intestinal schistosomiasis, Symmers' periportal fibrosis, has not been determined. Since abdominal ultrasonography has proven to be an effective tool in assessing the extent of Symmers' fibrosis in patients, we monitored the effect of chemotherapy, which involved the annual administration of praziquantel, on 48 Sudanese villagers having varying degrees of Symmers' fibrosis. Results indicate no significant differences in the fibrotic status of the 48 patients between 1986 and 1987, but test statistics (both the Wilcoxon signed rank test and Friedman's block/treatment test), indicated a significant decrease between the fibrotic status of the patients in 1986 and their fibrotic status in 1988 and 1989. Thus, after three years of therapy, 12 of the 48 patients no longer had detectable Symmers' fibrosis, while another 16 patients experienced a reduction in the amount of fibrosis in their livers. When coupled with our previous study, which demonstrated that annual treatment of children with praziquantel prevents the appearance of Symmers' fibrosis, it now appears that praziquantel may reverse this schistosomal-induced pathology.

Adolescent↗
PubMed1990

Antischistosomal action of mevinolin: evidence that 3-hydroxy-methylglutaryl-coenzyme a reductase activity in Schistosoma mansoni is vital for parasite survival.

The enzyme 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA reductase) plays an important role in regulating egg production in the human parasitic trematode Schistosoma mansoni. Infected mice treated with a low dose of mevinolin, an inhibitor of this enzyme, stopped egg production by the parasite and blocked the pathology associated with the infection. As an extension of this work, we exposed adult and developing schistosomes to higher doses of mevinolin for an extended period of time and observed the following: administration of 0.2% mevinolin in the diet of infected mice for 14 days resulted in 96-100% elimination of adult parasites. Administration of the same dose for 2 days before, and for 15 days after infection, resulted in 93-96% reduction of adult parasites. Co-administration of mevinolin with 0.5% mevalonate, but not 0.5% cholesterol, blocked the antischistosomal activity of mevinolin. We monitored schistosomal and mouse liver HMG-CoA reductase activity during the course of treatment and observed that HMG-CoA reductase activity was elevated in the liver but significantly reduced in the parasite. In vitro incubation of paired schistosomes with mevinolin, for up to 11 days, resulted in a dose- and time-dependent reduction of parasite motility and lactate production with eventual death. These in vitro effects were no evident when mevalonate was added to the culture system in place of, or in addition to, mevinolin. Collectively, the evidence suggests that mevalonate and/or metabolite not only plays a vital role in schistosome egg production, but is vital for survival of the parasite.

Animals↗
PubMed1990

Schistosoma mansoni: biochemical characteristics of the antischistosomal effects of Ro 15-5458.

We evaluated a variety of biochemical parameters in Schistosoma mansoni isolated from mice up to 4 days after dosing with 15 mg/kg Ro 15-5458. While no drug effect could be demonstrated in the utilization of media glucose, glycogen content, gut pigment, or ATP levels of the parasites, a significant reduction (P less than or equal to 0.05) in parasite weight and protein content was observed. Possible drug actions that may contribute to the loss in parasite protein and perhaps ultimately result in parasite death have been investigated. We noted significant reduction in the incorporation of leucine and thymidine into acid-insoluble fractions of the parasites. The reduction in the incorporation of leucine into parasite proteins was nonspecific and preceded the effect of the drug on the uptake of the amino acid. Parasite and host liver RNA isolated after dosing were translated in vitro in a rabbit reticulocyte system. Drug-treated parasite mRNA, but not that of the host, was less effective than control mRNA in directing the incorporation of [35S]methionine. We propose a hypothesis that attributes the loss in protein content to a defect in the biosynthesis of parasite proteins as a result of a drug-induced reduction in the quantity of mRNA in the parasites This effect of Ro 15-5458 on the parasite may provide the basis for its schistosomicidal action.

Acridines↗
PubMed1990

Studies on intrategumental pH and its regulation in adult male Schistosoma mansoni.

The intrategumental pH in adult male Schistosoma mansoni as measured with pH-sensitive microelectrodes is between 7.0 and 7.2, a value about one pH unit more alkaline than expected for equilibrium. This value is maintained for at least 4 h after isolation in media buffered with Hepes or CO2/HCO3- with or without serum present. CCCP (1 microM) and FCCP (10 microM) cause rapid acidification. DNP (1 mM) and Na-orthovanadate (1 mM) acidify but also cause significant depolarization of the tegument. NH4+ (20 mM) depolarizes and acidifies the tegument with no evidence of transient alkalinization. High K+ (25 mM) accelerates the acidification. Removal of the NH4+ causes a large transient further acidification with recovery requiring as long as 40 min. High K+ (25 mM) delays the early stage of the recovery. Presence of CO2/HCO3- in the incubation medium does not accelerate the recovery rate nor does SITS (100 microM) inhibit the recovery. Intrategumental Na+ is elevated after an acid load and amiloride (3 mM) as well as low Na+ medium interfere with recovery from the acid load indicating that a Na(+)-H+ exchanger may be present in the tegumental membrane.

Animals↗