Biomedical subjects

J Smit

Publications and source records attributed to J Smit.

At least 37 records · Page 2Linked to original sources

PubMed2000

Characterization of high density monolayers of the biofilm bacterium Caulobacter crescentus: evaluating prospects for developing immobilized cell bioreactors.

Caulobacters are biofilm-forming members of the natural flora of soil and aquatic environments, which exhibit several characteristics that make them attractive for development of high surface area microbial bioreactors or biosensors. Although caulobacters are well characterized genetically, little is known about their biofilm-forming characteristics as a monoculture, or their tolerance of bioreactor-like conditions. Here we investigated the ability of caulobacters to spontaneously form high-density monolayers on artificial surfaces under a variety of environmental conditions, using phase contrast image analysis to assess biofilm density, and epifluorescence with the vital stain DiBAC to assess viability. With adequate nutrition, extremely dense monolayers formed within 24-48 h, and maintained near 100% viability in experiments ranging up to 22 days. When areas were abraded to remove cells, repopulation occurred rapidly with characteristics similar to the population of a clean surface. When established monolayers were starved for nutrients, a significant fraction of the cells detached from the surface, and cells remaining on the surface no longer tested as viable. Within 4-6 h of nutrient restoration, however, cells in the monolayer again appeared normal and tested as 100% viable. This is the first demonstration that Caulobacter crescentus is stable and amenable to high density monolayer growth and resists starvation, though some cells may express a programmed response to detach from the surface under severe nutrient limitation.

Barbiturates↗

PubMed2000

Factors associated with an increased case-fatality rate in HIV-infected and non-infected South African gold miners with pulmonary tuberculosis.

SETTING: A gold mining company in the Free State Province, South Africa. AIM AND DESIGN: A retrospective cohort study to investigate factors associated with an increased case-fatality rate (CFR) at 6 months in human immunodeficiency virus (HIV) positive and negative tuberculosis (TB) patients. RESULTS: Between April 1993 and March 1997, there were 2236 men with culture-confirmed pulmonary TB in whom HIV status and treatment outcome were known. The overall CFR within the first 6 months of therapy was low (3.6%). After adjusting for confounding factors, HIV infection (OR 15.0, 95%CI 7.4-30.6), self-presentation compared to detection by the active radiological screening programme (OR 5.6, 95%CI 2.6-12.2) and presence of silicosis (OR 3.0, 95%CI 1.4-6.3) were significantly associated with an increased CFR. Opportunistic infections accounted for 56.2% (36/64) of deaths in HIV-positive men. Cryptococcal disease accounted for 75% (27/36) of deaths from opportunistic infections. CONCLUSION: HIV infection and silicosis are both powerful risk factors for TB and are associated with an increased risk of death. Strategies aimed at reducing these two risk factors within the workforce could reduce TB incidence and mortality. In settings with functional DOTS programmes and sufficient resources, expanding the DOTS programme to include active case detection should be explored as a means of reducing TB prevalence and mortality.

AIDS-Related Opportunistic Infections↗

PubMed1999

Translocations (X;10)(p22;q24) and (1;10)(q21;q11) in a follicular adenoma of the thyroid without apparent involvement of the RET protooncogene.

We report here the cytogenetic analysis of a follicular adenoma of the thyroid which revealed an abnormal clone with a t(X;10)(p22;q24) and a t(1;10)(q21;q11) together with normal cells. Fluorescence in situ hybridization (FISH) with YACs 273E3 and 344H4, which are located on 10q11.2 and are specific for the RET protooncogene, showed no abnormalities. It would therefore appear that this gene is not involved in the particular tumor, as has been reported in a number of papillary thyroid carcinomas. Several chromosomal aberrations have been suggested as been specific for follicular thyroid adenoma. However, until now, only a few such cases have been reported which involve structural abnormalities of chromosomes 10q11.2 and 10q24. We believe this to be the first report of a follicular thyroid adenoma with a t(X;10)and a t(1;10).

Adenoma↗

PubMed1999

Phylogeny and polyphasic taxonomy of Caulobacter species. Proposal of Maricaulis gen. nov. with Maricaulis maris (Poindexter) comb. nov. as the type species, and emended description of the genera Brevundimonas and Caulobacter.

The genus Caulobacter is composed of prosthecate bacteria often specialized for oligotrophic environments. The taxonomy of Caulobacter has relied primarily upon morphological criteria: a strain that visually appeared to be a member of the Caulobacter has generally been called one without challenge. A polyphasic approach, comprising 16S rDNA sequencing, profiling restriction fragments of 16S-23S rDNA interspacer regions, lipid analysis, immunological profiling and salt tolerance characterizations, was used to clarify the taxonomy of 76 strains of the genera Caulobacter. Brevundimonas, Hyphomonas and Mycoplana. The described species of the genus Caulobacter formed a paraphyletic group with Caulobacter henricii, Caulobacter fusiformis, Caulobacter vibrioides and Mycoplana segnis (Caulobacter segnis comb. nov.) belonging to Caulobacter sensu stricto. Caulobacter bacteroides (Brevundimonas bacteroides comb. nov.), C. henricii subsp. aurantiacus (Brevundimonas aurantiaca comb. nov.), Caulobacter intermedius (Brevundimonas intermedia comb. nov.), Caulobacter subvibrioides (Brevundimonas subvibrioides comb. nov.), C. subvibrioides subsp. albus (Brevundimonas alba comb. nov.), Caulobacter variabilis (Brevundimonas variabilis comb. nov.) and Mycoplana bullata belong to the genus Brevundimonas. The halophilic species Caulobacter maris and Caulobacter halobacteroides are different from these two genera and form the genus Maricaulis gen. nov. with Maricaulis maris as the type species. Caulobacter leidyia was observed to cluster with species of the genus Sphingomonas. Caulobacter crescentus is synonymous with C. vibrioides and C. halobacteroides is synonymous with Maricaulis maris as determined by these analyses and DNA-DNA hybridization. Biomarkers discerning these different genera were determined. The necessary recombinations have been proposed and a description of Maricaulis is presented.

Antigens, Bacterial↗

PubMed1999

Inhibition of various steps in the replication cycle of vesicular stomatitis virus contributes to its photoinactivation by AlPcS4 or Pc4 and red light.

Vesicular stomatitis virus (VSV) was used as a model virus to study the processes involved in photoinactivation by aluminum phthalocyanine tetrasulfonate (AlPcS4) or silicon phthalocyanine HOSiPcOSi(CH3)2(CH2)3N(CH3)2 (Pc4) and red light. Previously a very rapid decrease in the intracellular viral RNA synthesis after photodynamic treatment was observed. This decrease was correlated to different steps in the replication cycle. Binding of VSV to host cells and internalization were only slightly impaired and could be visualized by electron microscopy. The capability of the virus to fuse with membranes in an acidic endosomal environment was studied using both pyrene-labeled liposomes and a hemolysis assay as a model. These tests indicate a rapid decrease of fusion capacity after AlPcS4 treatment, which correlated with the decrease in RNA synthesis. For Pc4 treatment no such correlation was found. The fusion process is the first step in the replication cycle, affected by AlPcS4 treatment, but also in vitro RNA polymerase activity was previously shown to be inhibited. Inactivation of VSV by Pc4 treatment is apparently caused by damage to a variety of viral components. Photodynamic treatment of virus suspensions with both sensitizers causes formation of 8-oxo-7,8-dihydroguanosine in viral RNA as measured by HPLC with electrochemical detection. This damage might be partly responsible for inhibition of the in vitro viral RNA polymerase activity by photodynamic treatment.

Animals↗

PubMed1999

Current perspectives on the perioperative management of the latex-allergic patient.

The increasing incidence of latex allergy necessitates thorough preanesthetic screening for risk factors, which will be delineated in this article, that are associated with latex allergy. The pathophysiology, epidemiology, and testing procedures for latex allergy will be reviewed. This case report will illustrate the management of a patient who was found to be latex-sensitive during surgery and the management of intraoperative anaphylaxis is provided. Safe perioperative care can be provided for latex-sensitive patients if latex avoidance techniques are used consistently.

Adult↗

PubMed1998

The Caulobacter crescentus paracrystalline S-layer protein is secreted by an ABC transporter (type I) secretion apparatus.

Caulobacter crescentus is a gram-negative bacterium that produces a two-dimensional crystalline array on its surface composed of a single 98-kDa protein, RsaA. Secretion of RsaA to the cell surface relies on an uncleaved C-terminal secretion signal. In this report, we identify two genes encoding components of the RsaA secretion apparatus. These components are part of a type I secretion system involving an ABC transporter protein. These genes, lying immediately 3' of rsaA, were found by screening a Tn5 transposon library for the loss of RsaA transport and characterizing the transposon-interrupted genes. The two proteins presumably encoded by these genes were found to have significant sequence similarity to ABC transporter and membrane fusion proteins of other type I secretion systems. The greatest sequence similarity was found to the alkaline protease (AprA) transport system of Pseudomonas aeruginosa and the metalloprotease (PrtB) transport system of Erwinia chrysanthemi. The prtB and aprA genes were introduced into C. crescentus, and their products were secreted by the RsaA transport system. Further, defects in the S-layer protein transport system led to the loss of this heterologous secretion. This is the first report of an S-layer protein secreted by a type I secretion apparatus. Unlike other type I secretion systems, the RsaA transport system secretes large amounts of its substrate protein (it is estimated that RsaA accounts for 10 to 12% of the total cell protein). Such levels are expected for bacterial S-layer proteins but are higher than for any other known type I secretion system.

ATP-Binding Cassette Transporters↗

PubMed1997

Prejunctional histamine H3-receptors inhibit electrically evoked endogenous noradrenaline overflow in the portal vein of freely moving rats.

The effects of intra-arterial injection of different doses of the selective histamine H3-receptor agonist R-alpha-methylhistamine and the selective histamine H3-receptor antagonist thioperamide on basal and electrically evoked noradrenaline overflow in the portal vein as well as on mean arterial pressure (MAP) and heart rate (HR) were investigated in permanently instrumented freely moving rats. R-alpha-Methylhistamine (0.01, 0.1 and 1 mumol/kg) inhibited the evoked noradrenaline overflow up to 43%, the ED50 value being 0.013 mumol/kg. Thioperamide (0.1, 0.5 and 1.0 mumol/kg) antagonized the effect of 1.0 mumol/kg R-alpha-methylhistamine dose-dependently, evoked overflow returning to control values at 1.0 mumol/kg of the antagonist; thioperamide alone had no effect on electrically evoked noradrenaline overflow. Basal noradrenaline levels, blood pressure and heart rate were not at all influenced by R-alpha-methylhistamine and thioperamide, alone or in combination. The results clearly show the presence of prejunctional histamine H3-receptors inhibiting the electrically evoked noradrenaline overflow from vascular sympathetic nerve terminals in the portal vein of freely moving rats.

Animals↗

PubMed1997

An unusual cause of haemobilia: biliary ascariasis.

A case of haemobilia following biliary ascariasis is presented. Angiography confirmed an aneurysm of the left hepatic artery, which was successfully embolised. The patient made an uneventful recovery.

Aneurysm, False↗

PubMed1997

Low education is a genuine risk factor for accelerated memory decline and dementia.

A relatively high prevalence and incidence of dementia have been found in population strata with low levels of education in comparison to population strata with high levels of education. However, doubt remains whether this may be an artifact of education bias in the screening tests used. To investigate this matter, we analyzed results of two Dutch population surveys in which unbiased measures of memory decline were used. In the Longitudinal Aging Study Amsterdam (n = 1774) the percentage of words retained in a verbal learning test was found to be disproportionately low in the oldest age cohort (80-85 years) with less than 11 years of education. The Amsterdam Study of the Elderly (n = 4051) found a "dose-response" relationship between education and dementia prevalence. Cross-sectional and longitudinal results showed that, in less educated people, memory decline is faster and sets in at an earlier age. These findings indicate that the relationship between dementia and education is not just an artifact of case detection methods.

Aged↗

PubMed1997

Cell-surface display of a Pseudomonas aeruginosa strain K pilin peptide within the paracrystalline S-layer of Caulobacter crescentus.

The paracrystalline surface (S)-layer of Caulobacter crescentus is composed of a single secreted protein (RsaA) that interlocks in a hexagonal pattern to completely envelop the bacterium. Using a genetic approach, we inserted a 12 amino acid peptide from Pseudomonas aeruginosa strain K pilin at numerous semirandom positions in RsaA. We then used an immunological screen to identify those sites that presented the inserted pilin peptide on the C. crescentus cell surface as a part of the S-layer. Eleven such sites (widely separated in the primary sequence) were identified, demonstrating for the first time that S-layers can be readily exploited as carrier proteins to display 'epitope-size' heterologous peptides on bacterial cell surfaces. Whereas intact RsaA molecules carrying a pilin peptide could always be found on the surface of C. crescentus regardless of the particular insertion site, introduction of the pilin peptide at 9 of the 11 sites resulted in some proteolytic cleavage of RsaA. Two types of proteolytic phenomena were observed. The first was characterized by a single cleavage within the pilin peptide insert with both fragments of the S-layer protein remaining anchored to the outer membrane. The other proteolytic phenomenon was characterized by cleavage of the S-layer protein at a point distant from the site of the pilin peptide insertion. This cleavage always occurred at the same location in RsaA regardless of the particular insertion site, yielding a surface-anchored 26 kDa proteolytic fragment bearing the RsaA N-terminus; the C-terminal cleavage product carrying the pilin peptide was released into the growth medium. When the results of this work were combined with the results of a previous study, the RsaA primary sequence could be divided into three regions with respect to the location of a peptide insertion and its effect on S-layer biogenesis: (i) insertions in the extreme N-terminus of RsaA either produce no apparent effect on S-layer biogenesis or disrupt surface-anchoring of the protein; (ii) insertions in the extreme C-terminus either produce no apparent effect on S-layer biogenesis or disrupt protein secretion; and (iii) insertions more centrally located in the protein either have no apparent effect on S-layer biogenesis or result in proteolytic cleavage of RsaA. These data are discussed in relation to our previous assignment of the RsaA N- and C-terminus as regions that are important for surface anchoring and secretion respectively.

Bacterial Outer Membrane Proteins↗

PubMed1997

Functions of S-layers.

Although S-layers are being increasingly identified on Bacteria and Archaea, it is enigmatic that in most cases S-layer function continues to elude us. In a few instances, S-layers have been shown to be virulence factors on pathogens (e.g. Campylobacter fetus ssp. fetus and Aeromonas salmonicida), protective against Bdellovibrio, a depository for surface-exposed enzymes (e.g. Bacillus stearothermophilus), shape-determining agents (e.g. Thermoproteus tenax) and nucleation factors for fine-grain mineral development (e.g. Synechococcus GL 24). Yet, for the vast majority of S-layered bacteria, the natural function of these crystalline arrays continues to be evasive. The following review up-dates the functional basis of S-layers and describes such diverse topics as the effect of S-layers on the Gram stain, bacteriophage adsorption in lactobacilli, phagocytosis by human polymorphonuclear leukocytes, the adhesion of a high-molecular-mass amylase, outer membrane porosity, and the secretion of extracellular enzymes of Thermoanaerobacterium. In addition, the functional aspect of calcium on the Caulobacter S-layer is explained.

Abortion, Veterinary↗

PubMed1997

Factors controlling in vitro recrystallization of the Caulobacter crescentus paracrystalline S-layer.

The S-layer of Caulobacter is a two-dimensional paracrystalline array on the cell surface composed of a single protein, RsaA. We have established conditions for preparation of stable, soluble protein and then efficient in vitro recrystallization of the purified protein. Efficient recrystallization and long range order could not be obtained with pure protein only, though it was apparent that calcium was required for crystallization. Recrystallization was obtained when lipid vesicles were provided, but only when the vesicles contained the specific species of Caulobacter smooth lipopolysaccharide (SLPS) that previous studies implicated as a requirement for attaching the S-layer to the cell surface. The specific type of phospholipids did not appear critical; phospholipids rather different from those present in Caulobacter membranes or archaebacterial tetraether lipids worked equally well. The source of LPS was critical; rough and smooth variants of Salmonella typhimurium LPS as well as the rough form of Caulobacter LPS were ineffective. The requirement for calcium ions for recrystallization was further evaluated; strontium ions could substitute for calcium, and to a lesser extent, cobalt, barium, manganese and magnesium ions also stimulated crystallization. On the other hand, nickel and cadmium provided only weak crystallization stimulation, and zinc, copper, iron, aluminum ions, and the monovalent potassium, sodium, and lithium ions were ineffective. The recrystallization could also be reproduced with Langmuir-Blodgett lipid monolayers at an air-water interface. As with the vesicle experiments, this was only successful when SLPS was incorporated into the lipid mix. The best method for RsaA preparation, leading to apparently monomeric protein that was stable for many months, was an extraction with a low pH aqueous solution. We also achieved recrystallization, albeit at lower efficiency, using RsaA protein solubilized by 8 M urea, a method which allows retrieval of protein from inclusions, when expressed as heterologous protein in Escherichia coli or when retrieved as shed, precipitated protein from certain mutant caulobacters. In summary, the clarification of recrystallization methods has confirmed the requirement of SLPS as a surface attachment component and suggests that its presence in a membrane-like structure greatly stimulates the extent and quality of S-layer formation. The in vitro approach allowed the demonstration that specific ions are capable of participating in crystallization and now provides an assay for the crystallization potential of modified S-layer proteins, whether they were produced in or can be secreted by caulobacters.

Bacterial Outer Membrane Proteins↗

PubMed1997

Linker mutagenesis of the Caulobacter crescentus S-layer protein: toward a definition of an N-terminal anchoring region and a C-terminal secretion signal and the potential for heterologous protein secretion.

Linker insertion mutagenesis was used to modify the paracrystalline surface layer (S-layer) protein (RsaA) of the gram-negative bacterium Caulobacter crescentus. Eleven unique BamHI linker insertions in the cloned rsaA gene were identified; at the protein level, these linker insertions introduced 4 to 6 amino acids at positions ranging from the extreme N terminus to the extreme C terminus of the 1,026-amino-acid RsaA protein. All linker-peptide insertions in the RsaA N terminus caused the secreted protein to be shed into the growth medium, suggesting that the RsaA N terminus is involved in cell surface anchoring. One linker-peptide insertion in the RsaA C terminus (amino acid 784) had no effect on S-layer biogenesis, while another (amino acid 907) disrupted secretion of the protein, suggesting that RsaA possesses a secretion signal lying C terminal to amino acid 784, near or including amino acid 907. Unlike extreme N- or C-terminal linker-peptide insertions, those more centrally located in the RsaA primary sequence had no apparent effect on S-layer biogenesis. By using a newly introduced linker-encoded restriction site, a 3' fragment of the rsaA gene encoding the last 242 C-terminal amino acids of the S-layer protein was expressed in C. crescentus from heterologous Escherichia coli lacZ transcription and translation initiation information. This C-terminal portion of RsaA was secreted into the growth medium, confirming the presence of a C-terminal secretion signal. The use of the RsaA C terminus for the secretion of heterologous proteins in C. crescentus was explored by fusing 109 amino acids of an envelope glycoprotein from infectious hematopoietic necrosis virus, a pathogen of salmonid fish, to the last 242 amino acids of the RsaA C terminus. The resulting hybrid protein was successfully secreted into the growth medium and accounted for 10% of total protein in a stationary-phase culture. Based on these results and features of the RsaA primary sequence, we propose that the C. crescentus S-layer protein is secreted by a type I secretion system, relying on a stable C-terminal secretion signal in a manner analogous to E. coli alpha-hemolysin, the first example of an S-layer protein secreted by such a pathway.

Bacterial Outer Membrane Proteins↗

PubMed1997

Pilin-based anti-Pseudomonas vaccines: latest developments and perspectives.

Among the several adhesins produced by Pseudomonas aeruginosa (Pa), the type-4 pilus promotes the majority of the adherence capability of the bacterium to epithelial cells and it is a major virulence factor in an AB.Y/SnJ mouse infection model. Vaccines targeting the disulfide loop (DSL) adherence binding domain of the pilin protein should therefore provide an effective protection against initial colonization and infection with Pa. To selectively elicit adherence blocking antibodies, the pilin DSL domain was chosen as peptide antigen for the construction of recombinant protein and live vaccines. While synthetic peptide-carrier protein conjugates provided some strain-specific protection, chimeric proteins with N- or C-terminally fused pilin DSL peptides did not engender protective IgG titers mice. Integral fusions of the pilin DSL peptide with the minor coat protein of filamentous phage or surface exposed regions of an outer membrane protein resulted in a display of the peptide on the surface of the phage particles and bacterial cells respectively. However, in immunization studies neither of these live vaccines were effective immunogens. The paracrystalline S-layer of Caulobacter crescentus combines several advantages of an effective antigen surface display system. Recombinant S-layer proteins with singlecopy insertions of a pilin peptide did not engender significant IgG titers, whereas multiple tandem insertions of the same peptide increased the serum IgG response in mice a thousand times. Multiple insertions of DSL peptides from different frequent pilin prototypes may be an interesting alternative for a recombinant cross-protective anti-Pseudomonas vaccine.

Amino Acid Sequence↗

PubMed1996

Threshold effect of liver iron content on hepatic inflammation and fibrosis in hepatitis B and C.

BACKGROUND/AIMS: In hepatitis C, iron depletion may improve serum aminotransferases and the response to interferon, but it is not known whether inflammation and fibrosis correlate with hepatic iron content. Our aim was to establish whether hepatic iron content correlates with histological and serum indices of hepatic inflammation and fibrosis in hepatitis B and C. METHODS: Total hepatic iron was measured using computerized histomorphometry, and hepatic inflammation and fibrosis using a modified Knodell score, on histological slides from 31 patients with chronic hepatitis B and 38 with hepatitis C. RESULTS: Total hepatic iron was similar in the hepatitis B and C groups (0.82 +/- 1.72% and 0.56 +/- 1.12%; mean +/- SD). No iron was detectable in 11 patients with hepatitis B and 13 with hepatitis C. Alanine aminotransferase (85.96 +/- 67.1 vs 44.2 +/- 39.7 p < 0.05), aspartate aminotransferase (93.8 +/- 75.6 vs 47 +/- 33.5 IU/ml p < 0.05) and histological inflammatory score (9.33 +/- 3.51 vs 7.79 +/- 3.3 p = 0.07) were increased in those with stainable hepatic iron compared to those without. However, where iron was present, no association was found between the amount of hepatic iron and inflammatory or fibrosis scores. In hepatitis C, fibrosis was minimal in 77% of patients if iron was absent vs 24% with iron present, while marked fibrosis was present in 56% with iron vs 15% without iron (p < 0.01, Fisher's exact test). CONCLUSION: Hepatic iron is associated with increased hepatic inflammation in chronic hepatitis B and hepatitis C and with high fibrosis scores in hepatitis C. There is a threshold effect, and once present, increasing iron does not correlate with increasing inflammation or fibrosis.

Adolescent↗

PubMed1996

Influence of adrenodemedullation on beta 2- and beta 3-adrenoceptors mediating relaxation of oesophageal smooth muscle of spontaneously hypertensive rats.

1. In oesophageal smooth muscle strips from spontaneously hypertensive rats (SHR) of 8-10 and 22-24 weeks of age, respectively, beta-adrenoceptor-mediated relaxation was investigated, by use of the beta-agonists, (-)-isoprenaline and fenoterol (both in the absence and presence of the beta 2-selective antagonist ICI 118,551) and the selective beta 3-agonist, BRL 37,344. 2. In preparations from 8-10 week SHR, (-)-isoprenaline- and fenoterol-induced concentration-response curves (CRCs) were hardly antagonized by ICI 118,551 at concentrations up to 1 microM, indicating only a minor contribution of beta 2-adrenoceptors. pA2-values for ICI 118,551 of 5.30 ((-)-isoprenaline as agonist) and 5.46 (fenoterol as agonist), estimated from the shifts at the highest (10-100 microM) antagonist concentrations, are consistent with affinity at a beta 3-adrenoceptor, similar to that in Wistar rat oesophageal smooth muscle. 3. In 8-10 week SHR, adrenodemedullated at 4 weeks of age (SHR-ADM4) the potency of fenoterol was markedly increased and CRCs were shallow. In addition, ICI 118,551 (0.1 microM) now produced a clear rightward shift accompanied by a steepening of the CRC. A marked further shift was observed only at 100 microM of the antagonist. The data are compatible with the involvement of both beta 2- and beta 3-adrenoceptors. 4. In 22-24 week animals, the same differences between SHR and SHR-ADM4 were observed with fenoterol as in 8-10 week animals, though beta-adrenoceptor responsiveness was slightly decreased. The potency of ICI 118,551 at beta 3-adrenoceptors (pA2 = 5.11) was significantly different from the pA2 value of 5.46 obtained with the younger animals. 5. Responses to the beta 3-adrenoceptor agonist, BRL 37,344, were similar in Wistar rat and SHR preparations. In 8-10 week SHR, a small decrease in the maximal response was observed, which in animals of 22-24 weeks of age was accompanied by a small decrease in the pEC50 value as well. 6. The results clearly indicate that beta 2-adrenoceptors in SHR oesophageal muscularis mucosae are desensitized, whereas beta 3-adrenoceptor-mediated responses are unaffected and similar to the responses observed in the Wistar rat oesophagus. The functional presence of beta 2-adrenoceptor-responses in SHR-ADM4 suggests a major role for adrenal-derived adrenaline in the desensitization of the beta 2-adrenoceptor-population.

Adrenal Medulla↗

PubMed1996

Characterization of In Vivo Reporter Systems for Gene Expression and Biosensor Applications Based on luxAB Luciferase Genes.

Advances in genetic engineering methods have allowed the development of an increasing number of practical and scientific applications for bioluminescence with lux genes cloned from a variety of organisms. Bioluminescence derived from the shortened lux operon (luxAB genes) is a complex process, and applications seem to be proliferating in advance of an understanding of the underlying biochemical processes. In this report, we describe a two-phase kinetic behavior of the light emission which must be properly taken into account in any quantitative measurements of the bioluminescence signal. By using strains of Escherichia coli and Caulobacter crescentus, this behavior was characterized and interpreted in terms of the biochemistry underlying the bacterial luciferase mechanism. We show that the intensity profile of each of the two phases of the luminescence signal is responsive (and exhibits different sensitivities) to the concentration of added decanal and other components of the assay mix, as well as to the order of mixing and incubation times. This study illustrates the importance of appropriate protocol design, and specific recommendations for using the luxAB system as a molecular reporter are presented, along with versatile assay protocols that yield meaningful and reproducible signals.

Journal Article↗