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Biomedical subjects

J R Baker

Publications and source records attributed to J R Baker.

At least 37 records · Page 2Linked to original sources
PubMed2001

Exposure to heavy metals and infectious disease mortality in harbour porpoises from England and Wales.

We investigate whether long-term exposure to heavy metals, including immunosuppressive metals like mercury (Hg), is associated with infectious disease in a wild cetacean. Post-mortem investigations on 86 harbour porpoises, Phocoena phocoena, found dead along the coasts of England and Wales revealed that 49 of the porpoises were healthy when they died as a consequence of physical trauma (most frequently entrapment in fishing gear). In contrast, 37 porpoises died of infectious diseases caused by parasitic, bacterial, fungal and viral pathogens (most frequently pneumonia caused by lungworm and bacterial infections). We found that mean liver concentrations of Hg, selenium (Se), the Hg:Se molar ratio, and zinc (Zn) were significantly higher in the propoises that died of infectious disease compared to healthy porpoises that died from physical trauma. Liver concentrations of lead (Pb), cadmium (Cd), copper (Cu) and chromium (Cr) did not differ between the two groups. Hg, Se, and the Hg:Se molar ratio were also positively correlated with age. The association between Zn concentration and disease status may result from Zn redistribution in response to infection. Further work is required to evaluate whether chronic exposure to Hg may have presented a toxic challenge to the porpoises that succumbed to infectious disease.

Animals↗
PubMed2001

Identification of carmine allergens among three carmine allergy patients.

BACKGROUND: There have been several reports of carmine allergy; however, identification of the responsible carmine allergens has not been widely documented. METHODS: Three female patients presented with a history of anaphylaxis and/or urticaria/angioedema after ingestion of carmine-containing foods. All three patients had 4+ skin prick tests to carmine. Among them, two patients were confirmed to have carmine allergy by blinded, placebo-controlled food challenges to carmine. SDS-PAGE of cochineal insects and carmine, immunoblotting for IgE antibody with sera from all three patients, and immunoblotting inhibition with carmine were performed. RESULTS: SDS PAGE of minced cochineal insects revealed several protein bands of 23-88 kDa. Several of these bands were variably recognized by our three patients' sera, and this reactivity was inhibited by carmine. Although no protein bands could be visualized on SDS-PAGE of carmine in Coomassie brilliant blue staining, three protein bands were recognized by two of the three patients' serum. CONCLUSIONS: These results suggest that commercial carmine retains protein-aceous material from the source insects. These insect-derived proteins (possibly complexed with carminic acid) are responsible for IgE-mediated carmine allergy. Patient reactivity to these proteins may vary.

Administration, Oral↗
PubMed2001

Combination of electroporation and DNA/dendrimer complexes enhances gene transfer into murine cardiac transplants.

Electroporation is a new gene delivery method to increase gene transfer and expression in vivo. Starburst polyamidoamine dendrimers have been demonstrated to augment gene expression in vitro and in vivo. We hypothesized that the combination of electroporation and dendrimer could enhance the gene transfer and gene expression in cardiac transplants. After immersion in DNA/dendrimer complexes or intracoronary transfer of DNA/dendrimer complexes, both nonvascularized and vascularized syngeneic cardiac grafts, respectively, were subjected to serial electrical pulses before transplantation. beta-Galactosidase reporter gene expression in the graft was determined by X-Gal staining. Gene expression was enhanced 10- to 45-fold in grafts immersed in DNA/dendrimer complexes, or after intracoronary transfer of DNA/dendrimer complexes, and subjected to 20 square wave 25-ms pulses with a strength of 200 V/cm. The combination of electroporation and DNA/dendrimer complexes may provide a novel approach to enhance gene transfer and gene expression ex vivo.

Animals↗
PubMed2001

A novel surfactant nanoemulsion with a unique non-irritant topical antimicrobial activity against bacteria, enveloped viruses and fungi.

A novel non-ionic surfactant nanoemulsion designated 8N8 has been tested for its biocidal activity. One percent 8N8 produced effective bactericidal activity against Bacillus cereus, Bacillus subtilis, Haemophilus influenzae, Neisseria gonorrhoeae, Streptococcus pneumoniae, and Vibrio cholerae in 15 minutes. In contrast, most enteric gram-negative bacteria were resistant to 8N8. One percent 8N8 was also virucidal within 15 minutes for all tested enveloped viruses, including Herpes simplex type 1, influenza A and vaccinia viruses. One percent 8N8 also demonstrated fungistatic activity on Candida albicans. The rapid and non-specific inactivation of vegetative bacteria and enveloped viruses, in addition to its fungistatic activity and low toxicity in experimental animals, makes 8N8 a potential candidate for use as a topical biocidal agent.

Administration, Topical↗
PubMed2001

Susceptibility of thyroid cancer cells to 7-hydroxystaurosporine-induced apoptosis correlates with Bcl-2 protein level.

7-Hydroxystaurosporine (UCN-01) is a selective protein kinase C (PKC) inhibitor and is being developed as a novel anticancer agent. Because of reports that PKC may be involved in the pathogenesis of some forms of thyroid cancers, we examined four thyroid carcinoma lines (FRO, KAT5, NPA, and WRO). These cells were found to have different susceptibility to UCN-01 treatment, and there appeared to be a correlation between UCN-01-induced death and expression levels of endogenous Bcl-2. KAT5 cells, which normally express a low amount of Bcl-2, exhibited significantly higher sensitivity to UCN-01-induced death than the other cell lines. Of interest, susceptibility did not relate to PKC activity or its inhibition by UCN-01. In order to investigate the role of Bcl-2 in UCN-01-induced death, KAT5 cells were transfected to overexpress Bcl-2. KAT5/Bcl-2 cells were capable of conferring resistance to UCN-01-induced death. Furthermore, upregulating of Bcl-2 by 1alpha,25-dihydroxyvitamin D3 (VD3) could protect primary thyroid cell from death induced by UCN-01. Both in situ TUNEL staining and the flow cytometric analysis of cytokeratin-18 (CK18) cleavage confirmed that UCN-01 was indeed inducing apoptosis, and that this effect was inhibited by increased expression of Bcl-2. These results suggest that the Bcl-2 can block the UCN-01-activated cell death pathway and that the expression of Bcl-2 is inversely related to thyroid carcinoma cell susceptibility to UCN-01. Therefore, the analysis of the expression of apoptosis suppressors provides a basis for the use of UCN-01 in the treatment of thyroid cancer.

Alkaloids↗
PubMed2001

Characteristics of responding-, nonresponding- and refusing-parents in an adolescent lifestyle choice study.

The acquisition of a random sample is one of the many methodological problems that arise when conducting research with adolescent populations. Frequently, due to ethical considerations associated with collecting data from adolescents, active parental consent procedures are required. The current study examined characteristics of parents who consented, refused consent, or did not respond to an active consent request for their children to participate in a large-scale study of adolescent lifestyle behaviors. Results indicated nonresponding-parents were more likely to be employed than consenting-parents. Further, differences were found for a number of attitudinal variables and about the importance of adolescent research. There were significant differences between refusing-parents, and consenting- and nonresponding-parents who were similar in their attitudes toward adolescent research. The findings suggest that nonresponding-parents are characteristically more similar to consenting-parents than to refusing-parents, which supports the use of passive consent procedures as a reasonable alternative to requiring active parental consent in adolescent research.

Adolescent↗
PubMed2001

Changes in zinc and copper homeostasis in human livers and kidneys associated with exposure to environmental cadmium.

This study was undertaken to assess changes in zinc and copper homeostasis in human tissues that could be attributed to human exposure to environmental cadmium, using samples of lung, liver and kidney cortex of 61 Queensland residents, aged 2 to 89 years, who had died of accidental causes. None of the subjects were exposed to cadmium in the workplace. Levels of zinc in liver and kidney cortex samples showed inverse associations with donor age whereas zinc in lung only showed inverse association with gender. Lung zinc levels in females were 14% lower than in males. Zinc in liver and kidney cortex samples were found to exist in at least two pools; one was associated with cadmium that bound to metallothionein (MT) and the other was associated with non-MT bound copper. In liver, the amounts of zinc in the MT pool were smaller compared to those in non-MT pool given that only 7% of zinc variations were explained by cadmium whereas 22% of the liver zinc variations were accounted for by non-MT bound copper. In sharp contrast, larger amounts of zinc in kidney cortex samples were in the MT pool, compared to those in the non-MT pool given that cadmium was found to explain 69% of total zinc variation whereas copper explained only 17% of kidney zinc variations. The levels of copper in liver were found to be increased by 45-50% in subjects with high cadmium exposure level, compared to subjects of similar ages with medium exposure level. The levels of zinc and copper in kidney cortex samples in the subjects with high cadmium exposure were both found to be significantly elevated compared to those found in the medium-exposure group whereas copper contents were about 19-23% greater than in medium- as well as low-exposure groups. Taken together these results indicate increased sequestration of zinc and copper in liver and kidney cortex samples. The increases in metal sequestrations were observed in liver samples having cadmium contents of greater than 1 microg/g wet weight and in kidney cortex having cadmium contents of greater than 26 microg/g wet weight. Zinc and copper contents in lung of this sample group, however, were not associated with cadmium due probably to lower exposure levels compared to those of liver and kidney.

Adolescent↗
PubMed2001

Studies using recombinant fragments of human TSH receptor reveal apparent diversity in the binding specificities of antibodies that block TSH binding to its receptor or stimulate thyroid hormone production.

Patients with Graves' disease have autoantibodies that bind to the TSH receptor and stimulate the thyroid, leading to hyperthyroidism. Earlier studies have shown that the ectodomain of the glycosylated human TSH receptor contains epitopes that could adsorb these pathogenic antibodies. Further studies with mutated cDNAs, chimeric proteins, peptides, and antipeptide antibodies suggested that alterations in the conformation of the protein could lead to loss of reactivity, and that thyroid-stimulating antibodies interact with the N-terminal region of the TSH receptor. Although many of these studies provided valuable insights, they were somewhat inconclusive due to limitations inherent to each of the approaches. In an attempt to further define regions within the TSH receptor with which thyroid-stimulating antibodies interact, we expressed seven recombinant TSH receptor fragments in insect cells and tested them for their ability to neutralize TSH binding inhibitory Igs and thyroid-stimulating antibody activity in the sera of patients with Graves' disease. The fragments containing amino acids 22-305 were able to neutralize the TSH binding inhibitory Ig activity, whereas a fragment containing amino acids 54-254 was able to neutralize the thyroid-stimulating antibodies. Fragments containing additional amino acids, flanking residues 54-254, failed to neutralize the thyroid-stimulating antibody activity, suggesting that thyroid-stimulating antibody epitopes are masked. Our studies show that thyroid autoantibodies, with different functional properties, bind to distinct conformational epitopes on the TSH receptor.

Animals↗
PubMed2000

2-Methoxyestradiol, an endogenous estrogen metabolite, induces thyroid cell apoptosis.

The etiology of autoimmune thyroid diseases is unclear; however, the extreme female predominance suggests that sex hormones may have a pathogenic role. 2-Methoxyestradiol (2-ME) is present in the serum of women during the ovulatory and luteal phases of the menstrual cycle, and during pregnancy. We investigated the actions of 2-ME and estrogen on thyroid follicular cells. 2-ME induced dramatic changes in cell morphology and decreased the viability of the cells, as well as disrupted the structural integrity of cultured thyroid follicles. Flow cytometric analysis showed that 2-ME halted cell proliferation by arresting the cells in the G2/M cell-cycle compartment. Prolonged exposure to 2-ME led to apoptosis and to increased release of the autoantigen thyroid peroxidase (TPO). 17beta-estradiol failed to produce a similar effect even in 40-fold molar excess to 2-ME. Co-treatment with estrogen receptor antagonists did not alter the 2-ME effect, indicating that 2-ME was not operating through a classic nuclear estrogen receptor. In conclusion, this study indicates that 2-ME induces G2/M cycle arrest, apoptosis and the disruption of thyroid follicles. This process results in the release of thyroid antigens that may play a role in high incidence of thyroid autoantibodies and autoimmune thyroid disease in women.

2-Methoxyestradiol↗
PubMed2000

Phosphorylation of human glutamine:fructose-6-phosphate amidotransferase by cAMP-dependent protein kinase at serine 205 blocks the enzyme activity.

Glutamine:fructose-6-phosphate amidotransferase (GFAT) is the rate-limiting enzyme in glucosamine synthesis. Prior studies from our laboratory indicated that activation of adenylate cyclase was associated with depletion of O-GlcNAc modification. This finding and evidence that human GFAT (hGFAT) might be regulated by cAMP-dependent protein kinase (PKA) led us to investigate the role of PKA in hGFAT function. We confirmed that adenylate cyclase activation by forskolin results in diminished O-GlcNAc modification of several cellular proteins which can be overcome by exposure of the cells to glucosamine but not glucose, suggesting the PKA activation results in depletion of UDP-GlcNAc for O-glycosylation. To determine if GFAT is indeed regulated by PKA, we expressed the active form of the enzyme using a vaccinia virus expression system and showed that the activity of the enzyme was to decrease to undetectable levels by PKA phosphorylation. We mapped the PKA phosphorylation sites with the aid of matrix-assisted laser desorption ionization mass spectroscopy and showed that the protein was stoichiometrically phosphorylated at serine 205 and also phosphorylated, to a lesser extent at serine 235. Mutagenesis studies indicated that the phosphorylation of serine 205 by PKA was necessary for the observed inhibition of enzyme activity while serine 235 phosphorylation played no observable role. The activity of GFAT is down-regulated by cAMP, thus placing regulation on the hexosamine pathway that is in concert with the energy requirements of the organism. During starvation, hormones acting through adenylate cyclase could direct the flux of glucose metabolism into energy production rather than into synthetic pathways that require hexosamines.

Colforsin↗
PubMed2000

Intravascular and endobronchial DNA delivery to murine lung tissue using a novel, nonviral vector.

Gene transfer to the lung can be achieved via either the airway or the pulmonary vasculature. We evaluated gene transfer and expression by intravascular and endobronchial routes, using DNA complexed with G9 PAMAM dendrimer or naked plasmid DNA. Intravascular tail vein delivery of dendrimer-complexed pCF1CAT plasmid resulted in high levels of transgene expression in the lung at 12 and 24 hr, followed by a second peak of expression 3 to 5 days after administration. After intravenous administration of the complexes, CAT expression was never observed in organs other than the lung. There were only minimal levels of CAT protein expressed in the lung after intravenous administration of naked plasmid DNA. Repeated intravascular doses of the dendrimer-complexed plasmid, administered four times at 4-day intervals, maintained expression at 15-25% of peak concentrations achieved after the initial dose. Endobronchial delivery of naked pCF1CAT plasmid produced significant amounts of CAT protein in the lung. Comparison of intratracheal and intranasal routes resulted in similar expression levels of CAT in the lung and trachea. However, in juxtaposition to vascular delivery, intranasal delivery of dendrimer-complexed plasmid DNA gave lower levels of CAT expression than that observed with naked plasmid DNA. In situ localization of CAT enzymatic activity suggested that vascular administration seemed to achieve expression in the lung parenchyma, mainly within the alveoli, while endobronchial administration primarily targeted bronchial epithelium. Our results show that intravenously administered G9 dendrimer is an effective vector for pulmonary gene transfer and that transgene expression can be prolonged by repeated administration of dendrimer-complexed DNA.

Administration, Intranasal↗
PubMed2000

Characterization of the active site of group B streptococcal hyaluronan lyase.

Hyaluronan lyase is secreted by most strains of the human pathogen, group B streptococcus. Site-directed mutagenesis of the enzyme identified three amino acid residues important for enzyme activity, H479, Y488, and R542. These three residues are in close proximity in the putative active site of a homology model of group B streptococcal hyaluronan lyase. The homology model was based on the crystal structure of another related glycosaminoglycan lyase, chondroitin AC lyase, which exhibits different substrate specificity. Two asparagine residues in the active site groove, N429 and N660, were also found to be essential for enzyme activity. In addition, conversion of two adjacent tryptophan residues in the groove to alanines abolished activity. All amino acids found to be essential in GBS hyaluronan lyase are conserved in both enzymes. However, several amino acids in the active site groove of the two enzymes are not conserved. In the 18 cases in which one of these amino acids in GBS hyaluronan lyase was replaced with its corresponding amino acid in chondroitin AC lyase, no major loss of activity or change in substrate specificity was observed.

Calcium↗
PubMed2000

Pulmonary pathology of harbour porpoises (Phocoena phocoena) stranded in England and Wales between 1990 and 1996.

The pathological changes observed in the lungs of 197 freshly dead to moderately decomposed harbour porpoises (Phocoenaphocoena) stranded in England and Wales between October 1990 and December 1996 were reviewed. In 135 (69 per cent of the cases) macroscopic nematode infections of the bronchial tract with Pseudalius inflexus and Torynurus convolutus, either singly or in combination, were recorded, and 106 (54 per cent) also had P inflexus within the pulmonary blood vessels. All the macroscopically parasitised porpoises were adults or juveniles although two neonates had histological evidence of nematode infection. There were 62 cases of mild to severe, subacute to chronic bronchitis and bronchiolitis, 113 cases of mild to severe chronic granulomatous interstitial pneumonia, and 34 cases of mild to severe vasculitis or thrombovasculitis of pulmonary blood vessels attributable to these nematode infections. In 35 cases necropurulent or purulent (broncho)pneumonias were attributed either to secondary bacterial infections of the lungs or to septicaemias associated in seven cases with Streptococcus canis, in two cases with group B Salmonella species, in one case with Escherichia coli and in one case with Streptococcus lactis. The pulmonary lesions in 67 animals known or diagnosed to have been entrapped in fishing gear were non-specific and included persistent foam in the airways in 45 cases, diffuse congestion in 53, oedema in 50, and multifocal intra-alveolar haemorrhage in 33 cases. Seven cases of acute fibrinous or chronic fibrous pleuritis, seven cases of chronic necropurulent pneumonia associated with mycotic infections, four porpoises with traumatic lesions of the thorax and other parts of the body consistent with fatal attack from bottlenose dolphins (Tursiops truncatus), and one case of diffuse bronchointerstitial pneumonia associated with generalised morbillivirus infection were also recorded.

Animals↗
PubMed2000

Action pattern and substrate specificity of the hyaluronan lyase from group B streptococci.

The hyaluronan lyase of group B streptococci rapidly cleaves hyaluronan by an elimination mechanism to yield the unsaturated disaccharide 2-acetamido-2-deoxy-3-O-(beta-D-gluco-4-enepyranosyluronic acid)-D-glucose. Additionally, it has been shown that the enzyme has limited specificity for achondroitin sulphate and cleaves the chain at unsulphated sites [Baker,Yu, Morrison, Averett and Pritchard (1997) Biochem. J. 327,65-71]. In the present extension of that study it was found that 6-sulphated regions of chondroitin sulphate are also susceptible to cleavage by this hyaluronan lyase. Of the four 6- and/or 4-sulphated tetrasaccharides which can be isolated from testicular hyaluronidase digests of chondroitin sulphate, only those two tetrasaccharides with a6-sulphated disaccharide at the reducing end were cleaved. From thisand other data, a model is proposed for the cleavage specificity of hyaluronan lyase on a chondroitin sulphate. Evidence is presented in support of an action pattern for hyaluronan lyase which involves aninitial random endolytic cleavage followed by rapid exolytic and processive release of unsaturated disaccharide. Since the on lyoligosaccharides which tend to accumulate in near-complete digests of hyaluronan are unsaturated, it is argued that the processive cleavage occurs from the non-reducing to the reducing end of a hyaluronan chain. This detailed knowledge of substrate specificity contributes to our understanding of the enzyme's role in Group B streptococcal pathogenesis. In addition, the hyaluronan lyase may find application in sequence studies of chondroitin sulphates.

Animals↗
PubMed2000

DNA/dendrimer complexes mediate gene transfer into murine cardiac transplants ex vivo.

Starburst polyamidoamine dendrimers are synthetic polymers with unique structural and physical characteristics suitable for DNA gene transfer. Our previous studies demonstrated that Starburst dendrimers augment plasmid-mediated gene transfer efficiency in a nonvascularized, cardiac transplantation model. In this study, the fifth generation of ethylenediamine core dendrimer was investigated for its ability to enhance gene transfer and expression in a clinically relevant murine vascularized heart transplantation model. The plasmid pMP6A-beta-gal, encoding beta-galactosidase (beta-Gal), was incubated with dendrimers to form complexes. The complexes were perfused via the coronary arteries during donor graft harvesting, and reporter gene expression was determined by quantitative evaluation of X-Gal staining. The grafts infused with pMP6A-beta-gal/dendrimer complexes showed beta-Gal expression in myocytes from 7 to 14 days. A number of variables for transfer of the DNA/dendrimer complexes were tested, including DNA:dendrimer charge ratios, concentrations of DNA and dendrimer, preservation solutions, ischemic time, and enhancement of vascular permeability by serotonin, papaverine, and VEGF administration. The results showed that DNA/dendrimer complexes containing 20 microg of DNA and 260 microg of dendrimer (1:20 charge ratio) in a total volume of 200 microl resulted in highest gene expression in the grafts. The results also showed that prolonged incubation (cold ischemic time) to 2 h and pretreatment with serotonin further enhanced gene expression.

Animals↗
PubMed2000

Evidence for a synergistic interaction between cadmium and endotoxin toxicity and for nitric oxide and cadmium displacement of metals in the kidney.

This study was undertaken to examine changes in Zn and Cu homeostasis in the liver and kidney of rats caused by cadmium (Cd) or lipopolysaccharide (LPS) administration. Twenty-five male, 7- to 8-week-old Wistar rats were divided into five groups: saline only treatment, saline treatment and food deprivation, exposure to a single dose of Cd, exposure to LPS alone, and exposure to Cd + LPS. Changes in plasma nitrate concentrations and hepatic and renal Zn and Cu contents were measured together with urinary excretion rates for the metals and nitrate on 3 consecutive days: 24 h before treatment and 24 and 48 h after treatments. Cd exposure alone for 48 h caused a nearly 2-fold increase in plasma nitrate levels with no changes in urinary nitrate excretion whereas LPS treatment caused plasma nitrate levels to increase by 10-fold and urinary nitrate excretion to increase by 4-fold. Administration of LPS 24 h after Cd exposure caused a 10-fold increase in plasma nitrate concentrations and a 100-fold increase in urinary nitrate excretion compared to the rates prior to LPS administration. These results indicate a synergistic interaction between Cd and LPS toxicity. Cd exposure also caused a marked increase in hepatic Zn levels, but LPS did not cause any changes in hepatic Zn or Cu content. In sharp contrast, both Zn and Cu contents were decreased in the kidneys by 16 and 36% in animals exposed to Cd or LPS. A correlation analysis of measured variables reveals that renal Cu contents were inversely associated with plasma nitrate concentrations while urinary Cu excretion on day 3 showed a strong positive correlation with both urinary nitrate and Cd excretions on the same day. A linear regression analysis shows 20% of the variation in urinary Cu excretion was associated with urinary Cd excretion on the same day. It is concluded that reductions in renal Cu contents caused by Cd or LPS administration may be a result of Cd and NO displacement of Cu previously bound to metallothionein.

Animals↗