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Biomedical subjects

M J Schneider

Publications and source records attributed to M J Schneider.

At least 19 recordsLinked to original sources
PubMed2004

Comparison of a bioassay and a liquid chromatography-fluorescence-mass spectrometry(n) method for the detection of incurred enrofloxacin residues in chicken tissues.

Regulatory monitoring for most antibiotic residues in edible poultry tissues is often accomplished with accurate, although expensive and technically demanding, chemical analytical techniques. The purpose of this study is to determine if a simple, inexpensive bioassay could detect fluoroquinolone (FQ) residues in chicken muscle above the FDA established tolerance (300 ppb) comparable to a liquid chromatography-fluorescencemass spectrometry(n) method. To produce incurred enrofloxacin (ENRO) tissues (where ENRO is incorporated into complex tissue matrices) for the method comparison, 40-d-old broilers (mixed sex) were orally dosed through drinking water for 3 d at the FDA-approved dose of ENRO (50 ppm). At the end of each day of the 3-d dosing period and for 3 d postdosing, birds were sacrificed and breast and thigh muscle collected and analyzed. Both methods were able to detect ENRO at and below the tolerance level in the muscle, with limits of detection of 26 ppb (bioassay), 0.1 ppb for ENRO, and 0.5 ppb for the ENRO metabolite, ciprofloxacin (liquid chromatography-fluorescence-mass spectrometry(n)). All samples that had violative levels of antibiotic were detected by the bioassay. These results support the use of this bioassay as a screening method for examining large numbers of samples for regulatory monitoring. Positive samples should then be examined by a more extensive method, such as liquid chromatography-fluorescence-mass spectrometry(n), to provide confirmation of the analyte.

Animals↗
PubMed2001

Multiresidue analysis of fluoroquinolone antibiotics in chicken tissue using automated microdialysis-liquid chromatography.

An efficient procedure for the simultaneous extraction and analysis of six fluoroquinolone (FQ) antibiotics is developed using an automated microdialysis-liquid chromatographic (LC) system. In this method, samples extracted from chicken liver and muscle are further purified by microdialysis, separated on an LC column, and the FQs detected by their fluorescence. Recoveries from fortified chicken liver and muscle samples are at least 70% with limits of quantitation (microg/kg) for the FQs in liver (and muscle) as follows: 0.3 (0.4) for danofloxacin, 0.8 (0.2) for desethylene ciprofloxacin, 2 (1) for norfloxacin, 2 (0.8) for enrofloxacin, 3 (1) for ciprofloxacin, and 5 (2) for sarafloxacin. Enrofloxacin and ciprofloxacin are determined in enrofloxacin-incurred chicken liver and muscle samples using this method.

Animals↗
PubMed2001

Targeted disruption of the type 2 selenodeiodinase gene (DIO2) results in a phenotype of pituitary resistance to T4.

The type 2 deiodinase (D2), a selenoenzyme that catalyzes the conversion of T4 to T3 via 5'-deiodination, is expressed in the pituitary, brain, brown adipose tissue (BAT), and the reproductive tract. To examine the physiological role of this enzyme, a mouse strain lacking D2 activity was developed using homologous recombination. The targeting vector contained the Neo gene in place of a 2.6-kb segment of the Dio2 gene. This segment comprises 72% of the coding region and includes the TGA codon that codes for the selenocysteine located at the active site of the enzyme. Mice homologous for the targeted deletion [D2 knockout (D2KO)] had no gross phenotypic abnormalities, and development and reproductive function appeared normal, except for mild growth retardation (9%) in males. No D2 activity was observed in any tissue in D2KO mice under basal conditions, or under those that normally induce this enzyme such as cold-exposure (BAT) or hypothyroidism (brain, BAT, and pituitary gland). Furthermore, no D2 activity was present in cultured astrocytes, nor could it be induced by treatment of the cells with forskolin. Although D2 mRNA transcripts were detected in BAT RNA obtained from cold-exposed wild-type (WT) mice, none was detected in BAT RNA from comparably-treated D2KO mice. Levels of D1 in the liver, thyroid, and pituitary were the same in WT and D2KO animals, whereas D3 activity in D2KO cerebrum was twice that in WT cerebrum. Serum T3 levels were comparable in adult WT and D2KO mice. However, serum T4 and TSH levels were both elevated significantly (40% and 100%, respectively) in the D2KO mice, suggesting that the pituitary gland of the D2KO mouse is resistant to the feedback effect of plasma T4. This view was substantiated by the finding that serum TSH levels in hypothyroid WT mice were suppressed by administration of either T4 or T3, but only T3 was effective in the D2KO mouse. The data also suggest that the clearance of T4 from plasma was reduced in the D2KO mouse. In summary, targeted inactivation of the selenodeiodinase Dio2 gene results in the complete loss of D2 activity in all tissues examined. The increased serum levels of T4 and TSH observed in D2KO animals demonstrate that the D2 is of critical importance in the feedback regulation of TSH secretion.

Adipose Tissue, Brown↗
PubMed2001

Rating specific chiropractic technique procedures for common low back conditions.

OBJECTIVE: To rate specific chiropractic technique procedures used in the treatment of common low back conditions. DESIGN AND METHODS: A panel of chiropractors rated specific chiropractic technique procedures for their effectiveness in the treatment of common low back conditions, based on the quality of supporting evidence after systematic literature reviews and expert clinical opinion. Statements related to the rating process and clinical practice were then developed through a facilitated nominal consensus process. RESULTS: For most low back conditions presented in this study, the three procedures rated most effective were high-velocity, low- amplitude (HVLA) with no drop table (side posture), distraction technique, and HVLA prone with drop table assist. The three rated least effective were upper cervical technique, non-thrust reflex/low force, and lower extremity adjusting. The four conditions rated most amenable to chiropractic treatment were noncomplicated low back pain, sacroiliac joint dysfunction, posterior joint/subluxation, and low back pain with buttock or leg pain. CONCLUSIONS: The ratings for the effectiveness of chiropractic technique procedures for the treatment of common low back conditions are not equal. Those procedures rated highest are supported by the highest quality of literature. Much more evidence is necessary for chiropractors to understand which procedures maximally benefit patients for which conditions.

Acute Disease↗
PubMed1999

Simulation studies: a new method of prospective technology assessment and design.

In order to examine the social conditions and effects of a broad usage of telecooperation technology in the future, such as the development of design proposals, PROVET (Projektgruppe Verfassungsverträgliche Technikgestaltung) developed the method of simulation studies. The paper describes the problem of prospective technology design for telecooperation, a method to handle it, the simulation studies and the methological background. It demonstrates three simulation studies, in administration of justice, about office dossiers and for a reachability management. It presents results, particularly about the psychological effects of the digital signature as a technical-organizational measure to guarantee obligations between the cooperation partners. The conclusion discusses this research approach in relation to others.

Computer Simulation↗
PubMed1999

Isolation and characterization of the mouse gene for the type 3 iodothyronine deiodinase.

The type 3 iodothyronine deiodinase (D3) is a selenoenzyme that inactivates thyroid hormones by removing a iodine from the 5-position of the tyrosyl ring. D3 is highly expressed in many tissues during the early stages of development, and its activity is regulated by selected growth factors and various hormones. To gain further insights into the structure, functional role, and regulation of this enzyme, we screened a mouse liver genomic library with a rat D3 complementary DNA probe and isolated a 12-kb clone coding for the Dio3. Restriction analysis followed by Southern blotting and nucleotide sequencing demonstrated that the Dio3 contains a single exon, 1853 bp in length, that encodes the entire length of the messenger RNA expressed in murine placenta and neonatal skin. Primer extension experiments identified two potential transcriptional start sites located 77 and 60 nt upstream of the ATG translational start codon. The region immediately 5' to the start sites contains consensus TATA, CAAT, and GC elements. Furthermore, a 526-nucleotide genomic fragment from this region was demonstrated to efficiently drive a luciferase reporter construct when transfected into COS-7, XTC-2, or XL-2 cells or into primary cultures of rat preadipocytes derived from neonatal brown fat. In conclusion, D3 transcripts in the placenta and skin are encoded by the Dio3 gene from a single exon whose expression is regulated by an upstream region that contains several consensus promoter elements. Further characterization of this gene will provide new insights into the factors regulating the unique pattern of D3 expression during development.

Amino Acid Sequence↗
PubMed1999

Cloning of a 5.8 kb cDNA for a mouse type 2 deiodinase.

From studies with their cDNAs, the types 1 and 3 deiodinases (D1 and D3) have been shown unequivocally to be selenoproteins. Studies with recently cloned cDNAs for the mammalian type 2 deiodinase (D2) indicate that they also code for selenoproteins. However, these D2 cDNAs are not full length and they do not contain an essential selenocysteine insertion sequence (SECIS) in their 3'UTR; a heterologous SECIS had to be ligated to the coding region before expression of the D2 could be achieved. Thus their role as cDNAs for the native D2 is open to question. We now report the cloning of a 5.8 kb cDNA for the mouse D2. This cDNA contains a SECIS in its 3'UTR located more than 4.5 kb from the coding region. When the mRNA transcribed in vitro from this cDNA is injected into X. laevis oocytes, a deiodinase with characteristics of D2 is expressed.

3' Untranslated Regions↗
PubMed1997

The type 2 and type 3 iodothyronine deiodinases play important roles in coordinating development in Rana catesbeiana tadpoles.

In developing Rana catesbeiana tadpoles, the timing of the thyroid hormone (TH)-dependent metamorphic responses varies markedly among tissues. Yet at any one time these tissues are exposed to the same plasma concentration of TH, suggesting that TH action is regulated in part at the level of the peripheral tissues. A major factor in TH action is the intracellular level of the active TH, T3. This level is dependent not only on the plasma concentration of TH (mostly T4) but also on the intracellular activities of the type 2 5'-deiodinase (D2) and the type 3 5-deiodinase (D3), which are responsible, respectively, for generating and degrading T3. (D1 is not present in this species.) To determine whether differential expression of D2 and D3 among tissues could be a significant factor in the coordination of metamorphic events, the ontogenic profiles of the two enzyme activities and corresponding messenger RNA levels in most tissues of R. catesbeiana tadpoles have been documented. The profiles of D2 expression in tail, hindlimb, forelimb, intestine, skin, and eye differed markedly at both activity and messenger RNA levels, but it was notable that expression was invariably highest in a given tissue at the time of its major metamorphic change. D2 expression was very low in brain and heart and did not vary during development. D2 was not expressed in liver, kidney, or red blood cells. With the exception of red blood cells, D3 expression was detected in all tissues studied. Furthermore, it was evident that in tissues that expressed both deiodinase genes, the two expression profiles were comparable, indicating a potential for tight control of intracellular T3 levels. Direct evidence of the importance of the intracellular conversion of T4 to T3 for TH-dependent metamorphic events was obtained in tadpoles in which endogenous TH synthesis was blocked with methimazole, and the activities of D2 and D3 were inhibited by iopanoic acid. This treatment inhibited metamorphosis. The inhibition could be overcome by the concomitant administration of replacement levels of T3, but not T4. These results strongly support the view that coordinated development in amphibia depends in part on the tissue-specific expression patterns of the D2 and D3 genes, which ensure that the requisite level of intracellular T3 is attained in a given tissue, regardless of the current level of circulating TH, at the appropriate stage of metamorphosis.

Animals↗
PubMed1995

Cloning of a cDNA for the type II iodothyronine deiodinase.

Three types of iodothyronine deiodinase have been identified in vertebrate tissues. cDNAs for the types I and III have been cloned and shown to contain an inframe TGA that codes for selenocysteine at the active site of the enzyme. We now report the cloning of a cDNA for a type II deiodinase using a reverse transcription/polymerase chain reaction strategy and RNA obtained from Rana catesbeiana tissues. This cDNA (RC5'DII) manifests limited but significant homology with other deiodinase cDNAs and contains a conserved in-frame TGA codon. Injection of capped in vitro synthesized transcripts of the cDNA into Xenopus laevis oocytes results in the induction of deiodinase activity with characteristics typical of a type II deiodinase. The levels of RC5'DII transcripts in R. catesbeiana tadpole tail and liver mRNA at stages XII and XXIII correspond well with that of type II deiodinase activity but not that of the type III activity in these tissues. These findings indicate that the amphibian type II 5'-deiodinase is a structurally unique member of the family of selenocysteine-containing deiodinases.

Amino Acid Sequence↗
PubMed1995

Cloning and expression of a cDNA for a mammalian type III iodothyronine deiodinase.

The type III iodothyronine deiodinase metabolizes the active thyroid hormones thyroxine and 3,5,3'-triiodothyronine to inactive compounds. Recently, we have characterized a Xenopus laevis cDNA (XL-15) that encodes a selenoprotein with type III deiodinase activity (St. Germain, D.L., Schwartzman, R., Croteau, W., Kanamori, A., Wang, Z., Brown, D.D., and Galton, V.A. (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 7767-7771). Using the XL-15 as a probe, we screened a rat neonatal skin cDNA library. Among the clones isolated was one (rNS43-1) which contained a 2.1-kilobase pair cDNA insert that manifested significant homology to both the XL-15 and the G21 rat type I deiodinase cDNAs, including the presence of an in-frame TGA codon. Expression studies demonstrated that the rNS43-1 cDNA encodes a protein with 5-, but not 5'-, deiodinase activity that is resistant to inhibition by propylthiouracil and aurothioglucose. Northern analysis demonstrated a pattern of tissue expression in the rat consistent with that of the type III deiodinase and site directed mutagenesis confirmed that the TGA triplet codes for selenocysteine. We conclude that the rNS43-1 cDNA encodes the rat type III deiodinase and that the types I and III deiodinases present in amphibians and mammals constitute a family of conserved selenoproteins important in the metabolism of thyroid hormones.

Amino Acid Sequence↗
PubMed1995

The type III 5-deiodinase in Rana catesbeiana tadpoles is encoded by a thyroid hormone-responsive gene.

We have recently reported that the Xenopus laevis complementary DNA (cDNA), XL-15, encodes a selenoprotein that is a 5-deiodinase (5D). XL-15 represents a gene that is up-regulated by thyroid hormone in this species. We now report the isolation from a Rana catesbeiana (RC) cDNA library of a cDNA for the RC 5D (RC5D). RC5D a 1534-base pair cDNA that exhibits 78% identity to XL-15 in the coding region, including the TGA codon, which in XL-15 encodes selenocysteine, and a putative selenocysteine insertion sequence (SECIS) located in the 3'-untranslated region. Transcripts of RC5D, synthesized in vitro, induce 5D activity after their injection into Xenopus laevis oocytes. RC5D hybridizes to a 2.2-kilobase messenger RNA (mRNA) species in RC tissues. The expression of RC5D was studied in 10 tissues from untreated and T3-treated premetamorphic tadpoles. RC5D mRNA transcripts and 5D activity were detected in most tissues examined, and the levels of both were greatly increased in T3-stimulated tadpoles. Furthermore, the levels of RC5D mRNA transcripts correlated closely with 5D, but not with 5'-deiodinase (5'D), activity. 5'D was not enhanced by a 4-day exposure of the tadpoles to T3. The up-regulation of RC5D gene expression by T3 was demonstrable in tadpoles by stage V, but was found to be transient. The levels of RC5D transcripts and 5D activity were highest after 2-3 days and 5 days of T3 exposure, respectively, but had fallen by 10 days to levels comparable to those in unexposed tadpoles. It is concluded that RC5D encodes a selenoprotein that is a 5D, and it represents a gene that is up-regulated by T3. On the basis of the findings presented herein, it is suggested that the 5D system plays a major role in regulating intracellular T3 levels in developing tadpoles.

Amino Acid Sequence↗
PubMed1995

Effect of glucocorticoids on thyroid hormone action in cultured red blood cells from Rana catesbeiana tadpoles.

Metamorphosis in anuran amphibia requires thyroid hormone (TH) and can be induced prematurely by the administration of TH. There is also evidence that the developmental effects of TH in these forms are modified by other hormones. For example, PRL has been shown to retard and corticosterone (B) to accelerate some, but not all, components of TH-induced metamorphosis. Red blood cells (RBCs) of Rana catesbeiana tadpoles exhibit a 4- to 5-fold increase in thyroid hormone receptor (TR) number (sites per nucleus) in vivo during either spontaneous or TH-induced metamorphosis. In the present study this TH-induced effect on RBC TR number was examined in an in vitro culture system. RBC TR number was increased by T3 in vitro; the maximum effect (2-fold increase) was obtained after exposure to 0.3 nM T3 for 60 h. This T3-induced increase in TR number was completely abolished in the presence of either 34 nM B or 10 nM dexamethasone, whereas basal TR number was unaffected. The effect appears to be a specific effect of glucocorticoid (GC), because it was not mimicked by the sex steroid, testosterone, and it was not obtained when RU-486, a glucocorticoid antagonist, was included with B in the medium. Other experiments demonstrated that the T3-induced increase in RBC TR was associated with an increase in the TR alpha messenger RNA level. This increase in TR alpha messenger RNA was reduced, but not eliminated, in the presence of concentrations of GC that abolished the TH-induced increase in TR, suggesting that the effects of GC occur in part at a pretranslational level. Using a GC binding assay, tadpole RBCs were found to contain approximately 10(4) GC receptors/cell. These findings indicate that B may be a physiological modulator of TH action in tadpole RBCs. This inhibitory effect of GC contrasts with previous reports that GC accelerates some of the morphological effects of TH in developing tadpoles, indicating that the nature of this modulating effect on TH action is tissue specific.

Animals↗
PubMed1994

Cloning of a thyroid hormone-responsive Rana catesbeiana c-erbA-beta gene.

Two types of thyroid hormone receptor (c-erbA) gene have been identified in mammals and in lower species including chickens and the amphibian Xenopus laevis. The two genes are located on different chromosomes and have been named TR alpha and TR beta. We have described previously the cloning of a TR alpha cDNA from Rana catesbeiana (RC) tissues (RC15) and we now report the cloning of a TR beta cDNA from this species. The cloning strategy employed utilized the polymerase chain reaction (PCR), with primers based on the sequences of the X. laevis TR beta cDNA (XenTR beta) and an RCTR beta genomic clone, which, by analogy with XenTR beta, contains some of the 3' end of the open reading frame together with 3'-untranslated sequences. At the nucleotide and amino acid levels, respectively, the cloned RCTR beta cDNA is 90% and 98% homologous with XenTR beta, and 72% and 76% homologous with RC15. Following in vitro transcription and translation, the cDNA was shown to encode a 48 kilodalton protein which binds 3,5,3'-triiodothyronine (T3) with high affinity (mean Kd: 0.032 nM). Samples of total or poly(A) +RNA from tadpoles at different stages of metamorphosis and from adult frogs were analyzed for the presence of TR beta-specific transcripts by slot blot analysis using as probe a 258 bp section of the RCTR beta cDNA. This section of the cDNA does not hybridize to the corresponding section of RC15. In confirmation of previous findings, beta-specific transcripts were not detected in RNA from tadpole red blood cells (RBCs) and none was found in RBCs from adult frogs.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1993

Rana catesbeiana tadpole red blood cells express an alpha, but not a beta, c-erbA gene.

Thyroid hormone (TH) receptor number in red blood cells (RBCs) from Rana catesbeiana (RC) tadpoles increases 4-fold during both spontaneous and TH-induced metamorphosis, an effect that we have previously shown to be preceded by an increase in the level of c-erbA-related mRNA. The goals of the present study were to obtain an RC c-erbA alpha cDNA that contains the entire open reading frame for a putative TH receptor protein, to determine if this protein has characteristics typical of a TH receptor, and to assess its contribution to the developmentally related increase in TH receptor number. To accomplish this, the missing 5'-sequence of a previously isolated partial RC c-erbA alpha cDNA (RC12) was synthesized by polymerase chain reaction (PCR) and spliced to RC12 to yield a 1490-basepair cDNA (RC15) that contained the entire coding sequence of the receptor protein. Transcription of RC15 followed by translation of its mRNA in a rabbit reticulolysate system yielded a 50-kilodalton protein on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The protein binds T3 with high affinity (Kd, approximately 0.1 nM), and its affinity for T3 is at least 5 times that for T4. The results of cotransfection studies indicate that RC15 can function as a TH receptor; when COS cells were cotransfected with a construct consisting of RC15 cloned in the expression vector CMV4 and TK28 mult, a construct containing rat GH gene TH response element sequences up-stream of a chloramphenicol acetyltransferase reporter gene, chloramphenicol acetyltransferase activity is expressed in the presence, but not in the absence, of T3. To determine whether RBCs contain any c-erbA beta mRNA transcripts that might contribute to the developmentally related increase in the transcripts detected using RC c-erbA alpha cDNAs, alpha- and beta-specific cDNAs were synthesized by PCR and used as probes in a variety of hybridization assays. In all experiments using conditions in which c-erbA beta transcripts were detectable in other tissues, there was no evidence that tadpole RBCs contained such species. Lack of any beta-specific transcripts was confirmed by PCR, using as template cDNA prepared by reverse transcription of RC RBC RNA. It was also noted that the RBC at metamorphic climax is the tissue with the highest content of alpha-specific c-erbA transcripts. It is concluded that the c-erbA alpha gene encodes a TH receptor, and that only the alpha-gene is expressed in tadpole RBCs and subject to regulation during development and by TH.

Amino Acid Sequence↗
PubMed1991

The role of thyroid hormone in the regulation of hepatic carbamyl phosphate synthetase activity in Rana catesbeiana.

Both spontaneous and thyroid hormone (TH)-induced metamorphosis of Rana catesbeiana are accompanied by a marked increase in the activity of the urea cycle enzyme carbamyl phosphate synthetase (CPS). The increase induced by exogenous TH is de novo synthesis of enzyme and appears to be secondary to an increase in the CPS mRNA level resulting from the elevated plasma TH. Since endogenous TH levels rise sharply during spontaneous metamorphosis, a similar sequence of events would be anticipated. However, after midclimax, CPS activity continues to increase, while plasma TH levels steadily decline, suggesting that other factors are involved. To obtain insight into this problem, the changes in CPS mRNA level during spontaneous development were determined using a mammalian CPS cDNA probe and correlated with changes in CPS activity and plasma T3 concentration. CPS mRNA level and CPS activity were barely detectable until midprometamorphosis, but both increased rapidly during the latter half of this phase. CPS activity continued to rise, reaching a maximum in the adult frog. The CPS mRNA level, however, was highest during the first half of climax, but declined after midclimax and was relatively low in the adult frog. Studies were also performed in which the rise and fall in the plasma T3 concentration typical of metamorphic climax were induced by exposure of premetamorphic tadpoles to T3, followed by its withdrawal. Both CPS activity and CPS mRNA level were induced by T3, but when plasma T3 levels fell after removal of the exogenous T3, CPS mRNA level, but not CPS activity, also decreased. Additional studies indicated that the TH-induced increase in CPS mRNA was evident within 24 h, could be prevented by simultaneous injection of actinomycin-D, and could not be induced in tadpoles undergoing climax; in this phase the T3 receptors are fully occupied with endogenous TH. When premetamorphic tadpoles were immersed in T3-containing water (0-500 nM) for 6 days, CPS mRNA, CPS activity, and plasma T3 concentration increased in parallel, reaching a maximum at 50 nM. At 50 nM T3, the plasma T3 level was sufficient to saturate the receptors, and no additional increase in CPS mRNA level or CPS activity was obtained at higher concentrations of T3. These studies indicate that the CPS mRNA level during spontaneous development correlates with the plasma T3 concentration and suggest that it is a function of T3 receptor occupancy. The data are also consistent with an effect of TH on transcription of the CPS gene and with a relatively long half-life of its protein product, the CPS enzyme.

Animals↗