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Biomedical subjects

C J Reid

Publications and source records attributed to C J Reid.

At least 19 recordsLinked to original sources
PubMed2001

In vivo glycosylation of mucin tandem repeats.

The biochemical and biophysical properties of mucins are largely determined by extensive O-glycosylation of serine- and threonine-rich tandem repeat (TR) domains. In a number of human diseases aberrant O-glycosylation is associated with variations in the properties of the cell surface-associated and secreted mucins. To evaluate in vivo the O-glycosylation of mucin TR domains, we generated recombinant chimeric mucins with TR sequences from MUC2, MUC4, MUC5AC, or MUC5B, which were substituted for the native TRs of epitope-tagged MUC1 protein (MUC1F). These hybrid mucins were extensively O-glycosylated and showed the expected association with the cell surface and release into culture media. The presence of different TR domains within the chimeric mucins appears to have limited influence on their posttranslational processing. Alterations in glycosylation were detailed by fast atom bombardment mass spectrometry and reactivity with antibodies against particular blood-group and tumor-associated carbohydrate antigens. Future applications of these chimeras will include investigations of mucin posttranslational modification in the context of disease.

Amino Acid Sequence↗
PubMed2000

Primary, nonsyndromic vesicoureteric reflux and its nephropathy is genetically heterogeneous, with a locus on chromosome 1.

Primary vesicoureteric reflux (VUR) affects 1%-2% of whites, and reflux nephropathy (RN) causes up to 15% of end-stage renal failure in children and adults. There is a 30-50-fold increased incidence of VUR in first-degree relatives of probands, compared with the general population. We report the results of the first genomewide search of VUR and RN; we studied seven European families whose members exhibit apparently dominant inheritance. We initially typed 387 polymorphic markers spaced, on average, at 10 cM throughout the genome; we used the GENEHUNTER program to provide parametric and nonparametric linkage analyses of affected individuals. The most positive locus spanned 20 cM on 1p13 between GATA176C01 and D1S1653 and had a nonparametric LOD score (NPL) of 5.76 (P=.0002) and a parametric LOD score of 3.16. Saturation with markers at 1-cM intervals increased the NPL to 5.94 (P=.00009). Hence, VUR maps to a locus on chromosome 1. There was evidence of genetic heterogeneity at the chromosome 1 locus, and 12 additional loci were identified genomewide, with P<.05. No significant linkage was found to 6p, where a renal and ureteric malformation locus has been reported, or to PAX2, mutations of which cause VUR in renal-coloboma syndrome. Our results support the hypothesis that VUR is a genetic disorder.

Chromosome Mapping↗
PubMed1999

CFTR expression does not influence glycosylation of an epitope-tagged MUC1 mucin in colon carcinoma cell lines.

The cause of the mucus clearance problems associated with cystic fibrosis remains poorly understood though it has been suggested that mucin hypersecretion, dehydration of mucins, and biochemical abnormalities in the glycosylation of mucins may be responsible. Since the biochemical and biophysical properties of a mucin are dependent on O-glycosylation, our aim was to evaluate the O-glycosylation of a single mucin gene product in matched pairs of cells that differed with respect to CFTR expression. An epitope-tagged MUC1 mucin cDNA (MUC1F) was used to detect variation in mucin glycosylation in stably transfected colon carcinoma cell lines HT29 and Caco2. The glycosylation of MUC1F mucin was evaluated in matched pairs of Caco2 cell lines that either express wild-type CFTR or have spontaneously lost CFTR expression. The general glycosylation pattern of MUC1F was evaluated by determining its reactivity with a series of monoclonal antibodies against known blood group and tumor-associated carbohydrate antigens. Metabolic labeling experiments were used to estimate the gross levels of glycosylation and sulfation of MUC1F mucin in these matched pairs of cell lines. Expression of CFTR in this experimental system did not affect the gross levels of glycosylation or sulfation of the MUC1F mucin nor the types of carbohydrates structures attached to the MUC1F protein.

Caco-2 Cells↗
PubMed1999

Expression of the MUC 6 mucin gene in development of the human kidney and male genital ducts.

The MUC 6 mucin cDNA was isolated from a human stomach cDNA library and has been shown to be expressed in a number of other tissues in the gastrointestinal tract, including the gallbladder, pancreas, and parts of the ileum and colon. Here we establish that MUC 6 is expressed transiently in the nephrogenic zone of the kidney in the early mid-trimester of development. MUC 6 transcripts were detected in the epithelium of ureteric buds at 13 weeks and at lower levels from 17 to 23 weeks of gestation. Traces of MUC 6 mRNA were seen in the collecting ducts but not elsewhere in the developing kidney, and MUC 6 glycoprotein was detected in the epithelium of ureteric buds and collecting ducts. MUC 6 transcripts were absent from adult kidney. This pattern of expression of MUC 6 in the developing kidney suggests a role in epithelial organogenesis. MUC 6 transcripts were also present at low levels in mid-trimester epididymal epithelium.

Epididymis↗
PubMed1998

Localization of expression of human beta defensin-1 in the pancreas and kidney.

Defensins are antimicrobial peptides which play a key role in innate immunity. High levels of human beta defensin-1 (hBD-1) have previously been detected in the kidney and pancreas, but the cell-specific location of hBD-1 mRNA has not been determined. The expression of hBD-1 mRNA has been examined in fetal and adult pancreas and kidney by mRNA in situ hybridization. In fetal pancreas, hBD-1 expression was detected in the developing acini and in adult pancreas in the acini, but not in the pancreatic ducts. In both fetal and adult kidney, hBD-1 expression was detected in the collecting ducts and in the loops of Henle in adult kidney. Expression of hBD-1 mRNA in the pancreas and kidney from early development and in the acini of the adult pancreas, rather than in the pancreatic ducts, may indicate that in these tissues, hBD-1 fulfils physiological functions in addition to host defence.

Adult↗
PubMed1998

Roles of DctA and DctB in signal detection by the dicarboxylic acid transport system of Rhizobium leguminosarum.

The dctA gene, coding for the dicarboxylate transport protein, has an inducible promoter dependent on activation by the two-component sensor-regulator pair DctB and DctD. LacZ fusion analysis indicates that there is a single promoter for dctB and dctD. The dctA promoter is also induced by nitrogen limitation, an effect that requires DctB-DctD and NtrC. DctB alone is able to detect dicarboxylates in the absence of DctA and initiate transcription via DctD. However, DctA modifies signal detection by DctB such that in the absence of DctA, the ligand specificity of DctB is broader. dctAp also responds to heterologous induction by osmotic stress in the absence of DctA. This effect requires both DctB and DctD. A transposon insertion in the dctA-dctB intergenic region (dctA101) which locks transcription of dctA at a constitutive level independent of DctB-DctD results in improper signalling by DctB-DctD. Strain RU150, which carries this insertion, is defective in nitrogen fixation (Fix-) and grows very poorly on ammonia as a nitrogen source whenever the DctB-DctD signalling circuit is activated by the presence of a dicarboxylate ligand. Mutation of dctB or dctD in strain RU150 reinstates normal growth on dicarboxylates. This suggests that DctD-P improperly regulates a heterologous nitrogen-sensing operon. Increased expression of DctA, either via a plasmid or by chromosomal duplication, restores control of DctB-DctD and allows strain RU150 to grow on ammonia in the presence of a dicarboxylate. Thus, while DctB is a sensor for dicarboxylates in its own right, it is regulated by DctA. The absence of DctA allows DctB and DctD to become promiscuous with regard to signal detection and cross talk with other operons. This indicates that DctA contributes significantly to the signalling specificity of DctB-DctD and attenuates cross talk with other operons.

Bacterial Proteins↗
PubMed1998

Developmental expression of mucin genes in the human gastrointestinal system.

BACKGROUND AND AIMS: Mucin glycoproteins play a key role in the normal function of the epithelium lining the gastrointestinal tract. The expression of mucin genes, MUC 3, 4, 5AC, 5B, 6, 7, and 8 in human fetal tissues was examined to establish the localisation and age of onset of expression of each mucin gene during human development. METHODS: Mucin gene expression was assayed by mRNA in situ hybridisation. RESULTS: Expression of MUC3 was detected in the small intestine and colon from 13 weeks gestation onwards and at low levels in the main pancreatic duct at 13 weeks only. MUC4 expression was seen at a low level in the colonic epithelium from 13 weeks of gestation but not elsewhere in the gastrointestinal tract. MUC5AC mRNA was detected in the colon at 17 weeks and at high levels in the stomach at 23 weeks. MUC6 transcripts were evident in the pancreatic ducts from 13 weeks of gestation and at high levels in the stomach at 23 weeks. MUC5B, MUC7, and MUC8 transcripts were not detected. CONCLUSIONS: Mucin genes are expressed from the early mid-trimester of gestation in the developing human fetal gastrointestinal tract.

Colon↗
PubMed1997

Oligosaccharides expressed on MUC1 produced by pancreatic and colon tumor cell lines.

MUC1 is expressed at the apical surface of ductal epithelia of tissues, including breast, pancreas, airway, and the gastrointestinal tract, where its functions include lubrication and protection of the epithelia. In addition, roles for MUC1 have been suggested in both adhesive and antiadhesive properties of tumor cells, and extensive O-glycosylation of the MUC1 tandem repeat domain may contribute to these functions. Little information is available on the specific O-glycosylation of MUC1. One problem in identifying different MUC1 glycoforms has been that monoclonal antibodies raised against the MUC1 core protein recognize epitopes in the tandem repeat domain, which is often glycosylated to an extent that obscures these epitopes. We developed an epitope-tagged form of MUC1 that allowed the detection of multiple MUC1 glycoforms and established the presence of a number of important blood group and tumor-associated carbohydrate antigens on MUC1 expressed by two pancreatic tumor cell lines (Panc-1 and S2-013) and two colon tumor cell lines (Caco-2 and HT-29). Antigens detected include sialyl-Lewisa, sialyl-Lewisc, sialyl-Lewisx, and sialyl-Tn.

Antibodies↗
PubMed1997

The general amino acid permease of Rhizobium leguminosarum strain 3841 is negatively regulated by the Ntr system.

Cosmid-borne and chromosomal lacZ fusions to aapJ. aapQ and aapM were used to examine the nitrogen regulation of the general amino acid permease (Aap) of Rhizobium leguminosarum strain 3841. Transcription of the first gene of the operon (aapJ), which encodes the periplasmic binding protein, was 2-4-fold higher than aapQ and aapM, which encode the integral membrane proteins, under various growth conditions. This may be due to the presence of a putative stem loop in the intergenic region between aapJ and aapQ. All aap fusions were derepressed 3-5-fold after growth on glutamate as a nitrogen source, which effectively causes nitrogen limitation. An ntrC mutant was derepressed for transcription of the aap operon and had high rates of amino acid transport when grown on ammonia as the nitrogen source. Thus NtrC negatively regulates the aap operon, contrary to its usual role in positive gene activation. These results confirm that the aap-operon is subject to complex regulation involving both transcriptional and post-transcriptional factors.

Amino Acid Transport Systems↗
PubMed1997

Pulmonary thrombo-embolism in nephrotic syndrome treated with tissue plasminogen activator.

UNLABELLED: We describe the case of a boy with steroid sensitive nephrotic syndrome and left pulmonary artery thrombo-embolism. clinical presentation initially suggested sepsis and respiratory signs were minor. Treatment with tissue plasminogen activator infused into the pulmonary artery was successful. CONCLUSION: Pulmonary thrombo-embolism should be considered in unwell children with nephrotic syndrome.

Child, Preschool↗
PubMed1997

The cystic fibrosis transmembrane conductance regulator as a marker of human pancreatic duct development.

BACKGROUND & AIMS: The cystic fibrosis transmembrane conductance regulator (CFTR) protein is a small conductance adenosine 3',5'-cyclic monophosphate (cAMP)-activated chloride ion channel found in the apical membranes of epithelia within the pancreas, airway, intestine, bile duct, sweat gland, and male genital ducts. Pancreatic insufficiency is a feature of about 85% of patients with cystic fibrosis and is believed to be caused by pancreatic autolysis after pancreatic duct obstruction. The aim of this study was to investigate the expression of CFTR in the pancreas from early development to postnatal life to establish whether the CFTR plays a key role in development of the pancreatic duct epithelium. METHODS: Expression of CFTR from the start of the mid-trimester of human development through term to adult life by messenger RNA (mRNA) in situ hybridization was examined. RESULTS: CFTR mRNA is detected throughout the pancreatic duct epithelium and its pattern of expression follows the differentiation of the duct system. CONCLUSIONS: CFTR is a valuable marker of human pancreatic duct cell development and differentiation.

Aging↗
PubMed1997

Developmental expression of mucin genes in the human respiratory tract.

Mucin glycoproteins play a key role in the normal function of the airway epithelium. We examined the expression of mucin genes, MUC3, 4, 5AC, 5B, 6, 7, and 8 in human fetal tissues to establish the localization and age of onset of expression of each mucin gene during human development. We detected expression of MUC4, 5AC, 5B, and 7 in the mid-trimester airway epithelium but did not detect expression of MUC3, 6, or 8. MUC4 was expressed in the trachea and large airways in the majority of cells in the airway epithelium. Expression of MUC5AC was only seen in individual goblet cells in the trachea, while MUC5B was expressed in the surface epithelium of the trachea at 13 wk but was largely restricted to submucosal glands by 23 wk of gestation.

Base Sequence↗
PubMed1997

Cystic fibrosis of the pancreas: involvement of MUC6 mucin in obstruction of pancreatic ducts.

BACKGROUND: Cystic fibrosis (CF) is characterized by pancreatic destruction following the gradual obstruction of small pancreatic ducts, from the mid-trimester of gestation onwards. To date, the material causing the obstruction has not been identified. The MUC6 mucin cDNA was isolated from human stomach and has been shown to be expressed in a number of other tissues in the gastrointestinal tract, including the gall bladder and parts of the ileum and colon. MATERIALS AND METHODS: We have examined the expression of MUC6 mucin in the human pancreas, both during development and postnatally, by mRNA in situ hybridization and immunocytochemistry. RESULTS: In this report we establish that MUC6 transcripts are abundant in pancreatic epithelial cells and show a very similar pattern of expression in the epithelium lining small ducts and centroacinar cells to that shown by the cystic fibrosis transmembrane conductance regulator gene (CFTR). In addition, material obstructing the pancreatic ducts of CF pancreas was shown to contain MUC6 mucin. CONCLUSION: We have identified MUC6 mucin as a significant constituent of the material obstructing the small pancreatic ducts in CF.

Adolescent↗
PubMed1996

Aspartate transport by the Dct system in Rhizobium leguminosarum negatively affects nitrogen-regulated operons.

Amino acid uptake by the general amino acid permease (Aap) of Rhizobium leguminosarum strain 3841 was severely reduced by the presence of aspartate in the growth medium when glucose was the carbon source. The reduction in transport by the Aap appeared to be caused by inhibition of uptake and not by transcriptional repression. However, as measured with lacZ fusions, the Ntr-regulated gene glnII was repressed by aspartate. The negative regulatory effect on both the Aap and glnII was prevented by mutation of any component of the dicarboxylate transport (Dct) system or by the inclusion of a C4- dicarboxylate in the growth medium, including the non-metabolizable analogue 2-methylsuccinate. As measured by total uptake and with a dctA-lacZ fusion, aspartate was an efficient inducer of the Dct system, but slightly less so than succinate alone or succinate and aspartate together. Thus, aspartate does not cause overexpression of DctA leading to improper regulation of other operons. Transport measurements revealed that the Dct system has an apparent Km for succinate of 5 microM and an apparent Ki for aspartate inhibition of succinate uptake of 5 mM. These data imply that the Dct-mediated accumulation of aspartate causes an unregulated build-up of aspartate or a metabolic product of it in the cell. This accumulation of aspartate is prevented either by mutation of the dct system or by the presence of a higher affinity substrate that will reduce access of aspartate to the carrier protein. Elevation or disruption of the intracellular aspartate pool is predicted to disrupt N-regulated operons and nitrogen fixation.

Amino Acid Transport Systems↗