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Biomedical subjects

S Watkins

Publications and source records attributed to S Watkins.

At least 91 records · Page 5Linked to original sources
PubMed1994

Effect of secretory IgA on transepithelial passage of bacteria across the intact ileum in vitro.

BACKGROUND: Bacterial translocation is a process believed to result in nosocomial infections. Secretory IgA (sIgA) may have a role in the prevention of translocation by its ability to bind and aggregate bacteria, a function termed "immune exclusion." The present study was done to determine the effect of specific binding of sIgA to bacteria on the movement of these organisms across the intact epithelial membrane. STUDY DESIGN: Bacterial translocation across intact intestinal segments of rats were assessed in vitro using the Ussing model. Secretory IgA (0.25 mg per mL) from pooled human colostrum was added to the perfused segments of ileum in the Ussing system. Subsequently, the membranes were exposed to 5 x 10(9) cfu per mL Escherichia coli on their mucosal side. A second experiment tested the effect of human IgG when perfused with E. coli using the same preparation. All experiments had paired matched rats in a control group without immunoglobulin. The ability of sIgA and IgG to bind to E. coli was studied by an in vitro assay, as well as by transmission electron microscopy and immunofluorescence of random IgA/E. coli experiments. Measurements obtained in all experimental and control groups were the incidence and amount of bacterial passage and the potential difference generated by the intestinal segments (an index of viability). RESULTS: There were no differences in potential difference between control and experimental groups in either of the two experiments. Secretory IgA bound E. coli and completely prevented passage of E. coli as compared with rats in the control group. IgG bound E. coli; however, the incidence of passage was equal to that of rats in the control group. However, the presence of IgG resulted in a significantly reduced number of bacteria that passed when compared with controls (p < 0.05). Electron microscopic studies revealed intact surface morphology and immunofluorescence revealed aggregates of IgA and E. coli on the mucosal, but not submucosal, surface of the ileal membranes. CONCLUSIONS: This study provides direct evidence of immune exclusion by sIgA. When bound to bacteria, it prevents passage across a morphologically intact segment of viable intestinal tissue.

Animals↗
PubMed1994

Role of intestinal mucus in transepithelial passage of bacteria across the intact ileum in vitro.

BACKGROUND: Although gastrointestinal mucus is one of a number of putative host defense mechanisms that protect the gut barrier against microbial translocation, little experimental data are available to show its role in this process. The present study sought to determine the role of mucus depletion on the transepithelial passage of bacteria across viable segments of rat ileum mounted in Ussing chambers in vitro. METHODS: Intestinal mucus was depleted in 12 rats after injection with pilocarpine (160 mg/kg intraperitoneally) 45 minutes before intestinal harvest. The mucosal surfaces of the perfused gut segments mounted in the Ussing chamber were exposed to 5 x 10(9) CFU/ml Escherichia coli C-25. Viability was monitored by continuous measurements of the potential difference generated by the membranes. The electrical characteristics were unaltered by pilocarpine pretreatment or exposure to bacteria. RESULTS: Bacterial passage occurred in 100% of pilocarpine membranes as compared with 33.3% in controls (p < 0.05). Pilocarpine-treated membranes resulted in 19.9 +/- 7.5 mg of retrievable mucus as compared with 28.8 +/- 7.2 mg in controls (p < 0.05). Light and transmission electron microscopy revealed an intact epithelial surface in all membranes. There was a marked decrease in mucus on the surface of pilocarpine-treated membranes. CONCLUSIONS: Intestinal mucus secretion is a critical factor in the barrier function of the gut, and its depletion results in a dramatic increase in bacterial passage across the intact rat ileum.

Animals↗
PubMed1993

Protein and DNA sequence analysis of a 'private' genetic variant: albumin Ortonovo (Glu-505-->Lys).

Albumin Ortonovo is a slow moving variant of human serum albumin which has been found only in people coming from the small villages of Ortonovo and Nicola (Liguria, Italy) and reaches polymorphic frequency (> or = 1%) in the poorly admixed population group living in that area. This is the first report of a 'private' variant detected in a Caucasian population. It probably originated as a mutation in a founder individual many generations ago. Isoelectric focusing analysis of CNBr fragments from the purified variant localized the mutation in fragment CNBr VI (residues 447-548). This fragment was isolated on a preparative scale by reversed-phase HPLC and subjected to V8 proteinase digestion. Sequence analysis of the abnormal V8 peptide revealed that the variant arises from a previously unreported substitution at position 505 where glutamic acid has been replaced by lysine. The protein data were confirmed by DNA sequence analysis which indicated a single nucleotide change of GAA-->AAA in the corresponding codon of the structural gene. Since the amino acid substitution found in albumin Ortonovo accords with its electrophoretic mobility on cellulose acetate, residue 505 is probably exposed to the solvent. The clustering of the mutations in the intersubdomain connection linking subdomains IIIA and IIIB (residues 492-511) accords with the fact that this region lies on the molecular surface and is accessible to solvent.

Amino Acid Sequence↗
PubMed1993

cDNA and protein sequence of polymorphic macaque albumins that differ in bilirubin binding.

The rhesus monkey, Macaca mulatta, exhibits a geographically restricted polymorphism of serum albumins Mac A and Mac B that is recognized by electrophoresis and is associated with a difference in bilirubin-binding parameters. To identify the basis of the polymorphism, the cDNA and protein sequences of serum albumin from M. mulatta were determined. Screening of a lambda gt11 rhesus liver cDNA library yielded a 1988-bp cDNA sequence that encodes the complete amino acid sequence of mature albumin, the entire propeptide, and part of the prepropeptide. Isoelectric focusing and amino-terminal protein sequencing of CNBr fragments of albumin from A/A and B/B homozygotes were performed, and the structural difference was localized to a CNBr fragment (MCB3) spanning residues 124-264. Sequence analysis of lysyl endopeptidase peptides of MCB3 established that Mac A albumin has a glutamine residue at position 188 while the Mac B albumin has a glutamic residue at the same position. PCR amplification, subcloning, and DNA sequence analysis of clones from A/A and B/B homozygotes confirmed the protein sequence data and the codon difference of CAA versus GAA, respectively. Comparison of macaque and human serum albumin shows a 93.5% identity at the amino acid level. In human serum albumin, Glu188 is located close to the IIA binding pocket for ligands, probably including bilirubin. Derivatives of coumarin compete more efficiently with bilirubin for binding sites on the Mac A albumin than on the Mac B albumin. In regions where coumarin-containing plants are important food resources, Mac B albumin may confer a selective advantage because bilirubin is less readily displaced from it.

Amino Acid Sequence↗
PubMed1993

Screening procedures used to identify children with hearing loss.

This study focused on the hearing-screening procedures used to identify children who have hearing losses who were later enrolled in SKI*HI parent/infant programs throughout the country. Responses to a questionnaire for 1,404 children indicated that although auditory brainstem response (ABR) technology provided the lowest mean identification age, informal hearing-screening procedures (parental suspicion and referral) were the procedure of identification for 80% of the children. The data suggest that formal screening programs are not presently locating the vast majority of infants who have hearing losses. Recommendations for professional and parental education in the area of hearing-loss screening are provided.

Age Factors↗
PubMed1993

Community care. Working together for better health.

The purpose of community care is to promote privacy, dignity and independence and provide resources for living. It is a philosophy not a place, writes Stephen Watkins. In this extract from the Medical Practitioners' Union evidence to the house of commons health committee enquiry on community care, he sets out a new blueprint for the organisation and funding of a seamless health and social service based on joint, locality-based commissioning and practice-based case management.

Community Health Services↗
PubMed1992

Alloalbuminemia in Sweden: structural study and phenotypic distribution of nine albumin variants.

Plasma samples exhibiting alloalbuminemia on electrophoresis at pH 8.6 were requested from clinical laboratories throughout Sweden. Nine variants, each representing a different single point mutation, were found in 100 apparently unrelated Swedes. The overall prevalence of alloalbuminemia was estimated at 1:1700. Mutations were identified by protein-structural analysis followed by allele-specific DNA hybridization to verify the most common types. Slightly retarded (+1) mobility was seen in 80 cases. Of these, 71 had the Arg(-2)----Cys proalbumin variant previously called Malmö I proalbumin. Thirteen examples of the second most frequent type, the substitution Lys313----Asn and a mobility change of -1 charge unit, were found, as well as six cases of Glu570----Lys (albumin B) and a single case of Arg-1----Gln (proalbumin Christchurch). Five previously unreported types of alloalbuminemia were identified: four instances of Glu376----Gln, which is the second known mutation at this site; two examples of Asp550----Ala, the second mutation reported at this site; and one example each of Asp63----Asn, Gln268----Arg, and Asn318----Lys. Other mutations were identified among eight subjects of foreign descent. The high frequency and relatively uniform geographic distribution of the Arg-2----Cys mutation suggest that it may have occurred in a founder individual many generation ago in Sweden.

Amino Acid Sequence↗
PubMed1992

Endoplasmic reticulum resident protein of 90 kilodaltons associates with the T- and B-cell antigen receptors and major histocompatibility complex antigens during their assembly.

In the endoplasmic reticulum (ER), newly synthesized subunits of the T-cell antigen receptor (TCR), membrane-bound immunoglobulin (mIg), and major histocompatibility complex (MHC) class I antigens must fold correctly and assemble completely into multimeric protein complexes prior to transport to the cell surface. Although folding and assembly may occur spontaneously, the concept that molecular chaperones facilitate these events is emerging. Here, an intracellular protein of 90-kDa apparent molecular mass, denoted IP90, was shown to be an ER resident protein that associated with partial complexes of the TCR, mIg, and MHC class I proteins but was absent from fully assembled complexes. We speculate that IP90 might participate in folding and assembly processes of these and other multisubunit protein complexes during their transit through the ER.

Animals↗
PubMed1992

Cultured microvascular endothelial cells derived from the bovine corpus luteum possess NCAM-140.

Previously, five phenotypically different, stable types of microvascular endothelial cells (MVE) were isolated from the bovine corpus and cultured successfully. We found that three out of these five types of MVE express the neural cell adhesion molecule (NCAM). As shown by immunocytochemistry, weak NCAM immunoreactivity occurred mainly in the perinuclear area of cell type 1. Monolayers of types 2 and 5 revealed heavy NCAM immunoreactivity, which was localized predominantly at the lateral cell surface outlining the contact zones of adjacent cells. In contrast, cell types 3 and 4 were not NCAM immunoreactive. Western blot analyses substantiated these results: While cell type 1 showed a weak immunoreactive band, cell types 2 and 5 displayed strong NCAM-immunoreactive bands of a molecular weight of approximately 140 kDa (NCAM-140), which was absent in cell types 3 and 4. These results reveal for the first time that NCAM can be expressed by cultured MVE and may serve in mediating endothelial cell contacts. Since luteal cells also express NCAM-140, this adhesion molecule could in addition be involved in the interactions of luteal cells with MVE.

Animals↗
PubMed1992

Council on Education report: ART progression and long-range goals.

To answer the question of Accredited Record Technician long-range educational goals (including an indication for progression opportunities), the Council on Education (COE) of AHIMA in 1990 sent out a Request for Proposal for a needs assessment. Based on a proposal submitted in July 1990, a contract was awarded to The College of St. Scholastica. Shirley Eichenwald served as the project director. Kathleen LaTour was project associate and David Swenson was project consultant. A survey was developed and distributed with a deadline of February 1991. The findings from the survey were placed in a report entitled "Progression Potential and Educational Goals of ARTs: a Needs Assessment," dated April 1991, which was submitted to the Council on Education and reviewed at the August 1991 COE meeting. This article summarizes and highlights the key findings from this report.

Attitude of Health Personnel↗
PubMed1992

Development of a program for neonatal intensive care units managed by neonatal nurse practitioners.

Because of reduced medical coverage and changing educational experiences of the pediatric residents, the Department of Nursing Practice at the University of Utah Hospital in collaboration with the Department of Patient Care Services at Primary Children's Medical Center seized the opportunity to develop a nurse-managed patient care service within two tertiary-level nurseries in the State of Utah. Collaboration with nursing administrators and practitioners in both sites yielded joint plans for a successful NNP program. Initial efforts were to fund jointly and support the College of Nursing to develop an educational program, whereas the clinical program was developed before the reduction in medical coverage. Many issues were addressed because of the multidisciplinary and multi-institutional nature of the program. Work is ongoing in the recruitment of students and NNPs, development and review of clinical practice, and endorsement and privileging considerations.

Hospitals, Pediatric↗
PubMed1991

Purification and structural study of two albumin variants in an Irish population.

Two types of variant albumins were detected during routine electrophoresis on cellulose acetate on 34,000 sera from patients in a relatively stable Irish population. The fast type (IRE1) (relative mobility 1.05) had a heterozygote frequency of 1/3,780, and the slow type (IRE2) (relative mobility of 0.94) had a heterozygote frequency of 1/8,500. A method for purification of the two types of variants is described. Structural study of the fast variant established a single amino acid substitution 313 lysine----asparagine (313 Lys----Asn); this variant has been reported in several European populations and also in New Guinea indigenes. However, the slow variant has a new substitution, 479 glutamic acid----lysine (479 Glu----Lys). Because it appears to be uniquely Irish, the slow variant (formerly called IRE2) has been renamed albumin Dublin. Three other albumin variants most often reported in European populations (cumulative frequency only about 1/3,500) were not detected in this study. Because of the significance of albumin genetic variants for the study of protein evolution and as an aid in identification of drug-binding sites, clinical chemists are asked to be on the alert for cases of bisalbuminemia.

Adult↗
PubMed1991

Genetic variants of serum albumin in Americans and Japanese.

A collaborative search for albumin genetic variants (alloalbumins) was undertaken by cellulose acetate and agarose electrophoresis at pH 8.6 of the sera of patients at two major medical centers in the United States and of nearly 20,000 blood donors in Japan. Seventeen instances of alloalbuminemia were ascertained, and seven different alloalbumin types were characterized by structural study. Two previously unreported alloalbumin types were identified. In one type, which was present in a Caucasian family and designated Iowa City-1, aspartic acid at position 365 was replaced by valine (365 Asp----Val); this is the second reported mutation at this position. The other type present in a Japanese blood donor had the mutation 128 His----Arg. An unexpected finding was the presence in a single Japanese of a Naskapi-type alloalbumin (372 Lys----Glu), a variant that had previously been described only for certain Amerindian tribes in whom it occurs with a polymorphic frequency (greater than 1%) and in Eti Turks. An arginyl-albumin (-1 Arg, 1 Asp----Val) occurred in an American family. The other alloalbumin types identified were proalbumins Lille and Christchurch and albumin B that have a cumulative frequency of about 1:3500 in Caucasians probably because of the hypermutability of CpG dinucleotides at the mutated sites. All of the variants characterized in this study are point mutants, and the sites are spread throughout the albumin gene. However, about one-fourth of all known albumin mutations are clustered in the sequence segment from position 354 through 382.

Amino Acid Sequence↗
PubMed1991

A donor splice mutation and a single-base deletion produce two carboxyl-terminal variants of human serum albumin.

At least 35 allelic variants of human serum albumin have been sequenced at the protein level. All except two COOH-terminal variants, Catania and Venezia, are readily explainable as single-point substitutions. The two chain-termination variants are clustered in certain locations in Italy and are found in numerous unrelated individuals. In order to correlate the protein change in these variants with the corresponding DNA mutation, the two variant albumin genes have been cloned, sequenced, and compared to normal albumin genomic DNA. In the Catania variant, a single base deletion and subsequent frameshift leads to a shortened and altered COOH terminus. Albumin Venezia is caused by a mutation that alters the first consensus nucleotide of the 5' donor splice junction of intron 14 and the 3' end of exon 14, which is shortened from 68 to 43 base pairs. This change leads to an exon skipping event resulting in direct splicing of exon 13 to exon 15. The predicted Venezia albumin product has a truncated amino acid sequence (580 residues instead of 585), and the COOH-terminal sequence is altered after Glu-571. The variant COOH terminus ends with the dibasic sequence Arg-Lys that is apparently removed through stepwise cleavage by serum carboxypeptidase B to yield several forms of circulating albumin.

Amino Acid Sequence↗