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Biomedical subjects

L Pritchard

Publications and source records attributed to L Pritchard.

15 recordsLinked to original sources
PubMed2005

Autoimmunity to glutamic acid decarboxylase in the neurodegenerative disorder Batten disease.

The pathogenic mechanisms underlying Batten disease are unclear. Patients uniformly possess autoantibodies against glutamic acid decarboxylase (GAD) that are predominantly reactive with a region of GAD (amino acids 1 to 20) distinct from subjects with autoimmune type 1 diabetes or stiff-person syndrome. Batten patients did not possess autoantibodies against other type 1 diabetes-associated autoantigens and human leukocyte antigen genotypes revealed no specific associations with this disease.

Adolescent↗
PubMed2004

Genome sequence of the enterobacterial phytopathogen Erwinia carotovora subsp. atroseptica and characterization of virulence factors.

The bacterial family Enterobacteriaceae is notable for its well studied human pathogens, including Salmonella, Yersinia, Shigella, and Escherichia spp. However, it also contains several plant pathogens. We report the genome sequence of a plant pathogenic enterobacterium, Erwinia carotovora subsp. atroseptica (Eca) strain SCRI1043, the causative agent of soft rot and blackleg potato diseases. Approximately 33% of Eca genes are not shared with sequenced enterobacterial human pathogens, including some predicted to facilitate unexpected metabolic traits, such as nitrogen fixation and opine catabolism. This proportion of genes also contains an overrepresentation of pathogenicity determinants, including possible horizontally acquired gene clusters for putative type IV secretion and polyketide phytotoxin synthesis. To investigate whether these gene clusters play a role in the disease process, an arrayed set of insertional mutants was generated, and mutations were identified. Plant bioassays showed that these mutants were significantly reduced in virulence, demonstrating both the presence of novel pathogenicity determinants in Eca, and the impact of functional genomics in expanding our understanding of phytopathogenicity in the Enterobacteriaceae.

Base Sequence↗
PubMed2001

Comparison of Pkd1-targeted mutants reveals that loss of polycystin-1 causes cystogenesis and bone defects.

A high level of polycystin-1 expression is detected in kidneys of all patients with autosomal dominant polycystic kidney disease (ADPKD). Mice that overexpress polycystin-1 also develop renal cysts. Whether overexpression of polycystin-1 is necessary for cyst formation is still unclear. Here, we report the generation of a targeted mouse mutant with a null mutation in Pkd1 and its phenotypic characterization in comparison with the del34 mutants that carry a 'truncation mutation' in Pkd1. We show that null homozygotes develop the same, but more aggressive, renal and pancreatic cystic disease as del34/del34. Moreover, we report that both homozygous mutants develop polyhydramnios, hydrops fetalis, spina bifida occulta and osteochondrodysplasia. Heterozygotes also develop adult-onset pancreatic disease. We show further that del34 homozygotes continue to produce mutant polycystin-1, thereby providing a possible explanation for increased immunoreactive polycystin-1 in ADPKD cyst epithelia in the context of the two-hit model. Our data demonstrate for the first time that loss of polycystin-1 leads to cyst formation and defective skeletogenesis, and indicate that polycystin-1 is critical in both epithelium and chondrocyte development.

Animals↗
PubMed2001

Evaluation of a novel method for the identification of coevolving protein residues.

A novel method for the identification of correlated pairs in aligned homologous protein sequences is presented and evaluated against a model of simulated protein evolution incorporating covariation. Our method is shown to be capable of identifying all coevolutionary pairs of sites, with minimal interference by background correlations, in aligned sequence sets containing approximately 60 sequences with a tree depth of at least 30 accepted point mutations. This result is expected even in the presence of a large degree of neutral and non-correlated evolution. It is postulated that, since naturally occurring protein families may be subject to stronger selection pressures and a lesser degree of neutral evolution, this method of covariation analysis may be generally more robust than the model would indicate.

Amino Acid Sequence↗
PubMed2000

A human PKD1 transgene generates functional polycystin-1 in mice and is associated with a cystic phenotype.

Three founder transgenic mice were generated with a 108 kb human genomic fragment containing the entire autosomal dominant polycystic kidney disease (ADPKD) gene, PKD1, plus the tuberous sclerosis gene, TSC2. Two lines were established (TPK1 and TPK3) each with approximately 30 copies of the transgene. Both lines produced full-length PKD1 mRNA and polycystin-1 protein that was developmentally regulated, similar to the endogenous pattern, with expression during renal embryogenesis and neonatal life, markedly reduced at the conclusion of renal development. Tuberin expression was limited to the brain. Transgenic animals from both lines (and the TPK2 founder animal) often displayed a renal cystic phenotype, typically consisting of multiple microcysts, mainly of glomerular origin. Hepatic cysts and bile duct proliferation, characteristic of ADPKD, were also seen. All animals with two copies of the transgenic chromosome developed cysts and, in total, 48 of the 100 transgenic animals displayed a cystic phenotype. To test the functionality of the transgene, animals were bred with the Pkd1(del34) knockout mouse. Both transgenic lines rescued the embryonically lethal Pkd1(del34/del34) phenotype, demonstrating that human polycystin-1 can complement for loss of the endogenous protein. The rescued animals were viable into adulthood, although more than half developed hepatic cystic disease in later life, similar to the phenotype of older Pkd1(del34/+) animals. The TPK mice have defined a minimal area that appropriately expresses human PKD1. Furthermore, this model indicates that over-expression of normal PKD1 can elicit a disease phenotype, suggesting that the level of polycystin-1 expression may be relevant in the human disease.

Animals↗
PubMed2000

Do proteins learn to evolve? The Hopfield network as a basis for the understanding of protein evolution.

Correlations between amino-acid residues can be observed in sets of aligned protein sequences, and the analysis of their statistical and evolutionary significance and distribution has been thoroughly investigated. In this paper, we present a model based on such covariations in protein sequences in which the pairs of residues that have mutual influence combine to produce a system analogous to a Hopfield neural network. The emergent properties of such a network, such as soft failure and the connection between network architecture and stored memory, have close parallels in known proteins. This model suggests that an explanation for observed characters of proteins such as the diminution of function by substitutions distant from the active site, the existence of protein folds (superfolds) that can perform several functions based on one architecture, and structural and functional resilience to destabilizing substitutions might derive from their inherent network-like structure. This model may also provide a basis for mapping the relationship between structure, function and evolutionary history of a protein family, and thus be a powerful tool for rational engineering.

Animals↗
PubMed1999

Evolutionary trace analysis of the Kunitz/BPTI family of proteins: functional divergence may have been based on conformational adjustment.

The structural and functional evolution of the Kunitz/bovine pancreatic trypsin inhibitor (BPTI) family of proteins, which includes serine proteinase inhibitors and potassium channel blockers, was analysed with the evolutionary trace method. This method highlights sites in aligned primary sequences whose side-chain variation can be strongly linked with the past development of different functional classes or subgroups within the family. A total of 16 such "class-specific" positions distributed throughout the molecular fold were identified. On average, the side-chain chemistry at these positions had been more conserved and made greater contribution to molecular stability than the side-chain chemistry at remaining sites of variation. It was possible to use these 16 positions to describe the division of the Kunitz/BPTI family into general functional classes. According to known complexes of inhibitor variants with serine proteinases, only two of the 16 class-specific positions appear to be directly involved in intermolecular recognition via the "antiproteinase site". Instead, from various critical locations in the fold, the remainder seem to have been associated with various degrees of intramolecular conformational adjustment to the underlying framework of the antiproteinase site. It is, therefore, implied that functional diversification in this family has been founded upon both sustained evolutionary selection and conformational adjustment. The findings are important for protein engineers wishing to alter the binding selectivity of these molecules, because it appears that the issue of target recognition is dependent on the conformation of the chain segment to which the interactive side-chains are attached. To retarget members of this family towards potentially novel peptide binding sites, substitutions at certain structurally significant class-specific positions could be a good starting point.

Amino Acid Sequence↗
PubMed1999

Polycystin-1 expression in PKD1, early-onset PKD1, and TSC2/PKD1 cystic tissue.

BACKGROUND: The mutational mechanism responsible for cyst formation in polycystic kidney disease 1 gene (PKD1) remains controversial, with data indicating a two-hit mechanism, but also evidence of polycystin-1 expression in cystic tissue. METHODS: To investigate this apparent paradox, we analyzed polycystin-1 expression in cystic renal or liver tissue from 10 patients with truncating PKD1 mutations (including one early-onset case) and 2 patients with severe disease associated with contiguous deletions of TSC2 and PKD1, using monoclonal antibodies (mAbs) to both extreme N-(7e12) and C-terminal (PKS-A) regions of the protein. Truncation of the C-terminal epitope from the putative mutant proteins in each case allowed exclusive assessment of the nontruncated protein with PKS-A. RESULTS: In adult PKD1 tissue, the majority of cysts (approximately 80%) showed polycystin-1 expression, although staining was absent in a variable but significant minority (approximately 20%), in spite of the normal expression of marker proteins. Unlike adult PKD1, however, negative cysts were rarely found in infantile PKD1 or TSC2/PKD1 deletion cases. CONCLUSIONS: If a two-hit mutational mechanism is operational, these results suggest that the majority of somatic mutations in adult PKD1 are likely to be missense changes. The low level of polycystin-1-negative cysts in the three "early-onset" cases, however, suggests that a somatic PKD1 mutation may not always be required for cyst formation.

Adult↗
PubMed1995

OBRA '90: the impact on pharmacists and patients.

Two separate studies were conducted in Georgia for this investigation. The first study measured by observation, pharmacists' compliance with the mandates of the Omnibus Budget Reconciliation Act of 1990 (OBRA '90) before (December 1992) and after (June 1993) the law took effect in January 1993. The second assessed patient awareness, experience, and degree of cooperation with pharmacists in complying with OBRA '90. Eight chain and four independent pharmacies were recruited for the first study. For the second, a representative sampling of patients answered questions by telephone. Findings from the pharmacy study indicated more streamlined information gathering, more time devoted to prospective drug utilization review, and a 29% increase in counseling frequency in the post-OBRA '90 period. Some changes may be due to increased computer sophistication. The patient survey indicated a lower frequency of counseling than was reported in the pharmacy study. Patients appeared willing to supply information to pharmacists when patients viewed it as a way to improve the care they receive.

Georgia↗
PubMed1979

Histopathologic sequence of events in adult mice undergoing lethal graft-versus-host reaction developed across H-2 and/or non-H-2 histocompatibility barriers.

The sequence of histologic events in graft-versus-host reaction (GVHR) caused by major and/or minor histoincompatibilities was studied. It was discovered that GVHR may manifest itself in the form of two distinct multiphasic disease entities, depending on whether the donor cells are incompatible with the host for both major and minor histocompatibility antigens ("major GVHR") or for minor histocompatibility antigens alone ("minor GVHR"). The acute or major GVHR has four phases: 1) a transient phase of aplasia, 2) a repopulation phase, 3) a proliferative phase involving lymphoid, presumably immunocompetent, cells, and 4) a phase of acute organ rejection (terminal). The chronic or minor GVHR is characterized by six phases, namely: 1) a transient phase of aplasia, 2) a repopulation phase, 3) a phase of proliferation and tissue infiltration by lymphoid, presumably immunocompetent cells, 4) a phase of major immunologic injuries, 5) a phase of repair, and 6)a terminal phase with advanced sclerosis and proliferative glomerulonephritis. In acute or major GVHR the disease was manifested by the tissue reactions characteristic of acute organ rejection. Lesions were seen in the kidney, liver, bone marrow, lymph nodes, spleen, thymus, intestine, and skin. In the chronic or minor GVHR, tissue injuries were more widespread, affecting the collagen, vessel walls, adipose tissue, renal glomeruli, heart muscle, fascias of skeletal muscles, lymph nodes, spleen, thymus, bone marrow, intestine, skin, esophageal mucosa, and urinary tract. A pronounced plasma cell proliferation was a striking feature in the minor GVHR. Its evolution coincided with advanced thymic epithelial atrophy. It is suggested that the destruction of thymic epithelium resulted in depletion of suppressor T cells and, consequently, in an unopposed proliferation of plasma cells.

Animals↗