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Biomedical subjects

M A Walter

Publications and source records attributed to M A Walter.

At least 37 records · Page 2Linked to original sources
PubMed2000

Variation in residual PITX2 activity underlies the phenotypic spectrum of anterior segment developmental disorders.

The autosomal dominant disorders iris hypolasia (IH), iridogoniodysgenesis syndrome (IGDS) and Axenfeld-Rieger syndrome (ARS) are characterized by maldevelopment of the anterior segment of the eye associated with an increased risk of early-onset glaucoma. IH, IGDS and ARS are allelic disorders, as all three can result from mutations of the transcription factor PITX2. IH is the mildest of the three, whereas ARS exhibits the most severe ocular malformations. We hypothesize that varying amounts of residual PITX2 activity could underlie the severity of these phenotypes. Missense mutations of the PITX2 homeodomain identified in IH (Arg46Trp), IGDS (Arg31His) and ARS patients (Leu16Gln; Thr30Pro; Arg53Pro) were introduced into recombinant PITX2 cDNA by site-directed mutagenesis. PITX2 mutant proteins expressed in COS-7 cells were determined to be stable and localized to the nucleus; however, the Arg53Pro ARS mutant also displayed cytoplasmic staining. Our findings are consistent with the possibility of a novel nuclear localization signal (NLS) within helix 3 of the PITX2 homeodomain, homologous to the NLS of the related transcription factor PDX-1. Analysis of the five mutant PITX2 proteins by DNA-binding shifts and transactivation studies demonstrated reduced activity of the IH and IGDS mutant PITX2 proteins, with the IH mutant retaining the most activity in both studies, whereas the ARS mutant PITX2 proteins proved to be non-functional. In addition to providing insight into the etiological mechanism of IH, IGDS and ARS, these results are consistent with the hypothesis that mutant PITX2 proteins that retain partial function result in milder anterior segment aberrations.

Amino Acid Sequence↗
PubMed2000

Axenfeld-Rieger syndrome resulting from mutation of the FKHL7 gene on chromosome 6p25.

Mutations in the forkhead-like 7 (FKHL7) gene have been recently shown to cause juvenile glaucoma and anterior segment anomalies. We report on a three-generation family with Axenfeld-Rieger syndrome (ARS), harboring an alteration in the FKHL7 gene. Genetic linkage analyses excluded the ARS phenotype from chromosomes 4q25 and 13q14, the locations of the PITX2 and RIEG2 loci, respectively. Evidence of linkage was observed with markers at 6p25, near the FKHL7 gene. Direct sequencing of FKHL7 detected a C67T mutation that segregated with the ARS phenotype in this family, but was not detected in over 80 control chromosomes. This mutation is predicted to cause a nonsense mutation of the FKHL7 protein (Gln23Stop) upstream of the forkhead DNA-binding domain, and thus to generate a truncated FKHL7 protein product. This discovery broadly implicates FKHL7 in ocular, craniofacial, dental, and umbilical development.

Adolescent↗
PubMed2000

Isolation of a novel iris-specific and leucine-rich repeat protein (oculoglycan) using differential selection.

PURPOSE: To identify and characterize genes expressed in the iris. METHODS: A human adult iris cDNA library was constructed and subjected to a differential selection screen to identify genes preferentially expressed in iris or trabecular tissue versus those expressed in lymphoblasts. Selected cDNAs were partially sequenced. Novel cDNAs were chosen for further analysis. The cDNAs were localized within chromosomes using a radiation hybrid (RH) mapping panel. The tissue expression profile of each cDNA was found through computer-based searches. One novel cDNA was subjected to 5' rapid amplification of cDNA ends and Northern blot analysis. RESULTS: Of 24 differentially selected clones, 14 cDNAs had homology to known genes, whereas the other 10 were previously uncharacterized cDNA clones. IR185 was one novel iris cDNA identified. Northern blot analysis with IR185 indicated that it is expressed in human fetal liver as a 2.7-kb transcript and in adult iris as a 1.6-kb transcript. Computer-based searches of public databases and reverse transcription-polymerase chain reaction experiments have determined that IR185 is also expressed in retina. RH mapping experiments have localized IR185 to the chromosomal interval 1q31-q32, near the loci for age-related degeneration (1q25-q31) and retinitis pigmentosa 12 (1q31-q32), and IR185 is in the region for posterior column ataxia with retinitis pigmentosa (1q31-q32). It has a 996-bp open reading frame encoding a putative protein with homology to the small leucine-rich proteoglycan (SLRP) family. The IR185 gene has been tentatively named oculoglycan. CONCLUSIONS: Differential selection is a technique that has been useful in identifying genes specific to a variety of tissues. This is the first time this technique has been applied to the iris. Characterizing genes highly or uniquely expressed in the iris can assist in clarifying our understanding of iris function and lead to a better understanding of the molecular pathogenesis of ocular disease. IR185 is a tentative candidate for one eye disorder genetically localized to chromosome 1q31-q32.

Amino Acid Sequence↗
PubMed1999

Roles for the winged helix transcription factors MF1 and MFH1 in cardiovascular development revealed by nonallelic noncomplementation of null alleles.

The murine Mf1 and Mfh1 genes have overlapping patterns of expression in the embryo and encode forkhead/winged helix transcription factors with virtually identical DNA binding domains. Previous studies have shown that Mfh1 null mutants have severe cardiovascular defects, including interruptions and coarctations of the aortic arch and ventricular septal defects (Iida et al., Development 124, 4627-4638, 1997). Here, we show that Mf1(lacZ) homozygous null mutants also have a similar spectrum of cardiovascular abnormalities. Moreover, most embryos doubly heterozygous for Mfh1(tm1) and Mf1(lacZ) die before birth with interruptions and coarctations of the aortic arch, dysgenesis of the aortic and pulmonary valves, ventricular septal defects, and other cardiac anomalies. This nonallelic noncomplementation and the similar patterns of expression of the two genes in the mesenchyme and endothelial cells of the branchial arches, outflow tract, and heart suggest that Mf1 and Mfh1 play interactive roles in the morphogenesis of the cardiovascular system. Implications for the development of human congenital heart defects are discussed.

Alleles↗
PubMed1999

Histopathology and molecular basis of iridogoniodysgenesis syndrome.

Iridogoniodysgenesis is an autosomal dominant disorder in which there are abnormalities in the development of the iris stroma and trabecular meshwork tissues commonly resulting in glaucoma. The unoperated eye from an affected member of a family with iridogoniodysgenesis syndrome (IGDS) was removed shortly after death. Histopathological studies showed an incomplete, normally positioned line of Schwalbe and iris stromal hypoplasia. The molecular basis underlying the disorder is a missense mutation in the RIEG gene at 4q25, mutations of which have been previously shown to cause Axenfeld-Rieger syndrome (ARS). Coupled with another report of a missense mutation of the RIEG gene in a family with IGDS, we suggest that these mutations may interfere less with gene function and thereby may be responsible for a milder phenotype than occurs in the more characteristic ARS.

Congenital Abnormalities↗
PubMed1999

A computer program for conducting incinerator risk assessments.

In 1994, the United States Environmental Protection Agency (USEPA) developed a screening methodology for conducting indirect exposure risk assessments for combustion facilities. The United States Army Center for Health Promotion and Preventive Medicine currently utilizes this methodology in conjunction with other USEPA guidance documents to perform human health risk assessments (HHRAs). The HHRAs require the development of complex human health models using spreadsheet software packages which estimate various media concentrations of contaminants in the environment. Since the quality assurance/quality control procedures associated with verifying the model's results are extremely time consuming, a computer program was developed using Microsoft Excel to minimize the amount of time needed. This discussion describes the 6 steps taken in developing this computer program, which are: (1) understanding the problem; (2) establishing the structure of each table in the spreadsheets; (3) developing an algorithm to solve the problem; (4) writing code; (5) running the program; and (6) testing the results. The automated process of having the computer predict health risk and hazards for each potentially exposed individual saves a tremendous amount of time because each calculated value is placed in the correct spreadsheet cell location. In addition to the time needed to develop human health spreadsheets, this program also minimizes the potential for reducing human error.

Algorithms↗
PubMed1999

Analytical characterization of electrochemical biosensor test strips for measurement of glucose in low-volume interstitial fluid samples.

BACKGROUND: Minimally invasive interstitial fluid (ISF) sampling and glucose measurement technologies were integrated into a hand-held device for diabetic glucose monitoring investigations. METHODS: Conventional electrochemical test strip technology (Bayer Glucometer Elite) was adapted to measure glucose in small (0.5-2.0 microL) samples of ISF. Test strip glucose measurements were performed on a commercial potentiostat and were compared to various reference glucose methodologies (YSI 2300 analyzer, microhexokinase procedure, Bayer Glucometer Elite). Characterizations of the integrated ISF sampling-glucose test strip design included accuracy and precision in various sample media (saline, ISF surrogates, diabetic ISF samples), sample volume dependence, test strip sterilization studies (electron beam, gamma irradiation), and diabetic ISF sampling and glucose measurements. RESULTS: Glucose measurements were free from significant media effects. Sample volume variations (0.6-3.2 microL) revealed only modest dependence of glucose measurement bias on sample volume (-1.5% per microliter). Sterilization treatments had only a minor impact on glucose response and test strip aging and no significant impact on interferent responses of the glucose test strips. Diabetic subject testing under minimum fasting conditions of at least 2 h with integrated ISF sampling and glucose measurement gave low ISF glucose measurement imprecision (CV, 4%) and mean glucose results that were indistinguishable from reference (microhexokinase) ISF glucose measurements and from capillary blood glucose measurements (Glucometer Elite). CONCLUSIONS: Conventional single-use, electrochemical glucose test strip and ISF collection technologies can be readily integrated to provide real-time ISF sampling and glucose measurements for diabetic monitoring applications.

Biosensing Techniques↗
PubMed1998

The forkhead/winged helix gene Mf1 is disrupted in the pleiotropic mouse mutation congenital hydrocephalus.

Mf1 encodes a forkhead/winged helix transcription factor expressed in many embryonic tissues, including prechondrogenic mesenchyme, periocular mesenchyme, meninges, endothelial cells, and kidney. Homozygous null Mf1lacZ mice die at birth with hydrocephalus, eye defects, and multiple skeletal abnormalities identical to those of the classical mutant, congenital hydrocephalus. We show that congenital hydrocephalus involves a point mutation in Mf1, generating a truncated protein lacking the DNA-binding domain. Mesenchyme cells from Mf1lacZ embryos differentiate poorly into cartilage in micromass culture and do not respond to added BMP2 and TGFbeta1. The differentiation of arachnoid cells in the mutant meninges is also abnormal. The human Mf1 homolog FREAC3 is a candidate gene for ocular dysgenesis and glaucoma mapping to chromosome 6p25-pter, and deletions of this region are associated with multiple developmental disorders, including hydrocephaly and eye defects.

Amino Acid Sequence↗
PubMed1998

Chromosomal localization and genomic organization of the human retinal rod Na-Ca+K exchanger.

The retinal rod Na-Ca+K exchanger is a unique calcium extrusion protein found only in the outer segments of retinal rod photoreceptors. Rod Na-Ca+K exchanger cDNA (NCKX1) has been cloned from bovine and human retinas. Here, we have used fluorescent in situ hybridization and radiation hybrid mapping to localize the human NCKX1 gene to chromosome 15q22. We have determined the genomic organization of human rod NCKX1 and found one intron in the 5' untranslated region and eight introns within the coding region.

Amino Acid Sequence↗
PubMed1998

Mutations of the forkhead/winged-helix gene, FKHL7, in patients with Axenfeld-Rieger anomaly.

Genetic linkage, genome mismatch scanning, and analysis of patients with alterations of chromosome 6 have indicated that a major locus for development of the anterior segment of the eye, IRID1, is located at 6p25. Abnormalities of this locus lead to glaucoma. FKHL7 (also called "FREAC3"), a member of the forkhead/winged-helix transcription-factor family, has also been mapped to 6p25. DNA sequencing of FKHL7 in five IRID1 families and 16 sporadic patients with anterior-segment defects revealed three mutations: a 10-bp deletion predicted to cause a frameshift and premature protein truncation prior to the FKHL7 forkhead DNA-binding domain, as well as two missense mutations of conserved amino acids within the FKHL7 forkhead domain. Mf1, the murine homologue of FKHL7, is expressed in the developing brain, skeletal system, and eye, consistent with FKHL7 having a role in ocular development. However, mutational screening and genetic-linkage analyses excluded FKHL7 from underlying the anterior-segment disorders in two IRID1 families with linkage to 6p25. Our findings demonstrate that, although mutations of FKHL7 result in anterior-segment defects and glaucoma in some patients, it is probable that at least one more locus involved in the regulation of eye development is also located at 6p25.

Amino Acid Sequence↗
PubMed1998

Mutation in the RIEG1 gene in patients with iridogoniodysgenesis syndrome.

Axenfeld-Rieger syndrome (ARS) and iridogoniodysgenesis syndrome (IGDS) are clinically related autosomal dominant disorders which affect the anterior segment of the eye as well as non-ocular structures. ARS patients present with iris hypoplasia, a prominent Schwalbe line, adhesions between the iris stroma and the iridocorneal angle and increased intraocular pressure. IGDS is characterized by iris hypoplasia, goniodysgenesis and increased intraocular pressure. Each syndrome also presents with non-ocular features including maxillary hypoplasia, micro and anodontia, redundant periumbilical skin, hypospadius (in males), and each has been genetically linked to chromosome 4q25. RIEG1 , the gene responsible for the 4q25 ARS phenotype, recently has been cloned. RIEG1 encodes a novel member of the bicoid class of homeobox proteins known to be active as transcription factors. Mutational analysis has previously detected several mutations in this gene in ARS individuals. We have now detected a mutation in RIEG1 which segregates with the disease phenotype in a family with IGDS. This mutation is a G-->A transition altering an arginine residue to a histidine in a highly conserved location in the second helix of the homeobox of RIEG1. This mutation indicates that IGDS and ARS are allelic variants of the same disorder. This wide variability in clinical consequences of mutations at the RIEG1 4q25 locus implicates the RIEG gene broadly in ocular and craniofacial disorders.

Adolescent↗
PubMed1997

Identification of the human chromosomal region containing the iridogoniodysgenesis anomaly locus by genomic-mismatch scanning.

Genome-mismatch scanning (GMS) is a new method of linkage analysis that rapidly isolates regions of identity between two genomes. DNA molecules from regions of identity by descent from two relatives are isolated based on their ability to form extended mismatch-free heteroduplexes. We have applied this rapid technology to identify the chromosomal region shared by two fifth-degree cousins with autosomal dominant iridogoniodysgenesis anomaly (IGDA), a rare ocular neurocristopathy. Markers on the short arm of human chromosome 6p were recovered, consistent with the results of conventional linkage analysis conducted in parallel, indicating linkage of IGDA to 6p25. Control markers tested on a second human chromosome were not recovered. A GMS error rate of approximately 11% was observed, well within an acceptable range for a rapid, first screening approach, especially since GMS results would be confirmed by family analysis with selected markers from the putative region of identity by descent. These results demonstrate not only the value of this technique in the rapid mapping of human genetic traits, but the first application of GMS to a multicellular organism.

Chromosome Mapping↗
PubMed1996

A high-resolution whole genome radiation hybrid map of human chromosome 17q22-q25.3 across the genes for GH and TK.

We have constructed a whole genome radiation hybrid (WG-RH) map across a region of human chromosome 17q, from growth hormone (GH) to thymidine kinase (TK). A panel of 128 WG-RH hybrid cell lines generated by X-irradiation and fusion has been tested for the retention of 39 sequence-tagged site (STS) markers by the polymerase chain reaction. This genome mapping technique has allowed the integration of existing VNTR and microsatellite markers with additional new markers and existing STS markers previously mapped to this region by other means. The WG-RH map includes eight expressed sequence tag (EST) and three anonymous markers developed for this study, together with 23 anonymous microsatellites and five existing ESTs. Analysis of these data resulted in a high-density comprehensive map across this region of the genome. A subset of these markers has been used to produce a framework map consisting of 20 loci ordered with odds greater than 1000:1. The markers are of sufficient density to build a YAC contig across this region based on marker content. We have developed sequence tags for both ends of a 2.1-Mb YAC and mapped these using the WG-RH panel, allowing a direct comparison of cRay6000 to physical distance.

Animals↗
PubMed1996

Fibroblast growth factor-induced motor end plate regeneration in atrophic muscle.

OBJECTIVE: To determine whether fibroblast growth factor 1 implanted with viable nerve into atrophic muscle will stimulate formation of functional, acetylcholine producing motor end plates. DESIGN: Twelve male Lewis rats underwent predenervation of the hamstring muscle 8 weeks before implantation of the nerve at a site distant from the original motor end plate. Six animals underwent implantation of the tagged nerve ending into atrophic muscle with 50 microgram of fibroblast growth factor 1 in a fibrin adhesive carrier (group 1). Three animals underwent implantation with nerve, fibrin adhesive, and no fibroblast growth factor 1 (group 2); and three animals underwent implantation with fibroblast growth factor 1 and fibrin adhesive with no nerve (group 3). Animals were killed 9 weeks after implantation and nerve and muscle specimens were harvested. MAIN OUTCOME MEASURES: Histoenzymologic methods for acetylcholinesterase and silver impregnation of nerve fibers were performed 9 weeks after fibroblast growth factor 1-fibrin adhesive implantation. Variables included the number of motor end plates per highpower field and motor end plate length. RESULTS: Robust axonal sprouting and formation of multiple motor end plates were found arborized in serial fashion equidistant around the implanted nerve ending. Rare extrasynaptic staining occurred. End plate lengths were significantly shorter in the fibroblast growth factor 1-treated muscles (group 1) than in the specimens without fibroblast growth factor 1 (group 2) (31.2 vs 58.5 micron; P>001, paired t test). The arborization of motor end plates, rare extrasynaptic staining, and shorter end plate lengths seen in group 1 were all consistent with mature motor end plates. Controls (group 3) displayed limited motor end plate formation and extensive extrasynaptic staining typical of denervation. CONCLUSION: This study presents encouraging evidence that fibroblast growth factor 1 with fibrin adhesive carrier can facilitate the reinnervation of atrophied muscle by enhancing the formation or revitalization of motor end plates. Future studies will address muscle function and use of different carrier materials.

Animals↗
PubMed1996

Autosomal-dominant iridogoniodysgenesis and Axenfeld-Rieger syndrome are genetically distinct.

PURPOSE: To determine whether there is a locus for iridogoniodysgenesis (IGD)/ familial iris hypoplasia in the region of the known Axenfeld-Rieger syndrome (ARS) locus at 4q25 and to determine the ocular phenotype within the autosomal-dominant iris hypoplasia group of disorders. METHODS: Clinical examinations were performed on 27 members, with 11 affected from one family in which the IGD occurred in association with the nonocular features of ARS, and on 70 members with 30 affected from a second IGD family with ocular features only. Family members were genotyped for markers within the 4q25 region known to contain a locus for ARS. LOD scores were calculated with the MLINK option of the LINKAGE program. RESULTS: The iris hypoplasia in each IGD family was similar. In the IGD family with only ocular features (IGD anomaly), however, a majority of those affected had a goniodysgenesis with excess tissue in the angle and anomalous angle vascularity. These findings were absent in the IGD family with syndromic features (IGD syndrome). Linkage to the 4q25 region was excluded in the IGD anomaly family, whereas the family with IGD syndrome was found to be completely linked to the 4q25 region (peak LOD score with D4S407 of 7.827 at theta = 0.00). CONCLUSIONS: The authors' results suggest that mutations at the 4q25 locus can result in variable ocular features that also occur in combination with nonocular (dental and jaw) anomalies. Mutation of a separate locus must underlie IGD with ocular features only. A re-evaluation of the relation between the various forms of autosomal-dominant iris hypoplasia, therefore, may be warranted.

Abnormalities, Multiple↗
PubMed1996

A high-resolution radiation hybrid map of the region surrounding the Gorlin syndrome gene.

We have constructed a set of hybrid cell lines by irradiation of GM 64063 (human chromosome 9q only on hamster background) and fusion to hamster A23 Tk-, 109 independent lines were tested by PCR with 24 markers from chromosome 9q. The marker density is highest in the 9q22.3-q31 region containing the gene for Gorlin syndrome, a familial predisposition to basal cell carcinoma. The resolution of our map in this region is significantly higher than other published maps and will enable accurate placing of new markers and genes within the 9q22.3-q3.1 region. This is important since yeast artificial chromosomes from the region are likely to contain deletions.

Animals↗