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David P Witte

Publications and source records attributed to David P Witte.

36 records · Page 2Linked to original sources
PubMed2003

Analyses of temporal regulatory elements of the prosaposin gene in transgenic mice.

The expression of prosaposin is temporally and spatially regulated at transcriptional and post-translational levels. Transgenic mice with various 5'-flanking deletions of the prosaposin promoter fused to luciferase (LUC) reporters were used to define its temporal regulatory region. LUC expression in the transgenic mice carrying constructs with 234 bp (234LUC), 310 bp (310LUC) or 2400 bp (2400LUC) of the 5'-flanking region was analysed in the central nervous system and eye throughout development. For 310LUC and 2400LUC, low-level LUC activity was maintained until embryonal day 18 in brain, eye and spinal cord. The peak level of LUC activity was at birth, with return to a plateau (1/3 of peak) throughout adulthood. Deletion of the region that included the retinoic acid-receptor-related orphan receptor (ROR alpha)-binding site and sequence-specific transcription factor (Sp1) cluster sites (44-310 bp) suppressed the peak of activity. By comparison, the peak level for 234LUC was shifted 2 weeks into neonatal life in the brain, but not in the eye, and no peak of activity was observed in the spinal cord. The endogenous prosaposin mRNA in eye, spinal cord and cerebellum had low-level expression before birth and continued to increase into adulthood. In cerebrum, the endogenous mRNA showed similar expression profile to constructs 310LUC, 2400LUC and 234LUC, with the peak expression at 1 week and a decreased level in adult. In the brain of the newborn, 2400LUC was highly expressed in the trigeminal ganglion and brain stem regions when compared with the generalized expression pattern for endogenous prosaposin mRNA. These results suggest that the modifiers (ROR alpha- and Sp1-binding sites) residing within 310 bp of the 5'-flanking region mediate developmental regulation in the central nervous system and eye. Additional regulatory elements outside the 5' region of the 2400 bp promoter fragment appear to be essential for the physiological control of the prosaposin locus.

5' Flanking Region↗
PubMed2003

Dissection of experimental asthma with DNA microarray analysis identifies arginase in asthma pathogenesis.

Asthma is on the rise despite intense, ongoing research underscoring the need for new scientific inquiry. In an effort to provide unbiased insight into disease pathogenesis, we took an approach involving expression profiling of lung tissue from mice with experimental asthma. Employing asthma models induced by different allergens and protocols, we identified 6.5% of the tested genome whose expression was altered in an asthmatic lung. Notably, two phenotypically similar models of experimental asthma were shown to have distinct transcript profiles. Genes related to metabolism of basic amino acids, specifically the cationic amino acid transporter 2, arginase I, and arginase II, were particularly prominent among the asthma signature genes. In situ hybridization demonstrated marked staining of arginase I, predominantly in submucosal inflammatory lesions. Arginase activity was increased in allergen-challenged lungs, as demonstrated by increased enzyme activity, and increased levels of putrescine, a downstream product. Lung arginase activity and mRNA expression were strongly induced by IL-4 and IL-13, and were differentially dependent on signal transducer and activator of transcription 6. Analysis of patients with asthma supported the importance of this pathway in human disease. Based on the ability of arginase to regulate generation of NO, polyamines, and collagen, these results provide a basis for pharmacologically targeting arginine metabolism in allergic disorders.

Animals↗
PubMed2003

Developmental expression of Magmas in murine tissues and its co-expression with the GM-CSF receptor.

Magmas is a protein that is involved in GM-CSF signaling in a myeloid cell line. Its precise role in the signal transduction process is unclear. To accurately characterize Magmas expression in a variety of cells, mouse embryos and adult murine tissues were analyzed for both mRNA and protein content. Magmas expression was detected as early as the day 6.5 embryo. The level of expression was developmentally regulated. During embryogenesis, elevated Magmas was observed in several structures, including heart, liver, notochord, choroid plexus, cervical ganglion, and nasal mucosa. Muscle, pancreas, intestinal mucosa, and testes were among the adult tissues with high Magmas expression. Most cell types, including hepatocytes and skeletal, smooth, and cardiac myocytes, also expressed the GM-CSF receptor (GMR) but the relative tissue levels of GMR were not always proportional to Magmas. The expression patterns suggest that Magmas has a role in both developing and mature tissues.

Animals↗
PubMed2002

The Na,K-ATPase alpha 2 isoform is expressed in neurons, and its absence disrupts neuronal activity in newborn mice.

Na,K-ATPase is an ion transporter that impacts neural and glial physiology by direct electrogenic activity and the modulation of ion gradients. Its three isoforms in brain have cell-type and development-specific expression patterns. Interestingly, our studies demonstrate that in late gestation, the alpha2 isoform is widely expressed in neurons, unlike in the adult brain, in which alpha2 has been shown to be expressed primarily in astrocytes. This unexpected distribution of alpha2 isoform expression in neurons is interesting in light of our examination of mice lacking the alpha2 isoform which fail to survive after birth. These animals showed no movement; however, defects in gross brain development, muscle contractility, neuromuscular transmission, and lung development were ruled out. Akinesia suggests a primary neuronal defect and electrophysiological recordings in the pre-Bötzinger complex, the brainstem breathing center, showed reduction of respiratory rhythm activity, with less regular and smaller population bursts. These data demonstrate that the Na,K-ATPase alpha2 isoform could be important in the modulation of neuronal activity in the neonate.

Animals↗
PubMed2002

Lysosomal acid lipase deficiency: correction of lipid storage by adenovirus-mediated gene transfer in mice.

Lysosomal acid lipase (LAL) is the essential enzyme for hydrolysis of triglycerides (TGs) and cholesteryl esters (CEs) in lysosomes. Its deficiency produces two human phenotypes: Wolman disease (WD) and cholesteryl ester storage disease (CESD). The LAL null (lal(-/-)) mouse mimicks aspects of human WD and CESD. The potential for gene therapy of LAL deficiency was tested with first-generation adenoviral vectors containing human LAL cDNA (Ad-hLAL) by intravenous injection into lal(-/-) mice. Compared with phosphate-buffered saline-injected controls, the mice receiving Ad-hLAL had increased hepatic LAL activity, decreased hepatomegaly, and normalization of histopathology. hLAL protein and mRNA were detected by immunohistochemical staining and in situ hybridization in hepatic parenchymal and sinusoid lining cells, splenic sinusoidal cells, lung macrophages, and adrenal cortical cells. Mice showed TG reductions in liver, spleen, and small intestine of 68, 54, and 50%, respectively, and cholesterol reductions of 55, 52, and 34%, respectively, at 20 days postinjection. These studies provide the basis for the use of gene therapy, in the form of gene transfer via intravenously administered adenovirus, to correct deficiency states, such as WD and CESD, and histopathology of a variety of tissues.

Adenoviridae↗
PubMed2002

Distinction in genetic determinants for injury-induced neointimal hyperplasia and diet-induced atherosclerosis in inbred mice.

Five inbred strains of mice differing in susceptibility to diet-induced atherosclerosis were compared for neointimal hyperplasia after endothelial denudation with an epoxy resin-modified catheter probe. Results showed that all animals responded similarly to the arterial injury, with increased medial area and thickness after 14 days. In contrast, a significant strain-specific difference in neointimal formation after injury was observed. The atherosclerosis-susceptible C57L/J mice were also susceptible to injury-induced neointimal hyperplasia, and the C3H mice were resistant to both forms of vascular diseases. The 129/Sv mice, which displayed an intermediate level of diet-induced atherosclerosis, also displayed an intermediate level of injury-induced neointimal hyperplasia. Interestingly, the atherosclerosis-susceptible C57BL/6 mice were resistant to neointimal hyperplasia after endothelial denudation, whereas the atherosclerosis-resistant FVB/N mice were susceptible, displaying massive neointimal hyperplasia after arterial injury. All (C57L/JxC57BL/6)F1 hybrid mice were resistant to injury-induced neointimal hyperplasia. Moreover, N2 mice generated from backcrossing the F1 hybrid mice to the susceptible C57L/J mice displayed a range of arterial response to injury, spanning the most severe to the most resistant phenotype. These results indicate that injury-induced neointimal hyperplasia and diet-induced atherosclerosis are controlled by distinct sets of genes; the former appeared to be determined by recessive genes at > or =2 loci.

Animals↗
PubMed2002

DNA microarray analysis reveals novel gene expression profiles in collagen-induced arthritis.

Global gene expression was analyzed in early and late collagen-induced arthritis (CIA). Of 8734 cDNAs analyzed, 330 were induced and 55 downregulated greater than twofold in early or late disease. Hierarchical clustering of these 385 cDNAs demonstrated five distinct expression patterns differentiating early from late disease and correlating with histopathologic changes in the paw. Of the 385 cDNAs, 185 are known, characterized genes, the majority of which are not described as playing a role in arthritis. However, several of these genes are involved in pathological processes relating to arthritis, including apoptosis, inflammation, and cellular proliferation. One interesting gene, follistatin-like gene, is highly expressed along the margin of contact between inflammatory synovial pannus and eroding bone, suggesting a role in joint destruction. These results demonstrate that global gene expression profiles distinguish early and late CIA and reveal several genes novel to arthritis the further characterization of which will advance our understanding of arthritis.

Animals↗
PubMed2002

Targeted inactivation of the mouse guanylin gene results in altered dynamics of colonic epithelial proliferation.

Heat-stable enterotoxin (STa), elaborated by enterotoxigenic Echerichia coli, is a worldwide cause of secretory diarrhea in infants and travelers. Both STa and guanylin, a peptide structurally similar to STa, increase intracellular cGMP levels after binding to the same intestinal receptor, guanylate cyclase C (GC-C). Distinct from its role as an intestinal secretagogue, guanylin may also have a role in intestinal proliferation, as guanylin expression is lost in intestinal adenomas. To determine the function of guanylin in intestinal epithelia, guanylin null mice were generated using a Cre/loxP-based targeting vector. Guanylin null mice grew normally, were fertile and showed no signs of malabsorption. However, the levels of cGMP in colonic mucosa of guanylin null mice were significantly reduced. The colonic epithelial cell migration rate was increased and increased numbers of colonocytes expressing proliferating cell nuclear antigen (PCNA) were present in crypts of guanylin null mice as well. The apoptotic index was similar in guanylin null mice and littermate controls. We conclude from these studies that loss of guanylin results in increased proliferation of colonic epithelia. We speculate that the increase in colonocyte number is related to decreased levels of cGMP and that this increase in proliferation plays a role in susceptibility to intestinal adenoma formation and/or progression.

Animals↗
PubMed2002

Microarray analysis of novel cell lines representing two stages of metanephric mesenchyme differentiation.

Clonal cell lines representing different developmental stages of the metanephric mesenchyme were made from transgenic mice with the Simian Virus 40 T-antigen (SV40 Tag) gene driven by the Hoxa 11 promoter. The resulting mK3 cell line represented early metanephric mesenchyme, prior to induction by the ureteric bud. These cells showed a spindle-shaped, fibroblast morphology. They expressed genes characteristic of early mesenchyme, including Hoxa 11, Hoxd 11, collagen I, and vimentin. Moreover, the mK3 cells displayed early metanephric mesenchyme biological function. In organ co-culture experiments they were able to induce growth and branching of the ureteric bud. Another cell line, mK4, represented later, induced metanephric mesenchyme undergoing epithelial conversion. These cells were more polygonal, or epithelial in shape, and expressed genes diagnostic of late mesenchyme, including Pax-2, Pax-8, Wnt-4, Cadherin-6, Collagen IV, and LFB3. To better define the gene expression patterns of kidney metanephric mesenchyme cells at these two stages of development, RNAs from the mK3 and mK4 cells were hybridized to Affymetrix GeneChip probe arrays. Over 4000 expressed genes were identified and thereby implicated in kidney formation. Comparison of the mK3 and mK4 gene expression profiles revealed 121 genes showing greater than a ten-fold difference in expression level. Several are known to be expressed during metanephric mesenchyme differentiation, but most had not been previously associated with this process. In situ hybridizations were used to confirm that selected novel genes were expressed in the developing kidney.

Animals↗
PubMed2002

Microarray analysis of novel cell lines representing two stages of metanephric mesenchyme differentiation.

Clonal cell lines representing different developmental stages of the metanephric mesenchyme were made from transgenic mice with the Simian Virus 40 T-antigen (SV40 Tag) gene driven by the Hoxa 11 promoter. The resulting mK3 cell line represented early metanephric mesenchyme, prior to induction by the ureteric bud. These cells showed a spindle-shaped, fibroblast morphology. They expressed genes characteristic of early mesenchyme, including Hoxa 11, Hoxd 11, collagen I, and vimentin. Moreover, the mK3 cells displayed early metanephric mesenchyme biological function. In organ co-culture experiments they were able to induce growth and branching of the ureteric bud. Another cell line, mK4, represented later, induced metanephric mesenchyme undergoing epithelial conversion. These cells were more polygonal, or epithelial in shape, and expressed genes diagnostic of late mesenchyme, including Pax-2, Pax-8, Wnt-4, Cadherin-6, Collagen IV, and LFB3. To better define the gene expression patterns of kidney metanephric mesenchyme cells at these two stages of development, RNAs from the mK3 and mK4 cells were hybridized to Affymetrix GeneChip probe arrays. Over 4000 expressed genes were identified and thereby implicated in kidney formation. Comparison of the mK3 and mK4 gene expression profiles revealed 121 genes showing greater than a ten-fold difference in expression level. Several are known to be expressed during metanephric mesenchyme differentiation, but most had not been previously associated with this process. In situ hybridizations were used to confirm that selected novel genes were expressed in the developing kidney.

Animals↗
PubMed2002

Prosaposin: threshold rescue and analysis of the "neuritogenic" region in transgenic mice.

Prosaposin is the precursor of four glycoprotein activators (saposins) for lysosomal hydrolases. Intact prosaposin also has lipid transfer properties in vitro as well as neuritogenic effects ex vivo and in vivo. Such "neuritogenic" effects of saposin C were evaluated in vivo using transgenic mice with prosaposin cDNAs having normal (PS-N) or mutated neuritogenic region. The mutant prosaposin cDNA (PS-CBC) encoded a chimeric saposin C that contained the non-neuritogenic sequence of saposin B, but retained acid beta-glucosidase (GCase) activation effects. When driven by the PGK (3-phosphoglycerate kinase) promoter, transgene expression was highest in the cerebrum for any of the transgenes (range from 15% to 42% of wild-type). Low levels were in visceral tissues. Prosaposin knock-out (PS-/-) mice expressing N or CBC transgenes, even at low levels, had delayed onset of neurologic signs and neuropathology, and significant lengthening of life span (from 1.7- to 7-fold) with age dependent partial correction of GlcCer and LacCer accumulation in the brain. Neuropathologic progression and neuronal glycosphingolipid storage were related directly to the transgene expression levels in the brain. Purkinje cell loss was age dependent. Gross brain and neuronal organizations were indistinguishable in PS-/- mice with or without the various transgenes, albeit the phenotype appeared later in the mice with transgenes. These studies show the degree of neuropathologic manifestations in each transgenic line depended on expression level rather than on the nature of the transgene. These studies also show in vivo localization of the GCase activation region to the carboxy terminal half of saposin C and the lack of a significant gross trophic effect of saposin C on CNS organization in vivo.

Animals↗
PubMed2002

Placental transfer of ganciclovir in a woman with acquired immunodeficiency syndrome and cytomegalovirus disease.

A pregnant woman with AIDS developed cytomegalovirus (CMV) retinitis and pneumonitis, requiring intravenous ganciclovir. At 34 weeks gestation the woman delivered a 1.4-kg girl. Examination of the placenta revealed transplacental passage of CMV. Low concentrations of ganciclovir were detected in the neonate's plasma. The neonate had mild anemia but no other signs of congenital CMV infection.

Acquired Immunodeficiency Syndrome↗
PubMed2002

Apolipoprotein J/clusterin prevents a progressive glomerulopathy of aging.

Apoliprotein J (apoJ)/clusterin has attracted considerable interest based on its inducibility in multiple injury processes and accumulation at sites of remodeling, regression, and degeneration. We therefore sought to investigate apoJ/clusterin's role in kidney aging, as this may reveal the accumulated effects of diminished protection. Aging mice deficient in apoJ/clusterin developed a progressive glomerulopathy characterized by the deposition of immune complexes in the mesangium. Up to 75% of glomeruli in apoJ/clusterin-deficient mice exhibited moderate to severe mesangial lesions by 21 months of age. Wild-type and hemizygous mice exhibited little or no glomerular pathology. In the apoJ/clusterin-deficient mice, immune complexes of immunoglobulin G (IgG), IgM, IgA, and in some cases C1q, C3, and C9 were detectable as early as 4 weeks of age. Electron microscopy revealed the accumulation of electron-dense material in the mesangial matrix and age-dependent formation of intramesangial tubulo-fibrillary structures. Even the most extensively damaged glomeruli showed no evidence of inflammation or necrosis. In young apoJ/clusterin-deficient animals, the development of immune complex lesions was accelerated by unilateral nephrectomy-induced hyperfiltration. Injected immune complexes localized to the mesangium of apoJ/clusterin-deficient but not wild-type mice. These results establish a protective role of apoJ/clusterin against chronic glomerular kidney disease and support the hypothesis that apoJ/clusterin modifies immune complex metabolism and disposal.

Aging↗
PubMed2002

Posterior distal femoral and proximal tibial metaphyseal stripes at MR imaging in children and young adults.

PURPOSE: To determine the frequency and distribution of the hyperintense stripe seen along the posterior surface of distal femoral and proximal tibial metaphyses at magnetic resonance (MR) imaging. MATERIALS AND METHODS: One hundred forty-two MR imaging studies obtained in 139 children and young adults were reviewed. The authors recorded the presence and distribution of posterior distal femoral and proximal tibial metaphyseal stripes. Presence of stripe was correlated with patient age and sex and with patency of the adjacent physis. Fifty-nine studies of adults were reviewed similarly. Two-way analysis of variance was performed to compare mean patient age for sex among four different categories that were based on stripe presence and physeal patency. Orthogonal contrasts were used to determine whether a linear trend across the categories existed. In one cadaveric femur, imaging and histologic analysis were performed. RESULTS: A metaphyseal stripe was seen in all patients with a completely or partially open physis (110 femora, 102 tibiae) and in 56 femora and 60 tibiae in the patients with fused physes. Thirty-five femora and 35 tibiae showed no stripe; all patients were skeletally mature. Correlations between metaphyseal stripe visualization and physeal patency were significant (P <.001). Differences in mean patient age among the four categories were significant for both (femoral and tibial) locations (P <.001), and a linear trend with age (P <.001) was demonstrated. This linear trend was also observed in both sexes (P <.001). Histologic analysis revealed highly vascular loose fibrous tissue. CONCLUSION: A posterior metaphyseal stripe is seen at MR imaging of the skeletally immature knee and likely reflects normal bone growth.

Adolescent↗
PubMed2002

The role of the receptor tyrosine kinase Ron in nickel-induced acute lung injury.

Acute lung injury (ALI), a severe respiratory syndrome, develops in response to numerous insults and responds poorly to therapeutic intervention. Recently, cDNA microarray analyses were performed that indicated several pathogenic responses during nickel-induced ALI, including marked macrophage activation. Macrophage activation is mediated, in part, via the receptor tyrosine kinase Ron. To address the role of Ron in ALI, the response of mice deficient in the cytoplasmic domain of Ron (Ron tk-/-) were assessed in response to nickel exposure. Ron tk-/- mice succumb to nickel-induced ALI earlier, express larger, early increases in interleukin-6, monocyte chemoattractant protein-1, and macrophage inflammatory protein-2, display greater serum nitrite levels, and exhibit earlier onset of pulmonary pathology and augmented pulmonary tyrosine nitrosylation. Increases in cytokine expression and cellular nitration can lead to tissue damage and are consistent with the differences between genotypes in the early onset of pathology and mortality in Ron tk-/- mice. These analyses indicate a role for the tyrosine kinase receptor Ron in ALI.

Androstadienes↗
PubMed2002

Rescue of prothrombin-deficiency by transgene expression in mice.

Prothrombin has diverse biological functions in addition to its well established role in blood coagulation. In order to study these functions in more detail mouse model systems are needed. Since deficiency of prothrombin in mice results in partial embryonic lethality and neonatal death, alternative approaches are required to study the biology of prothrombin in the adult mouse. The liver is the major site of synthesis of prothrombin and therefore liver-specific promoters were used to express prothrombin in transgenic mice. Mice generated from crosses with these transgenic mice and mice hemizygous for the knock-out allele were used to test whether liver-specific expression is sufficient to correct the phenotype of null mice and whether liver-specific expression is sufficient for the development and survival of mice to adulthood. The mouse albumin promoter/enhancer was used initially for transgene expression without success in obtaining transgene positive, endogenous prothrombin null mice. Two lines of transgene positive, endogenous prothrombin deficient mice were obtained using the mouse transthyretin (TTR) promoter/enhancer driving expression of a human prothrombin cDNA. One line was able to rescue both the embryonic and the neonatal lethality while the other line was only able to correct the embryonic lethality. Expression of prothrombin was restricted to the liver and stomach in one line and to the liver, pancreas, stomach and kidney in the other line of mice. Thrombin activity for one line was determined to be at 5-10% of wildtype levels. These mice developed normally and did not have spontaneous bleeding events unless traumatized. Therefore, transgenic expression of human prothrombin is sufficient for the rescue of the lethality found for prothrombin deficiency in mice.

Albumins↗