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Biomedical subjects

M A Goldberg

Publications and source records attributed to M A Goldberg.

At least 19 recordsLinked to original sources
PubMed2001

Identification and characterization of a low oxygen response element involved in the hypoxic induction of a family of Saccharomyces cerevisiae genes. Implications for the conservation of oxygen sensing in eukaryotes.

An organism's ability to respond to changes in oxygen tension depends in large part on alterations in gene expression. The oxygen sensing and signaling mechanisms in eukaryotic cells are not fully understood. To further define these processes, we have studied the Delta9 fatty acid desaturase gene OLE1 in Saccharomyces cerevisiae. We have confirmed previous data showing that the expression of OLE1 mRNA is increased in hypoxia and in the presence of certain transition metals. OLE1 expression was also increased in the presence of the iron chelator 1,10-phenanthroline. A 142-base pair (bp) region 3' to the previously identified fatty acid response element was identified as critical for the induction of OLE1 in response to these stimuli using OLE1 promoter-lacZ reporter constructs. Electromobility shift assays confirmed the presence of an inducible band shift in response to hypoxia and cobalt. Mutational analysis defined the nonameric sequence ACTCAACAA as necessary for transactivation. A 20-base pair oligonucleotide containing this nonamer confers up-regulation by hypoxia and inhibition by unsaturated fatty acids when placed upstream of a heterologous promoter in a lacZ reporter construct. Additional yeast genes were identified which respond to hypoxia and cobalt in a manner similar to OLE1. A number of mammalian genes are also up-regulated by hypoxia, cobalt, nickel, and iron chelators. Hence, the identification of a family of yeast genes regulated in a similar manner has implications for understanding oxygen sensing and signaling in eukaryotes.

Base Sequence↗
PubMed2001

MGA2 is involved in the low-oxygen response element-dependent hypoxic induction of genes in Saccharomyces cerevisiae.

Eukaryotes have the ability to respond to changes in oxygen tension by alterations in gene expression. For example, OLE1 expression in Saccharomyces cerevisiae is upregulated under hypoxic conditions. Previous studies have suggested that the pathway regulating OLE1 expression by unsaturated fatty acids may involve Mga2p and Spt23p, two structurally and functionally related proteins. To define the possible roles of each of these genes on hypoxia-induced OLE1 expression, we examined OLE1 expression under normoxia, hypoxia, and cobalt treatment conditions in Deltamga2 or Deltaspt23 deletion strains. The results of OLE1 promoter-lacZ reporter gene and Northern blot analyses showed that hypoxia- and cobalt-induced OLE1 expression was dramatically decreased in a Deltamga2 strain but not in a Deltaspt23 strain. Further analyses using low-oxygen response element (LORE)-CYC1-lacZ fusion reporter assays and electrophoretic mobility shift assays (EMSAs) demonstrated that MGA2 significantly affects the LORE-dependent hypoxic induction pathway of gene expression. When MGA2 was supplied by a plasmid, the LORE-dependent hypoxia-inducible reporter expression was recovered, as was the hypoxia-inducible complex in EMSAs in the S. cerevisiae Deltamga2 strain. Supershift analysis of EMSAs using crude extracts containing mycMga2p indicated that Mga2p is a component of the LORE-binding complex. Another LORE-dependent, hypoxia-inducible gene, ATF1, was similarly affected in the Deltamga2 strain. These results indicate that MGA2 is required for the LORE-dependent hypoxic gene induction in S. cerevisiae.

Cell Hypoxia↗
PubMed2000

Angiogenesis is induced in a rabbit model of hindlimb ischemia by naked DNA encoding an HIF-1alpha/VP16 hybrid transcription factor.

BACKGROUND: Hypoxia-inducible factor-1 (HIF-1) is a heterodimeric transcription factor that regulates expression of genes involved in O(2) homeostasis, including vascular endothelial growth factor (VEGF), a potent stimulator of angiogenesis. We sought to exploit this native adaptive response to hypoxia as a treatment for chronic ischemia. METHODS AND RESULTS: A hybrid protein consisting of DNA-binding and dimerization domains from the HIF-1alpha subunit and the transactivation domain from herpes simplex virus VP16 protein was constructed to create a strong, constitutive transcriptional activator. After transfection into HeLa, C6, and Hep3B cells, this chimeric transcription factor was shown to activate expression of the endogenous VEGF gene, as well as several other HIF-1 target genes in vitro. The bioactivity of HIF-1alpha/VP16 hybrid gene transfer in vivo was examined in a rabbit model of hindlimb ischemia. Administration of HIF-1alpha/VP16 was associated with significant improvements in calf blood pressure ratio, angiographic score, resting and maximal regional blood flow, and capillary density (all P:<0.01). CONCLUSIONS: The HIF-1alpha/VP16 hybrid transcription factor is able to promote significant improvement in perfusion of an ischemic limb. These results confirm the feasibility of a novel approach for therapeutic angiogenesis in which neovascularization may be achieved indirectly by use of a transcriptional regulatory strategy.

Angiography↗
PubMed2000

Effect of topical povidone-iodine versus topical ofloxacin on experimental Staphylococcus keratitis.

PURPOSE: To compare the antibacterial effect of povidone-iodine (PI) with that of ofloxacin in an experimental model of bacterial keratitis. METHODS: Staphyloccocal keratitis was induced in 21 eyes of Dutch Belted rabbits by intrastromal inoculation of approximately 280 organisms of Staphylococcus aureus. Six hours later, the animals were divided in four groups treated topically with saline 0.9%, Betadine 10%, Betadine 0.5% or Ofloxacin 0.3% (2 gtt every 30 min for 8 h). The central 8-mm cornea was excised, washed and homogenized. Colony counts were performed on serial 10-fold dilutions plated on blood and brain infusion agar and incubated overnight. RESULTS: Colony-forming units per cornea were 7.4x10(7) for the saline group compared to 8.2x10(7) for PI 10% (P>0.5), 4.3x10(7) for PI 0.5% (P<0.01) and no organisms for ofloxacin 0.3%. CONCLUSIONS: Betadine 0.5% demonstrates a statistically significant bactericidal effect compared with untreated staphyloccocal keratitis in our experimental model. Ofloxacin has superior antibacterial effect under the conditions studied. Further improvements in the povidone-iodine formulation are warranted prior to consideration for human keratitis.

Animals↗
PubMed1999

Inhibition of hypoxia-inducible factor 1 activation by carbon monoxide and nitric oxide. Implications for oxygen sensing and signaling.

It has been proposed that cells sense hypoxia by a heme protein, which transmits a signal that activates the heterodimeric transcription factor hypoxia-inducible factor 1 (HIF-1), thereby inducing a number of physiologically relevant genes such as erythropoietin (Epo). We have investigated the mechanism by which two heme-binding ligands, carbon monoxide and nitric oxide, affect oxygen sensing and signaling. Two concentrations of CO (10 and 80%) suppressed the activation of HIF-1 and induction of Epo mRNA by hypoxia in a dose-dependent manner. In contrast, CO had no effect on the induction of HIF-1 activity and Epo expression by either cobalt chloride or the iron chelator desferrioxamine. The affinity of CO for the putative sensor was much lower than that of oxygen (Haldane coefficient, approximately 0.5). Parallel experiments were done with 100 microM sodium nitroprusside, a nitric oxide donor. Both NO and CO inhibited HIF-1 DNA binding by abrogating hypoxia-induced accumulation of HIF-1alpha protein. Moreover, both NO and CO specifically targeted the internal oxygen-dependent degradation domain of HIF-1alpha, and also repressed the C-terminal transactivation domain of HIF-1alpha. Thus, NO and CO act proximally, presumably as heme ligands binding to the oxygen sensor, whereas desferrioxamine and perhaps cobalt appear to act at a site downstream.

Aryl Hydrocarbon Receptor Nuclear Translocator↗
PubMed1998

Cerebral toxoplasmosis and lymphoma in AIDS: perfusion MR imaging experience in 13 patients.

PURPOSE: To evaluate the perfusion magnetic resonance (MR) imaging characteristics of cerebral toxoplasmosis and lymphoma in patients with acquired immunodeficiency syndrome (AIDS). MATERIALS AND METHODS: Perfusion MR imaging was performed prospectively in 13 patients with AIDS who had contrast material-enhancing focal brain lesions (six with active lymphoma, five with toxoplasmosis, one with treated lymphoma in remission, and one with toxoplasmosis plus lymphomatoid granulomatosis). Regional cerebral blood volume (rCBV) was determined by using dynamic echo-planar MR imaging during bolus injection of a gadolinium chelate. RESULTS: The rCBV was decreased (44% +/- 24 [standard deviation] of rCBV in the contralateral regions) throughout the toxoplasmosis lesions and in the surrounding edema of both lesion types, whereas all active lymphomas displayed areas of increased rCBV (258% +/- 99). These differences were significant (P < .005). CONCLUSION: Reduced rCBV i toxoplasmosis lesions is probably due to a lack of vasculature within the abscess; increased rCBV in lymphomas is probably due to hypervascularity in foci of active tumor growth; and decreased rCBV in the edema is probably due to vasoconstriction associated with increased interstitial pressure. Perfusion MR imaging is a rapid, noninvasive tool that may allow differentiation between cerebral lymphoma and toxoplasmosis in patients with AIDS.

AIDS-Related Opportunistic Infections↗
PubMed1997

Sequencing of the human vascular endothelial growth factor (VEGF) 3' untranslated region (UTR): conservation of five hypoxia-inducible RNA-protein binding sites.

Vascular endothelial growth factor (VEGF) is a potent angiogenic factor whose mRNA expression is induced by hypoxia. This induction is due in large part to an increase in the stability of its mRNA. The RNA sequences and cognate proteins responsible for this increased stability with hypoxia are not well understood. In order to identify regions of functional importance in the 3'UTR of VEGF mRNA, we have sequenced the human VEGF 3'UTR and compared it to the rat sequence. Overall sequence homology was 82% with complete conservation of all four potential polyadenylation signals and both nonameric instability elements. Five hypoxia-inducible RNA protein-binding (HI-RPB) sites were identified by RNA electromobility shift assay (EMSA) in the human and rat genes. EMSA and competition studies suggest that these sites bind a similar or related protein complex. On average, the five sites were 95% conserved at the nucleotide level between the rat and corresponding human sequence. This conservation taken together with several previously described, independent correlations between the presence of these RNA-protein complexes and an increase in VEGF mRNA stability suggest an important functional role for these sites in mediating hypoxia-inducible VEGF mRNA stability.

Animals↗
PubMed1997

Coexistent Acanthamoeba keratitis and herpetic keratitis.

OBJECTIVE: To describe a series of patients with proved herpes simplex virus keratitis (herpetic keratitis) who also had documented Acanthamoeba keratitis. METHODS: Herpetic keratitis was documented with viral cultures, immunologic stains, or histopathologic examination for multinucleated giant cells in the corneal epithelium. Acanthamoeba organisms were identified using confocal microscopy and epithelial biopsy with hematoxylin-eosin staining. Biopsy of the stroma and epithelium was used to identify Acanthamoeba organisms in 1 case. RESULTS: Cultures for herpes simplex virus were positive in 6 of the 9 cases. Immunologic stains were positive in an additional 2 cases, and in 1 case multinucleated giant cells were present in the epithelium consistent with the diagnosis of herpes simplex virus keratitis. Tandem scanning confocal microscopic findings were positive for Acanthamoeba in 8 of the 9 cases, and all of them demonstrated Acanthamoeba organisms in epithelial scrape biopsy specimens. In 1 case, which was not evaluated with confocal microscopy, Acanthamoeba was detected using a stromal and epithelial biopsy. Two of the 9 patients had a history of contact lens use. CONCLUSION: Acanthamoeba keratitis may be present as a secondary or opportunistic infection in patients with herpetic keratitis.

Acanthamoeba↗
PubMed1997

Image data compression.

The tutorial will expand on all of the topics outlined above and will provide additional information on other compression techniques. The practical implications of data compression and the important considerations in choosing a compression scheme will also be discussed. Development of new compression algorithms remains an active area of investigation and will continue to increase the degree of compression possible.

Algorithms↗
PubMed1997

Lack of diurnal variation in vision, refraction, or keratometry after excimer laser photorefractive keratectomy.

PURPOSE: To examine whether diurnal variation in visual acuity, spherical equivalent, keratometry, or intraocular pressure occurs in patients who have undergone photorefractive keratectomy. METHODS: Twenty-nine post-photorefractive keratectomy patients underwent morning and evening measurements of visual acuity, spherical equivalent, keratometry, and intraocular pressure at intervals between 5 and 13 months after photorefractive keratectomy. RESULTS: There was no statistically significant post-photorefractive keratectomy diurnal variation in visual acuity, spherical equivalent, or keratometry in the group studied. Diurnal variation in post-photorefractive keratectomy intraocular pressure was not significantly different between patients' treated and untreated eyes. CONCLUSION: Photorefractive keratectomy does not appear to result in diurnal variation in visual acuity, spherical equivalent, or keratometry post-operatively, nor does it seem to effect normal diurnal variation in intraocular pressure.

Adult↗
PubMed1997

FDG PET characterization of renal masses: preliminary experience.

OBJECTIVE: This study was undertaken to evaluate the potential efficacy of fluorine-18 2-fluoro-2-deoxy-D-glucose positron emission tomography (FDG PET) to detect renal tumours and to characterize indeterminate renal cysts. SUBJECTS AND METHODS: Twenty-six PET scans were performed in 21 patients (14 PET scans in 10 patients with malignant renal tumours and 12 PET scans in 11 patients with Bosniak type 3 indeterminate renal cysts). Pathological proof was obtained in 18 of 21 patients (10 with solid neoplasms, eight with indeterminate cysts). Imaging was performed 1 h after injection of 5-10 mCi of FDG with IV administration of Lasix (10 mg) 20 mins after injection. Two consecutive 9.7-cm image segments were scanned to cover the entire renal areas. RESULTS: PET accurately depicted solid neoplasms as areas of increased uptake in nine of 10 patients. Bilateral renal cell carcinomas were missed in one diabetic patient. All but one indeterminate renal cysts were correctly classified as benign (photopenic areas), but an indeterminate cyst with a 4-mm papillary neoplasm was wrongly classified as benign. There were no false positive PET interpretations. The mean tumour-to-kidney ratio was 3.0 for malignant lesions. CONCLUSION: We conclude that FDG PET scanning shows promise in the evaluation of indeterminate renal cysts. A positive PET scan in the appropriate clinical setting obviates the need for cyst aspiration. A negative PET scan in conjunction with a negative cyst aspiration offers confirmatory evidence of benignity. Our preliminary results are encouraging and further work is ongoing.

Adult↗
PubMed1997

Focal hepatic lesions: effect of three-dimensional wavelet compression on detection at CT.

PURPOSE: To evaluate the effect of three-dimensional, wavelet-based compression on the detection of focal hepatic lesions at computed tomography (CT). MATERIALS AND METHODS: CT images obtained in 69 patients with focal hepatic lesions were studied (35 consecutive cases and 34 cases selected to be difficult on the basis of lesion size or contrast). Image data were compressed by means of a three-dimensional, wavelet-based algorIthm at ratios of 10:1, 15:1, and 20:1. Normal and abnormal sections (on original and compressed images) were reviewed by using an interactive workstation. Four readers rated the presence or absence of a lesion with a five-point scale. RESULTS: At receiver operating characteristic analysis, no statistically significant difference was detected for all cases considered together. Differences approached but did not reach statistical significance for the diagnostic performance of one reader with compressed images (15:1, P = .054; 20:1, P = .051). For the subset of difficult cases, a significant difference was observed with 20:1 compressed images for one reader (P = .026). Diagnostic performance of readers was less certain in normal than in abnormal cases with both original and compressed images (difference was significant for 15:1 [P = .035] and 20:1 [P < .0001] compressed images). CONCLUSION: Three-dimensional wavelet compression is satisfactory at ratios of at least 10:1. Additional studies with a larger sample would help confirm findings with higher ratios.

Cysts↗
PubMed1997

Regulation of vascular endothelial growth factor expression by insulin-like growth factor I.

Insulin-like growth factor I (IGF-I) and vascular endothelial growth factor (VEGF) levels are correlated with retinal ischemia-associated intraocular neovascularization in humans. Since VEGF is required for iris and retinal neovascularization in animal models of retinal ischemia, we tested whether IGF-I could act as an indirect angiogenic factor by increasing VEGF gene expression. IGF-I increased retinal pigment epithelial (RPE) cell VEGF mRNA in a concentration-dependent manner with an EC50 of 7 nmol/1 (53.6 ng/ml). RPE and bovine smooth muscle cells exposed to 50 nmol/l (383 ng/m1) IGF-I achieved peak VEGF mRNA expression within 2 h. IGF-I-treated RPE cells increased VEGF protein levels in conditioned media and stimulated capillary endothelial cell proliferation. Blockade of the IGF-I receptor with a neutralizing antibody abrogated the VEGF increases in RPE cells. Further, hypoxia-mediated and IGF-I-mediated increases in VEGF mRNA and protein levels were additive in RPE cells, and the hypoxia-induced VEGF increases were independent of endogenous IGF-I. VEGF promoter activity was enhanced by IGF-I in RPE cells, but VEGF transcript half-life was unaltered. In summary, the supplementation of RPE and smooth muscle cell cultures with IGF-I at 5-100 nmol/l increased VEGF mRNA and secreted protein levels. The VEGF increases in RPE cells occurred primarily through enhanced transcription of the VEGF gene and via the IGF-I receptor. Elevated IGF-I levels may promote neovascularization through increased retinal VEGF gene expression.

Animals↗