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Biomedical subjects

I R Brown

Publications and source records attributed to I R Brown.

At least 55 records · Page 3Linked to original sources
PubMed1995

Expression of heat shock protein 90 (hsp90) in neural and nonneural tissues of the control and hyperthermic rabbit.

Tissue-specific differences were apparent in the constitutive level of hsp90 in various body tissues of the unstressed rabbit. Western blotting with monoclonal antibody 29A revealed very low levels in muscle and highest levels in neural regions (cerebellum, cerebral hemispheres, and retina) and in testes and thymus. Intermediate levels were apparent in other tissues such as liver, kidney, heart, and small intestine. Following hyperthermia, induction of hsp90 was not detected with 1-D Western blotting in tissues which demonstrated high constitutive levels; however, elevations were noted in tissues which showed lower constitutive amounts of the protein, such as kidney, heart, and muscle. Immunocytochemical studies revealed that hsp90 is preferentially localized to neuronal cell populations in the rabbit brain and that this pattern does not alter following hyperthermic conditions which result in glial induction of hsp70. In kidney, where constitutive levels of hsp90 are lower than in brain, an induction of hsp90 was noted in renal tubules following hyperthermia.

Animals↗
PubMed1995

Expression of low-molecular-weight neurofilament (NF-L) mRNA during postnatal development of the mouse brain.

A regional Northern blot analysis demonstrated that the highest levels of NF-L mRNA in the adult mouse brain are present in brain stem followed by mid-brain, with lower levels found in neocortex, cerebellum, and hippocampus. The study was extended to the cellular level over course of postnatal development using in situ hybridization. This developmental analysis revealed that the expression of NF-L mRNA closely follows the differentiation pattern of many large neurons during postnatal neurogenesis. Neurons which differentiate early such as Purkinje, mitral, pyramidal, and large neurons of brain stem and thalamic nuclei, expressed high levels of NF-L mRNA at postnatal day 1. Early expression of NF-L mRNA may be required for the maintenance of the extensive neurofilament protein networks that are detected within the axons of larger neurons. Smaller neurons which differentiate later, such as dentate gyrus granule cells, small pyramidal and granule cells of the neocortex, and granule cells of the cerebellum, exhibit a delayed expression of NF-L mRNA.

Aging↗
PubMed1995

A child with an intra-abdominal testicular teratoma: a case report and review of prepubertal cryptorchid germ cell tumors.

OBJECTIVES: We report the case of a tumor in an intra-abdominal cryptorchid testis of a 7-month-old male infant. Torsion of a testicular teratoma was confirmed by pathologic examination. A review was undertaken to identify and characterize other reports of prepubertal cryptorchid germ cell neoplasms. METHODS: Cases of testicular germ cell neoplasms in association with cryptorchidism in phenotypically normal males were identified through a MEDLINE search of the English literature and review of published bibliographies. RESULTS: A total of 14 cases were identified with testis location provided in 12 patients. Half of the neoplasms were located in abdominal testis, and all of these were associated with torsion, although 2 of 6 patients were asymptomatic. Two patients had tumors in the contralateral normally descended testis. Teratoma was the most frequently encountered tumor type. CONCLUSIONS: Characteristics of prepubertal cryptorchid testicular germ cell tumors reflect those seen in post-pubertal cryptorchid tumors in location but differ in histologic type. Possible pathogenesis is discussed. Follow-up was not provided in all cases although outcome appears to be good. We believe that these lesions are likely underreported.

Abdominal Neoplasms↗
PubMed1994

Deficient induction of human hsp70 heat shock gene transcription in Y79 retinoblastoma cells despite activation of heat shock factor 1.

One of the basic features of the inducible heat shock response is the activation of heat shock factor which results in the rapid transcriptional induction of the heat shock genes. Although it is widely considered that the heat shock response is ubiquitous, several reports have indicated that the transcriptional response can vary in both intensity and kinetics and often in a tissue-specific manner. Of interest have been studies on the expression of heat shock genes in the brain, particularly observations that certain cultured neuronal cells exhibit a diminished heat shock response. We demonstrate that transcription of the gene encoding a 70-kDa heat shock protein (hsp70) is diminished upon heat shock in Y79 human retinoblastoma cells (which are of neuronal origin) despite both the activation of heat shock factor 1 and induced transcription of another heat shock gene, hsp90 alpha. This uncoupling of stress-induced transcription of the hsp70 and hsp90 alpha genes, which are typically coordinately regulated in response to stress, appears to be due to the selective inability of trans-acting factors, including heat shock factor 1, to bind in vivo to the hsp70 promoter as the result of a chromatin-mediated effect.

DNA-Binding Proteins↗
PubMed1994

Induction of heat shock (stress) protein 70 and its mRNA in the normal and light-damaged rat retina after whole body hyperthermia.

In situ hybridization and immunocytochemistry were used to investigate the distribution of the 70 kDa heat shock or stress protein (hsp70) and its mRNA in specific layers of the retina of adult rats at 0, 4, 18, and 48 or 50 hr after a brief whole body hyperthermic treatment. Induction of hsp70 mRNA was noted in the photoreceptor layer of the retina within 4 hr after hyperthermia. Pronounced accumulation of inducible hsp70 immunoreactivity was observed in cytoplasmic extensions of the photoreceptor cells, especially the inner segment zone which attained peak levels at the 18 hr time point. Selective destruction of photoreceptors by light damage prior to hyperthermia inhibited the post-hyperthermic rise in newly synthesized retinal hsp70. Our results suggest that the photoreceptor cell layer is the primary site of synthesis of hsp70 in the rat retina and that the greatest increase in hsp70 immunoreactivity following such a hyperthermic stress occurs in that layer. This stress response of the photoreceptors is discussed in relation to their location and function in the retina.

Animals↗
PubMed1994

Developmental expression in the rat cerebellum of SC1, a putative brain extracellular matrix glycoprotein related to SPARC.

In the nervous system, extracellular matrix (ECM) molecules have been shown to have effects on cell migration, process outgrowth and the survival of neurons. Recently we have described the molecular cloning of SC1, a putative brain extracellular matrix glycoprotein, showing partial similarity to the ECM glycoprotein SPARC/osteonectin. We have now examined the expression of SC1 during the development of the rat cerebellum at both the protein and mRNA levels. Our results indicate that SC1 is both temporally and spatially regulated during this process. Bergmann glial cells express SC1 mRNA and the resultant protein is deposited along the length of their radial fibres during the process of granule cell migration in the developing cerebellum. SC1 mRNA and protein is also found in the adult cerebellum, concentrated in the Bergmann glial cells and their radial processes, indicating that this putative ECM molecule continues to play roles in the central nervous system after migration and proliferative events have ceased.

Activated-Leukocyte Cell Adhesion Molecule↗
PubMed1994

Expression of the gene encoding the extracellular matrix glycoprotein SPARC in the developing and adult mouse brain.

The pattern of expression of the SPARC gene was examined during postnatal development of the mouse brain using in situ hybridization. At postnatal day 3 (P3), a strong signal representing SPARC mRNA was apparent in boundary layers such as the pia mater and the lining of the ventricles. By P12, increased levels of SPARC mRNA were noted in the cerebellum, midbrain and brain stem with a lower signal in more frontal areas, a pattern which was retained in the adult. This pronounced caudal versus frontal difference in SPARC mRNA levels was confirmed by Northern blot analysis. At P3, SPARC mRNA was detected in developing blood vessels in the cerebral cortex, suggesting a role for SPARC in angiogenesis. During development of the cerebellum, expression of SPARC mRNA became highly restricted to the Purkinje cellular layer and in the adult was localized to Bergmann glial cells rather than Purkinje neurons.

Aging↗
PubMed1993

Temporal and spatial distribution of heat shock mRNA and protein (hsp70) in the rabbit cerebellum in response to hyperthermia.

We have previously investigated the expression of hsp70 genes in the hyperthermic rabbit brain at the mRNA level by Northern blot and in situ hybridization procedures. Our studies have now been extended to the protein level utilizing Western blot and immunocytochemistry. Using an antibody which is specific to inducible hsp70, a prominent induction of hsp70 protein in glial cells of hyperthermic animals was noted. In particular, Bergmann glial cells in the cerebellum are strongly immunoreactive while adjacent Purkinje neurons are immunonegative. Extension of our in situ hybridization studies to a time course analysis revealed that the initial glial induction events were followed by a delayed accumulation of inducible hsp70 mRNA in Purkinje neurons at 10 hr post-heat shock. In control animals, high levels of constitutively expressed hsc70 mRNA and protein were observed in Purkinje neurons. Similar hsc70 and hsp70 mRNA observations were also made in neurons of the deep cerebellar nuclei and in motor neurons of the spinal cord. Our results suggest that these neuronal cell types accumulate hsp70 mRNA in response to hyperthermic treatment; however, the response is delayed when compared to the rapid response seen in glial cells. The high constitutive levels of hsc70 in certain neuronal cell types may play a role in the initial dampening of the hsp70 induction response in these cells.

Animals↗
PubMed1993

Modulation of a neuronal calmodulin mRNA species in the rat brain stem by reserpine.

Reserpine evokes transsynaptic impulse activity by depleting catecholaminergic neurotransmitters in the rat brain. Previous studies suggest a relationship between catecholaminergic activity and calmodulin concentration. In this report we employ Northern blot analysis to examine the effect of a single subcutaneous injection of reserpine on levels of calmodulin mRNA species which are preferentially expressed in neurons of the rat brain. Regional differences in mRNA levels were also investigated by in situ hybridization and drug-induced changes were noted particularly in specific regions of the rat brain stem. The riboprobe used in the in situ hybridization study recognized a 4.0 kilobase neuronal calmodulin mRNA species (NGB1), which was derived from the rat CaM1 gene. A calmodulin radio-immunoassay was utilized to demonstrate a drug-induced increased in calmodulin protein levels in a region which included the brain stem.

Animals↗
PubMed1992

Expression of heat shock genes (hsp70) in the rabbit spinal cord: localization of constitutive and hyperthermia-inducible mRNA species.

We have previously reported that hyperthermia induces the expression of a heat shock gene in the rabbit brain (Sprang and Brown, Mol Brain Res 3:89-93, 1987). Striking regional and cell type differences in the pattern of induction of the hsp70 mRNA were noted. Tissue injury also induces the rapid induction of hsp70 mRNA in the mammalian brain (Brown et al., Neuron 2:1559-1564, 1989). In the present study, in situ hybridization with 35S-labelled riboprobes specific for constitutive and inducible hsp70 mRNA species was employed to investigate the effect of fever-like temperatures on hsp70 gene expression in the rabbit spinal cord. Expression of constitutive hsp70 mRNA was detected in large motor neurons of both control and hyperthermic animals. Within 1 hr after hyperthermia, a massive induction of inducible hsp70 mRNA was noted in fibre tracts of the spinal cord, a pattern consistent with a strong glial response to heat shock. Induction was not observed in the large motor neurons.

Animals↗
PubMed1992

Interaction of multiple nuclear proteins with the promoter region of the mouse 68-kDa neurofilament gene.

Four brain-specific, DNase I hypersensitive sites (HSS) have been mapped to the 5' flanking region of the mouse 68-kDa neurofilament gene. These sites are contained within a 1.7-kb sequence that confers neuronal specificity of expression of this gene in transgenic mice. To identify DNA sequences that might be involved in gene regulation, the HSS situated near the promoter region has been analyzed by gel mobility shift assays and DNase I footprinting to investigate protein binding sequences. Of particular interest are two footprints localized to a 9-nucleotide sequence that flanks both the light and medium neurofilament gene in mouse and to a sequence that demonstrates partial homology to several promoter regions, including element-1, a motif required for neuron specificity in Drosophila. A prominent footprint was also detected at a sequence that contains a near-perfect palindrome centered at a PstI restriction site.

Animals↗
PubMed1992

Molecular cloning of a novel mRNA using an antibody directed against synaptic glycoproteins.

It has been suggested by a number of investigators that glycoproteins may play a role in the development and/or maintenance of synapses in the mammalian CNS. For many synaptic glycoproteins, however, little precise structural or functional information is available. In an effort to isolate probes specific to individual glycoproteins, we have screened a rat brain cDNA expression library with a mixed polyclonal antibody directed against concanavalin A-binding synaptic junctional glycoproteins. Using this approach, we have previously reported the cloning of SC1, a putative extracellular matrix glycoprotein found in adult brain (Johnston et al., Neuron 2:165-176, 1990). We now report the cloning and characterization of a second novel cDNA, which has been designated SC2. Northern blots show that this cDNA recognizes a 1.2-kb mRNA that is present throughout postnatal development in the rat. It is expressed at high levels in brain and is also found at lower levels in several other tissues. In situ hybridization suggests that the SC2 mRNA is strongly expressed by many types of neurons. Sequence data reveals a single open reading frame in the cDNA, encoding a putative hydrophobic protein with a calculated molecular weight of 36.1 kDa. Sequence analysis reveals some similarity between SC2 and 5 alpha-reductase, a microsomal membrane protein important in testosterone metabolism.

Amino Acid Sequence↗
PubMed1992

Effect of a discrete dorsal forebrain lesion in the rat on the expression of neuronal and glial-specific genes: induction of calmodulin, NF-L, SC1, and GFAP mRNA.

The influence of a localized tissue injury on the expression of genes encoding specific neuronal and glial proteins was examined using in situ hybridization. A pronounced induction of neuron-specific calmodulin (CaM) mRNA was evident within cells proximal to the wound site by 2 hours following a cortical lesion in rats. By 12 hours postlesion, intense signal corresponding to CaM mRNA was found to extend 1 mm from the wound site. Changes in the expression of mRNA encoding two additional neuronal proteins, the 68 kilodalton neurofilament protein and the extracellular matrix protein, SC1, were also evident at 12 hours following the cortical injury. Of the two glial proteins examined, a dramatic elevation in levels of mRNA for glial fibrillary acidic protein (GFAP) was observed at the wound site by 12 hours postlesion. This intense labeling corresponding to GFAP mRNA was evident in the ipsilateral glial limitans and hippocampus as well as in the contralateral glial limitans. In contrast, the pattern of labeling for the beta-subunit of the S100 protein did not differ from that of control animals at either of the postlesion intervals examined. This study identifies four genes, CaM, GFAP, SC1, and NF-L, which are induced following a localized cortical injury and which encode mRNA species enriched in specific cell-types in the central nervous system.

Activated-Leukocyte Cell Adhesion Molecule↗
PubMed1992

Developmental expression of neuronal calmodulin mRNA species in the rat brain analyzed by in situ hybridization.

The temporal and spatial distribution of calmodulin mRNAs which are preferentially expressed in neurons was determined during postnatal development of rat central nervous system. Expression of these mRNAs was strongly detected in the developing neocortex, hippocampus, and cerebellum. Differences in the pattern of expression of a 1.8 and 4.0 kb neuronal calmodulin mRNA species were identified in the developing cerebellum. Expression of the smaller mRNA appeared to correlate with proliferating and developing cerebellar granule neurons while the larger mRNA was present in the mature granule neuron population. A transient elevation in the neuronal calmodulin mRNA species was observed in the superior and inferior colliculus and in the thalamus at postnatal days 5 and 10.

Aging↗
PubMed1992

Distribution of constitutive- and hyperthermia-inducible heat shock mRNA species (hsp70) in the Purkinje layer of the rabbit cerebellum.

In previous studies we have analyzed the effect of hyperthermia on the expression of hsp70 genes in the rabbit cerebellum using an hsp70 riboprobe which hydridized to both constitutively expressed and stress-inducible transcripts. These studies have now been extended utilizing riboprobes which are able to discriminate hyperthermia-inducible hsp70 mRNA of size 2.7 kb and constitutively expressed mRNA of size 2.5 kb. In situ hybridization with the inducible specific riboprobe revealed a prominent induction of the 2.7 kb species 1 hr after a 2-3 degrees C increase in body temperature in the following cerebellar cell types: i) Bergmann glial cells in the Purkinje layer, ii) glial cells in deep white matter fiber tracts and iii) granule neurons. The inducible transcript was not detected in the cerebellum of control animals. The constitutive specific riboprobe detected the 2.5 kb transcript in several neuronal cell types of the cerebellum such as Purkinje and granule neurons with little increase in signal in hyperthermic animals compared to controls.

Animals↗
PubMed1992

Molecular cloning of calmodulin mRNA species which are preferentially expressed in neurons in the rat brain.

A cDNA clone designated NGB, which was isolated from a rat brain expression library, detected two mRNA species of 1.8 and 4.0 kb which are highly enriched in brain tissue. cDNAs NGB1 and NGB2 corresponding to these two mRNAs have been isolated and characterized. Sequence data showed that both mRNA species contain the same open reading frames but differ in their 3' untranslated regions. The open reading frame encodes a calmodulin protein of 148 amino acids. Both mRNA species are derived from the rat CaMI gene by utilization of different polyadenylation addition sites. Analysis of the 3' untranslated sequence which is unique to the larger mRNA species revealed a putative AU-rich 'destabilizer' sequence which is thought to be involved in mechanisms of selective mRNA breakdown. In situ hybridization studies revealed that the two calmodulin mRNAs are expressed strongly in neuronal cells in the adult rat brain. Levels of the two mRNA species increased during early postnatal development.

Aging↗