Search PubMed⌕ Search

Biomedical subjects

J G Joshi

Publications and source records attributed to J G Joshi.

At least 19 recordsLinked to original sources
PubMed1998

Iron metabolism and human ferritin heavy chain cDNA from adult brain with an elongated untranslated region: new findings and insights.

Ferritin is a ubiquitous protein which plays a major role in iron sequestration, detoxification and storage. In this paper we highlight the role of ferritin in iron homeostasis and describe factors and diseases that affect its expression. We also describe new studies which further characterize the structure and expression of a novel form of ferritin heavy (H) chain mRNA that was identified in brain and discuss possible implications of these findings. Human fetal and adult brain cDNA libraries previously were screened with cDNA for well-characterized liver ferritin H. In addition to 'liver-like' brain ferritin H cDNA, novel ferritin H cDNAs with an additional 279 nucleotide sequence at the 3'untranslated region (UTR) were identified in both libraries (see refs. 1 and 2; Dhar, M., Chauthaiwale, V., and Joshi, J. G., Gene, 1993, 126, 275 and Dhar, M., and Joshi, J. G., J. Neurochem., 1993, 61, 2140). However, relative to liver ferritin H cDNA, these novel cDNAs were incomplete at their 5'ends [see ref. 3; Joshi, J. G., Fleming, J. T., Dhar, M. S., and Chauthaiwale, V., J. Neurol Sci., 1995, 134, (Suppl.), 52]. In the present paper, by sequencing of cDNAs using reverse transcriptase polymerase chain reaction, we show that the 279 nt 3'UTR sequence, a coding sequence identical to that in human liver ferritin H, and a full-length 5'UTR that includes one mRNA regulatory iron-response element sequence, co-exist in at least one species of ferritin H transcript in six normal human adult and six late-onset, sporadic Alzheimer disease (AD) brains. This sequence is the same in the normal and AD brains. Dot-blot analysis of poly A+ RNAs from different human tissues indicates that relative to the coding sequence of ferritin H, expression of the 279 nt 3'UTR sequence varies among different tissues, is highest in the adult brain, and is very low in fetal brain. In normal adult hippocampus, ferritin H RNA with the novel 279 nt sequence localizes strongly to small non-neuronal cells, capillary endothelial cells, and to selected populations of neurons (granule cells of the dentate gyrus). Significant homology was observed between a region in the 279 nt 3'UTR segment of ferritin H RNA and the 3'UTR of cyclooxygenase-2 mRNA (an inducible iron-containing enzyme involved in prostaglandin synthesis). Possible functions for ferritin H protein derived from the novel message and for the elongated 3'UTR and 5'UTR are discussed.

Adult↗
PubMed1996

Can the mechanisms of aluminum neurotoxicity be integrated into a unified scheme?

Regardless of the host, the route of administration, or the speciation, aluminum is a potent neurotoxicant. In the young adult or developmentally mature host, the neuronal response to Al exposure can be dichotomized on morphological grounds. In one, intraneuronal neurofilamentous aggregates are formed, whereas in the other, significant neurochemical and neurophysiological perturbations are induced without neurofilamentous aggregate formation. Evidence is presented that the induction of neurofilamentous aggregates is a consequence of alterations in the posttranslational processing of neurofilament (NF), particularly with regard to phosphorylation state. Although Al has been reported to impact on gene expression, this does not appear to be critical to the induction of cytoskeletal pathology. In hosts responding to Al exposure without the induction of cytoskeletal pathology, impairments in glucose utilization, agonist-stimulated inositol phosphate accumulation, free radical-mediated cytotoxicity, lipid peroxidation, reduced cholinergic function, and altered protein phosphorylation have been described. The extent to which these neurochemical modifications correlate with the induction of a characteristic neurobehavioral state is unknown. In addition to these paradigms, Al is toxic in the immediate postnatal interval. Whether unique mechanisms of toxicity are involved during development remains to be determined. In this article, the mechanisms of Al neurotoxicity are reviewed and recommendations are put forth with regard to future research. Primary among these is the determination of the molecular site of Al toxicity, and whether this is based on Al substitution for divalent metals in a number of biological processes. Encompassed within this is the need to further understand the genesis of host- and developmental-specific responses.

Adult↗
PubMed1996

Can the controversy of the role of aluminum in Alzheimer's disease be resolved? What are the suggested approaches to this controversy and methodological issues to be considered?

Aluminum (Al) is unquestionably neurotoxic in both experimental animals and certain human diseases. Minute quantities injected intracerebrally into rabbits will induce severe neurological symptoms and neuropathological features of neurodegeneration. Hyper-aluminemia often develops in patients with renal failure being treated with intermittent hemodialysis on a chronic basis, and in severe cases results in an encephalopathy. Uremic adults and premature infants not on dialysis treatment also can develop encephalopathy due to Al toxicity, as is the case when large amounts of alum are used as a urinary bladder irrigant. There are many other examples of Al-induced neurotoxicity; however, the question as to whether Al presents a health hazard to humans as a contributing factor to Alzheimer's disease is still the subject of debate. Several lines of evidence are presented that have formed the basis of the debate concerning the possible pathogenic role for Al in Alzheimer's disease. Important evidence for an Al-Alzheimer's causal relationship is the observation by laser microprobe mass analysis (LMMS) of the presence of Al in neurofibrillary tangles, although there are conflicting data on the extent of the Al deposition. The relatively poor sensitivity of some of the analytical instruments available for these challenging in situ microanalyses could explain the discrepant results, although LMMS and perhaps secondary ion mass spectrometry (SIMS) appear to be sufficiently sensitive. Harmonization of the techniques is an essential next step. There is new evidence that exposure to Al from drinking water might result in cognitive impairment and an increased incidence of Alzheimer's disease. However, these epidemiological studies have inherent problems that must be scrutinized to determine if an association really does exist. An understanding of a possible enhanced bioavailability of Al in this type of exposure, versus other exposures such as antacid intake or industrial exposure, needs to be considered and explored. There has been one promising clinical trial of the treatment of Alzheimer's disease patients with the Al chelator desferrioxamine (DFO). Further studies are needed, and if confirmation is forthcoming then such data could also support an Al-Alzheimer's disease link as well as suggesting that DFO offers potential as a therapeutic agent. The possibility that iron might be the offending agent needs to be considered since DFO is a very strong iron chelator. The significance of Al-induced neurofibrillary degeneration in experimental animals should be assessed especially in light of new data showing that this model exhibits abnormally phosphorylated tau protein structures in the neuronal perikarya. Thus the key questions that must be answered before it can be asserted that Al possesses causal relationship to Alzheimer's disease, are as follows and are addressed in this present discussion: (1) Are there elevations of the concentration of Al in the brains of Alzheimer's disease patients? (2) Is there a relationship between environmental exposure to Al, particularly in drinking water, and an increased risk of Alzheimer's disease? (3) Is treatment with DFO a potentially useful therapeutic approach and to what extent might beneficial effects of DFO implicate Al in the etiology of Alzheimer's disease? (4) Are there similarities between the experimental animal studies and Alzheimer's disease particularly in the development of abnormal forms of tau seen in neurofibrillary tangles? (5) Does Al promote the deposition of the A beta peptide in Alzheimer's disease? (6) Does hyperaluminemia associated with long-term hemodialysis treatment induce neurofibrillary degeneration? If the answer to each of these six questions is yes, then does this assert that Al possesses a causal relationship to Alzheimer's disease? On the other hand, must all six be met to be able to make this assertion?

Aluminum↗
PubMed1995

A novel ferritin heavy chain messenger ribonucleic acid in the human brain.

In the aging human brain, the concentrations of iron and its major storage protein, ferritin, rise but the distribution of metal and protein remains non-uniform. More ferritin could be isolated from the brains of humans who died of Alzheimer's disease (AD) than from age- and sex-matched controls. Also, brain ferritin of rats chronically exposed to aluminum chloride in their drinking water contained more aluminum and iron. Based on these earlier observations, a more detailed study of human brain ferritin was initiated. The results showed that ferritin is a component of neuritic (senile) plaques in AD. Ferritin obtained from normal or AD brains is composed of 24 subunits (70% heavy (H) chain; 30% light (L) chain). With high performance liquid chromatography, the subunits resolved into a cluster of four H-chain peaks and one major L-chain peak. Western blot analysis confirmed the identity of H- and L-fractions. The techniques of molecular biology revealed the presence of an additional ferritin messenger ribonucleic acid (mRNA) species for the H subunit which was more abundant in the brain than in other human tissues. It contained the entire sequence of 919 nucleotides of H chain mRNA from liver but also an additional segment of 279 nucleotides in the 3'-untranslated region. The two mRNA seemed to arise by the use of an alternate polyadenylation site of the same primary transcript. Ribonuclease protection assays revealed that the concentrations of the longer mRNA in the normal hippocampus and the hippocampus of patients with AD brains were similar.

Aluminum↗
PubMed1995

Localization of a new ferritin heavy chain sequence present in human brain mRNA to chromosome 11.

Two types of ferritin heavy (H) chain clones have been isolated from cDNA libraries of human fetal and adult brain: one corresponds to the ferritin H chain mRNA that is abundant in liver and is called "liver-like" brain cDNA; the other contains an additional 279 nucleotide (nt) sequence in the 3' untranslated region and is called brain ferritin H chain cDNA. To map the 279-nt sequence, polymerase chain reaction (PCR) amplification was carried out using DNA from rodent x human hybrid cell lines containing single human chromosomes as templates, and oligomeric primers homologous to the 3' end of the 279-nt sequence (primer A) and to a coding sequence just 5' to the 279-nt sequence. Significant PCR product of the size expected from analysis of the brain ferritin H chain cDNA clones and a genomic ferritin H chain clone (487 bp) was generated only from hybrid-cell DNA containing human chromosome 11. This PCR product and the "liver-like" brain cDNA (lacking the 279-nt sequence) both hybridized to chromosome 11 fragments that are known to define the well-characterized functional liver ferritin H chain gene and a putative pseudogene. Preliminary data indicate that primer A (and thus the 279-nt sequence) maps to the functional ferritin H chain gene fragments, but binding to the pseudogene has not been ruled out.

Adult↗
PubMed1994

Iron and aluminum homeostasis in neural disorders.

The brain is the most compartmentalized organ. It is also highly aerobic. Because nerve cells grow but do not regenerate, the brain is the organ best suited for the accumulation of metabolic errors colocalized in specific areas of the brain over an extended period. Alzheimer's disease (AD) is primarily a neurological disorder of the elderly. It is suggested that this disorder results from the accumulation of such errors, and that AD onset aluminum and iron contribute to but do not necessarily initiate the onset of the disease. In vitro and in vivo evidence summarized here suggests that this is effected by interfering in the utilization of glucose and glucose-6-phosphate, and sequestration of iron by ferritin. beta-amyloid precusor proteins (beta-APPs) are normal components of the human brain and some other tissues. Proteolysis of these, presumably by serine proteases, generates a 39 to 42 amino acid long peptide, the alpha-amyloid (beta-AP). In AD brains, beta-AP aggregates into plaque, the hallmark of AD brains. Some of the alpha-APPs also contain a 56 amino acid long segment which inhibits serine proteases. We show that in vitro, at pH 6.5, aluminum activates beta-chymotrypsin 2-fold and makes it dramatically resistant to protease inhibitors such as bovine pancreatic trypsin inhibitor (bPTI) or its mimic present in the beta-amyloid precursor proteins (beta-APPs). Iron and oxygen are reported to favor cross-linking of beta-AP in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)

Aluminum↗
PubMed1994

Detection and quantitation of the novel ferritin heavy chain message in human tissues.

We reported that in the fetal brain use of an alternate polyadenylation site in the premRNA for ferritin heavy chain generates two mature mRNAs of different lengths. The larger mRNA contains an additional 279 nucleotide sequence at the 3' untranslated region. Here we use Northern blot analysis and show that this mRNA is also present in other human tissues. Its relative concentration is in the order: brain > kidney > lung > skeletal muscle > pancreas > heart = placenta = liver.

Blotting, Northern↗
PubMed1993

The human and bovine 14-3-3 eta protein mRNAs are highly conserved in both their translated and untranslated regions.

14-3-3 proteins form a highly conserved protein family whose members have been shown to activate tyrosine and tryptophan hydroxylases, inhibit protein kinase C and possess phospholipase A2 activity in vitro. We have isolated and analyzed a 14-3-3 protein cDNA clone (H14-3-3) from a human fetal brain cDNA library and found it to possess a high level of sequence identity with the bovine 14-3-3 eta protein cDNA in both the translated and untranslated regions, suggesting the presence of cis-regulatory elements in the untranslated regions of these mRNAs. The proteins encoded by these two cDNAs are 98.4% identical. Two different sized RNA species, approx. 1.9 and 3.5 kb in size that are expressed in a variety of tissues hybridize with this cDNA. However, only the 1.9 kb RNA is detected in the fetal brain. Northern blot analysis of poly(A)+ RNA isolated from eight different human tissues shows that 14-3-3 protein mRNAs are expressed in many tissues in the body. In agreement with previous reports, the highest abundance of RNA hybridizing with this cDNA is seen in the brain.

14-3-3 Proteins↗
PubMed1993

Sequence of a cDNA encoding the ferritin H-chain from an 11-week-old human fetal brain.

A cDNA library in lambda Charon BS(-) from 11-week-old human fetal brain (FB) was screened using a human liver ferritin (FTH)-encoding cDNA as a probe. The complete sequence of the positive clone, cFB1, showed that the coding region and a part of the 5' and 3' untranslated regions (UTR) are identical to the corresponding published sequence of the liver cDNA. However, a particularly noteworthy difference is the presence of 279 bp of additional sequence in the FB 3'-UTR. Northern blot analysis of FB poly(A)+RNA showed it to be a part of the FTH transcript. Comparison of the 279-bp sequence with the GenBank and EMBL databases showed it to be 94.1, 62.5, and 58.9% similar to segments from human, mouse and rat FTH genomic sequences, respectively. However, in all these cases, only a part of this 279-bp sequence has been found in the nontranscribed region. We therefore conclude that in FB, the 279-bp sequence is a part of the mature FTH mRNA. Sequence analysis also suggests a differential poly(A) site selection in the production of FTH mRNA in FB and liver.

Amino Acid Sequence↗
PubMed1993

Regulation of serine protease activity by aluminum: implications for Alzheimer disease.

The brain of Alzheimer disease patients contains plaques that are diagnostic for the disease. The plaques also contain beta-amyloid peptide, alpha 1-antichymotrypsin, and the element aluminum. We present indirect evidence that can relate all three components of plaques to each other in such a way as to suggest their involvement in the etiology of the disease. The beta-amyloid peptide is derived by proteolytic processing from beta-amyloid precursor proteins and some of these proteins contain a domain that is highly homologous to bovine pancreatic trypsin inhibitor. Bovine pancreatic trypsin inhibitor also inhibits alpha-chymotrypsin and we show that aluminum affects both the activity and the inhibition of this enzyme. At pH 6.5, in the presence of aluminum, the enzyme activity is doubled, and the inhibitor is only 1% as effective as in the absence of the metal ion. The inhibition by BX-9, a protease inhibitor prepared from protein components of amyloid plaques, is also reduced by aluminum; so too is that by alpha 1-antichymotrypsin but to a lesser degree. In the Alzheimer brain, we propose that aluminum may accelerate proteolytic processing of the beta-amyloid precursor protein by suppression of the inhibitor domain. Thus, the beta-amyloid peptide may accumulate and initiate plaque formation.

Aluminum↗
PubMed1993

Differential processing of the ferritin heavy chain mRNA in human liver and adult human brain.

Northern blot analyses of the poly(A)+ RNAs from human brain and liver, using a human brain ferritin heavy chain (FTH) cDNA as the probe, shows the presence of two transcripts of 1.4 and 1.1 kb. The larger, 1.4-kb RNA, is expressed predominantly in the brain, whereas the smaller, 1.1 kb, is expressed abundantly in the liver. Screening of two normal human brain cDNA libraries yielded two types of human brain FTH cDNAs. One type corresponds to the previously characterized 1.1-kb RNA from liver and lymphocytes. The other is also identical to the previously characterized FTH cDNA except that it contains an additional 279-bp sequence at the 3' untranslated region. This additional sequence shows 94.1%, 62.5%, and 58.9% identity to the 3' flanking sequence of the human liver and mouse and rat FTH genomic clones, respectively. A fragment of a genomic clone containing the 279-bp sequence was also isolated and sequenced. These data suggest that differential processing of the primary transcript for the FTH mRNA in human brain and liver could generate two mature mRNAs of 1.4 and 1.1 kb. This could be due to the use of alternative polyadenylation sites in the pre-mRNA.

Adult↗
PubMed1991

Neurochemical hypothesis: participation by aluminum in producing critical mass of colocalized errors in brain leads to neurological disease.

1. Aluminum is an established neurotoxin. Prolonged exposure to even low levels of aluminum permit its chelation and subsequent transport to brain where it is non-uniformly distributed. 2. Available evidence suggests that (i) aluminum interferes with glucose metabolism by inhibiting hexokinase and glucose-6-phosphate dehydrogenase; (ii) it binds to calmodulin and affects numerous phosphorylation-dephosphorylation reactions; (iii) it binds to transferrin and ferritin, affects the function of these proteins which in turn affect iron metabolism. 3. Thus accumulation of aluminum-induced metabolic errors colocalized in specific areas of the brain may lead to neurological disorders.

Aluminum↗
PubMed1990

Ferritin is a component of the neuritic (senile) plaque in Alzheimer dementia.

A strong immunoreactivity for ferritin was observed in the neuritic (senile) plaques in Alzheimer's disease hippocampus. The ferritin accumulation was almost exclusively associated with the microglia, which appeared to have proliferated greatly. These cells were also positive for HLA-DR, a putative marker for reactive microglia. In contrast, in the diffuse plaques, which were without neuritic pathology, the ferritin-stained microglia appeared to be normal. Microglia were seen frequently in contact with neurons undergoing neurofibrillary changes but only the tangles in the extracellular space were ferritin positive. No ferritin was detected, by Western blots, in paired helical filaments isolated from Alzheimer's disease brain, suggesting that ferritin was most likely weakly associated with and was not a constituent of these fibrils. No correlation between increased ferritin/microglia activity and blood-brain barrier leakage was detected. Ferritin, an iron-storage protein, might have a role in the formation of amyloid through the action of free radicals generated during the release of iron from the ferritin molecule. Alternatively, the ferritin/microglia system might be secondarily involved in the removal and processing of the amyloid.

Aged↗
PubMed1990

Characterization of glucose-6-phosphate dehydrogenase isozymes from human and pig brain.

Homogenates of human and pig brain in 10 mM Tris-HCl, pH 8.0 were centrifuged at 25,400 x g for 1 h. The supernatants were electrophoresed in polyacrylamide gels were stained for glucose-6-phosphate dehydrogenase (EC 1.1.1.49) activity. Five distinct bands were visible. Isozymes corresponding to two of those bands were purified from human and pig brain. The isozymes were electrophoretically homogeneous. The native proteins, Mr, 220,000, dissociated in sodium dodecyl sulphate-polyacrylamide gels into a 57,000 Mr subunit. Therefore, the native isozymes are tetramers. None of the isozymes required additional metal ions for activity. At 1 mM concentration Mg2+ and Ca2+, independently or together, activated the isozymes 1.5-fold. The isozymes were NADP(+)-specific. Kmapp values of the G6PD isozymes were similar for NADP+ (6-8 microM), but different for G6P (56-180 microM). The specific activities of the isozymes varied from 50 to 210 units per mg of protein. All isozymes were inhibited by NADPH. The inhibition was competitive with respect to NADP+ and non-competitive with respect to G6P. NADH did not affect any of the isozymes. ATP inhibited the isozymes competitively with respect to G6P and non-competitively with respect to NADP+. Palmitoyl-CoA dissociated the active tetramers into enzymatically inactive dimeric forms. This treatment also abolished the 6-phosphogluconate activity of the isozyme II from both sources. High performance liquid chromatography peptide maps of the tryptic digest and amino acid analyses of the isozymes showed extensive homologies between the corresponding isozymes from the two species. Interestingly, only the isozyme II in human and pig brain was active with 6-phosphogluconate as a substrate (Kmapp = 864 and 279 microM). The specific activities of the isozyme II with 6-phosphogluconate (14 and 48 unit per mg of protein for human and pig brain isozyme II, respectively) was four times less than those with G6P. It is therefore suggested that isozyme II is a bifunctional enzyme.

Amino Acids↗
PubMed1990

Aluminum, a neurotoxin which affects diverse metabolic reactions.

Experimental evidence is summarized to support the hypothesis that chronic exposure to low levels of aluminum may lead to neurological disorders. These disorders result from defective phosphorylation--dephosphorylation reactions, reduced glucose utilization and site-specific damage inflicted by free radicals produced by altered iron metabolism. The brain is a highly compartmentalized organ. Therefore, a co-localization of critical mass of metabolic errors rather than a single event may be essential to precipitate a neural disease. Aluminum appears to participate in formulating this critical mass. Patients with dialysis dementia get partial relief by desferroxamine which chelates aluminum. However, it also chelates iron and therefore limits its applicability. While the specific chelator for aluminum is yet to be made available, exercising a caution in aluminum intake appears prudent.

Aluminum↗
PubMed1990

Concanavalin A is not a ferritin.

Contrary to recent claims, in vitro evidence has been obtained to establish that Concanavalin A (Con A) is not a ferritin. Four techniques including immunoprecipitation, gel filtration, sucrose density gradient ultracentrifugation and CsCl centrifugation were employed. None of them showed that Con A is a ferritin.

Centrifugation, Density Gradient↗
PubMed1989

Inactivation of bakers' yeast glucose-6-phosphate dehydrogenase by aluminum.

Preincubation of yeast glucose-6-phosphate dehydrogenase (G6PD) with Al(III) produced an inactive enzyme containing 1 mol of Al(III)/mol of enzyme subunit. None of the enzyme-bound Al(III) was dissociated by dialysis against 10 mM Tris-HCl, pH 7.0, containing 0.2 mM EDTA at 4 degrees C for 24 h. Citrate, NADP+, EDTA, or NaF protected the enzyme against the Al(III) inactivation. The Al-(III)-inactivated enzyme, however, was completely reactivated only by citrate and NaF. The dissociation constant for the enzyme-aluminum complex was calculated to be 4 x 10(-6)M with NaF, a known reversible chelator for aluminum. Modification of histidine and lysine residues of the enzyme with diethyl pyrocarbonate and acetylsalicylic acid, respectively, inactivated the enzyme. However, the modified enzyme still bound 1 mol of Al(III)/mol of enzyme subunit. Circular dichroism studies showed that the binding of Al(III) to the enzyme induced a decrease in alpha-helix and beta-sheet and an increase in random coil. Therefore, it is suggested that inactivation of G6PD by Al(III) is due to the conformational change induced by Al(III) binding.

Aluminum↗