Biomedical subjects

K Iqbal

Publications and source records attributed to K Iqbal.

At least 109 records · Page 6Linked to original sources

PubMed1994

Alzheimer disease: correlation of cerebro-spinal fluid and brain ubiquitin levels.

Neurofibrillary degeneration is one of the histopathological hallmarks of Alzheimer disease (AD). Previous studies have shown an association of ubiquitin with the cytoskeletal protein pathology in AD. In the present study, we report (i) the measurement of ubiquitin levels in cerebrospinal fluid (CSF) from histopathologically confirmed AD and control cases, using a new rapid immunoassay, the competitive enzyme-linked immunoflow assay (CELIFA), (ii) the determination of ubiquitin levels in brain tissue taken from the same cases, using a competitive enzyme-linked immunosorbent assay (ELISA), and (iii) an evaluation of the correlation between levels of ubiquitin in CSF and in brain tissue. Ubiquitin levels in CSF of AD and neurological control groups are significantly higher than those of non-neurological aged controls. Ubiquitin levels in brain homogenates of the AD group are significantly higher than those of both non-neurological aged and neurological control groups. The source of this increase in brain ubiquitin in AD is the particulate fraction, because ubiquitin levels in the brain cytosol fraction are the same among the three groups. In AD and non-neurological aged controls, there is a significant positive correlation between ubiquitin levels in CSF and in homogenate of cerebral white matter. In contrast, the correlation in the non-AD neurological control group has a negative tendency. These studies suggest that in AD, elevated levels of ubiquitin in the CSF reflect the increased amount of the protein in the brain and, therefore, can serve as a biomarker of the neuropathology in this disease.

Alzheimer Disease↗

PubMed1994

Comparison of the phosphorylation of microtubule-associated protein tau by non-proline dependent protein kinases.

Microtubule-associated protein tau from Alzheimer brain has been shown to be phosphorylated at several ser/thr-pro and ser/thr-X sites (Hasegawa, M. et al., J. Biol. Chem. 267, 17047-17054, 1992). Several proline-dependent protein kinases (PDPKs) (MAP kinase, cdc2 kinase, glycogen synthase kinase-3, tubulin-activated protein kinase, and 40 kDa neurofilament kinase) are implicated in the phosphorylation of the ser-thr-pro sites. The identity of the kinase(s) that phosphorylate the ser/thr-X sites are unknown. To identify the latter kinase(s) we have compared the phosphorylation of bovine tau by several brain protein kinases. Stoichiometric phosphorylation of tau was achieved by casein kinase-1, calmodulin-dependent protein kinase II, Gr kinase, protein kinase C and cyclic AMP-dependent protein kinase, but not with casein kinase-2 or phosphorylase kinase. Casein kinase-1 and calmodulin-dependent protein kinase II were the best tau kinases, with greater than 4 mol and 3 mol 32P incorporated, respectively, into each mol of tau. With the sequential addition of these two kinases, 32P incorporation approached 6 mol. Peptide mapping revealed that the different kinases largely phosphorylate different sites on tau. After phosphorylation by casein kinase-1, calmodulin-dependent protein kinase II, Gr kinase, cyclic AMP-dependent protein kinase and casein kinase-2, the mobility of tau isoforms as detected by SDS-PAGE was decreased. Protein kinase C phosphorylation did not produce such a mobility shift. Our results suggest that one or more of the kinases studied here may participate in the hyperphosphorylation of tau in Alzheimer disease.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1994

N-methyl-2-pyrrolidone as a cosolvent: relationship of cosolvent effect with solute polarity and the presence of proton-donating groups on model drug compounds.

N-Methyl-2-pyrrolidone (methylpyrrolidone), a cosolvent which has been used in veterinary medicine and in transdermal delivery devices, was investigated as a cosolvent for model drug compounds of widely varying polarity. These compounds were digoxin, sulfamethoxazole, hydrocortisone acetate, theophylline, phenytoin, and reserpine. Methylpyrrolidone was found to be an extremely efficient cosolvent for low solubility polar drugs such as digoxin or drugs containing multiple proton-donating groups such as phenytoin. The increase in solubility observed in aqueous solutions of digoxin and phenytoin to which 0.2 volume fraction of methylpyrrolidone was added was 500x and 65x, respectively. Significant deviations from log-linear solubilization were observed with digoxin, sulfamethoxazole, phenytoin, and reserpine, indicating significant water-solute-cosolvent interactions.

Chemical Phenomena↗

PubMed1994

Rabbit IgG cross-reacts with Alzheimer neurofibrillary tangles.

Biochemical studies have demonstrated that the paired helical filaments (PHF) of Alzheimer neurofibrillary tangles are mostly made up of tau and to a lesser degree of ubiquitin and other proteins. In addition, immunocytochemical labeling of tangles with antibodies to various other neuronal proteins has been shown previously. We report here the labeling of the locations of PHF, i.e., Alzheimer neurofibrillary tangles, neuropil threads and plaque neurites in tissue sections with a goat antiserum to rabbit IgG (GAR-T). The labeling is comparable in strength and distribution to that of tau and ubiquitin antibodies. The PHF-staining antibodies could be removed by absorption with native rabbit IgG but not with human IgG, IgG-depleted rabbit serum, rabbit IgG heavy chains or light chains eluted from nitrocellulose membranes. Furthermore, the PHF reactivity was obliterated by absorption with brain homogenate and a fraction enriched in soluble abnormally phosphorylated tau, but not with purified bovine tau or SDS-washed preparations of the relatively insoluble population of PHF. On immunoblots of both normal human tau and Alzheimer abnormally phosphorylated tau-enriched preparations, GAR-T labeled a set of three to five polypeptides in the tau region. Some of these polypeptides co-migrated with the tau bands. These results indicate (i) that PHF in Alzheimer's disease brain cross-react with a structural epitope/s present on native rabbit IgG, and (ii) that the cross-reactivity with PHF is probably due to tau.

Aged↗

PubMed1994

Dephosphorylation of Alzheimer's disease abnormally phosphorylated tau by protein phosphatase-2A.

Microtubule-associated protein tau is abnormally hyperphosphorylated in the brain of patients with Alzheimer's disease, and is the major protein subunit of paired helical filaments. There is also a significant pool of non-paired helical filament abnormally phosphorylated tau in Alzheimer's disease brain. In the present study, the site-specific dephosphorylation of this Alzheimer's disease abnormally phosphorylated tau by protein phosphatase-2A was studied and compared with that by protein phosphatase-2B. The dephosphorylation was detected by its interaction with several phosphorylation-dependent antibodies to various abnormal phosphorylation sites. Protein phosphatase-2A was able to dephosphorylate the abnormally phosphorylated tau at Ser-46, Ser-199, Ser-202, Ser-396 and Ser-404, but not at Ser-235 (the amino acids are numbered according to the largest isoform of human tau, tau441). Two major types of protein phosphatase-2A, protein phosphatase-2A1 and -2A2, dephosphorylated the abnormally phosphorylated tau at approximately the same rate. After the abnormally phosphorylated tau was dephosphorylated by protein phosphatase-2A, its relative mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis increased. The dephosphorylation of the abnormal tau by protein phosphatase-2A1 and -2A2 was markedly stimulated by Mn2+. These results suggest that tau dephosphorylation is catalysed by protein phosphatase-2A in addition to protein phosphatase-2B. A deficiency of either protein phosphatase-2A or -2B, or both, may be involved in abnormal phosphorylation of tau in Alzheimer's disease.

Aged↗

PubMed1994

Alzheimer's disease abnormally phosphorylated tau is dephosphorylated by protein phosphatase-2B (calcineurin).

Abnormally hyperphosphorylated tau is the major protein subunit of paired helical filaments in Alzheimer brains. We have examined its site-specific dephosphorylation by different protein phosphatases. Dephosphorylation of tau was monitored by its interaction with several phosphorylation-dependent antibodies. Alzheimer tau was dephosphorylated by brain protein phosphatase-2B at the abnormally phosphorylated sites Ser46, Ser199, Ser202, Ser235, Ser396, and Ser404, and its relative mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis shifted to that of normal tau. Protein phosphatases-1 and -2A could dephosphorylate only some of the above six phosphorylation sites. These results indicate that protein phosphatase-2B might be involved in hyperphosphorylation of tau in Alzheimer's disease.

Alzheimer Disease↗

PubMed1994

Pathological profile in chronic suppurative otitis media--the regional experience.

Peroperative findings in 145 consecutive cases of chronic suppurative otitis media, operated at Civil Hospital, Karachi were recorded. The mean age was 24 years. More than half of the patients (51%) had subtotal perforations and majority had damage of more than one ossicle. Involvement of all the three ossicles was seen in 30 (40%) cases. The frequency of damaged malleus along with incus was higher than other series. Granulations and subtotal perforations were commoner and could account for it. Extensive cholesteatoma was present in 30 cases.

Adolescent↗

PubMed1993

Microtubule-associated protein tau. Abnormal phosphorylation of a non-paired helical filament pool in Alzheimer disease.

The major protein subunit of the paired helical filaments (PHF) of Alzheimer disease (AD) is the microtubule-associated protein tau. Tau is a family of phosphopolypeptides that are abnormally phosphorylated in PHF. In this study, a non-PHF pool of tau abnormally phosphorylated at Ser-199/202, and tau not phosphorylated at this site (AD P-tau and AD tau, respectively) were isolated from the 27,000 x g to 200,000 x g fraction of AD brain homogenate by extraction in 8 M urea, followed by dialysis against Tris buffer. AD P-tau and AD tau were further purified and separated from each other by acid precipitation, glial fibrillary acidic protein affinity chromatography, and phosphocellulose chromatography. The resulting AD P-tau and AD tau preparations were free of cytoskeletal proteins, ubiquitin, and beta-amyloid peptide. Immunochemical and morphological analysis of AD P-tau preparations revealed that most of the protein was of non-PHF origin. The AD P-tau was about 3-4-fold (approximately 8 mol P04/mol protein, M(r) 41,318) more phosphorylated than cytosolic tau from AD and control brains. Unlike PHF, the AD P-tau lacked ubiquitin. In AD brain the levels of cytosolic tau were about half of those in control aged cases. These findings suggest that the abnormal phosphorylation of tau in AD occurs in the cytosol.

Aged↗

PubMed1993

Peptide compositions of the cerebrovascular and senile plaque core amyloid deposits of Alzheimer's disease.

The pathological findings of Alzheimer's disease include amyloid deposition in cerebral blood vessels and in senile plaques. Both deposits are known to include peptides that contain a common sequence. Both forms of amyloid were isolated and their peptide compositions were determined. The peptides were resolved by size-exclusion chromatography in 70% formic acid, and reverse-phase chromatography in 60% formic acid, 0-40% acetonitrile. Senile plaque amyloid cores contain about 25% protein, about 70% of which is composed of peptides containing the beta-amyloid sequence. Amino-terminal sequencing of the core amyloid peptides (CAPs) revealed extensive amino-terminal heterogeneity, with variable amounts of blocked amino termini. Matrix-assisted, laser-desorption-time-of-flight mass spectrometry of the CAP mixture revealed an array of peptides the molecular weights of which corresponded to peptides beginning with each of the first 11 amino acids of the beta-peptide sequence and ending with Ala-42 of that sequence. The carboxyl-terminal residues were identified by tandem mass spectrometry of chymotrypsin digests. CAP possessed a minor degree of carboxyl-terminal heterogeneity. Cerebrovascular amyloid peptides (CVAPs) possessed minor degrees of both amino- and carboxyl-terminal heterogeneity. The major CVAP commenced at Asp-1 and ended at Val-40. Minor components of CAP possessed masses of 8000-9000 Da and the same amino-terminal residues as the major components of CAP. They may be precursors to the smaller CAPs. The differences in amino termini and carboxyl termini of CAPs and CVAPs suggest that the two types of amyloid form by different pathways, on which they encounter different proteases.

Alzheimer Disease↗

PubMed1993

Alzheimer neurofibrillary tangles contain 2.1 nm filaments structurally identical to the microtubule-associated protein tau: a high-resolution transmission electron microscope study of tangles and senile plaque core amyloid.

Alzheimer neurofibrillary tangles (NFT) and senile plaque core amyloid (SPCA) isolated from the brain of patients with Alzheimer's disease were freeze-dried and replicated with a new platinum-carbon (Pt-C) vertical deposition method for high-resolution transmission electron microscopy (TEM). The resolution of this vertical Pt-C replication method is superior to either of the more conventional 20 degrees rotary replication or 45 degrees unidirectional replication methods and is dependent on the Pt-C film thickness coating the specimen. The paired helical filaments (PHF) observed within the tangles were right-handed helices with a fairly regular twist period averaging 79.3 +/- 5.9 nm and a fairly regular maximum width averaging 14.9 +/- 1.0 nm. The PHF regions of minimum width were not regular and fell into three size categories: 2.4 +/- 0.3 nm, 4.9 +/- 0.6 nm and 9.6 +/- 1.4 nm. In addition to the PHF found in the tangles, a new filament was found within all the tangles. These 2.1 +/- 0.2 nm diameter filaments were triple-stranded left helices with 1.0 +/- 0.2 nm diameter strands with a structure identical to bovine tau. Like bovine tau polymer a number of filaments (130 nm to 238 nm) were longer than a fully stretched tau monomer of 96 nm. Images of neuritic senile plaque core amyloid (SPCA) showed that amyloid had a more solid appearance than the NFT and its branched filament structures were unlike the approximately 2.1 nm diameter filaments or the PHF found in NFT.

Alzheimer Disease↗

PubMed1993

The organization of the microtubule associated protein tau in Alzheimer paired helical filaments.

The structural relationship of the microtubule associated protein tau to paired helical filaments (PHF) was examined by high resolution transmission electron microscopy (TEM) without treatment with any chemical fixatives. Neurofibrillary tangles (NFT) were isolated in the absence of detergent from Alzheimer diseased brains, were freeze-dried, and were vertically platinum-carbon replicated for TEM. The PHF we observed made one helical turn (L) in 74 +/- 8.5 nm and had a wide region (W) of 14.8 +/- 0.6 nm similar to PHF previously modeled with a periodic morphology. The PHF thin region (T) measured approximately 2.4 nm, approximately 4.9 nm, approximately 7.4 nm and approximately 9.7 nm and the most often observed width was approximately 2.4 nm. No surface features regularly divide the PHF into two filaments. Morphologically the PHF are thin helical ribbons with an often observed thickness of approximately 2.4 nm. At high magnification, approximately 1.0 nm and some approximately 0.4 nm strands identical to normal and denatured tau monomer covered PHF surfaces and were aggregated in non-periodic fashion. Bovine tau polymer approximately 2.1 nm diameter filaments, trapped on a filter, were partially heat denatured and showed some of the morphological features of PHF.

Alzheimer Disease↗

PubMed1993

Stability and control of a frontal four-link biped system.

A conceptual model for studying the involvement of the central nervous system (CNS) in the performance of lateral swaying movements is described. The model is based on a four-link planar biped that approximates gross human locomotion in the frontal plane. The viscoelastic function of the musculoskeletal system provides a linear controller for the system. Such an intrinsic controller can effectively duplicate simple well-learned tasks in the absence of higher level CNS feedback. This hypothesis is supported by comparing the proposed controller with two neurophysiologically involved linear decoupling schemes. Reference trajectories for swaying commands are recorded from experiments conducted in the Gait Analysis Laboratory of the Ohio State University Hospitals. These reference trajectories are inputs to all three controllers. The viability of intrinsic feedback scheme in the execution of swaying tasks is demonstrated via comparison of responses from the three controllers.

Central Nervous System↗

PubMed1993

Phosphoprotein phosphatase activities in Alzheimer disease brain.

Microtubule-associated protein tau is known to be hyperphosphorylated in Alzheimer disease brain and this abnormal hyperphosphorylation is associated with an inability of tau to promote the assembly of microtubule in the affected neurons. Our previous studies demonstrated that abnormally phosphorylated tau could be dephosphorylated after treatment with alkaline phosphatase, thereby suggesting that the abnormal phosphorylation of tau might in part be the result of a deficiency of the phosphoprotein phosphatase system in patients with Alzheimer disease. In the present study we used 32P-labeled phosphorylase kinase and poly(Glu, Tyr) 4:1 as substrates to measure phosphoprotein phosphatase activities in Alzheimer disease and control brains. The activities of phosphoseryl/phosphothreonyl-protein phosphatase types 1, 2A, 2B, and 2C and of phosphotyrosyl-protein phosphatase in frontal gray and white matters from 13 Alzheimer brains were determined and compared with those from 12 age-matched control brains. The activities of type 1 phosphatase and phosphotyrosyl phosphatase in gray matter and of type 2A phosphatase in both gray and white matters were significantly lower in Alzheimer disease brains than in controls. These findings suggest that the hyperphosphorylation of tau in Alzheimer disease brain could result from a protein dephosphorylation defect in vivo. The decrease in the phosphatase activities in Alzheimer disease might also be involved in the formation of beta-amyloid by augmenting the amyloidogenic pathway processing of beta-amyloid precursor protein.

Adult↗

PubMed1993

Molecular pathology of Alzheimer neurofibrillary degeneration.

The most characteristic brain lesion of Alzheimer disease is the accumulation of paired helical filaments (PHF) in the affected neurons. Based on solubility in detergents there are two general populations of PHF, the readily soluble (PHF I) and the sparingly soluble (PHF II) types. The major polypeptides of PHF are the microtubule associated protein tau. Tau in PHF is present in abnormally phosphorylated forms. In addition to the PHF, the abnormal tau is also present in unpolymerized form in the AD brain. Small amounts of ubiquitin (%) are associated with PHF II but neither with PHF I nor with the unpolymerized abnormally phosphorylated tau in AD brain. Furthermore, the pretangle neurons can readily be immunolabeled for abnormally phosphorylated tau but not for ubiquitin. The level of tau in neocortex is several-fold higher than in AD aged control cases, but this increase is in the form of the abnormally phosphorylated protein. The microtubule associated proteins from AD brain do not promote the assembly of microtubules in vitro, whereas the in vitro dephosphorylated PHF polypeptides stimulate the binding of GTP to the exchangeable site of tubulin and the assembly of microtubules. In vitro the phosphate groups in PHF are less accessible than those of tau to alkaline phosphatase. It is suggested that a defect in the protein phosphorylation/dephosphorylation system leads to hyperphosphory-lation of tau. The altered tau contributes to a microtubule assembly defect and consequently compromises the axoplasmic flow and leads to neuronal degeneration.

Alzheimer Disease↗

PubMed1992

Tau and ubiquitin in the human hypothalamus in aging and Alzheimer's disease.

Immunocytochemical staining of hypothalamic cell groups with four antibodies to Alzheimer paired helical filaments (PHF) (i.e., anti-PHF serum 60e and monoclonal antibody (mAb) Alz-50, both directed against normal and abnormally phosphorylated tau; mAb tau-1, which recognizes tau; and mAb 3-39 to PHF, which recognizes the carboxy terminal domain of ubiquitin) revealed a clear distinction between 12 Alzheimer's disease (AD) patients and seven controls in the hypothalamus. Dystrophic neurites, which appeared to be the most specific components in AD, were most conspicuous after Alz-50 staining. However, Alz-50 also stained neuronal cytoplasm and normal, thin, beaded neurites in the paraventricular nucleus (PVN) of controls, even of young cases. This staining was clearly distinct from the staining of cytoplasm and dystrophic neurites in the PVN of Alzheimer patients. The abundant staining of dystrophic neurites and cell bodies in the nucleus tuberalis lateralis (NTL) in AD, in which no neuronal loss is observed, suggests that alterations in cytoskeletal markers do not necessarily indicate impending cell death. Moreover, the cytoskeletal changes in the NTL, sexually dimorphic and suprachiasmatic nuclei in AD indicate that this condition is not restricted to cortical areas or nuclei projecting to the cortex. Consequently, the pathophysiological implications of cytoskeletal staining in AD are at present far from clear. The human hypothalamus may not only provide a better insight into the pathogenesis of Alzheimer's disease, but could also be of help in the neuropathological diagnosis of this condition.

Aged↗

PubMed1992

Effect of buffer constituents on the determination of therapeutic proteins by capillary electrophoresis.

Capillary electrophoresis has proved to be a versatile method for the determination of proteins, peptides and amino acids in pharmaceutical formulations. For quantification of the capillary electrophoresis data, however, significant errors may result if the analysis is performed using improper separation conditions. The peak area response for protein analytes, which is generally low in conventional UV detection, may also vary dramatically depending on the nature of the buffer used in the separation. This paper describes the effects of various buffer constituents and analytical conditions on the capillary electrophoretic separation and quantification of a humanized monoclonal antibody in bulk form and in a typical therapeutic formulation. For optimum peak area response and reproducibility, protein derivatization with an appropriate chromophore (e.g., fluorescamine) and separation in the presence of a moderate ionic strength buffer containing lithium chloride, tetramethylammonium chloride or trimethylammonium propylsulfonate is recommended. General guidelines for the determination of proteins by capillary electrophoresis and a rationale for the use of internal standards to improve the quantification of data are also discussed.

Antibodies, Monoclonal↗

PubMed1992

Assay of protein drug substances present in solution mixtures by fluorescamine derivatization and capillary electrophoresis.

A method is described to enhance the resolution and detection sensitivity of proteins, peptides, and amino acids in capillary electrophoretic analysis of solution mixtures. The method consists of derivatizing the analytes with fluorescamine, which is normally used as a fluorogenic reagent for compounds containing a reactive primary amine functional group, and then using the derivative as an ultraviolet chromophore to enhance detection sensitivity (measured at 280 nm) in capillary electrophoresis. The results demonstrated a significant improvement in the separation and detection sensitivity of the derivatized analytes as compared to their underivatized counterparts. The use of chromophores, such as fluorescamine, in capillary electrophoresis facilitates the analysis of components of solution mixtures, such as pharmaceutical formulations, that could not be resolved and/or detected by conventional capillary electrophoresis procedures.

Antibodies, Monoclonal↗

PubMed1992

Detection of point mutations in codon 331 of mitochondrial NADH dehydrogenase subunit 2 in Alzheimer's brains.

Point mutations in codon 331 of mitochondrial NADH dehydrogenase subunit 2 (ND2) were detected in 10 of 19 Alzheimer's brains but not in 11 normal brains. The same mutations were also detected in 2 of 6 patients with amyotrophic lateral sclerosis (ALS). However, neurofibrillary tangles and neuritic plaques characteristic of Alzheimer's disease were found histologically in the brain of one ALS patient who was positive of the mutation. The finding suggests that a point mutation in ND2 is a potential risk factor for Alzheimer's disease.

Adult↗