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Biomedical subjects

S A Ahmed

Publications and source records attributed to S A Ahmed.

At least 19 recordsLinked to original sources
PubMed2000

Induced minor malformations in the neocortex of normal mice do not alter immunological functions.

The interactive relationship between the CNS and the immune system is well established. Major lesions in the brain have been shown to affect immune response. However, whether minor, focal lesions (ectopias), as seen in autoimmune mice, may induce alterations in the immune system is unknown. To address this point, ectopic lesions in the neocortex were induced in neonatal DBA/2 mice (Induced minor malformations; IMM) and their immune capabilities were assessed at adulthood. Serum was collected from each animal and analyzed for the presence of autoantibodies. In addition, splenic lymphocytes and thymocytes were collected to ascertain proliferative capabilities and to assess for possible phenotypic changes in lymphocyte subsets. Mice with IMM did not manifest IgG autoantibodies against cardiolipin, dsDNA or brain membrane antigens. Total lymphocyte cellularity was not affected. The induction of cerebrocortical ectopias did not impair the ability of splenic and thymic lymphocytes to proliferate in response to anti-CD3 antibodies or Concanavalin-A (Con-A) as determined by non-radioactive (Alamar Blue) and radioactive (3H-thymidine) assays. Moreover, no difference in proliferation of unstimulated and anti-CD3-stimulated splenic lymphocytes exposed to rIL-2 or rIL-7 was observed. Flow cytomeric analysis of a variety of cell surface antigens, indicated that there was no difference in lymphocyte subsets between control and IMM groups. Therefore, we conclude that induced IMM lesions in the CNS of normal DBA/2 mice do not alter immune functions.

Animals↗
PubMed1999

Comparison of multiple assays for kinetic detection of apoptosis in thymocytes exposed to dexamethasone or diethylstilbesterol.

BACKGROUND: Techniques to measure apoptosis are used to study a wide spectrum of conditions, from acquired immune deficiency syndrome (AIDS) to cancer to autoimmune diseases. Therefore, a critical comparison of common assays for apoptosis is warranted. METHODS: The kinetics of apoptosis induction in dexamethasone (DEX)-exposed thymocytes was examined after 2, 4, 8, 12, 26-28, and 48-50 h of culture. An additional aim was to ascertain whether a similar thymic atrophy-inducing hormone, diethylstilbestrol (DES), also directly induces thymocyte apoptosis. Apoptosis was evaluated by flow cytometric examination of cells stained with propidium iodide (PI), 7-aminoactinomycin D (7-AAD), or fluorescein isothiocyante (FITC)-annexin; by forward-and side-scatter (FS, SS) analysis, cell-size analyzer; and through cytopathologic examination. RESULTS: After 4 h of DEX exposure, apoptosis was evident by 7-AAD, annexin, and cytopathological assays, but no cells with sub-diploid DNA content were evident by PI analysis. Maximal apoptosis was evident by all the above flow cytometric techniques at 12 h after DEX exposure. The 7-AAD and annexin assays, which allow discrimination between early apoptosis and late apoptosis/necrosis, were comparable and identified similar apoptotic populations. Appearance of a FSlow/SSincreased population was evident only after 12 h of DEX exposure. Apoptosis could not be detected by any of the above assays in thymocytes exposed to various doses of DES. CONCLUSION: Two of the six assays, 7-AAD and annexin, were similar in detecting apoptosis at an early kinetic time point. Results of both assays were comparable at all time points studied. Our studies imply that DEX and DES induce thymic atrophy, in vivo, by different mechanisms.

Animals↗
PubMed1999

Species, interindividual, and tissue specificity in endocrine signaling.

The activity of endocrine-active agents exhibits specificity at many levels. Differential responsiveness to these agents has been observed between different species and extends to interindividual differences within a species and between different tissues as well. In cases where they have been identified, the biologic and molecular mechanisms underlying this specificity are quite diverse. Determinants of species specificity include differences that exist in receptor binding, gene transcription, and cellular responses to endocrine-active compounds between species. Interindividual differences in responsiveness may be determined at the level of genetic polymorphisms in hormone-metabolizing enzymes, hormone receptors, and in those genes that are transactivated by these receptors, as well as during changing windows of susceptibility that occur as a function of age, such as prenatal and postmenopausal exposures. Extrinsic factors such as diet can also impact individual susceptibility to endocrine-active agents. Tissue-specific determinants of susceptibility are well documented, but little is known regarding the mechanisms underlying these different responses. Differences in the expression of accessory proteins for steroid hormone receptors and different patterns of receptor expression, estrogen receptor alpha and estrogen receptor beta; for example, may contribute to tissue specificity, as may differences in the pattern of expression of other genes such as hormone-metabolizing enzymes. The use of animal model systems and development of appropriate mathematical models has the potential to yield additional valuable information for elucidating the role of these determinants of specificity at low-dose exposures and for improved risk assessments for the adverse health effects of endocrine-active compounds.

Age Factors↗
PubMed1999

Gender and risk of autoimmune diseases: possible role of estrogenic compounds.

A striking common feature of many autoimmune diseases in humans and experimental animals, despite differences in pathology, is that females are highly susceptible to autoimmune conditions compared to males. In several animal models, estrogens promote, whereas androgens abrogate, B-cell-mediated autoimmune diseases. To understand mechanisms by which estrogens regulate autoimmunity, it is first necessary to decipher estrogen effects on the normal immune system. Estrogen treatment of nonautoimmune mice diminished lymphocyte numbers in both developmental and mature lymphoid organs. Estrogen dysregulated T- and B-cell balance by inducing selective T-cell hypoactivity and B-cell hyperactivity. Even though estrogen did not alter the relative percentages of splenic T-cell subsets, splenic lymphocytes had a reduced proliferative response to T-cell stimulants and were refractory to rescue from activation-induced apoptosis compared to cells from placebo-treated mice. In contrast, estrogen induced B-cell hyperactivity (promoted autoantibodies to double-stranded DNA and phospholipids, increased numbers of plasma cells, and increased autoantibody yield per B cell). Note that treatment of normal mice with estrogen can alter T- and B-cell regulation and overcome B-cell tolerance to result in autoimmunity in normal individuals. Could environmental estrogens promote some human autoimmune disorders? Is there a link between environmental estrogens and autoimmune disorders, especially since these disorders are reported possibly more frequently? These provocative questions warrant investigation. Our findings on immunomodulatory effects may serve as a benchmark to examine whether endocrine-disrupting chemicals will have similar immunologic effects.

Animals↗
PubMed1999

Role of cortisol in the deterioration of glucose tolerance in Sudanese pregnant women.

OBJECTIVE: To study the state of insulin resistance which occurs in pregnancy and the possible role of cortisol in augmentation of this state. DESIGN: Case-control study. SETTING: Khartoum teaching hospital, Khartoum north hospital, Soba hospital, Ibrahim Malek hospital, maternity hospital and Fath-Elbasheer referral centre from January to August 1996. SUBJECTS: Thirty pregnant women with gestational diabetes mellitus(GDM) and thirty pregnant women with impaired glucose tolerance(IGT) were compared with thirty control pregnant women with normal glucose tolerance. RESULTS: The GDM and the IGT groups were found to have significantly higher levels of serum cortisol than that of the control group [937.2 +/- 79.4 nmol/l (mean +/- S.E.) and 794.2 +/- 60.5 vs 597.3 +/- 30.9 respectively, p < 0.0006]. CONCLUSION: Cortisol has a great role to play in the deterioration of glucose tolerance in pregnancy.

Adult↗
PubMed1998

Estrogen increases the number of plasma cells and enhances their autoantibody production in nonautoimmune C57BL/6 mice.

The immunological consequences of chronic estrogen exposure in normal individuals are not known, particularly in relation to B cells. In this study, by employing ELIspot, image cytometry, flow cytometry, cytology, and ELISA, we show that long-term exposure of normal mice to estrogen activates B cells to produce higher numbers of not only immunoglobulin-producing cells, but also autoantibody-producing cells. Estrogen promoted a decrease in B220(+) splenic lymphocytes, but resulted in a 10-fold increase in plasma cells. Further, the output of immunoglobulins including autoantibodies from individual plasma cells from estrogen-exposed mice was markedly increased, suggesting B cell hyperactivity. Importantly, our findings show that treatment of normal mice, solely with estrogen, can override B cell tolerance and promote autoreactive B cells in normal individuals.

Animals↗
PubMed1998

Benzo[a]pyrene-induced hypocellularity of the pronephros in tilapia (Oreochromis niloticus) is accompanied by alterations in stromal and parenchymal cells and by enhanced immune cell apoptosis.

Numerous reports indicate that carcinogenic polycyclic aromatic hydrocarbons (PAH) are mammalian immunotoxicants. These environmental contaminants are widely distributed in both freshwater and costal marine ecosystems where they have been found to bioaccumulate in aquatic species, yet limited information exists regarding potential adverse effects of specific PAH on fish immune function. In the present report, Oreochromis niloticus fish (tilapia) were exposed by intraperitoneal injection to 5, 25, or 50 mg/kg of the PAH, benzo[a]pyrene (B[a]P). Histopathologic evaluation of the primary hematopoietic compartment of fish, the pronephros, demonstrated increased vacuolation of both stromal and parenchymal cells, reduction of lymphoid elements, and immune cell apoptosis. Total pronephros cell counts were diminished in a dose-dependent manner by the chemical exposure. The oxidative metabolic burst in phorbol myristate acetate (PMA)-simulated macrophages isolated from the pronephros was significantly inhibited by B[a]P, but only at the highest dose level employed. The phagocytic capacity of pronephros macrophages was not altered by the chemical treatment.

Animals↗
PubMed1998

Mutation of an active site residue of tryptophan synthase (beta-serine 377) alters cofactor chemistry.

To better understand how an enzyme controls cofactor chemistry, we have changed a tryptophan synthase residue that interacts with the pyridine nitrogen of the pyridoxal phosphate cofactor from a neutral Ser (beta-Ser377) to a negatively charged Asp or Glu. The spectroscopic properties of the mutant enzymes are altered and become similar to those of tryptophanase and aspartate aminotransferase, enzymes in which an Asp residue interacts with the pyridine nitrogen of pyridoxal phosphate. The absorption spectrum of each mutant enzyme undergoes a pH-dependent change (pKa approximately 7.7) from a form with a protonated internal aldimine nitrogen (lambdamax = 416 nm) to a deprotonated form (lambdamax = 336 nm), whereas the absorption spectra of the wild type tryptophan synthase beta2 subunit and alpha2 beta2 complex are pH-independent. The reaction of the S377D alpha2 beta2 complex with L-serine, L-tryptophan, and other substrates results in the accumulation of pronounced absorption bands (lambdamax = 498-510 nm) ascribed to quinonoid intermediates. We propose that the engineered Asp or Glu residue changes the cofactor chemistry by stabilizing the protonated pyridine nitrogen of pyridoxal phosphate, reducing the pKa of the internal aldimine nitrogen and promoting formation of quinonoid intermediates.

Aspartic Acid↗
PubMed1998

Mutations in the contact region between the alpha and beta subunits of tryptophan synthase alter subunit interaction and intersubunit communication.

Interaction between the alpha and beta subunits of tryptophan synthase leads to mutual stabilization of the active conformations and to coordinated control of the activities of the two subunits. To elucidate the roles of specific residues in the interaction site between the alpha and beta subunits, mutant alpha and beta subunits were constructed, and the effects of mutation on subunit interaction and intersubunit communication were determined. Mutation of either alpha subunit Asp56 (alphaD56A) or beta subunit Lys167 (betaK167T), residues that interact in some crystal structures of the tryptophan synthase alpha2beta2 complex, decreases the ability of the alpha subunit to activate the beta subunit and alters the reaction and substrate specificity of the beta subunit. Partial conformational repair is provided by alpha-glycerol 3-phosphate, a ligand that binds to the alpha subunit, or by Cs+ or NH4+, ligands that bind to the beta subunit. Mutation of beta subunit Arg175 (betaR175A), a residue that interacts with alpha subunit Pro57 in some structures, has much smaller effects on activity but results in a 15-fold increase in the apparent Kd for dissociation of the alpha and beta subunits. Replacement of the single tryptophan in the beta subunit by phenylalanine (W177F) has only small effects on activity but increases the apparent subunit dissociation constant approximately 10-fold. The most important conclusions of this investigation are that interaction between alphaAsp56 and betaLys167 is important for intersubunit communication and that mutual stabilization of the active conformations of the two subunits is impaired by mutation of either residue.

Amino Acid Substitution↗
PubMed1998

Learning in year-old female autoimmune BXSB mice.

BXSB/ MpJ-Yaa and NZB/BINJ mice have been used as animal models for both developmental learning disability and systemic autoimmune disease. Approximately 40-60% of these animals show ectopic clusters of neurons in Layer I of cortex similar to those found in postmortem analyses of human dyslexics, and all exhibit an autoimmune condition similar to systemic lupus erythematosus (SLE) in humans. The expression of immune disease in the BXSB strain, unlike in humans, is more severe in males than females. Most previous studies have examined the behavioral sequelae of neocortical ectopias at a relatively young age, when the BXSB females (unlike the male BXSB and female and male NZBs) are not yet showing high titers of autoantibodies associated with their lupus-like form of autoimmune disease. This study examined the behavior of BXSB females at an age subsequent to autoimmune disease onset. When contrasted with younger animals, year-old BXSB females showed good learning behavior, with no differences in Lashley maze learning and remarkably good performance in a visual discrimination learning task. These results are consistent with other data which indicate that many types of learning behavior are apparently unperturbed by systemic autoimmune disease. Results also showed significant interactions between a measure of lateral paw preference and the presence or absence of ectopias in Lashley maze learning. Animals without ectopias that exhibited a right lateral paw preference showed the greatest number of errors on a number of test measures. These findings support previous results indicating that behavioral effects associated with ectopias may vary based upon the behavioral laterality of affected animals.

Age Factors↗
PubMed1998

Water management practices in rural and urban homes: a case study from Bangladesh on ingestion of polluted water.

Although Bangladesh has achieved remarkable success in extending the availability of hand pumped and piped water, unsafe water is still ingested. This brief study attempted to assess water management practices in rural and urban homes in Bangladesh so as to establish the routes by which unsafe water is ingested, to examine methods of collection and storage, and determine why unsafe water sources are used when unsafe supplies are available. Forty-eight rural and forty-five urban slum households were studied. Observations, interviews and water quality investigations were conducted. The results show that the respondents were aware that hand pump/tap water is safe and took care to use these safe sources for drinking purposes. However, they continued to use surface water for non-drinking activities such as bathing, washing and rinsing their mouths. Reasons were given that it was a traditional practice to bathe in surface water and was more enjoyable. One of the reasons given for not using hand pumped water to wash clothing and food was that such groundwater caused staining. Bacteriological results from such ingested water showed the quality, especially in rural areas, to be poor. Results also showed the internal surfaces of the base of storage containers to be heavily contaminated with bacteria. This showed that water that was safe when it was first drawn would became contaminated during storage. This study had a limited scope; much further research is needed to find what determines and how water becomes contaminated in containers. These factors include how to reduce contamination of water, in particular the relationship between growth of bacteria in stored water and the material from which the container is made and how to improve the overall quality of water. On the management side, studies could be conducted as to how to improve people's understanding of the use of water for domestic purposes and its safe management.

Adult↗
PubMed1997

A dye-based lymphocyte proliferation assay that permits multiple immunological analyses: mRNA, cytogenetic, apoptosis, and immunophenotyping studies.

Alamar Blue in the microenvironment of activated cells, undergoes color change and also becomes fluorescent. By using the Alamar Blue dye, we have reported a non-radioactive colorimetric assay to indirectly determine proliferation of murine lymphocytes. We further show that the pattern of mitogen-induced proliferation assessed fluorometrically was comparable to the 3H-thymidine incorporation assay (3H-Tdr assay). Of practical importance is that the color/fluorescence changes were stable at 4 degrees C in the dark for 3-4 weeks. In immunological studies, it is important to further analyze lymphocytes that have undergone activation and/or proliferation. This is not possible with the standard 3H-Tdr assay, which requires lysis of cells. In contrast, the Alamar Blue-based non-radioactive assay does not require cell lysis. We therefore tested the hypothesis that further analysis of lymphocytes is possible, after assessing the proliferation using Alamar Blue. Following assessment of proliferation in a 72-h culture, the Alamar Blue dye was washed-off and cells were re-utilized to perform additional immunological analysis. Short-term exposure of lymphocytes to Alamar Blue was not detrimental to lymphocytes, as assessed by trypan blue exclusion and the propidium iodide (PI) assays. Exposure of dexamethasone-treated cells to Alamar Blue did not interfere with the performance of apoptosis assays, such as flow cytometric analysis of PI-stained cells and microscopic examination of ethidium bromide/acridine orange-stained cells. In addition, prior exposure of lymphocytes to Alamar Blue did not affect the analysis of chromosomal aberrations or the visualization of cell surface antigens by flow cytometry. Further, the expression of cytokine mRNA in lymphocytes previously exposed to Alamar Blue was similar to unexposed cells. Together, a notable advantage of this assay is that it now enables the investigator to maximize information by following or correlating proliferation with other immunologic events in the same cells.

Animals↗
PubMed1997

Crystal structures of a mutant (betaK87T) tryptophan synthase alpha2beta2 complex with ligands bound to the active sites of the alpha- and beta-subunits reveal ligand-induced conformational changes.

Three-dimensional structures are reported for a mutant (betaK87T) tryptophan synthase alpha2beta2 complex with either the substrate L-serine (betaK87T-Ser) or product L-tryptophan (betaK87T-Trp) at the active site of the beta-subunit, in which both amino acids form external aldimines with the coenzyme, pyridoxal phosphate. We also present structures with L-serine bound to the beta site and either alpha-glycerol 3-phosphate (betaK87T-Ser-GP) or indole-3-propanol phosphate (betaK87T-Ser-IPP) bound to the active site of the alpha-subunit. The results further identify the substrate and product binding sites in each subunit and provide insight into conformational changes that occur upon formation of these complexes. The two structures having ligands at the active sites of both alpha- and beta-subunits reveal an important new feature, the ordering of alpha-subunit loop 6 (residues 179-187). Closure of loop 6 isolates the active site of the alpha-subunit from solvent and results in interaction between alphaThr183 and the catalytic residue alphaAsp60. Other conformational differences between the wild type and these two mutant structures include a rigid-body rotation of the alpha-subunit of approximately 5 degrees relative to the beta-subunit and large movements of part of the beta-subunit (residues 93-189) toward the rest of the beta-subunit. Much smaller differences are observed in the betaK87T-Ser structure. Remarkably, binding of tryptophan to the beta active site results in conformational changes very similar to those observed in the betaK87T-Ser-GP and betaK87T-Ser-IPP structures, with exception of the disordered alpha-subunit loop 6. These large-scale changes, the closure of loop 6, and the movements of a small number of side chains in the alpha-beta interaction site provide a structural base for interpreting the allosteric properties of tryptophan synthase.

Binding Sites↗
PubMed1997

Importance of conserved and variable C-terminal residues for the activity and thermal stability of the beta subunit of tryptophan synthase.

To assess the functional roles of helix 13 and of the conserved and variable residues in the C-terminal region (residues 378-397) of the tryptophan synthase beta subunit, we have constructed four C-terminal truncations and 12 point mutations. The effects of these mutations on kinetic and spectroscopic properties and thermal stability are reported here. The mutant beta subunits all form stable alpha2beta2 complexes that have been purified to homogeneity. The mutant alpha2beta2 complexes are divided into two classes on the basis of activity in the reaction of L-serine with indole to form tryptophan. Class I enzymes, which have mutations at Arg-379 or Asp-381 or truncations (384-397 or 385-397), exhibit significant activity (1-38% of wild type). Class II enzymes, which have mutations at Lys-382 or Asp-383 or truncations (382-397 or 383-397), exhibit very low activity (<1% of wild type). Although Class II enzymes have drastically reduced activity in the reaction of L-serine with indole and an altered distribution of enzyme-substrate intermediates in the reaction of L-serine with beta-mercaptoethanol, they retain activity in the reaction of beta-chloro-L-alanine with indole. Correlation of the results with the three-dimensional structure of the alpha2beta2 complex suggests that Lys-382 and Asp-383 serve important roles in a proposed "open" to "closed" conformational change that occurs in the reactions of L-serine. Because mutant beta subunits having C-terminal truncations (383-397 or 384-397) undergo much more rapid thermal inactivation at 60 degrees C than the wild type beta subunit, the C-terminal helix 13 stabilizes the beta subunit.

Amino Acid Sequence↗
PubMed1997

Manipulation of behavioral disorders in autoimmune mice via prolactin.

Autoimmune mice perform poorly in two-way active avoidance tasks, and the extent of this performance deficit appears to be related to the extent of autoimmunity following developmental manipulations. In the current study, the pituitary hormone prolactin, which has immune-enhancing effects, was used to manipulate this behavioral disorder in adulthood. Prolatinergic manipulation may be achieved by the use of dopaminergic drugs. In two experiments, autoimmune NZB X NZW F1 (BW) mice received either pimozide (PIM; a D2 antagonist) or bromocriptine (CB154; a dopamine agonist) in their drinking water. Control subjects received plain water. Following treatment, subjects were tested in an activity monitor, and active avoidance learning. Circulating PRL levels, as measured by RIA, were significantly increased by PIM and significantly decreased by CB154. Neither drug affected circulating levels of autoantibodies to DNA or cardiolipin, a phospholipid. In Experiment 1, in which mice were tested at 12 weeks of age, after 6 weeks of drug treatment, PIM treated animals of both sexes showed significantly more failures to escape the shock in avoidance conditioning, while CB154 did not have significant effects. In Experiment 2, in which mice were tested at 16 weeks of age, after 12 weeks of drug treatment, CB154 treated females (males were not tested) showed significantly fewer failures to escape, while PIM did not have significant effects. The effects of PRL on behavior, and its relation to immune system function, are discussed.

Animals↗
PubMed1997

Dissociation equilibria of the tryptophan synthase alpha 2 beta 2 complex in saline buffer and guanidine isothiocyanate, as studied by sedimentation equilibrium.

The dissociation equilibria of Salmonella typhimurium tryptophan synthase alpha 2 beta 2 complex were studied via centrifugation of the complex to sedimentation equilibrium in neutral saline buffers containing 0 to 137 mM guanidine isothiocyanate (GuSCN). The resulting concentration gradients were analyzed in the context of an equilibrium model for sequential dissociation of two alpha subunits from a stable beta 2 subunit. Under the conditions of these experiments, the first dissociation constant alone could be evaluated at GuSCN concentrations < or = 100 mM, and the second dissociation constant alone could be evaluated at GuSCN = 137 mM. At intermediate GuSCN, both dissociation constants were sufficiently well defined to rule out the presence of a large equilibrium cooperative effect in the stepwise dissociation of the alpha subunits.

Bacterial Proteins↗