Biomedical subjects

Ashok K Singh

Publications and source records attributed to Ashok K Singh.

18 recordsLinked to original sources

PubMed2006

Glomerular epithelial cells transform to myofibroblasts: early but not late removal of TGF-beta1 reverses transformation.

Studies were carried out to determine whether epithelial-mesenchymal transformation (EMT), well described in renal tubular epithelial cells, also occurs in glomerular epithelial cells and whether it is reversible. To this effect, cultured glomerular epithelial cells were incubated with TGF-beta(1) and their transformation into myofibroblasts was studied. At 4 days, the cells altered their phenotype, as shown by a change in shape, an increase in intracellular staining for alpha-smooth muscle actin (alpha-SMA), a decrease in membrane staining for cytokeratin, and an increase in matrix deposition. Changing the medium after 4 days by excluding TGF-beta(1) and adding fetal bovine serum (FBS) [as a source of epidermal growth factor (EGF) and other growth factors] caused the cells to revert to their original epithelial phenotype. By contrast, when the medium was changed in the same manner after 8 days of exposure to TGF-beta(1), the cells did not revert but remained myofibroblastic. Staining the cells for expression of EGF receptor before and after exposure to TGF-beta(1) caused this receptor, originally present on the plasma membrane, to become partly intracellular after 4 days of TGF-beta(1) exposure and completely intracellular after 8 days of TGF-beta(1) exposure. Kidney sections from 2 models of renal mass reduction were stained. Loss of the epithelial marker (podocalyxin) staining and the acquisition of alpha-SMA staining was observed in the glomeruli. It is concluded that EMT takes place in glomerular epithelial cells in vivo and in vitro. In cultured glomerular epithelial cells, the process can be reversed by early, but not late intervention. It seems that TGF-beta(1) exposure progressively downregulates the EGF receptor on the membrane, rendering the cell refractory to EGF signals critical for maintaining the epithelial phenotype.

Actins↗

PubMed2006

Lipoprotein glomerulopathy: a new apolipoprotein E mutation with enhanced glomerular binding.

We describe a case of lipoprotein glomerulopathy, the second ever reported from the United States, in a Mexican man with a hitherto undescribed mutation in the apolipoprotein E gene (substitution of proline for arginine at position 147 [Arg147Pro]). In this patient, glomerular basement membranes showed double contours and circumferential mesangial extensions, suggesting that deposition of lipids could be injurious to endothelial cells. Immunofluorescence staining of thrombi was positive for apolipoprotein E and B. To study the reason for lipid deposition in glomeruli, we incubated normal human kidney sections with serum from the patient and a healthy control. Apolipoprotein E from the patient's serum showed binding to the glomerular capillary wall, but the control did not, suggesting enhanced binding of the mutated apolipoprotein E to glomerular capillaries. Apolipoprotein E genotyping by means of restriction endonuclease digestion of polymerase chain reaction-amplified genomic DNA showed it to be of the wild-type E3/E3.

Adult↗

PubMed2005

Effect of high glucose on superoxide in human mesangial cells: role of angiotensin II.

BACKGROUND/AIMS: Reactive oxygen species, and especially superoxide (O2*),have been implicated in diabetic nephropathy. O2* accumulation in cells is dependent on O2* production (by NADH/NADPH oxidase) as well as scavenging by superoxide dismutase (SOD) activity. This study was designed to investigate the effects of high glucose (HG) on O2* accumulation and SOD activity in human mesangial cells (HMC) and to determine if these effects are mediated by angiotensin II (Ang II). METHODS: HMC were incubated in media containing 10 mM glucose (control, C), 30 mM glucose (HG), 10 mM glucose + either 20 mM 2-deoxy-D-glucose (2-DG) or 20 mM mannitol (high mannitol, HM) (osmotic controls), or Ang II (10(-5) M). Ang II action was antagonized by employing 10(-4) M of Ang II receptor antagonists (losartan or irbesartan) or 10(-4) M of NADH/NADPH oxidase inhibitors [diphenyleneiodonium chloride (DPI) or apocynin]. Superoxide and total SOD activity were assayed using chemiluminescence of lucigenin. RESULTS: Incubation of HMC in HG resulted in a 1.6-fold increase in Ang I (p < 0.05) and a 1.4-fold increase in Ang II levels (p < 0.05) in cell lysates. These changes were accompanied by a >2-fold increase in O2* accumulation (p < 0.01), which was inhibited by losartan and irbesartan. Exogenous Ang II increased net O2* accumulation by 2.7-fold (p < 0.01), which was normalized by losartan and irbesartan. DPI and apocynin blocked the HG and Ang II-induced increases in O2* (p < 0.01). HG but not exogenous Ang II inhibited total SOD activity by 30%, which was not affected by losartan. CONCLUSION: High glucose increases O2* accumulation in HMC primarily by increasing its production via the Ang II-NADH/NADPH oxidase system.

Angiotensin II↗

PubMed2005

A novel mechanism for angiotensin II formation in streptozotocin-diabetic rat glomeruli.

Recent evidence suggests that the intrarenal renin-angiotensin system (RAS) may play an important role in the development of glomerular changes associated with diabetic nephropathy. In this study, the glomerular RAS was examined in male Sprague-Dawley rats made diabetic with streptozotocin (STZ), and the findings compared with those obtained in control nondiabetic rats. In diabetic rat glomerular extracts, angiotensinogen and angiotensin II (ANG II) levels were increased significantly by 2.2- and 1.9-fold, respectively, compared with nondiabetic controls. No significant differences in ANG I and angiotensin-converting enzyme (ACE) levels were observed between these groups. The HPLC analysis of the glomerular extracts demonstrated that exogenous ANG I was converted into various ANG peptides including ANG II, ANG1-9, and ANG1-7. A significant increase in formation of ANG II from exogenous ANG I was observed in STZ rats compared with control rats. Preincubation of glomerular extracts with captopril resulted in a 20-30% decrease in ANG II conversion from exogenous ANG I in diabetic and control rats. The possible role of ANG1-9 in formation of ANG II was examined by HPLC. Exogenous ANG1-9 in glomerular extracts was converted into ANG II, this conversion being significantly higher in STZ rats than in control rats. These findings provide new information that ANG1-9 is produced in rat glomerular extracts, can be converted to ANG II, and that this conversion is also stimulated in diabetic rat glomeruli. Thus this study demonstrates that in diabetic rats, glomerular ANG II levels are increased due to an increase in angiotensinogen and an increase in the formation of ANG II.

Angiotensin I↗

PubMed2005

Antagonism of vascular endothelial growth factor results in microvessel attrition and disorganization of wound tissue.

Vascular endothelial growth factor (VEGF) is a potent growth factor that is indispensable for the development of blood vessels in the fetus and for wound healing in adults. VEGF likely plays a role in maintaining the blood vessels once they have been formed. It is not clear, however, whether a low tissue VEGF (caused either by disease or by systemic administration of VEGF antagonists) can cause abnormalities in preexisting blood vessels, especially of wound tissue that requires high local levels of VEGF for healing. The present study investigated the effect of VEGF antagonism on blood vessels of foreign-body granulomas (a model of wound-healing tissue). Granulomas were induced by implanting perforated polyvinyl tubes into the subcutaneous tissue of rats and allowed to develop for 14 days, at which time the implanted tubes were completely encapsulated by the subcutaneous tissue. The encapsulated granulomas consisted of 3 distinct histological layers, of which the middle layer was well perfused by a rich supply of microvessels. Morphologically, the granuloma remained "stable" after developing for 14 days. At 1 week, VEGF levels in the granuloma fluid, which is in equilibrium with the interstitial fluid, were 25 times higher than in the plasma. VEGF levels in the granuloma fluid continued to increase for up to 3 weeks, reflecting the high dependence of the wound tissue on ambient VEGF levels. After injection of the VEGF receptor antagonist in the fully formed granuloma, the preexisting blood vessels in the middle layer regressed and underwent apoptosis, accompanied by expansion of the extracellular matrix (predominately collagen I) into areas normally devoid of matrix. We conclude that wound tissue is sensitive to ambient VEGF levels, and that a low VEGF condition resulting from VEGF receptor antagonism can disrupt the healing of wound tissue.

Animals↗

PubMed2004

Quantitative analysis of phytoestrogens in kudzu-root, soy and spiked serum samples by high-pressure liquid chromatography.

A sensitive and reliable HPLC method that allows simultaneous quantification of phytoestrogens extracted from kudzu-root and soy preparations, and serum samples has been developed. Kudzu-root and soy preparations were mixed with 5 microg flavone and 15 microg rutin (internal standards) and the phytoestrogens were extracted by using solid-phase (C18) extraction cartridges. Blank or spiked serum samples were extracted by using either C18 cartridges or trichloroacetic acid-methanol extraction. The extracts were analyzed by the HPLC equipped with a reverse-phase (250 x 4 mm, C18) column and UV, diode-array or MS detector. A linear gradient of acetic acid and acetonitrile provided excellent separation of glycoside and aglycone-phytoestrogens from kudzu root and soy preparations. The C18 cartridge extraction of serum yielded excellent recovery of both glycoside- and aglycone-phytoestrogens, while the trichloroacetic acid-methanol extraction yielded excellent recovery of glycoside but poor recovery of aglycone compounds. UV and MS detectors were suitable for phytoestrogen analysis in plant and serum samples, while the diode-array detector was suitable for generating the UV absorbance curve for phytoestrogens.

Chromatography, High Pressure Liquid↗

PubMed2004

Vascular factors altered in glucose-treated mesangial cells and diabetic glomeruli. Changes in vascular factors impair endothelial cell growth and matrix.

We studied the effect of a high glucose (HG) environment on the vascular factors that are secreted by mesangial cells, and regulate endothelial growth and mesangial matrix deposition. To this effect, we measured the vascular factors in the glomeruli of streptozotocin-induced diabetic kidneys and in mesangial cells exposed to a HG concentration. We then transferred the media of mesangial cells previously exposed to high glucose to cultured endothelial cells to study the effects on endothelial growth, matrix formation, and in vitro capillary proliferation. In 1-week diabetic kidneys, glomerular vascular endothelial growth factor (VEGF) and angiopoietin-1 were inhibited by 38 and 57%, respectively, but angiopoietin-2 was increased by 318%. We found similar results in mesangial cells exposed to HG. There was a decrease of VEGF (50% by enzyme immunoassay, 27% by mRNA), decrease of angiopoietin-1 (65% by mRNA), and a much greater increase of angiopoietin-2 (280% by immunoassay, 523% by mRNA). Compared to controls, the media of mesangial cells previously exposed to HG impaired endothelial cell growth by 61%, increased extracellular matrix by 100%, and decreased capillary formation by 90%. We conclude that high ambient glucose alters the secretion of vascular factors elaborated by mesangial cells, resulting in an expansion of the endothelial cell matrix and disruption of capillary structure.

Angiogenic Proteins↗

PubMed2004

Differential activation of NF kappa B/RelA-p50 and NF kappa B/p50-p50 in control and alcohol-drinking rats subjected to carrageenin-induced pleurisy.

BACKGROUND: Carrageenin (CAR) injection into the pleural cavity causes local inflammation called carrageenin-induced pleurisy (CAR-IP). Inflammation onset is characterized by an activation of pro-inflammatory NFkappaB, RelA-p50, while inflammation resolution is characterized by an activation of an anti-inflammatory NFkappaB, p50-p50, that re-establishes homeostasis, an essential process for an organism's survival. Although chronic alcohol intake disrupts inflammation, the mechanism behind the development of inflammatory disorder in alcoholics is not yet known. Therefore, the aim of this investigation was to study the effects of ethanol intake on CAR-IP and NFkappaB activation in pleural fluid neutrophils in P rats. METHODS: Alcohol-preferring, P rats were given free choice of alcohol (15% ethanol) and water or water alone (for control) for 15 days. Then, each rat was injected with 0.2 ml of 2% CAR into the pleural cavity under light ether anesthesia. At different time intervals after the CAR injection, rats were anesthetized and their blood and pleural fluid samples were collected. Pleural fluid inflammatory cells were identified with Turk's or Wright-Giemsa staining. Different cell types were sorted using a fluorescence-activated cell sorter. Pleural fluid neutrophils were examined for apoptosis and activation of the two NFkappaB subspecies. RESULTS: In control rats, fluid began to accumulate in the pleural cavity 0.5 h after, which peaked 24 h after, CAR injection. Then, the values declined gradually. The increase in pleural fluid correlated with RelA-p50 activation, while the decline in pleural fluid correlated with p50-p50 activation and apoptosis in neutrophils. In alcohol-drinking rats, pleural fluid remained elevated for up to 6 days after CAR injection. Neutrophils from alcohol-drinking rats exhibited suppressed apoptosis, augmented RelA-p50 activation, and suppressed p50-p50 activation. CONCLUSIONS: Alcohol intake prolonged inflammation in P rats. An alcohol-induced upregulation of RelA-p50 activation and downregulation of p50-p50 activation may be causally related to the alcohol-induced inflammation dysregulation.

Alcohol Drinking↗

PubMed2004

Effects of purified puerarin on voluntary alcohol intake and alcohol withdrawal symptoms in P rats receiving free access to water and alcohol.

Alcohol preferring (P) rats, given "free choice" of water, exhibited daily intake of 60-75 g of water/kg of body weight. When given "free choice" of water and 15% ethanol, P rats consumed 7-13 g of alcohol/kg. Their water intake decreased proportionally to the alcohol intake, but total fluid intake did not differ significantly. Alcohol withdrawal after 50 days of alcohol drinking caused withdrawal symptoms such as hypersensitivity, poor coordination, and tremors. A daily 50 mg/kg dose of puerarin (PU) caused approximately 50% suppression in alcohol intake, but did not affect body weight and food and total fluid intake in P rats receiving "free choice" of water and 15% ethanol. Alcohol ingestion gradually returned to the control level despite consistent PU intake. However, alcohol intake following alcohol withdrawal was suppressed in PU-fed P rats. PU suppressed the severity of alcohol withdrawal symptoms. Thus, withdrawal symptoms do not occur in PU-fed rats even though their alcohol ingestion is comparable to that in control P rats. Brain, plasma, and liver samples were analyzed for the presence of kudzu root isoflavones, which are mostly PU (>90% of total isoflavones) and a trace amount of daidzin. Liver samples obtained from PU-fed P rats contained 20-30 microg/g of PU. An important observation was that plasma or brain samples obtained from PU-fed or alcohol + PU-fed rats did not contain PU. This study indicated that PU feeding transiently suppressed alcohol intake and abolished withdrawal symptoms at a time when alcohol intake had returned to the control level. The absence of PU in plasma and brain indicates the possibility that some nonspecific mechanism may be involved in the anti-alcoholism effects of PU in P rats.

Alcohol Drinking↗

PubMed2004

Kudzu root extract suppresses voluntary alcohol intake and alcohol withdrawal symptoms in P rats receiving free access to water and alcohol.

Alcohol-preferring (P) rats, given free choice to drink water or 15% alcohol, drank 7-10 g of alcohol/kg/day, giving blood alcohol values ranging from 16 to 24 mg/dL. Body weight and food and total fluid intake values in control and alcohol-drinking P rats did not differ significantly, while water intake was inversely related to the amount of alcohol consumed. Alcohol withdrawal after 50 days of alcohol drinking caused withdrawal symptoms such as hypersensitivity, poor landing coordination, and tremors. A daily 0.5, 0.75, and 1.0 g/kg dose of kudzu root (KdR) did not affect body weight and food and water intake values in control (no alcohol) P rats. Subchronic feeding of relatively higher KdR doses (0.75 and 1.0 g/kg) caused a 25-30% reduction in weight gain. The 0.5 g/kg KdR dose caused a 50-60% reduction in alcohol consumption, abolished the development of alcohol withdrawal symptoms, but did not affect blood alcohol levels. The higher KdR doses did not further reduce alcohol consumption. Alcohol suppressed the weight-reducing effects of KdR. The KdR extract used in this study contained 150 mg/g of puerarin, 13 mg/g of daidzin, 4 mg/g of daidzein, 3 mg/g of genistin, 0.2 mg/g of genistein, and 1 mg/g of glycetin. Blood and liver samples contained mostly puerarin and a trace amount of daidzein that may have been formed by the hydrolysis of daidzin by liver enzymes. An important observation was that brain samples obtained from KdR-fed or alcohol + KdR-fed rats did not contain any of the KdR isoflavones. Thus, KdR isoflavones suppressed alcohol drinking and withdrawal symptoms without entering the brain.

Alcohol Drinking↗

PubMed2004

Behavioral and developmental effects of neem extracts on Clavigralla scutellaris (Hemiptera: Heteroptera: Coreidae) and its egg parasitoid, Gryon fulviventre (Hymenoptera: Scelionidae).

Extracts of neem, Azadirachta indica A. Juss, negatively affected feeding and development of Clavigralla scutellaris (Westwood), a coreid pest of pigeonpea, Cajanus cajan (L.) Millspaugh. Labial dabbing, pod wall penetration, and seed damage by fifth instars were significantly reduced on beans, Phaseolus vulgaris (L.), that had been dipped in aqueous, methanolic, or hexane extracts of neem seed kernel. When fourth instars were dipped directly into aqueous extract, developmental abnormalities of the wings occurred at all levels tested and fecundity dropped to zero at concentrations above 0.3125%. The LC50 value was 3.14% (220 ppm azadirachtin) at 8 d. The scelionid wasp Gryon fulviventre (Crawford) is an important natural enemy of Clavigralla spp.; egg mortality from this parasitoid ranged from 37 to 85% during the fall cropping season. Feeding by newly emerged wasps was dramatically reduced when honey was mixed with aqueous neem suspension, but 6-d survivorship of adults did not differ significantly from that of the control. Wasp oviposition behavior was altered slightly when coreid eggs were treated with neem: the period of antennation was significantly extended, but time for drilling, oviposition, and marking was unaffected. Neem-dipped eggs were accepted for oviposition and progeny emerged successfully from these treated eggs. Exposure of already parasitized eggs to neem did not interfere with progeny emergence, longevity, or sex ratio. Thus, neem extract and egg parasitoids seem to be compatible and promising control strategies for C. scutellaris. Our results suggest that use of neem against pod-sucking bugs will not interfere with natural control provided by G. fulviventre.

Animals↗

PubMed2004

Degradation of albumin by the renal proximal tubule cells and the subsequent fate of its fragments.

BACKGROUND: In view of recent reports of large amounts of albumin fragments present in normal urine we investigated the mechanism of albumin handling by the proximal tubule. METHODS: We injected (125)I-albumin intravenously in rats and measured the excretion of intact and degraded (125)I-albumin in the urine by trichloroacetic acid (TCA) precipitation. The excretion rate of intact (125)I-albumin was compared to that obtained by routine radioimmunoassay (RIA). Human proximal tubular HK-2 cells were used to characterize the albumin receptor and study the degradation of albumin to peptides, establish their size by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and gel filtration chromatography, and determine the direction in which the degradation products are removed from the cell. RESULTS: Following injection of (125)I-albumin intravenously into rats we recovered large quantities of (125)I-albumin fragments in urine and determined that 98% was in a highly degraded form and only 2% was intact. Only the intact albumin could be detected by RIA. We observed similar results in the urine of ex vivo kidneys perfused with (125)I-albumin. We found that (125)I-albumin was taken up by HK-2 cells via a receptor, degraded in the lysosomes and the peptides exocytosed to both the apical and basolateral sides of the cells. CONCLUSION: We conclude that normally the kidney degrades large amounts of albumin and that the degradation fragments appear in the urine. These findings are in sharp contrast with the established view that degraded albumin is completely reabsorbed into the blood stream.

Animals↗

PubMed2003

Ovipositional responses of Chilo partellus (Swinhoe) (Lepidoptera: pyralidae) to natural products from leaves of two maize (Zea mays L.) Cultivars.

Ovipositional responses of Chilo partellus (Swinhoe) (Lepidoptera: Pyralidae) to hexane extracts of leaves of two maize (Zea mays L.) cultivars, one resistant (Kisan) and one susceptible (Basilocal), were studied in two-choice bioassays. Gravid females laid a significantly higher percentage of eggs on substrates smeared with extract of Basilocal leaves (HEBL) (69%) than on those smeared with extracts of Kisan leaves (HEKL) (31%). Several chemicals were isolated from HEKL, three of which were characterized as dotriacontanol, heptadecanol, and nonadecanol. These chemicals were either absent or were present in very small amounts in HEBL, but in HEKL they were detected in much larger amounts. Each isolated chemical was tested for its effect on C. partellus oviposition in two-choice bioassays. Maximum ovipositional deterrence (90%) was observed for the compound MR-22a, followed in decreasing order by nonadecanol, MR-7, and heptadecanol. The identity of the remaining compounds is being investigated. The results indicate that the relative resistance of Kisan maize compared to Basilocal is partly due to the presence of certain ovipositional deterrents in its leaves.

Animals↗

PubMed2003

Mechanism of increased angiotensin II levels in glomerular mesangial cells cultured in high glucose.

Previous studies have shown that glucose increases angiotensin II (AngII) levels in rat glomerular mesangial cells and that AngII mediates the inhibitory effects of high glucose on matrix degradation in these cells. The present study addresses the following questions: (1) What are the mechanisms for the generation of AngII in mesangial cells? (2) What are the effects of glucose on AngII generation by these mechanisms? Experiments employed primary mesangial cells from normal Sprague-Dawley rats. The levels of immunoreactive angiotensinogen (AGT), angiotensin I (AngI), and angiotensin II (AngII) were measured by ELISA. AGT mRNA expression was determined by Northern blot analysis. Incubation of cells for 24 h in high glucose (30 mM) increased AGT levels by 1.5-fold and increased AGT mRNA expression; this was accompanied by a 1.5-fold increment in AngI and 1.7-fold increment in AngII levels. Renin activity (measured as AngI generation in the presence of excess AGT) and ACE levels and activity were not altered by high glucose. In further experiments, the effect of high glucose on formation of Ang peptides from exogenous AngI in mesangial cell extracts was examined using HPLC. Exogenous AngI was converted into various Ang peptides, including AngII, Ang(1-9), Ang(1-7), and Ang(3-8). A significant increase in formation of AngII from AngI was observed in cells incubated in high glucose. In addition, AngII production from exogenous Ang(1-9) in cell extracts was also stimulated by high glucose. These findings demonstrate that glucose increases mesangial AngII levels via an increase in AGT and AngI. In addition, this study provides new information that Ang(1-9) is produced by mesangial cells, can be converted to AngII, and that this conversion is also stimulated under high-glucose conditions.

Angiotensin I↗

PubMed2002

Handling of low-density lipoprotein by the renal tubule: release of fragments due to incomplete degradation.

Because the mechanism by which lipoproteins are processed and modified in the renal tubule in patients with nephrosis is not completely understood, we studied the handling of low-density lipoprotein (LDL) in perfused rat kidneys made permeable by protamine. Protamine pretreatment increased the clearance of 125(I) LDL 25-fold compared to controls, thereby simulating a proteinuric kidney. Similar studies were also conducted in kidneys of rats made proteinuric by the induction of passive Heymann nephritis. Of the perfused iodinated LDL, 5% was localized in the cortex and lesser amounts in the medulla and urine. In the cortex and medulla, iodinated LDL was present mainly in the intact form (90%); just 10% was present in the degraded form. Using horseradish peroxidase conjugated to LDL, we demonstrated specific staining in the proximal tubules, suggesting that specific LDL receptors were present in that location. Although LDL in the tissue was present mostly in the intact form, it was 95% degraded in urine, and the degradation was inhibited by chloroquine, indicating that the lysosomes were the site of LDL metabolism. Gel chromatography and electrophoresis of iodinated LDL in the urine showed the presence of fragments in the range of 5 to 15 kD. We conclude that renal degradation of LDL is incomplete and that the incompletely degraded fragments released into the urine may be toxic to the kidney by virtue of their lipid side-chains.

Animals↗

PubMed2002

Composition, red blood cell uptake, and serum protein binding of phytoestrogens extracted from commercial kudzu-root and soy preparations.

Kudzu-root and soy phytoestrogens have been associated with anti-cancer and anti-intoxication activities. Sales of capsules containing kudzu-root and soy extracts through health food stores and the Internet are unregulated. To compare efficacy, the amount of phytoestrogens present in commercial preparations and their fate in biological samples must be determined. In this study, the levels and composition of phytoestrogens in kudzu-root and soy extracts were studied using high-performance liquid chromatography with ultraviolet light detection. The bioavailability of phytoestrogens was studied by measuring red blood cell (RBC) uptake and serum protein binding ability. Phytoestrogen levels in acidified kudzu-root samples were 5- to 10-fold greater than those in nonacidified samples. Puerarin accounted for 80% of total phytoestrogens in kudzu-root. In soy extract, puerarin was absent while genistin, glycetein, and daidzin or daidzein were the major phytoestrogens. The RBC uptake depended on the phytoestrogen's polarity and molecular length. When serum was dialyzed with phytoestrogen standards in a buffer, the protein binding of phytoestrogens correlated negatively with their polarity. However, when serum was dialyzed with kudzu-root or soy extract, almost all of the phytoestrogens present in the extract bound to serum protein. Therefore, this study suggests differences in the bioavailability of phytoestrogens when they are ingested as purified compounds compared with crude plant extract. The differential composition of phytoestrogens in kudzu-root and soy may account for the differences in their therapeutic activities.

Biological Availability↗

PubMedDate not supplied

Immunotoxicity of acute acephate exposure in control or IL-1-challenged rats: correlation between the immune cell composition and corticosteroid concentration in blood.

Corticosterone concentration and the immune cell composition were measured in rats exposed by intraperitoneal (i.p.) injection to different doses (10-500 mg kg(-1)) of acephate (Ace) and 250 micro g kg(-1) of interleukin 1 (IL-1), either alone or in combination. Two different combination protocols were used: IL-1 and Ace were administered simultaneously; and IL-1 was injected 60 min after Ace administration (sequential exposure). Ace, in a dose- and time-dependent manner, inhibited blood and brain acetylcholinesterase (AChE) activities, increased blood corticosterone concentrations, suppressed blood CD4, CD8, B cell and monocyte contents and increased blood neutrophil counts. The Ace-induced changes lasted for up to 24 h after Ace exposure. Interleukin 1 increased blood corticosterone concentrations without affecting blood or brain AChE activities. The IL-1-induced corticosterone concentration returned to the basal level within 3-10 h after IL-1 exposure. The CD4, CD8, B cell and monocyte counts increased significantly at 10 min after IL-1 exposure. The cell counts decreased gradually thereafter and returned to the basal level within 30 min after IL-1 exposure. Simultaneous exposure of rats to Ace and IL-1 partially suppressed the IL-1-induced increase in the immune cell counts and decreased the immune cell numbers below the basal values. Sequential injection of Ace and IL-1 blocked the IL-1-induced increase in the immune cell numbers. Thus, Ace exposure would impair the normal distribution of immune cells and deregulate the IL-1 response in rats. This study therefore suggests that Ace would suppress the immune cell numbers in blood, thus decreasing an organism's immunity. Ace exposure occurring concurrent with injury would augment the acute-phase response, which would augment the toxic effects of IL-1 and other cytokines, and Ace exposure occurring prior to the injury would suppress or abolish the initial stimulatory effects of IL-1, which would decrease an organism's ability to combat infection or injury.

Acetylcholinesterase↗

PubMedDate not supplied

Enzyme-linked immunosorbent assay for ultratrace determination of antibiotics in aqueous samples.

Two commercially available enzyme-linked immunosorbent assay (ELISA) kits that are commonly used for tylosin or tetracycline residues in meat and milk were adapted for ultratrace analysis of these antibiotics in surface and ground waters. These two antibiotics are commonly fed to swine, turkeys, and cattle at subtherapeutic doses for growth promotion purposes. Both ELISA techniques were found to be highly sensitive and selective for the respective antibiotics with detection limits of 0.10 and 0.05 microg L(-1) for tylosin and tetracycline, respectively. The recovery of both tylosin and tetracycline from spiked samples of lake waters, runoff samples, soil saturation extracts, and nanopure water was close to 100%. Tetracycline ELISA was highly specific for tetracycline and chlortetracycline but not for other forms of tetracycline (oxytetracycline, demeclocycline, and doxycycline). Analysis of a few liquid swine manure samples by liquid chromatography-mass spectrometry (LC-MS) showed lower concentrations for chlortetracycline as compared with concentrations obtained using ELISA. However, the concentrations of tylosin from ELISA were comparable with that of LC-MS. The lower concentrations of chlortetracycline obtained by LC-MS in manure samples indicate the presence of other similar or transformed compounds that were detected by ELISA but not determined by LC-MS. These results indicate that both ELISA kits can be useful tools for low-cost screening of tylosin, tetracycline, and chlortetracycline in environmental waters. Furthermore, both ELISA procedures are rapid, portable, and easily adaptable for testing of multiple samples simultaneously.

Animals↗