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T W Rademacher

Publications and source records attributed to T W Rademacher.

At least 19 recordsLinked to original sources
PubMed2002

Insulin reduces serum glycosylphosphatidylinositol phospholipase D levels in human type I diabetic patients and streptozotocin diabetic rats.

The enzyme glycosylphosphatidylinositol phospholipase D has a postulated role in the insulin-mimetic signaling pathway of glycosylphosphatidylinositol compounds. We have investigated enzyme activity in the serum of human type I diabetic patients and plasma and tissues of streptozotocin-induced diabetic rats following insulin administration. In the human diabetic patients serum enzyme activity fell by an average of 10.6% (SEM = 2.7; P = 0.008; n = 20) following administration of insulin. In addition serum enzyme activity appeared to be depleted by 27% (SEM = 8.8; P = 0.011; n = 10) compared to nondiabetic controls. In untreated diabetic rats plasma enzyme activity gradually increased 0.3-fold over a 6-week period (P < 0.001; n = 8), this increase was reversed and activity normalized when these animals were treated with insulin. Cloning of the rat glycosylphosphatidylinositol phospholipase D cDNA enabled confirmation of the liver as the principal organ of synthesis. Analysis of mRNA levels in the livers of the diabetic rats showed that gene expression was reduced in the insulin-treated animals compared to the noninsulin-treated controls by 0.7-fold (P = 0.004; n = 4). Tissue enzyme activity was also reduced in the insulin-treated rats; in skeletal muscle enzyme activity was 0.3-fold lower (P = 0.001; n = 4). Insulin therefore decreases glycosylphosphatidylinositol phospholipase D synthesis in diabetic animals resulting in decreased serum enzyme levels, suggesting a relationship between this enzyme and the function of insulin.

Animals↗
PubMed2002

Placentally derived prostaglandin E2 acts via the EP4 receptor to inhibit IL-2-dependent proliferation of CTLL-2 T cells.

A number of immunomodulatory molecules are present in the placenta, including cytokines, prostaglandins, progesterone and indoleamine 2,3-dioxygenase. An undefined factor capable of down-regulating T-cell activity has recently been reported [1] as being produced by short-term cultures of placental fragments. By careful repetition of these studies we have confirmed that chorionic villi isolated from term placenta produce a low molecular weight, heat stable factor capable of inhibiting the IL-2-dependent proliferation of mouse CTLL-2 cells. This activity was not due, however, to a previously unknown immunosuppressive molecule, but rather to prostaglandin E2 (PGE2). Expression of cyclooxygenase (COX)-2 was detected in the syncytiotrophoblast of chorionic villi explants using immunohistochemistry. Culture of the explants in the presence of the COX-1/COX--2 inhibitors indomethacin and diclofenac, or with the COX-2-selective inhibitor DFP, blocked the production of the immunosuppressive factor. The immunosuppressive activity was restored by adding PGE2 to the supernatants obtained from diclofenac-inhibited explants. A number of different receptors are involved in mediating the biological effects of prostaglandins. By utilizing selective antagonists of individual receptors, we have established that the immunosuppressive effect of PGE2 on CTLL-2 cells is exerted via the EP4 receptor. Thus, addition of an EP4-selective antagonist, but not of EP1 or EP3 antagonists, abolished the immunosuppressive effect of PGE2 on CTLL-2 cells. This may have implications for attempts to selectively manipulate T-cell responses.

Animals↗
PubMed2001

Reversal of type 2 diabetes in mice by products of malaria parasites. II. Role of inositol phosphoglycans (IPGs).

We have previously shown that infection with Plasmodium yoelii malaria or injection of extracts from malaria-parasitized red cells induces hypoglycemia in normal mice and normalizes the hyperglycemia in mice made moderately diabetic with streptozotocin. Inositol phosphoglycans (IPGs) are released outside cells by hydrolysis of membrane-bound glycosylphosphatidylinositols (GPIs), and act as second messengers mediating insulin action. The C57BL/Ks-db/db and C57BL/6J-ob/ob mice offer good models for studies on human obesity and Type 2 diabetes. In the present study, we show that a single iv injection of IPG-A or IPG-P extracted from P. yoelii significantly (P < 0.02) lowers the blood glucose in STZ-diabetic, db/db, and in ob/ob mice for at least 4--6 h. Using rat white adipocytes, IPG-P increased lipogenesis by 20--30% in the presence and absence of maximal concentrations of insulin (10(-8) M) (P < 0.01) and stimulated pyruvate dehydrogenase (PDH) phosphatase in a dose-related manner. Both IPG-A and IPG-P inhibited c-AMP-dependent protein kinase (PKA) in a dose-related manner. Compositional analysis of IPGs after 24 h hydrolysis revealed the presence of myo-inositol, phosphorus, galactosamine, glucosamine, and glucose in both IPG-A and IPG-P. However, hydrolysis of IPGs for 4 h highlighted differences between IPG-A and IPG-P. There are some functional similarities between P. yoelii IPGs and those previously described for mammalian liver. However, this is the first report of the hypoglycemic effect of IPGs in murine models of Type 2 diabetes. We suggest that IPGs isolated from P. yoelii, when fully characterized, may provide structural information for the synthesis of new drugs for the management of diabetes mellitus.

Animals↗
PubMed2000

Structure and expression of the human glycosylphosphatidylinositol phospholipase D1 (GPLD1) gene.

Here we report the structure of the human glycosylphosphatidylinositol phospholipase D1 gene, which covers at least 80 kb on chromosome 6p22. The gene comprises 25 exons and encodes a 5.8 kb mRNA, which was detected only in the liver. Southern blot analysis shows that the human genome contains only one GPLD gene and we could only detect one of the two previously reported cDNAs.

Base Sequence↗
PubMed2000

Inositolphosphoglycan mediators structurally related to glycosyl phosphatidylinositol anchors: synthesis, structure and biological activity.

The preparation of the pseudopentasaccharide 1a, an inositol-phosphoglycan (IPG) that contains the conserved linear structure of glycosyl phosphatidylinositol anchors (GPI anchors), was carried out by using a highly convergent 2+3-block synthesis approach which involves imidate and sulfoxide glycosylation reactions. The preferred solution conformation of this structure was determined by using NMR spectroscopy and molecular dynamics simulations prior to carrying out quantitative structure--activity relationship studies in connection with the insulin signalling process. The ability of 1a to stimulate lipogenesis in rat adipocytes as well as to inhibit cAMP dependent protein kinase and to activate pyruvate dehydrogenase phosphatase was investigated. Compound 1a did not show any significant activity, which may be taken as a strong indication that the GPI anchors are not the precursors of the IPG mediators.

Animals↗
PubMed2000

Inositol phosphoglycans and signal transduction systems in pregnancy in preeclampsia and diabetes: evidence for a significant regulatory role in preeclampsia at placental and systemic levels.

Measurements have been made of the urinary content of inositol phosphoglycans IPG P-type and IPG A-type, putative insulin second messengers, in preeclampsia, in type I insulin-treated diabetic pregnant women and their matched control subjects, and nonpregnant women of child-bearing age. The content of IPG P-type and IPG A-type was also measured in the placenta from preeclamptic patients and from normal pregnancies. Pregnancy was associated with an increase, approximately twofold, in urinary output of IPG-P-type relative to nonpregnant controls (P<0.01). The 24-h output of IPG P-type in urine in preeclamptic women was significantly higher (2- to 3-fold) than in pregnant control subjects matched for age, parity, and stage of gestation (P<0.02). In contrast, insulin-dependent diabetic pregnant women did not show any significant change in urinary output of IPG P-type or IPG A-type relative to pregnant control subjects. Evidence for a possible relationship and correlation between the urinary excretion of IPG P-type and markers of preeclampsia, including proteinuria (r = 0.720, P<0.01), plasma aspartate transaminase (r = 0.658, P<0.05), and platelet counts (r = 0.613, P<0.05) is presented. A high yield of IPG P-type was extracted from human placenta, in preeclampsia some 3-fold higher (P = 0.03) than the normal value, whereas no IPG A-type (with lipogenic-stimulating activity) was found. Low concentrations of placental IPG A-type were detected relative to IPG P-type using assay systems dependent upon the effect of this mediator on cAMP-dependent protein kinase or on a proliferation assay using thymidine incorporation into DNA of EGFR T17 fibroblasts. It is postulated that the high urinary excretion IPG P-type in preeclampsia reflects high placental levels and relates to the accumulation of glycogen in the placenta. The paracrine effects of placental IPG P-type (stimulation off other endocrine glands and/or endothelial cells) could contribute to the pathogenesis of the maternal syndrome. A possible theoretical link between elevated placental IPG P-type and apoptosis is proposed.

Adult↗
PubMed2000

Inositol phosphoglycans and the regulation of the secretion of leptin: in vitro effects on leptin release from adipocytes and the relationship to obesity.

The ratio of two families of inositol phosphoglycans (IPG-A:IPG-P), insulin second messengers, is raised in non-insulin-dependent diabetes mellitus (NIDDM) and obesity. It is shown here that IPG A type inhibits leptin release from adipocytes, contrasting with the action of insulin (stimulation) and IPG P type (no effect). The significance of inhibitory effects of IPG A type on leptin release is important in relation to obesity and NIDDM in view of the action of leptin in promoting Lep expression and fat oxidation in muscle, in addition to its effects on satiety. Energy conservation and oxidation via interorgan regulation by leptin could be compromised by a rise in the IPG-A:IPG-P ratio.

Adipocytes↗
PubMed1999

Inositol phosphoglycans in diabetes and obesity: urinary levels of IPG A-type and IPG P-type, and relationship to pathophysiological changes.

Measurements have been made, in adult male diabetic patients and control subjects, of the urinary content of inositol phosphoglycans (IPGs), the IPG A-type and IPG P-type forms, which, among other actions, regulate pathways of glucose utilization, lipogenesis, triglyceride formation, and pyruvate dehydrogenase (PDH) activity. Urine samples from the entire diabetic group showed a 2- to 3-fold increase in IPG A-type, and a fall in the IPG P-type:IPG A-type ratio relative to the control group. Subdivision of the diabetic patients into lean IDDM and obese NIDDM groups revealed significant differences in the IPG P-type:IPG A-type ratio between these groups, this ratio decreasing with increases in the body mass index (BMI). Analysis of the relationships among IPGs and HbA1, blood pressure, and BMI indicated that a fall in the IPG P-type:IPG A-type ratio correlated with a rise in the HbA1 (indicative of impaired glycemic control), with increased systolic blood pressure and increased obesity, all factors linked to Syndrome X. There was a parallism between the profile of the IPG P-type:IPG A-type ratio and the well-established pattern of insulin resistance and BMI. In vitro studies of the effects of alterations in the IPG P-type:IPG A-type ratio on the activation of the pyruvate dehydrogenase complex (PDH complex) at the PDH phosphatase reaction demonstrated that IPG A-type forms antagonized the stimulation of the PDH phosphatase by IPG P-type forms, thus having a negative effect on the conversion of PDH to the active, dephosphorylated, form. This observation could provide a mechanism whereby the shifts in the IPG P-type:IPG A-type ratio reported above could change the metabolic pattern from one directed to glucose oxidation to one more directed toward energy conservation and lipid storage.

Adult↗
PubMed1998

Recent advances in glycoconjugate analysis and glycobiology.

Glycoconjugate chemistry and glycobiology were rapidly evolving scientific disciplines in the 1980s. Their impact, however, on understanding fundamental biological processes was not immediately forthcoming. Within the past 12-18 months, there has been a resurgence in the field with major discoveries leading to powerful new insights into the complex role of glycoconjugates in biological processes.

Carbohydrate Sequence↗
PubMed1998

Correlation between IgG anti-type II collagen levels and arthritic severity in murine arthritis.

This study examines the relationship between the serum levels of IgG antibodies to heterologous and homologous type II collagen and the subsequent arthritic severity in a collagen induced arthritis model. Arthritis was induced in DBA-1 mice using intradermal injections of heterologous type II collagen in Freunds complete adjuvant, the time of arthritis onset was noted and the severity was monitored regularly. Serum samples were taken and IgG levels of anti-heterologous and homologous type II collagen were analyzed both pre and post arthritis onset. We observed that post induction/pre arthritic serum IgG anti-heterologous and homologous type II collagen levels showed a significant correlation (both p < 0.01) with the severity of the arthritis that subsequently developed. Mice with early arthritis showed a highly significant correlation (p < 0.002) between sera IgG anti-homologous type II collagen levels and arthritic severity, a lesser correlation was also apparent between anti-heterologous type II collagen titres and arthritic severity (p < 0.05). The high levels of correlation observed in this study between anti-type II collagen titres and arthritic severity before actual onset of arthritis, clearly suggest that the magnitude of the initial humoral response to type II collagen plays a crucial role in determining the resultant arthritic severity. This observation is only apparent due to the use of an arthritis susceptible inbred mouse strain, which removes variables such as H-2 restriction, antigen processing/presentation and possible complement deficiencies, and the early time scale of the analysed sera samples.

Animals↗
PubMed1997

Reduction in the incidence and severity of collagen-induced arthritis in DBA/1 mice, using exogenous dehydroepiandrosterone.

OBJECTIVE: This study examined the effect of exogenous dehydroepiandrosterone (DHEA) on the onset, incidence, and severity of collagen-induced arthritis (CIA). METHODS: DHEA was administered subcutaneously prior to arthritis induction in DBA/1 mice, and the severity of the subsequent arthritis was monitored. Serum levels of total IgG and IgG isotype-specific anti-murine type II collagen were measured. RESULTS: Repeated administration of DHEA during arthritis induction delayed the onset and decreased the severity of arthritis in male and female DBA/1 mice. DHEA failed to have an observable effect on established arthritis. IgG isotype autoantibody levels were found to be decreased in the sera of DHEA-treated mice. CONCLUSION: Administration of exogenous DHEA offered protection against the development of CIA. These data support the results of human studies in which low DHEA levels have been identified as a potential risk factor for the development of rheumatoid arthritis. These findings also highlight DHEA as a potential therapy worthy of further investigation.

Animals↗
PubMed1997

Isolation and partial characterisation of insulin-mimetic inositol phosphoglycans from human liver.

Extracts of human liver were found to contain activities which copurified and coeluted with the two major subtypes of mediators (type A and type P) isolated from insulin-stimulated rat liver. The putative type A mediator from human liver inhibited cAMP-dependent protein kinase from bovine heart, decreased phosphoenolypyruvate carboxykinase mRNA levels in rat hepatoma cells, and stimulated lipogenesis in rat adipocytes. The putative type P mediator stimulated bovine heart pyruvate dehydrogenase phosphatase. Both fractions were able to stimulate proliferation of EGFR T17 fibroblasts and the type A was able to support growth in organotypic cultures of chicken embryo cochleovestibular ganglia. Both activities were resistant to Pronase treatment and the presence of carbohydrates, phosphate, and free-amino groups were confirmed in the two fractions. These properties are consistent with the structure/ function characteristics of the type A and P inositolphosphoglycans (IPG) previously characterized from rat liver. Further, the ability of the human-derived mediators to interact with rat adipocytes and bovine-derived metabolic enzymes suggests similarity in structure between the mediators purified from different species. Galactose oxidase-susceptible membrane-associated glycosylphosphatidylinositols (GPI) have been proposed to be the precursors of IPG. GPI was purified from human liver membranes followed by treatment with galactose oxidase and reduction with NaB3H4. Serial t.l.c. revealed three radiolabeled bands which comigrated with the putative GPI precursors found in rat liver. These galactose-oxidase-reactive lipidic compounds, however, were only partially susceptible to hydrolysis with phosphatidylinositol-specific phospholipase C from Bacillus thuringiensis and were resistant to glycosylphosphatidylinositol-specific phospholipase C from Trypanosoma brucei. These data indicate that IPG molecules with insulin-like biological activities are present in human liver.

Adult↗
PubMed1996

Bias in murine IgG isotype immobilisation. Implications for IgG glycoform analysis ELISA procedures.

This study investigates immobilisation of murine IgG in various ELISA procedures. Monoclonal murine IgG isotypes and polyclonal IgG from sera were studied. Similar binding curves to plastic were found for all four individual murine IgG isotypes. Single isotypes displayed different affinities for both protein A and protein G, in particular IgG1 was poorly and IgG3 strongly bound to both of these proteins. When mixtures of the isotypes were bound to either plastic, protein A or protein G, competition was observed in which IgG3 was dominant. Paradoxically, studies on the binding rates of single isotypes direct to plastic revealed that IgG3 had the slowest binding rate. Heating of bound IgGs resulted in significant but isotypically non-selective losses from the plates. The data demonstrate that despite obtaining equivalent individual IgG isotype binding curves, mixtures of IgG isotypes behave very differently, with competition for binding occurring even on plastic. The IgG isotype levels of murine sera were measured for individual mice, and the capture efficiency of each IgG isotype by protein A determined at different serum dilutions. Comparisons were made between the observed capture levels of IgG isotypes and their known serum levels. At all dilutions tested, greater than expected binding of IgG3, IgG2b and IgG2a was observed. At a serum dilution of 1/100 the binding of these three isotypes was increased 16-, 2.9- and 0.4-fold, respectively. These increases were balanced by a decrease in IgG1 binding which was the most prevalent serum IgG isotype. The results described above suggest that capture techniques are biased and unlikely to provide a coating of IgG isotypes that accurately reflects that of the serum. This bias is derived from the specificity of the individual isotypes for either protein A or protein G, and the errors further compounded by direct competition between isotypes whatever the capture surface. Induced coalescence of IgG3 may explain the latter observations.

Animals↗
PubMed1996

The glycosylation of Bowes melanoma tissue plasminogen activator: lectin mapping, reaction with anti-L2/HNK-1 antibodies and the presence of sulphated/glucuronic acid containing glycans.

The glycosylation of tissue plasminogen activator (t-PA) obtained from the Bowes melanoma cell line was re-examined using methods of serial lectin affinity chromatography coupled with Bio-Gel P-4 gel filtration chromatography and exoglycosidase sequencing. This study clarified an earlier discrepancy in the literature and confirmed that the major complex N-linked glycans on Bowes t-PA that carry sialic acid as their sole charged group are bi-antennary, core fucosylated, with terminal N-acetylgalactosamine residues. We also report the characterization of a series of related and previously unidentified sialylated glycans. Further we show that Bowes t-PA expresses glucuronic acid/sulphate containing N-linked glycans and is recognized by anti-carbohydrate L2/HNK-1 monoclonal antibodies. The presence on Bowes t-PA of glycans associated primarily with the nervous system is consistent with its expression in a cell line of neuroectodermal origin.

Antibodies, Monoclonal↗
PubMed1996

Analysis of murine IgG isotype galactosylation in collagen-induced arthritis.

The galactosylation status of IgG from both control and arthritic DBA-1 mice was determined by exoglycosidase sequencing and an anti-GlcNAc monoclonal based ELISA. Two to three weeks after arthritis onset mice with collagen-induced arthritis (CIA) showed a modest increase in IgG anti-GlcNAc reactivity against controls (0.644 +/- 0.080 and 0.530 +/- 0.087, respectively, mean absorbance +/- SEM n = 4) consistent with previous literature reports. However, the authors were unable to detect any significant changes in the galactosylation of purified IgG from arthritic and control mice using direct sequencing techniques (percentage G0 of 34.5% +/- 1.9 and 37.7% +/- 2.4, respectively, mean +/- SEM n = 4). It has been demonstrated that a correlation exists between percentage G0 and anti-GlcNAc reactivity for purified human IgG and sera samples, respectively; no such correlation was found for murine IgG and sera. The galactosylation of purified polyclonal murine IgG isotypes was analysed on a separate group of arthritic mice selected for their high anti-GlcNAc reactivities. No significant differences were observed between control and arthritic mice. Immunoglobulin G isotype specific differences were found, with IgG1 exhibiting the highest percentage G0 (45-48%) followed by IgG2a (27-37%), IgG3 (20-32%) and IgG2b the lowest (13-17%). The percentage G0 and anti-GlcNAc reactivity of purified IgG1 and IgG2b showed a narrow range of values when compared to those of IgG2a and IgG3 samples. Pooled sera from both arthritic and control mice was used to purify large quantities of IgG3. Which on analysis revealed a fourfold increase in anti-GlcNAc reactivity in the arthritic sample compared to control. Paradoxically these same IgG3 samples contained similar percentage G0 levels as determined by direct sequencing. The results suggest that IgG1 and IgG2b exhibit isotype selective oligosaccharide processing as little sample heterogeneity could be observed. These two purified IgG isotypes displayed a good correlation between percentage G0 and anti-GlcNAc reactivity; this was in contrast to IgG2a and IgG3. Immunoglobulin G3 from arthritic mice may have G0 oligosaccharides selectively paired together with no net increase in percentage G0. This observation is discussed in detail as is the role of agalactosyl IgG in murine type II collagen-induced arthritis.

Animals↗
PubMed1996

Structural similarities among malaria toxins insulin second messengers, and bacterial endotoxin.

Malaria toxin causes hypoglycemia and induction of tumor necrosis factor. Extracts of parasitized erythrocytes which were coeluted and copurified with one of the two subtypes of mammalian insulin-mimetic inositolphosphoglycans similarly induced fibroblast proliferation in the absence of serum. In addition, induction of tumor necrosis factor in macrophages by malaria toxin and by lipopolysaccharide from Escherichia coli was enhanced by pretreatment of these toxins with alpha-galactosidase. Thus, parasitized erythrocytes contain both soluble inositolphosphoglycan-like insulin second messengers and endotoxin-like lipidic molecules.

Animals↗
PubMed1996

Malaria: a tumour necrosis factor inhibitor from parasitized erythrocytes.

The excessive production of tumour necrosis factor (TNF) is associated with the pathology of blood-stage malaria and phosphatidylinositol-containing phospholipid antigens from parasitized erythrocytes stimulate its secretion by macrophages, thus acting as toxins. This brief report describes some properties of an inhibitor present in lysates from erythrocytes infected with malarial parasites that blocked the detection of recombinant TNF in an enzyme-linked immunosorbent assay and diminished or abolished the cytotoxicity of TNF. It was not found in control lysates of normal erythrocytes. Its addition to macrophage cultures stimulated by toxic malarial preparations or by bacterial lipopolysaccharide also blocked the detection of TNF. These findings may explain the contradictory results obtained from different assays for TNF, and emphasize the need for caution when interpreting the results of a single assay system. If released when parasitized erythrocytes rupture in vivo, the inhibitor could help protect both parasite and host from the damaging effects of TNF.

Animals↗