Acute renal failure and liver dysfunction after subcutaneous injection of 3-sn-phosphatidylcholine (Lipostabil®)-case report.

INTRODUCTION: Drug-induced tubulointerstitial nephritis and acute tubular necrosis are common, and are often caused by drugs especially antibiotics or non-steroidal anti-inflammatory drugs. Drug-induced liver dysfunction and renal failure after subcutaneous injection of phosphatidylcholine was not reported so far. 3-sn-Phosphatidylcholine has been described as a cell lysis reaction-inducing drug. Its in vitro data indicated a relevant toxicity potential. In particular human cell types such as fibroblast-like preadipocytes, vascular and skeletal muscle cells, or renal epithelial cells react more sensitive than other human cell types. CASE REPORT: We present a 28-year-old woman who received 3.5 g (70 mL) of 3-sn-phosphatidylcholine (Lipostabil®) at once subcutaneously (s. c.) in both gluteal regions. The drug was originally introduced to prevent fat embolism. Nevertheless, its off-label use in aesthetic therapy for treatment of localized fat deposits through subcutaneous administration is becoming increasingly common. Three hours after injection the patient suffered from severe nausea and emesis. Within 24 hours a dramatic increase of liver enzymes and a beginning liver dysfunction were observed. Subsequently, renal function deteriorated two days later making a temporary haemodialysis necessary. Hepatic improvement was observed after three days of treatment. Renal function was fully recovered after two weeks. CONCLUSION: To the best of our knowledge, this is the first reported patient presenting with acute liver dysfunction and renal failure after subcutaneous injection of 3-sn-phosphatidyl-choline (Lipostabil®) indicating the risk of an off-label use of this drug.

BlotBase: a northern blot database.

With the availability of high-throughput gene expression analysis, multiple public expression databases emerged, mostly based on microarray expression data. Although these databases are of significant biomedical value, they do hold significant drawbacks, especially concerning the reliability of single gene expression profiles obtained by microarray data. Simultaneously, reliable data on an individual gene's expression are often published as single northern blots in individual publications. These data were not yet available for high-throughput screening. To reduce the gap between high-throughput expression data and individual highly reliable expression data, we designed a novel database "BlotBase", a freely and easily accessible database, currently containing approximately 700 published northern blots of human or mouse origin (http://www.medicalgenomics.org/Databases/BlotBase). As the database is open for public data submission, we expect this database to quickly become a large expression profiling resource, eventually providing higher reliability in high-throughput gene expression analysis. Realizing BlotBase, Pubmed was searched manually and by computer based text mining methods to obtain publications containing northern blot results. Subsequently, northern blots were extracted and expression values of different tissues calculated utilizing Image J. All data were made available through a user friendly web front end. The data may be searched by either full text search or list of available northern blots of a specific tissue. Northern blot expression profiles were displayed by three expression states as well as a bar chart, allowing for automated evaluation. Furthermore, we integrated additional features, e.g. instant access to the corresponding RNA sequence or primer design tools making further expression analysis more convenient. Finally, through a semiautomatic submission system this database was opened to the bioinformatics community.

Feasibility study for co-digestion of sewage sludge with OFMSW on two wastewater treatment plants in Germany.

In the past years, wastewater treatment plants (WWTP) in Germany have often been enlarged or expanded. However, it has become evident that the prognosticated increase in wastewater amount has not become a reality and thus free capacities, particularly in the sewage sludge digesters, are available. A possibility for the use of these available capacities is the fermentation of sewage sludge together with organic waste. A feasibility study for two different wastewater treatment plants in Germany was done in order to estimate if fermentation of the organic fraction of municipal solid waste (OFMSW) affects the wastewater treatment plant operation. In this study, the technical, economic and ecological aspects of co-digestion were investigated for the plants selected.

Effects of doxazosin on atherosclerosis in cholesterol-fed rabbits.

Doxazosin was administered to rabbits fed diets enriched in cholesterol and peanut oil for 7.5 or 12 weeks, in 2 separate experiments. Doxazosin suppressed the accumulation of cholesterol and formation of atherosclerotic plaques in the aortas of treated rabbits and prevented a diet-induced increase in aortic collagen and wall mass. Doxazosin was more effective in the thoracic and abdominal segments of the aorta than in the aortic arch. Pharmacokinetic analysis indicated that treated rabbits were exposed to concentrations of doxazosin, integrated over 24 h, which were consistent with the therapeutic range of doxazosin measured in patients treated for hypertension. Doxazosin did not alter serum levels of cholesterol or triglycerides, nor were there any consistent effects on glucose, free fatty acid or ketone levels. Hypotheses of the mechanism of action of doxazosin are discussed, including the possible involvement of alpha 1-adrenergic receptors in recruitment of smooth muscle cells by subintimal macrophages and nonadrenergic mechanisms of inhibition of lipid infiltration.

Antinuclear antibody profile in UCD line 200 chickens: a model for progressive systemic sclerosis.

The fully blown disease of human progressive systemic sclerosis (PSS, scleroderma) is serologically associated with the emergence of several types of autoantibodies, some of them regarded as more specific for scleroderma (e.g. Scl-70, anti-centromere) and some common also to other connective tissue diseases (e.g. anti-ssDNA). Since most patients suffering from PSS are not under medical control until clinical manifestations are fully established, only scarce data are available on the dynamics and clinical significance of autoantibodies in the very early stages of this systemic fibrotic disease. The University of California at Davis line 200 (UCD 200) of chickens spontaneously develop a PSS-like disorder with an acute inflammatory stage around 60 days after hatching, leading to fibrosis with fast progression. In order to address a possible correlation between the occurrence and titer of autoantibodies and the initial disease activity, we have chronologically investigated the presence and titer of autoantibodies directed against several human nuclear antigens in this animal strain. In UCD-200 chickens, we found a progressive increase in autoantibodies to histones, to ssDNA and--to a lesser degree--dsDNA with peaks at the age of 60 and 120 days, to poly(I) and poly(G) with a peak at 120 days and an elevation in anti-cardiolipin antibodies. Total immunoglobulin concentrations, anti-Ro, anti-La and anti-Sm showed no significant differences as compared to negative results in healthy normal controls. Our data reveal parallels in the antinuclear antibody (ANA) spectrum between UCD-200 chickens and human autoimmune collagen diseases, but do not reflect the typical ANA spectrum found in the foudroyant form of diffuse scleroderma.(ABSTRACT TRUNCATED AT 250 WORDS)

Substituted dihydrobenzopyran and dihydrobenzofuran thiazolidine-2,4-diones as hypoglycemic agents.

A series of dihydrobenzofuran and dihydrobenzopyran thiazolidine-2,4-diones (compounds 3-26) was synthesized from the corresponding aryl aldehydes 1 in two steps. These compounds represent conformationally restricted analogues of the novel hypoglycemic ciglitazone. The series was evaluated by hypoglycemic effects in vitro by measuring stimulation of 2-deoxyglucose uptake in L6 myocytes and stimulation of expression of the glucose transporter protein in 3T3-L1 adipocytes. In vivo hypoglycemic effects were evaluated in the genetically obese ob/ob mouse, and structure-activity relationships are discussed. On the basis of this in vivo potency, we have selected the 2(R)-benzylbenzopyran derivative to be further studied in a clinical setting.

Actions of novel antidiabetic agent englitazone in hyperglycemic hyperinsulinemic ob/ob mice.

The effects of CP 68722 (racemic englitazone) were examined in ob/ob mice, in adipocytes and soleus muscles from ob/ob mice, and in 3T3-L1 adipocytes. Administration of englitazone at 5-50 mg.kg-1.day-1 lowered plasma glucose and insulin dose dependently without producing frank hypoglycemia in either the diabetic or nondiabetic lean animals. The glucose-lowering effect in ob/ob mice preceded the reduction in hyperinsulinemia. On cessation of drug, plasma insulin returned to untreated levels within 48 h, whereas plasma glucose rose slowly over 5 days. Englitazone (50 mg/kg) for 11 days lowered plasma glucose (22.2 +/- 1.4 to 14.0 +/- 1.9 mM), insulin (7.57 +/- 0.67 to 1.64 +/- 0.60 nM), nonesterified fatty acids (1813 +/- 86 to 914 +/- 88 microM), glycerol (9.20 +/- 0.98 to 4.94 +/- 0.03 mM), triglycerides (1.99 +/- 0.25 to 1.03 +/- 0.11 g/L), and cholesterol (6.27 +/- 0.96 to 3.87 +/- 0.57 mM), but no effects were observed 3 h after a single dose. Basal and insulin-stimulated lipogenesis were enhanced in adipocytes from ob/ob mice treated with 50 mg/kg englitazone for 11 days compared with lipogenesis in cells from vehicle-treated controls. Treatment of ob/ob mice with 50 mg/kg englitazone reversed the defects in insulin-stimulated glycolysis (from [3-3H]glucose) and glycogenesis and basal glucose oxidation (from [1-14C]glucose) in isolated soleus muscles. Englitazone (30 microM) stimulated 2-deoxy-D-glucose transport in 3T3-L1 adipocytes from 0.37 +/- 0.03 to 0.65 +/- 0.06 and 1.53 nmol.min-1.mg-1 protein at 24 and 48 h, respectively. Thus, englitazone has 1) insulinomimetic and insulin-enhancing actions in vitro and 2) glucose-, insulin-, triglyceride-, and cholesterol-lowering properties in an animal model of non-insulin-dependent diabetes mellitus (NIDDM) in which sulfonylureas have little or no effect. Thus, this new agent may have beneficial effects including a reduced risk of hypoglycemia in patients with NIDDM.

Serum monoclonal antibodies derived from patients with multiple myeloma react with mycobacterial phosphoinositides and nuclear antigens.

The sera of 46 patients with multiple myeloma were examined for the presence of anti-tuberculosis (TB) glycolipids antibodies. In positive sera samples, anti-polynucleotides, anti-histones, anti-cardiolipin, anti-Sm and anti-RNP activities were sought. Antibodies against three different mycobacterial glycolipids were detected in 10 of the 46 sera. Three (P429, P557, P5) showed high titres of antibodies against all three different TB glycolipids. Of the 10 reactive samples, two antibodies were purified (P429, P557). P557 reacted with various polynucleotides and other antigens tested (Sm, RNP, cardiolipin, histones). One immunoglobulin (P207) which showed high activity against Sm and RNP had no activity against TB glycolipids and was employed as a control. The cross-reactivity between mycobacterial glycolipids and the nuclear antigens was further established by bi-directional competition assays with P429 and P557. Our study shows a high incidence (22%) of anti-TB glycolipids antibodies in sera of patients with monoclonal gammopathies, some of which show anti-DNA and other anti-nuclear antigens activities. This is additional evidence for the mycobacterial-nuclear antigen cross-reactivity which may suggest a possible role of infection (e.g., tuberculosis) in autoimmune diseases.

Autoantibody production by patients infected with Leishmania.

Sera from 29 patients with visceral leishmaniasis and 14 patients with cutaneous leishmaniasis were tested against a panel of nine nuclear antigens employing an enzyme-linked immunosorbent assay (ELISA). Anti- Sm, RNP, SS-A and SS-B antibodies were present in high titres in 83, 86, 36 and 73 per cent of the patients with visceral leishmaniasis and in 7, 14, 25 and 25 per cent of the patients with cutaneous leishmaniasis. One serum from a patient with visceral leishmaniasis which reacted strongly with Sm, RNP, SS-A and SS-B was examined by immunoblotting on extractable nuclear antigen from Hela cells. This serum binds to nine different antigenic bands (16, 23, 29, 30, 40, 50, 58, 100 and 115 kD). These same antigens were recognized by serum from a patient with systemic lupus erythematosus. The binding of visceral leishmaniasis serum antibodies to ribonucleoproteins was inhibited by prior incubation of serum with either leishmanial membrane antigens, from four different species of Leishmania, or intact cells of Leishmania donovani, implying molecular resemblance between common leishmanial antigens and ribonuclear antigens. It seems that appearance of autoantibodies to ribonucleoproteins in sera of patients infected with Leishmania is not only due to simply polyclonal activation of lymphocytes, but is also the result of a molecular mimicry between leishmanial antigens and ribonucleoproteins.

The detection of anti-Sm-RNP activity in sera of patients with monoclonal gammopathies.

The sera of 141 patients with monoclonal gammopathies were examined for the presence of anti-RNP and anti-Sm activities. An enzyme-linked immunosorbent assay (ELISA) was employed. Thirty-two sera were found to bind RNP while 16 bound to Sm. The anti-Sm and anti-RNP antibodies were found in sera of patients with IgG, IgM and IgA gammopathies. The activity against RNP and/or Sm was confirmed further by using purified immunoglobulins, employing competition assays and immunoblotting. Anti-Sm antibodies are regarded as being highly specific for SLE, yet none of the patients whose serum was found to contain high titres of anti-Sm/anti-RNP antibodies presented with symptoms related to SLE or other rheumatic diseases. Our results of a high incidence of anti-ribonucleoproteins activity in the serum of patients with monoclonal gammopathies support previous reports of autoantibody properties characteristic of these immunoglobulins.

Antinuclear and related autoantibodies in sera of healthy subjects with IgA deficiency.

The sera of 49 healthy IgA-deficient (SIgAD) subjects were evaluated for the presence of autoantibodies directed against 10 different nuclear and cytoskeletal antigens, as well as for the presence of the common lupus anti-DNA idiotype (16/6 Id). Twenty-nine sera were from IgG subclass-deficient subjects (4 = IgG2, 25 = IgG3), and 25 from normal healthy subjects, used as controls. The incidence of antinuclear but not anti-cytoskeletal antibodies were found to be significantly greater in the SIgAD group, as compared to the IgG-deficient subjects and the normal controls. Overall, 39% of SIgAD sera demonstrated polyreactivity, namely reactivity against more than one nuclear antigen. The incidence of specific antibody detection ranged from 37% against cardiolipin to 12% against RNP in the IgA-deficient group, albeit not with statistical significance in all cases when compared to the control group. Isotype evaluation of the antinuclear and related antibodies in the SIgAD group showed a greater tendency towards IgG. This increased incidence of autoantibody production in SIgAD may preceed the development of an overt autoimmune disease in the future.

Effects of doxazosin and other antihypertensives on serum lipid levels and lipoprotein lipase in the C57BR/cdJ mouse.

Doxazosin has been shown to lower serum cholesterol levels in the cholesterol-fed (0.75% in a synthetic diet that contains sucrose and cholic acid) C57BR/cdJ mouse. These studies show that the drug's main effect is to lower low-density lipoprotein (LDL) cholesterol and leave high-density lipoprotein (HDL) cholesterol levels unchanged. The drug had cholesterol-lowering effects in this model at doses down to 3 mg/kg. In order to determine if these effects are unique to selective alpha 1-inhibitors, other antihypertensives including hydralazine, papaverine, and captopril were investigated. None of the drugs has any effects on the plasma lipid metabolite levels. The effects of propranolol and polythiazide on plasma lipid levels were also examined in these mice. Propranolol had no effect, whereas the diuretic increased plasma cholesterol levels. Both propranolol and polythiazide increased plasma triglycerides. Doxazosin has been shown to inhibit cGMP phosphodiesterase in the laboratory. The effects of zaprinast, a cGMP phosphodiesterase inhibitor, were tested in order to determine if this property of the drug could be responsible for its lipid-lowering activity. The data show that there are no effects on plasma lipids in zaprinast-treated animals. Doxazosin treatment increased heparin-releasable lipoprotein lipase in fasted chow-fed mice. The drug was without effect on the activity of hepatic lipase present in the plasma after heparin release. No effects were observed on the tissue levels of either hepatic or lipoprotein lipases (heart or adipose tissue).

Dexamethasone stimulates insulin receptor synthesis in cultured rat hepatocytes.

The ability of the glucocorticoid dexamethasone to modulate the insulin receptor was examined directly in primary cultures of hepatocytes prepared from adult male rats. Hepatocytes were cultured in a defined medium in the presence and absence of dexamethasone, 0.1 microM. The exposure of hepatocytes to dexamethasone resulted in a time-dependent (steady state by 32 h) increase in insulin binding in both intact hepatocytes and Triton X-100-soluble extracts (total insulin receptor content). The enhanced insulin binding found in soluble extracts of dexamethasone-treated hepatocytes was the result of an increase in insulin receptor number without a change in receptor affinity. In order to assess the mechanism by which dexamethasone "up-regulates" the insulin receptor, the heavy isotope density-shift technique was used to analyze insulin receptor turnover in control and dexamethasone-treated hepatocytes. Hepatocytes were initially cultured for 32 h in standard culture media containing only "light" (14C, 12C, 1H) amino acids. In hepatocytes exposed to dexamethasone, a 417% increase in insulin binding in Triton X-100-soluble extracts was observed. After 32 h, when steady state binding is achieved in dexamethasone-treated cultures, parallel cultures of hepatocytes incubated in the absence and presence of dexamethasone were washed and subsequently cultured in media containing "heavy" amino acids (15N, 13C, 2H). The time-dependent disappearance of light insulin receptor (receptor degradation) and appearance of heavy insulin receptor (receptor synthesis) were monitored using CsCl gradients to resolve the two density species of receptor. At steady state, the rate of receptor synthesis (k8) was 2.94 and 0.62 fmol of insulin bound h-1 in dexamethasone-treated and control hepatocytes, respectively. In contrast to this large increase in the rate of receptor synthesis observed in dexamethasone-treated cells, the first order rate constant for decay (k d) was the same in dexamethasone-treated (0.074 h-1) and in control (0.077 h-1) hepatocytes. We therefore conclude that glucocorticoid-induced up-regulation of the insulin receptor in the liver is due to stimulation of insulin receptor synthesis.

Down-regulation of epidermal growth factor receptor correlates with plasminogen activator activity in human A431 epidermoid carcinoma cells.

Human A431 epidermoid carcinoma cells in culture exhibit epidermal growth factor (EGF)-induced "down-regulation" of cell-surface and total cellular (Triton X-100 extractable) EGF receptors caused entirely by an enhanced rate (4-fold) of receptor inactivation [Krupp, M. N., Connolly, D. T. & Lane, M. D. (1982) J. Biol. Chem. 257, 11489-11496]. The following observations show that this enhanced rate of EGF receptor inactivation is closely correlated with an increased cellular activity of plasminogen activator (PA), a serine protease. First, EGF-induced down-regulation of cell-surface and total cellular EGF receptors and the concomitant increase in cellular PA activity occur with identical kinetics, the t 1/2 for both processes being 3-3.5 hr. Second, the EGF dose-response curves for down-regulation of total cellular EGF receptor and increased PA activity are similar. The EGF concentrations for half-maximal responses of both processes are 10-15 nM and 20 nM, respectively. Third, the removal of EGF from previously down-regulated cells results in the recovery of total cellular EGF binding activity with a concurrent loss of cellular PA activity. Fourth, blocking PA synthesis or activity with cycloheximide or dexamethasone prevents down-regulation of the EGF receptor. Fifth, the addition of leupeptin, an inhibitor of PA and plasmin action, blocks EGF-induced receptor down-regulation as well as the increase of PA activity. That EGF receptor down-regulation is independent of plasminogen per se in the culture medium suggests that PA-mediated events may initiate the rapid inactivation of the EGF receptor that occurs during down-regulation.

Synthesis, turnover, and down-regulation of epidermal growth factor receptors in human A431 epidermoid carcinoma cells and skin fibroblasts.

Epidermal growth factor (EGF) receptors extracted with Triton X-100 from human skin fibroblasts and A431 epidermoid carcinoma cells rapidly lose EGF-binding activity precipitable with polyethylene glycol. The presence of concanavalin A which can cross-link and, thereby, aggregate the receptors, allowed quantitative recovery of the lost EGF-binding activity. Scatchard analysis of EGF binding of Triton X-100-solubilized receptors showed that A431 cells and skin fibroblasts possess approximately 1.5 X 10(6) and 7 X 10(4) EGF-binding sites/cell, respectively, which exhibit similar affinities for the ligand. The heavy isotope density-shift method was employed to determine whether differences in rates of receptor synthesis or decay account for the large difference in number of receptors/cell between the two cell types. After shifting cells to medium containing heavy (15N, 13C, and 2H) amino acids, light and heavy receptors, solubilized from total cellular membranes, were resolved by isopycnic banding on density gradients and then quantitated. It was demonstrated that A431 cells synthesize EGF receptors at a rate 12 times faster than skin fibroblasts and that the half-life for receptor decay of A431 cells is somewhat longer (t1/2 = 16 h) than that (t1/2 = 9 h) of fibroblasts. Down-regulation of cell surface and total cellular EGF-binding capacity in A431 cells occurs with a t1/2 of 2-3 h and results in a 70-83% decrease in receptor level in 12 h. Scatchard analysis revealed that these changes in EGF binding were due to an alteration of receptor number and not EGF-binding affinity. Rates of EGF receptor synthesis and inactivation/decay were determined by the heavy isotope density-shift method. No change in the rate of receptor synthesis occurred as a consequence of EGF receptor down-regulation. Down-regulation, however, caused a decrease in receptor half-life from 16 to 4.5 h. These results indicate that EGF-dependent regulation of EGF receptor level in A431 cells involves an alteration of the rate of receptor inactivation.