Ruud Brands

Toxicity of organotin compounds. II. Comparative in vivo and in vitro studies with various organotin and organolead compounds in different animal species with special emphasis on lymphocyte cytotoxicity.

A series of organotin and organolead compounds were fed to male and female weanling rats for 2 weeks at dietary levels of 0, 50, or 150 ppm, in order to evaluate their toxic effects, with special emphasis on the thymus and peripheral lymphoid organs. A dose-related reduction in the weight of thymus, spleen, and popliteal lymph node, associated with lymphocyte depletion in the cortex of the thymus and the thymus-dependent areas in the peripheral lymphoid organs, was observed in animals fed di-n-octyltin dichloride (DOTC) and di-n-butyltin dichloride (DBTC). As shown previously, these effects were not related to stress. Similar, but less pronounced, lymphoid organ changes occurred in animals fed diets containing diethyltindichloride (DETC) and di-n-propyltindichloride. In contrast, other dialkyltin compounds (dimethyltindichloride; di-n-dodecyltindibromide; and di-n-octadecyltindibromide), as well as mono-n-octyltintrichloride (MOTC), tri-n-octyltinchloride, and tetraoctyltin did not cause atrophy of lymphoid organs. Of the dialkyllead compounds tested, di-n-butylleaddiacetate and di-n-hyxylleaddiacetate, the latter caused distinct lymphoid atrophy, but only when associated with severe growth retardation. A similar structure-activity relationship regarding thymus atrophy by dialkyltin compounds was observed after iv application. DOTC and DBTC were effective at a dose of 1 mg/kg. A dose-related reduction of thymus weight, cell count, and cell viability occurred in rats after a single iv injection of 0, 1, 2, 4, or 8 mg of DOTC/kg. The effect of DOTC was completely reversible. In contrast to rats, lymphoid atrophy did not occur in mice, guinea pigs, or Japanese quail fed DOTC or DBTC. In vitro incubation of thymocytes and bone marrow cells with DBTC and DOTC revealed the same selectivity regarding cell type and species origin of cells as was found in vivo. A dose-related decrease in the survival of rat thymocytes was observed, whereas rat bone marrow cells were not affected by DBTC and DOTC at levels up to 50 μg/ml. In contrast, the number and viability of mouse and guinea pig thymocytes exposed to DBTC were the same as the controls. As the survival of human thymocytes was markedly decreased by DBTC, it possibly acts in the same manner in man and rat. In agreement with in vivo results, the survival of rat thymocytes was not significantly decreased by dimethyltindichloride and diethyltindichloride.

Toxicity of organotin compounds. III. Suppression of thymus-dependent immunity in rats by di-n-butyltindichloride and di-n-octyltindichloride

To evaluate the functional significance of di-n-butyl (DBTC)- and di-n-octyltin dichloride (DOTC)-induced lymphoid depletion various immune function studies were carried out. The delayed type hypersensitivity reaction, a parameter of cell-mediated immunity was decreased in rats fed 50 or 150 ppm of DOTC for 6 weeks. This decrease was dose-related. Allograft rejection, another cellular immune response, was significantly delayed by DBTC and DOTC. The antibody response against E. coli LPS, probably a T cell-independent antigen, was not affected by DBTC. However, the humoral immune response against sheep red blood cells (SRBC), which needs the co-operation of T helper cells and B cells, was distinctly depressed by DBTC. Hemagglutination and hemolysin titers as well as the number of direct plaque-forming cells against SRBC per spleen were decreased in a dose-related manner by DBTC. The phagocytic capacity of macrophages of rats was not affected by DOTC as was shown in the carbon clearance test. Altered immune functions were never found in mice or guinea pigs exposed to DBTC or DOTC. From this study it is concluded that both DBTC and DOTC induce immune suppression in rats by a selective inhibition of T-lymphocyte activity. Immune suppression was most pronounced in animals exposed to the chemicals during the developmental phase of the lymphoid system.

Bovine Intestinal Alkaline Phosphatase Attenuates the Inflammatory Response in Secondary Peritonitis in Mice

ABSTRACT Lipopolysaccharide (LPS) contributes importantly to morbidity and mortality in sepsis. Bovine intestinal alkaline phosphatase (BIAP) was demonstrated to detoxify LPS through dephosphorylation. LPS injection combined with BIAP reduced inflammation and improved survival in various experimental settings. In this study, single-dose intravenous administration of BIAP (0.15 IU/g) was applied in a murine cecal ligation and puncture (CLP) model of polymicrobial sepsis. Saline was given as control (S group). Treatment with BIAP prior to CLP (prophylaxis; BIAP-P group) or shortly after (early treatment; BIAP-ET group) reduced cytokine concentrations in plasma and peritoneal lavage fluid (PLF). Tumor necrosis factor-alpha peak levels decreased from 170 pg/ml (S) to 57.5 (BIAP-P) and 82.5 (BIAP-ET) in plasma and in PLF from 57.5 pg/ml (S) to 35.3 (BIAP-P) and 16.8 (BIAP-ET) (all, P < 0.05). Peak interleukin-6 levels in plasma decreased from 19.3 ng/ml (S) to 3.4 (BIAP-P) and 11.5 (BIAP-ET) and in PLF from 32.6 ng/ml (S) to 13.4 (BIAP-P) and 10.9 (BIAP-ET) (all, P < 0.05). Macrophage chemoattractant protein 1 peak levels in plasma decreased from 2.0 ng/ml (S) to 1.0 (BIAP-P) and 0.7 (BIAP-ET) and in PLF from 6.4 (S) to 2.3 (BIAP-P) and 1.3 ng/ml (BIAP-ET) (all, P < 0.05). BIAP-treated groups showed decreased transaminase activity in plasma and decreased myeloperoxidase activity in the lung, indicating reduced associated hepatocellular and pulmonary damage. Survival was not significantly altered by BIAP in this single-dose regimen. In polymicrobial secondary peritonitis, both prophylactic and early BIAP treatment attenuates the inflammatory response both locally and systemically and reduces associated liver and lung damage.

Lymphocytotoxicity and immunosuppression by organotin compounds. Suppression of graft-versus-host reactivity, blast transformation, and E-rosette formation by Di-n-butyltindichloride and Di-n-Octyltindichloride

Di-n-butyltindichloride (DBTC) and di-n-octyltindichloride (DOTC) represent a new group of organometallic compounds with antilymphocytic properties. In rats they induce lymphocyte depletion in thymus and thymus-dependent areas of spleen and peripheral lymph nodes without signs of myelotoxicity or a generalized toxicity. The number and viability of cells isolated from thymus and peripheral lymphoid organs was severely decreased, whereas the number and viability of bone marrow cells was not reduced. Immunosuppressive properties of DBTC and DOTC are indicated, in this study, by a severe decrease of the graft-versus-host response and the response to the T-cell mitogens phytohemagglutinin and concanavalin A. The T-cell selectivity of these compounds is discussed. In vitro DBTC and DTOC are extremely cytotoxic. Blast transformation of human as well as rat thymocytes was already inhibited at concentrations as low as 0.02 micrograms DBTC (or 0.1 micrograms DOTC) ml medium. Also the E-rosette formation was inhibited at very low drug levels. The similarity of effects upon rat and human lymphocytes suggests that DBTC and DOTC acts in the same manner in rat and man and offers the possibility of a therapeutic use of these compounds.

Intestinal alkaline phosphatase contributes to the reduction of severe intestinal epithelial damage

Inflammatory bowel disease is characterized by chronic inflammation of the intestine and is accompanied by damage of the epithelial lining and by undesired immune responses towards enteric bacteria. It has been demonstrated that intestinal alkaline phosphatase (iAP) protects against the induction of inflammation, possibly due to dephosphorylation of lipopolysaccharide (LPS). The present study investigated the therapeutic potential of iAP in intestinal inflammation and epithelial damage. Intestinal epithelial damage was induced in C57BL/6 mice using detran sulfate sodium (DSS) and iAP was administered 4days after initial DSS exposure. Loss in body weight was significantly less in iAP-treated mice and accompanied with reduced colon damage (determined by combination of crypt loss, loss of goblet cells, oedema and infiltrations of neutrophils). Treatment with iAP was more effective in case of severe inflammation compared to situations of mild to moderate inflammation. Rectal administration of LPS into a moderate inflamed colon did not aggravate inflammation. Furthermore, soluble iAP did not lower LPS-induced nuclear factor-kappaB activation in epithelial cells in vitro but induction of cellular AP expression by butyrate resulted in decreased LPS response. In conclusion, the present study shows that oral iAP administration has beneficial effects in situations of severe intestinal epithelial damage, whereas in moderate inflammation endogenous iAP may be sufficient to counteract disease-aggravating effects of LPS. An approach including iAP treatment holds a therapeutic promise in case of severe inflammatory bowel disease.

A novel hypothesis for an alkaline phosphatase 'rescue' mechanism in the hepatic acute phase immune response.

The liver isoform of the enzyme alkaline phosphatase (AP) has been used classically as a serum biomarker for hepatic disease states such as hepatitis, steatosis, cirrhosis, drug-induced liver injury, and hepatocellular carcinoma. Recent studies have demonstrated a more general anti-inflammatory role for AP, as it is capable of dephosphorylating potentially deleterious molecules such as nucleotide phosphates, the pathogenic endotoxin lipopolysaccharide (LPS), and the contact clotting pathway activator polyphosphate (polyP), thereby reducing inflammation and coagulopathy systemically. Yet the mechanism underlying the observed increase in liver AP levels in circulation during inflammatory insults is largely unknown. This paper hypothesizes an immunological role for AP in the liver and the potential of this system for damping generalized inflammation along with a wide range of ancillary pathologies. Based on the provided framework, a mechanism is proposed in which AP undergoes transcytosis in hepatocytes from the canalicular membrane to the sinusoidal membrane during inflammation and the enzymes expression is upregulated as a result. Through a tightly controlled, nucleotide-stimulated negative feedback process, AP is transported in this model as an immune complex with immunoglobulin G by the asialoglycoprotein receptor through the cell and secreted into the serum, likely using the receptors State 1 pathway. The subsequent dephosphorylation of inflammatory stimuli by AP and uptake of the circulating immune complex by endothelial cells and macrophages may lead to decreased inflammation and coagulopathy while providing an early upstream signal for the induction of a number of anti-inflammatory gene products, including AP itself.

Endotoxin release in cardiac surgery with cardiopulmonary bypass: pathophysiology and possible therapeutic strategies. An update

Cardiac surgery with cardiopulmonary bypass provokes a systemic inflammatory response syndrome caused by the surgical trauma itself, blood contact with the non-physiological surfaces of the extracorporeal circuit, endotoxemia, and ischemia. The role of endotoxin in the inflammatory response syndrome has been well investigated. In this report, we reviewed recent advances in the understanding of the pathophysiology of the endotoxin release during cardiopulmonary bypass and the possible therapeutic strategies aimed to reduce the endotoxin release or to counteract the inflammatory effects of endotoxin. Although many different strategies to detoxify endotoxins were evaluated, none of them were able to show statistically significant differences in clinical outcome.

Anti-Inflammatory Effects of Alkaline Phosphatase in Coronary Artery Bypass Surgery with Cardiopulmonary Bypass

Laboratory and clinical data have implicated endotoxin as an important factor in the inflammatory response to cardiopulmonary bypass. Alkaline phosphatase prevents endotoxin-induced systemic inflammation in animals and humans. We assessed the effects of the administration of bovine intestinal alkaline phosphatase on surgical complications in patients undergoing coronary artery bypass grafting. In a double blind, randomized, placebo-controlled study, a total of 63 patients undergoing coronary artery bypass grafting were enrolled. Bovine intestinal alkaline phosphatase or placebo was administered as an intravenous bolus followed by continuous infusion for 36 hours. The primary endpoint was reduction of post-surgical inflammation. No significant safety concerns were identified. The overall inflammatory response to coronary artery bypass grafting with cardiopulmonary bypass was low in both placebo and bovine intestinal alkaline phosphatase patient group. Five patients in the placebo group displayed a significant TNFalpha response followed by an increase in plasma levels of IL-6 and IL-8. Such a TNFalpha response was not observed in the bovine intestinal alkaline phosphatase group, suggesting anti-inflammatory activity of bovine intestinal alkaline phosphatase. Other variables related to systemic inflammation showed no statistically significant differences. Bovine intestinal alkaline phosphatase can be administered safely in an attempt to reduce the inflammatory response in coronary artery bypass grafting patients with a low to intermediate EuroSCORE. The anti-inflammatory effects might be more pronounced in patients developing more fulminant postoperative inflammatory responses. This will be investigated in a further trial with inclusion of patients undergoing complicated cardiac surgery, demanding extended cardiopulmonary bypass and aortic cross clamp time. In this review article some recent patents related to the field are also discussed.

Prophylactic treatment with alkaline phosphatase in cardiac surgery induces endogenous alkaline phosphatase release.

Introduction Laboratory and clinical data have implicated endotoxin as an important factor in the inflammatory response to cardiopulmonary bypass. We assessed the effects of the administration of bovine intestinal alkaline phosphatase (bIAP), an endotoxin detoxifier, on alkaline phosphatase levels in patients undergoing coronary artery bypass grafting. Methods A total of 63 patients undergoing coronary artery bypass grafting were enrolled and prospectively randomized. Bovine intestinal alkaline phosphatase (n=32) or placebo (n=31) was administered as an intravenous bolus followed by continuous infusion for 36 hours. The primary endpoint was to evaluate alkaline phosphatase levels in both groups and to find out if administration of bIAP to patients undergoing CABG would lead to endogenous alkaline phosphatase release. Results No significant adverse effects were identified in either group. In all the 32 patients of the bIAP-treated group, we found an initial rise of plasma alkaline phosphatase levels due to bolus administration (464.27±176.17 IU/L). A significant increase of plasma alkaline phosphatase at 4–6 hours postoperatively was observed (354.97±95.00 IU/L) as well. Using LHA inhibition, it was shown that this second peak was caused by the generation of Tissue Non Specific Alkaline Phosphatase (TNSALP-type alkaline phosphatase). Conclusions Intravenous bolus administration plus 8 hours continuous infusion of alkaline phosphatase in patients undergoing coronary artery bypass grafting with cardiopulmonary bypass results in endogenous alkaline phosphatase release. This endogenous alkaline phosphatase may play a role in the immune defense system.

An alkaline phosphatase transport mechanism in the pathogenesis of Alzheimer's disease and neurodegeneration.

Systemic inflammation is associated with loss of blood-brain barrier integrity and neuroinflammation that lead to the exacerbation of neurodegenerative diseases. It is also associated specifically with the characteristic amyloid-β and tau pathologies of Alzheimers disease. We have previously proposed an immunosurveillance mechanism for epithelial barriers involving negative feedback-regulated alkaline phosphatase transcytosis as an acute phase anti-inflammatory response that hangs in the balance between the resolution and the progression of inflammation. We now extend this model to endothelial barriers, particularly the blood-brain barrier, and present a literature-supported mechanistic explanation for Alzheimers disease pathology with this system at its foundation. In this mechanism, a switch in the role of alkaline phosphatase from its baseline duties to a stopgap anti-inflammatory function results in the loss of alkaline phosphatase from cell membranes into circulation, thereby decreasing blood-brain barrier integrity and functionality. This occurs with impairment of both amyloid-β efflux and tau dephosphorylating activity in the brain as alkaline phosphatase is replenished at the barrier by receptor-mediated transport. We suggest systemic alkaline phosphatase administration as a potential therapy for the resolution of inflammation and the prevention of Alzheimers disease pathology as well as that of other inflammation-related neurodegenerative diseases.