Mohamed B. Abou-Donia

Organophosphorus ester-induced chronic neurotoxicity.

Organophosphorus compounds are potent neurotoxic chemicals that are widely used in medicine, industry, and agriculture. The neurotoxicity of these chemicals has been documented in accidental human poisoning, epidemiological studies, and animal models. Organophosphorus compounds have 3 distinct neurotoxic actions. The primary action is the irreversible inhibition of acetylcholinesterase, resulting in the accumulation of acetylcholine and subsequent overstimulation of the nicotinic and muscarinic acetylcholine receptors, resulting in cholinergic effects. Another action of some of these compounds, arising from single or repeated exposure, is a delayed onset of ataxia, accompanied by a Wallerian-type degeneration of the axon and myelin in the most distal portion of the longest tracts in both the central and peripheral nervous systems, and is known as organophosphorus ester-induced delayed neurotoxicity (OPIDN). In addition, since the introduction and extensive use of synthetic organophosphorus compounds in agriculture and industry half a century ago, many studies have reported long-term, persistent, chronic neurotoxicity symptoms in individuals as a result of acute exposure to high doses that cause acute cholinergic toxicity, or from long-term, low-level, subclinical doses of these chemicals. The author attempts to define the neuronal disorder that results from organophosphorus ester-induced chronic neurotoxicity (OPICN), which leads to long-term neurological and neurobehavioral deficits. Although the mechanisms of this neurodegenerative disorder have yet to be established, the sparse available data suggest that large toxic doses of organophosphorus compounds cause acute necrotic neuronal cell death in the brain, whereas sublethal or subclinical doses produce apoptotic neuronal cell death and involve oxidative stress.

DEPLETED AND NATURAL URANIUM: CHEMISTRY AND TOXICOLOGICAL EFFECTS

Depleted uranium (DU) is a by-product from the chemical enrichment of naturally occurring uranium. Natural uranium is comprised of three radioactive isotopes: 238U, 235U, and 234U. This enrichment process reduces the radioactivity of DU to roughly 30% of that of natural uranium. Nonmilitary uses of DU include counterweights in airplanes, shields against radiation in medical radiotherapy units and transport of radioactive isotopes. DU has also been used during wartime in heavy tank armor, armor-piercing bullets, and missiles, due to its desirable chemical properties coupled with its decreased radioactivity. DU weapons are used unreservedly by the armed forces. Chemically and toxicologically, DU behaves similarly to natural uranium metal. Although the effects of DU on human health are not easily discerned, they may be produced by both its chemical and radiological properties. DU can be toxic to many bodily systems, as presented in this review. Most importantly, normal functioning of the kidney, brain, liver, and heart can be affected by DU exposure. Numerous other systems can also be affected by DU exposure, and these are also reviewed. Despite the prevalence of DU usage in many applications, limited data exist regarding the toxicological consequences on human health. This review focuses on the chemistry, pharmacokinetics, and toxicological effects of depleted and natural uranium on several systems in the mammalian body. A section on risk assessment concludes the review.

Antimuscarinic drugs for overactive bladder and their potential effects on cognitive function in older patients.

Antimuscarinic agents are the predominant pharmacological treatment for patients with overactive bladder (OAB). These drugs are thought to act primarily through antagonism at muscarinic M3 receptors located at neuromuscular junctions in the human bladder detrusor muscle. Several of these drugs have been shown to be efficacious in ameliorating the symptoms of OAB in older patients, but most currently available agents lack selectivity for the M3 receptor subtype, and interaction with other muscarinic receptor subtypes throughout the body may adversely affect a variety of physiological functions and result in unwanted side effects, including cognitive dysfunction. With the recent availability of antimuscarinic agents that show increased selectivity for M3 receptors relative to other muscarinic subtypes, an invitational expert panel meeting was convened to review not only the mechanisms by which antimuscarinic agents could affect cognitive function, but also the published literature on cognitive adverse events. A review of the literature shows that the cholinergic system in the central nervous system (CNS) exerts a major influence on cognitive processes, in particular memory via M1 cholinergic receptors. In addition, recent evidence suggests a role for M2 receptors in mediating cognitive function. Thus, cognitive dysfunction (including memory loss) during treatment with nonselective antimuscarinic agents for OAB is of growing concern, particularly in older patients and those with mild cognitive impairment or dementia. Increased blood–brain barrier permeability, which can occur with advanced age and certain comorbidities, may also facilitate CNS access of antimuscarinic agents (regardless of their physiochemical properties) and add to antimuscarinic burden. On the basis of available evidence, antimuscarinic agents with selectivity for M3 over M1 and M2 receptors, limited CNS penetration, or both may therefore offer a favorable balance of efficacy in treating OAB together with a reduced risk of adverse cognitive events in the older population.

Physiological effects and metabolism of gossypol

Gossypol1 is a yellow coloring matter which occurs in various parts of the cotton plant. Longmore (1886) first isolated it as a crude pigment from cottonseed oil “foots,” a material which results from the refinement of crude cottonseed oil with sodium hydroxide and contains free fatty acids, phospholipids, gossypol, and other pigments. Marchlewski (1899) crystallized the acetic acid derivative of a compound from the same source, which he named “gossypol” to designate its origin, genus Gossypium (family Malvaceae), and its chemical nature as a phenol. Gossypol is found in the pigment “glands” (ovispherical bodies about 100 to 400 µm long) of the seed, leaf, stem, laproot bark, and roots of the cotton plant (Royce et al. 1941, Smith 1962). Studies of changes in the gossypol content of cottonseed during different stages of development of the bolls showed that the greatest increase in gossypol occurs between the time of maturity of the boll and the time it is about to open (Adams et al. 1960). Gossypol synthesis in all tissues of the cotton plant was induced in response to such irritants as pathogens, metabolic inhibitors, and cupric and mercuric ions (Bell 1967). Gossypol constitutes 20 to 40% of the weight of the pigment glands in cottonseed (Boatner 1948). Cottonseed usually contains 0.4 to 1.7% gossypol. The total world production of cottonseed is about 25 million tons/year (containing approximately 78,000 tons of gossypol) and about three to six million tons are produced in the United States (Decossas et al. 1968). An average ton of cottonseed gives about 335 pounds of oil and 945 pounds of meal (Altschul et al. 1958). If refined 60% protein cottonseed flour is prepared for human use, 300 to 400 pounds/ton is produced; the remainder can be used for animal feed. It is estimated that only onefourth of the cottonseed flour potentially available could satisfy the present worldwide shortage of protein (Gillham 1969).

Splenda Alters Gut Microflora and Increases Intestinal P-Glycoprotein and Cytochrome P-450 in Male Rats

Splenda is comprised of the high-potency artificial sweetener sucralose (1.1%) and the fillers maltodextrin and glucose. Splenda was administered by oral gavage at 100, 300, 500, or 1000 mg/kg to male Sprague-Dawley rats for 12-wk, during which fecal samples were collected weekly for bacterial analysis and measurement of fecal pH. After 12-wk, half of the animals from each treatment group were sacrificed to determine the intestinal expression of the membrane efflux transporter P-glycoprotein (P-gp) and the cytochrome P-450 (CYP) metabolism system by Western blot. The remaining animals were allowed to recover for an additional 12-wk, and further assessments of fecal microflora, fecal pH, and expression of P-gp and CYP were determined. At the end of the 12-wk treatment period, the numbers of total anaerobes, bifidobacteria, lactobacilli, Bacteroides, clostridia, and total aerobic bacteria were significantly decreased; however, there was no significant treatment effect on enterobacteria. Splenda also increased fecal pH and enhanced the expression of P-gp by 2.43-fold, CYP3A4 by 2.51-fold, and CYP2D1 by 3.49-fold. Following the 12-wk recovery period, only the total anaerobes and bifidobacteria remained significantly depressed, whereas pH values, P-gp, and CYP3A4 and CYP2D1 remained elevated. These changes occurred at Splenda dosages that contained sucralose at 1.1–11 mg/kg (the US FDA Acceptable Daily Intake for sucralose is 5 mg/kg). Evidence indicates that a 12-wk administration of Splenda exerted numerous adverse effects, including (1) reduction in beneficial fecal microflora, (2) increased fecal pH, and (3) enhanced expression levels of P-gp, CYP3A4, and CYP2D1, which are known to limit the bioavailability of orally administered drugs.

Biomarkers of apoptosis: release of cytochrome c, activation of caspase-3, induction of 8-hydroxy-2'-deoxyguanosine, increased 3-nitrotyrosine, and alteration of p53 gene.

Biomarkers rely on biochemical, histological, morphological, and physiological changes in whole organisms. Their use is becoming an important tool to examine changes at cellular and molecular levels, especially in nucleic acids and proteins. Biomarkers are used to measure exposure to a toxic agent, to detect severity of any toxic response, and to predict the possible outcome. Information on the mechanisms of action of toxicants can allow the development of potential biomarkers of effect and thus improvement of the risk assessment processes. Use of biomarkers as a tool to predict induction of apoptosis allows identification of biological signs that may indicate increased risk for disease. In cells undergoing apoptosis, the release of cytochrome c from the mitochondria to the cytoplasm and the activation of caspase-3, a key enzyme to execution stage of apoptotic pathway, have been studied as biomarkers of cell death (apoptosis). Products of DNA fragmentation that either accumulate in the cellular tissues or are excreted in the urine are useful markers of DNA damage. The induction level of urinary or cellular level of 8-hydroxy-2-deoxyguanosine and 3-nitrotyrosine has been used as a marker to measure extent of DNA oxidative damage. Furthermore, alteration or overexpression of the p53 gene was considered an indication of apoptosis. This article reviews some of the aspects of biomarkers of apoptosis, indicating relevance of their uses to predict apoptosis following exposure to environmental toxicants.

Gossypol: Uncoupling of respiratory chain and oxidative phosphorylation☆

Abstract Gossypol produced adverse effects in vitro on rat liver mitochondira. It stimulated mitochondrial respiration at low concentrations, inhibited it at high concentrations; abolished ADP/O and respiration control ratios; reversed inhibition caused by oligomycin; stimulated adenosine triphosphatase activity at low concentrations and inhibited it at high concentrations; and its effect on this enzyme was pH dependent. The possibility that gossypol may exert its toxic effect on poultry and animals by uncoupling respiratory chain-linked phosphorylation is discussed.

Disruption of the blood-brain barrier and neuronal cell death in cingulate cortex, dentate gyrus, thalamus, and hypothalamus in a rat model of Gulf-War syndrome.

We investigated the effects of a combined exposure to restraint stress and low doses of chemicals pyridostigmine bromide (PB), N, N-diethyl-m-toluamide (DEET), and permethrin in adult male rats, a model of Gulf-War syndrome. Animals were exposed daily to one of the following for 28 days: (i) a combination of stress and chemicals (PB, 1.3 mg/kg/day; DEET, 40 mg/kg/day; and permethrin, 0.13 mg/kg/day); (ii) stress and vehicle; (iii) chemicals alone; and (iv) vehicle alone. All animals were evaluated for: (i) the disruption of the blood-brain barrier (BBB) using intravenous horseradish peroxidase (HRP) injections and endothelial barrier antigen (EBA) immunostaining; (ii) neuronal cell death using H&E staining, silver staining, and glial fibrillary acidic protein (GFAP) immunostaining; and (iii) acetylcholinesterase (AChE) activity and m2-muscarinic acetylcholine receptors (m2-AChR). Animals subjected to stress and chemicals exhibited both disruption of the BBB and neuronal cell death in the cingulate cortex, the dentate gyrus, the thalamus, and the hypothalamus. Other regions of the brain, although they demonstrated some neuronal cell death, did not exhibit disruption of the BBB. The neuropathological changes in the above four brain regions were highly conspicuous and revealed by a large number of HRP-positive neurons (21-40% of total neurons), a decreased EBA immunostaining (42-51% reduction), a decreased number of surviving neurons (27-40% reduction), the presence of dying neurons (4-10% of total neurons), and an increased GFAP immunostaining (45-51% increase). These changes were also associated with decreased forebrain AChE activity and m2-AchR (19-25% reduction). In contrast, in animals exposed to stress and vehicle or chemicals alone, the above indices were mostly comparable to that of animals exposed to vehicle alone. Thus, a combined exposure to stress and low doses of PB, DEET, and permethrin leads to significant brain injury. The various neurological symptoms reported by Gulf-War veterans could be linked to this kind of brain injury incurred during the war.

Stress and Combined Exposure to Low Doses of Pyridostigmine Bromide, DEET, and Permethrin Produce Neurochemical and Neuropathological Alterations in Cerebral Cortex, Hippocampus, and Cerebellum

Exposure to a combination of stress and low doses of the chemicals pyridostigmine bromide (PB), DEET, and permethrin in adult rats, a model of Gulf War exposure, produces blood–brain barrier (BBB) disruption and neuronal cell death in the cingulate cortex, dentate gyrus, thalamus, and hypothalamus. In this study, neuropathological alterations in other areas of the brain where no apparent BBB disruption was observed was studied following such exposure. Animals exposed to both stress and chemical exhibited decreased brain acetylcholinesterase (AChE) activity in the midbrain, brainstem, and cerebellum and decreased m2 muscarinic acetylcholine (ACh) receptor ligand binding in the midbrain and cerebellum. These alterations were associated with significant neuronal cell death, reduced microtubule-associated protein (MAP-2) expression, and increased glial fibrillary acidic protein (GFAP) expression in the cerebral cortex and the hippocampal subfields CA1 and CA3. In the cerebellum, the neurochemical alterations were associated with Purkinje cell loss and increased GFAP immunoreactivity in the white matter. However, animals subjected to either stress or chemicals alone did not show any of these changes in comparison to vehicle-treated controls. Collectively, these results suggest that prolonged exposure to a combination of stress and the chemicals PB, DEET, and permethrin can produce significant damage to the cerebral cortex, hippocampus, and cerebellum, even in the absence of apparent BBB damage. As these areas of the brain are respectively important for the maintenance of motor and sensory functions, learning and memory, and gait and coordination of movements, such alterations could lead to many physiological, pharmacological, and behavioral abnormalities, particularly motor deficits and learning and memory dysfunction.

The cytoskeleton as a target for organophosphorus ester-induced delayed neurotoxicity (OPIDN)

Although the immediate action of organophosphorus esters is the inhibition of acetylcholinesterase, some of these compounds also produce a neurodegenerative disorder known as organophosphorus ester-induced delayed neurotoxicity (OPIDN). Tri-o-cresyl phosphate (TOCP) first produced this condition in humans and later in sensitive animal species. OPIDN is characterized by a delay period prior to onset of ataxia and paralysis. The neuropathologic lesions are Wallerian-type degeneration of the axon and myelin in the distal parts of the large tracts in both the central and peripheral nervous systems. In the past decade we have demonstrated that the pathognomonic features of OPIDN are an aberrant increase in autophosphorylation of calcium/calmodulin kinase II (CaM kinase II) and an increase in phosphorylation of cytoskeletal proteins, i.e., MAPs, tubulin, neurofilament triplet proteins, and myelin basic protein. Protein kinase-mediated phosphorylation of cytoskeletal proteins plays a critical role in regulating the growth and maintenance of the axon. We hypothesize that, in OPIDN, hyperphosphorylation of cytoskeletal proteins and axonal swelling are causally linked. Hyperphosphorylation of cytoskeletal proteins decreases their transport rate down the axon relative to their rate of entry into the axon, thus leading to their accumulation. Consistent with this hypothesis is our finding of the anomalous accumulation of phosphorylated neurofilament aggregates in the central and peripheral axons of hens treated with TOCP.