Showing papers in "Advances in Experimental Medicine and Biology in 1996"

Fumonisins: history, world-wide occurrence and impact.

Abstract: The history, toxicological effects, world-wide natural occurrence and impact of the fumonisins, food-borne carcinogenic mycotoxins produced by Fusarium moniliforme, are reviewed from the original description of the fungus in 1881 to the present. Following the isolation and characterization of fumonisin B1 and B2 and the publication of the first 3 papers on fumonisins by South African researchers in 1988, the interest in these compounds increased dramatically during 1989 and 1990 because of numerous outbreaks of mycotoxicoses in animals associated with the 1989 corn crop in the USA. Major advances made during this period were published in approximately 49 papers from 1988 to 1991. During the period 1992 to 1994, there was an explosion in the literature on fumonisins and at least 212 papers were published. The information contained in the more than 260 papers on fumonisins published to date is reviewed with respect to toxicological effects, association with animal and human diseases, and world-wide natural occurrence in corn and corn-based feeds and foods. Impact of the fumonisins is addressed with respect to their implications for human and animal health, risk assessment and establishment of tolerance levels.

Structure-function analysis of Bcl-2 family proteins. Regulators of programmed cell death.

Abstract: The Bcl-2 protein blocks a distal step in an evolutionarily conserved pathway for programmed cell death and apoptosis. To gain better understanding of how this protein functions, we have undertaken a structure-function analysis of this protein, focusing on domains within Bcl-2 that are required for function and for interactions with other proteins. Four conserved domains are present in Bcl-2 and several of its homologs: BH1 (residues 136–155), BH2 (187–202), BH3 (93–107) and BH4 (10–30). Deletion of the BH1, BH2, or BH4 domains of Bcl-2 abolishes its ability to suppress cell death in mammalian cells and prevents homodimerization of these mutant proteins, though these mutants can still bind to the wild-type Bcl-2 protein. These mutants also fail to bind to BAG-1 and Raf-1, two proteins that we have shown can associate with protein complexes containing Bcl-2 and which cooperate with Bcl-2 to suppress cell death. Deletion of either BH1 or BH2 nullifies the ability of Bcl-2 to: (a) suppress death in mammalian cells; (b) block Bax-induced lethality in yeast; and (c) heterodimerize with Bax. In contrast, deletion of the BH4 domain of Bcl-2 nullifies anti-apoptotic function and homodimerization, but does not impair binding to the pro-apoptotic protein Bax. Taken together, the data suggest the possibility that both Bcl-2/Bcl-2 homodimerization and Bcl-2/Bax heterodimerization are necessary but insufficient for the anti-apoptotic function of the Bcl-2 protein. Homodimerization of Bcl-2 with itself involves a head-to-tail interaction, in which an N-terminal domain where BH4 resides interacts with the more distal region of Bcl-2 where BH1, BH2, and BH3 are located. In contrast, Bcl-2/Bax heterodimerization involves a tail-to-tail interaction, that requires the portion of Bcl-2 where BH1, BH2, and BH3 reside and a central region in Bax where the BH3 domain is located. The BH3 domain of Bax is also required for Bax/Bax homodimerization and pro-apoptotic function in both yeast and mammalian cells. Thus, Bcl-2 may suppress cell death at least in part by binding to Bax via the BH3 domain and thereby preventing formation of Bax/Bax homodimers. Further studies however are required to delineate the full significance of Bcl-2/Bcl-2, Bcl-2/Bax, and Bax/Bax dimers and the biochemical mechanisms by which Bcl-2 family proteins ultimately control cell life and death.

The Neuronal Ceroid Lipofuscinoses (Batten Disease)

Abstract: The neuronal ceroid lipofuscinoses (NCL), also called Batten disease, are a group of neurodegenerative lysosomal storage diseases affecting humans and other animals They are inherited as autosomal recessive traits Affected children start life normally and then develop increasing dementia, blindness and seizures, culminating in premature death There are three major forms of human NCL, the infantile form (INCL or CLN1) which occurs most frequently in Finland, and the late infantile (LINCL or CLN2) and juvenile (JNCL or CLN3) forms which occur worldwide A number of other forms have been described including a rare adult form, Kufs disease (CLN4) and a Finnish late infantile variant (CLN5) (Dyken, 1988) Each form is distinguished by the age of onset and the clinical course of the disease, and a confusing number of eponyms have been used to describe them (Dyken, 1988; Kohlschutter et al, 1993) Partly because of this, the collective incidence of these diseases is unknown However they are relatively common and an estimate of about 1 in 12,500 live births world-wide has been made (Rider and Rider, 1988)

Host range restricted, non-replicating vaccinia virus vectors as vaccine candidates

Abstract: The use of a recombinant virus containing a heterologous gene of another microorganism as a live vaccine was suggested more than 10 years ago (Mackett et al, 1982; Panicali and Paoletti, 1982). Vaccinia virus was considered for such a purpose because of its success as a smallpox vaccine and ease and economy of production, distribution and administration (Fermer et al., 1988). The extensive experimental use of recombinant vaccinia viruses was facilitated by the construction of plasmid transfer vectors containing a vaccinia virus promoter, one or more convenient restriction endonuclease sites for inserting a foreign gene, flanking DNA sequences for homologous recombination into a non-essential site of the vaccinia virus genome and for selection and/or screening of recombinant viruses (Chakrabarti et al., 1985; Mackett et al., 1984). Humoral and cell mediated immune responses to an expressed foreign protein and protection of experimental animals against challenge with the corresponding pathogen were demonstated in a variety of animal model systems (Cox et al., 1992; Moss, 1991).

Fas-mediated apoptosis.

Abstract: Homeostasis in vertebrates is tightly regulated by cell death as well as by cell proliferation. The death of cells during embryogenesis, metamorphosis, endocrine-dependent tissue atrophy, and normal tissue turnover is “programmed cell death”, mediated by a process called “apoptosis”. Cytotoxic T lymphocytes and natural killer cells kill the target cells by inducing apoptosis. Apoptosis can be distinguished from necrosis, which occurs as a result of injury, complement attack, severe hypoxia and hyperthermia. Morphological and biochemical analyses of the apoptotic cell death process indicated that apoptosis is accompanied by condensation of cytoplasm, loss of plasma membrane microvilli, segmentation of nucleus, and extensive degradation of chromosomal DNA into oligomers of 180 bp. Cellular proliferation and differentiation are mediated by a family of proteins called cytokines. Our studies on the Fas ligand and Fas have indicated that apoptosis is also mediated by a cytokine and its receptor in some cases. Here, I summarize the current status of the Fas death factor system.

Soy Isoflavonoids and Cancer Prevention

Abstract: The isoflavonoids in soy, genistein and daidzein, have been proposed to contribute an important part of the anti-cancer effect of soy. Although there have been many interesting studies on the effects of isoflavones on biochemical targets in tissue culture experiments, in most cases the concentrations used by investigators have exceeded 10 μM. However, based on simple pharmacokinetic calculations involving daily intake of isoflavones, absorption from the gut, distribution to peripheral tissues, and excretion, it is unlikely that blood isoflavone concentrations even in high soy consumers could be greater than 1–5 μM. Experiments designed to evaluate these pharmacological principles were carried out in anesthetized rats with indwelling biliary catheters and in human volunteers consuming soy beverages. The data from these experiments indicate that genistein is efficiently absorbed from the gut, taken up by the liver and excreted in the bile as its 7-O-β-glucuronide. Re-infused genistein 7-O-β-glucuronide was also well absorbed from the gut, although this occurred in the distal small intestine. In human subjects fed a soy beverage for a period of two weeks, plasma levels of genistein and daidzein, determined by HPLC-mass spectrometry, ranged from 0.55–0.86 μM, mostly as glucuronide and sulfate conjugates. In summary, genistein is well absorbed from the small intestine and undergoes an enterohepatic circulation. Although the plasma genistein levels achievable with soy food feeding are unlikely to be sufficient to inhibit the growth of mature, established breast cancer cells by chemotherapeutic-like mechanisms, these levels are sufficient to regulate the proliferation of epithelial cells in the breast and thereby may cause a chemopreventive effect.

Brain Serotonin, Carbohydrate-craving, obesity and depression.

Abstract: Serotonin in the brain, and perhaps elsewhere, is involved in mechanisms that affect macronutrient selection, generate feelings of satiety, and, if malfunctioning, predispose to obesity. Drugs which increase the quantities of serotonin present within synapses can cause weight reduction. Such drugs presently include those that release the neurotransmitter by a direct action on nerve terminals (e.g., dexfenfluramine’s metabolite dexnorfenfluramine) or by activating serotoninergic neurons (e.g., nicotine); by activating post-synaptic serotonin receptors (e.g., dexnorfenfluramine); or by prolonging serotonin’s existence within synapses by blocking its reuptake (e.g., dexfenfluramine; fluoxetine, sertraline; paroxetine). Additional ways are known by which intrasynaptic serotonin levels can be augmented (e.g. increasing its synthesis with tryptophan; inhibiting its destruction by monoamine oxidase), and it can be anticipated that drugs acting at these loci will also become candidates for treating obesity.

Treatment of Acyclovir-Resistant Herpes Simplex and Varicella Zoster Virus Infections

Abstract: Acyclovir-resistant HSV (ACV-R-HSV) infections were first described in 1982.1 To date, over 100 patients with ACV-R-HSV infection have been identified. Although in vitro resis10 tance to acyclovir has been demonstrated in isolates recovered from immunocompetent hosts,2 for practical purposes only the severely immunocompromised patient develops clinically significant ACV-R-HSV (e.g. patients with AIDS, hematologic malignancies, congenital immune deficiencies, or those who have undergone organ transplantation3–9). The single exception to date is a report by Kost et al. of an HIV-negative immunocompetent man with increasingly frequent recurrences of genital HSV despite the use of maximal doses of oral acyclovir for suppression or for treatment.10 This patient, however, appeared to have acquired ACV-R-HSV infection through direct transmission, having had in the recent past 3 new sexual partners of whom 2 had HIV infection.

Production of Influenza Virus in Cell Cultures for Vaccine Preparation

Abstract: Influenza virus strains of different types for use as an inactivated vaccine have been successfully grown in different cell lines. Increasing titres were obtained with BHK-21/BRS, VERO and MDCK cells. Cultures in stationary flasks, in spinner cultures or in large bioreactor systems were tested and the optimal conditions were studied. MDCK cells grown in serum-free medium before and during the virus production phase were found to yield high titres in the presence of trypsin. Satisfactory results were obtained with egg-adapted strains of human and equine origin as well as with strains just isolated from human patients without any further passages in eggs or cell culture.

The importance of superoxide in nitric oxide-dependent toxicity: evidence for peroxynitrite-mediated injury.

Abstract: The potent vasodilator effects of pharmacological agents such as nitroglycerin have been known for over 100 years. In the last 20 years evidence has accumulated suggesting that nitric oxide is the species responsible for the biological effects of nitro-vasodilators. However, only within the last 10 years has the importance of earlier studies become clear; nitric oxide is an enzymatically-produced second messenger. Work of atmospheric chemists would indicate that nitrogen oxides like nitric oxide are harmful, yet we live out our lives constantly generating nitric oxide in endothelial cells and neurons and pharmacologic doses of nitro-vasodilators are devoid of overt toxicity. Furthermore, at physiologically relevent concentrations, no direct toxicity of nitric oxide itself has been demonstrated. Nontheless, endogenous production of nitric oxide has been implicated in reoxygenation injury following ischemia (1,2), glutamate-mediated neuronal toxicity (2,3), inflammation (4–6) and graft versus host disease (7–9). How then, can nitric oxide be injurious?

Hepatotoxicity and -carcinogenicity of the fumonisins in rats. A review regarding mechanistic implications for establishing risk in humans.

Abstract: Cancer induction by the non-genotoxic mycotoxin, fumonisin B1, has been investigated by studying the mechanisms involved during cancer initiation and promotion in rat liver. Cancer initiation is effected through a toxic-proliferative response while the inhibitory effect on hepatocyte cell proliferation appears to be a key aspect determining cancer promotion. Dose-response effects of the fumonisins on the induction of early neoplastic lesions in both long- and short-term animal experiments have been established. The biphasic response of FB1 on hepatocyte proliferation will be discussed in relation to the known mechanisms of cancer induction by the genotoxic hepatocarcinogens. Recent investigations regarding the effect of the fumonisins on lipid biosynthesis and its inhibitory effect on hepatocyte growth stimulatory responses in vitro will be highlighted. Integration of our current knowledge regarding the carcinogenic potential of the fumonisins in setting a realistic and applicable risk assessment model for this non-genotoxic carcinogen will finally be addressed.

The mycotoxin fumonisin induces apoptosis in cultured human cells and in livers and kidneys of rats.

Abstract: Fumonisin B1 is a mycotoxin produced by Fusarium moniliforme, a fungus that infects corn and other grains in the U.S. Fumonisin ingestion causes a variety of effects including equine leukoencephalomalacia and porcine pulmonary edema, and has been associated epidemiologically with human esophageal cancer. Fumonisin B1 produces growth inhibition and increased apoptosis in primary human keratinocyte cultures and in HET-1A cells. In order to set the doses for a 2-year tumor bioassay, male and female F344 rats were fed fumonisin B1 (99, 163, 234, and 484 ppm) for 28 days and the organs examined histologically. There was a dose dependent decrease in liver and kidney weights in the rats. The liver weight loss was accompanied by the induction of apoptosis and hepatocellular and bile duct hyperplasia in both sexes, with the female rats being more responsive at lower doses. The induction of tubular epithelial cell apoptosis was the primary response of the kidneys to dietary fumonisin B1. Apoptosis was present at all doses in the kidneys of the male rats, and occurred in the females only at 163, 234, and 484 ppm fumonisin B1. These results demonstrate that fumonisin B1 treatment causes a similar increase in apoptosis both in vivo and in vitro.

Cathepsin B expression in human tumors

Abstract: Cathepsin B has been linked to tumor progression through observations that its activity, secretion or membrane association are increased. The most malignant tumors, and specifically the cells at the invasive edge of those tumors, express the highest activity. Cathepsin B may facilitate invasion directly by dissolving extracellular matrix barriers like the basement membrane, or indirectly by activating other proteases capable of digesting the extracellular matrix. Cathepsin B also might play a role in tumor growth and angiogenesis. Cathepsin B activity is the result of several levels of regulation: transcription, post-transcrip-tional processing, translation and glycosylation, maturation and trafficking, and inhibition. The majority of reports on cathepsin B expression in tumors have focused on measurements of activity or protein staining. In some tumors, e.g., gliomas, a correlation between the amounts of cathepsin B mRNA, protein and activity and tumor progression has been established. Regulation of cathepsin B at the transcriptional and post-transcriptional levels is still poorly understood. Although the putative promoter regions have characteristics of housekeeping-type promoters, cathepsin B mRNA expression varies depending on the cell type and state of differentiation. We have evidence that more than one promoter could direct expression of human cathepsin B. Multiple transcript species have been detected, resulting from alternative splicing in the 5′- and 3′-untranslated regions, and possibly the use of alternative promoter regions. The existence of transcript variants indicates a potential for post-transcriptional control of expression. In support of this, ras-transformation of MCF-10A human breast epithelial cells results in an increase in protein levels without a concomitant increase in mRNA levels. Cathepsin B mRNA species with distinct 5′-or 3′-untranslated regions may differ in their stability and translatability. Variations in the coding region may also alter cathepsin B properties. We and Frankfater’s group have observed transcript species that would encode a truncated protein, lacking the prepeptide and about half of the propeptide. This truncated protein, if synthesized in cells, would be expected to be cytosolic; therefore its function is unclear. Once the several mechanisms of regulation of cathepsin B expression and activity are better understood, they could provide us with new strategies to specifically reduce cathepsin B activity in tumors.

Reactive dopamine metabolites and neurotoxicity: implications for Parkinson's disease.

Abstract: Parkinson’s disease occurs in approximately one percent of individuals over the age of 55. It is characterized by the presence of tremor, rigidity, and bradykinesia. Pathologically, the hallmark of Parkinson’s disease is the progressive degeneration of the dopamine (DA)-containing neurons of the nigrostriatal projection. Neurological deficits associated with the loss of DA neurons do not appear until the degeneration is extensive, presumably due to compensatory properties of the remaining DA neurons and their targets (Bernheimer et al., 1973; Zigmond et al., 1984). The mechanism responsible for the degenerative process is not known, although factors such as genetic predisposition and environmental toxins have been proposed to play a role (Agid et al., 1993). There is also growing evidence that endogenous factors such as oxidative stress and DA itself may contribute to the neurodegenerative process (Cohen, 1983; Agid et al., 1993; Zigmond et al., 1992).

Biological Effects of Feed and Forage Saponins and Their Impacts on Animal Production

Abstract: Saponins occur in numerous forages and crop plants used to provide feed for domestic animals. The objective of this paper is to review the significance of saponins to animal production, with particular emphasis on new findings. In some cases, saponins are being revisited after a period of many years of inactivity. This applies, for example, to the roles of saponins in bloat and photosensitization.

Effects of oral taurine supplementation on lipids and sympathetic nerve tone.

Abstract: To assess effects of oral taurine supplementation on lipids and sympathetic nerve tone in healthy young men on experimental high fat and cholesterol diets.

Effect of saponins on the growth and activity of rhizosphere bacteria

Abstract: Saponins are a group of steroid or triterpenoid glycosides and related chemicals (sapogenins: non-glycosylated) found in roots, shoots, seeds, and flowers of many plant species. Saponins are of agronomic interest because of allelopathic interference with plant growth (Oleszek and Jurzysta, 1987; Waller et al., 1993). Saponins can be released into the soil by secretion from roots and/or leaching from living or decaying plant material (Mishustin and Naumova, 1955; Oleszek and Jurzysta, 1987). Conservation management practices designed to maintain plant residues on the soil surface such as the use of cover crops and reduced tillage are rapidly being adopted by growers; thus the impact of these compounds and other natural products on crop productivity and soil and rhizosphere microbial ecology warrants study.

Neurogenic inflammation. A model for studying efferent actions of sensory nerves.

Abstract: Several lines of evidence suggest that sensory nerves of the carotid body have an efferent function in addition to their afferent function of conducting chemoreceptive impulses to the brain. However, it has been difficult to document the release of substances from sensory nerve terminals on glomus cells and to determine whether such an efferent function plays a role in chemoreception. By comparison, the phenomenon of neurogenic inflammation has been relatively easy to study in rats and guinea pigs and has proven to be an informative model system for analyzing efferent actions of sensory nerves. The main characteristic of neurogenic inflammation is plasma leakage. Chemical irritants that activate unmyelinated sensory nerves cause plasma leakage in the skin, respiratory tract, and other organs by triggering the release of substances from sensory nerve fibers. Substance P, which is synthesized and released by some sensory neurons, appears to be the main active mediator, although other tachykinins, calcitonin gene-related peptide, and perhaps other peptides may also participate. Neurogenic inflammation results from the action of substance P on NK1 receptors, as demonstrated by selective NK1 receptor agonists and antagonists. The NK1 receptors involved in plasma leakage are located on the endothelial cells of postcapillary venules and collecting venules. Within seconds of the activation of NK1 receptors by substance P, gaps form in the endothelium of target vessels. The endothelial gaps are transient, and the leak normally ends in a few minutes. However, the magnitude of the response can increase in pathological conditions such as Mycoplasma pulmonis infection in rats, which results in a chronic inflammatory disease of the respiratory tract. The infected airway mucosa becomes abnormally vascular as a result of angiogenesis, and the endothelial cells of the newly formed vessels express increased numbers of NK1 receptors and thus are abnormally sensitive to substance P. Studies of neurogenic inflammation not only have helped to understand the efferent actions of sensory nerves but also have given insight into the mechanism and consequences of inflammatory changes in endothelial cells and in the plasma leakage that follows.

Structural aspects of autophagy.

Abstract: As a first step towards isolation of autophagic sequestering membranes (phagophores), we have purified autophagosomes from rat hepatocytes. Lysosomes were selectively destroyed by osmotic rupture, achieved by incubation of hepatocyte homogenates with the cathepsin C substrate glycyl-phenylalanyl-naphthylamide (GPN). Mitochondria and peroxisomes were removed by Nycodenz gradient centrifugation, and cytosol, microsomes and other organelles by rate sedimentation through metrizamide cushions. The purified autophagosomes were bordered by dual or multiple concentric membranes, suggesting that autophagic sequestration might be performed either by single autophagic cisternae or by cisternal stacks.

Effect of processing on fumonisin content of corn.

Abstract: Fumonisins (FBs) are a family of mycotoxins produced by Fusarium moniliforme and F. proliferatum, predominant corn pathogens, and are found in most corn-containing foods. The FBs are heat stable, resistant to ammoniation, and unlike most mycotoxins, are water-soluble. The levels in corn and corn-containing foods will be presented ranging from 2 ppm. Washing of contaminated FB-corn with water did not reduce the measured FB levels of significantly. The traditional processing step to make tortilla flour, nixtamalization [Ca(OH)2 cooking] to produce masa, reduced FB levels but produced hydrolyzed FB which was almost as toxic as FB. Retorting sweet corn in brine apparently produced hydrolyzed FB. Fermentation of corn to ethanol did not alter FB levels but distillation yielded FB-free ethanol. Attempts to enzymatically modify FB with several enzymes were unsuccessful. Reactions between FB and reducing sugars (glucose or fructose) to produce Schiff’s bases yielded products that were not toxic. The effects of these processing treatments must be evaluated both chemically and biologically.

Carotid chemoreflex. Neural pathways and transmitters.

Abstract: The chemoreceptor projection site encompasses a midline area (0-0.5 mm caudal, 0-0.5 mm lateral and 0.3-0.5 deep with respect to the calamus scriptorius) which includes the commissural subnucleus of NTS. The chemosensitive neurons located in this site fire tonically, without any rhythmic relation to the phrenic nerve bursts, and are not responsive to baroreceptor stimulation. These neurons may serve as a pre-central pattern generator for respiration. The mechanisms which mediate respiratory responses (i.e., PN responses) to carotid chemoreceptor stimulation are as follows. Activation of carotid chemoreceptors results in the release of an excitatory amino acid (probably glutamate) in the chemoreceptor projection site; both NMDA and non-NMDA receptors in this site mediate the responses to chemoreceptor stimulation. The chemosensitive neurons send projections (which have not been identified yet) to respiratory neurons. Unlike in the neonatal rat, the inspiratory drive to the phrenic motoneurons is mediated by both NMDA and non-NMDA receptors. The PN response to chemoreceptor stimulation are also mediated by NMDA and non-NMDA receptors located in the phrenic motor nucleus. Cardiovascular responses to chemoreceptor stimulation are mediated by neural pathways connecting the chemosensitive neurons to the rostral ventrolateral medullary pressor area. These projections are glutamatergic. Activation of the sympatho-excitatory neurons in the rostral ventrolateral medulla results in pressor and tachycardic responses characteristic of chemoreceptor activation.

Fumonisin toxicity and sphingolipid biosynthesis

Abstract: Fumonisins are inhibitors of sphinganine (sphingosine) N-acyltransferase (ceramide synthase) in vitro, and exhibit competitive-type inhibition with respect to both substrates of this enzyme (sphinganine and fatty acyl-CoA). Removal of the tricarballylic acids from fumonisin B1 reduces the potency by at least 10 fold; and fumonisin A1 (which is acetylated on the amino group) is essentially inactive. Studies with diverse types of cells (hepatocytes, neurons, kidney cells, fibroblasts, macrophages, and plant cells) have established that fumonisin B1 not only blocks the biosynthesis of complex sphingolipids; but also, causes sphinganine to accumulate. Some of the sphinganine is metabolized to the 1-phosphate and degraded to hexadecanal and ethanolamine phosphate, which is incorporated into phosphatidylethanolamine. Sphinganine is also released from cells and, because it appears in blood and urine, can be used as a biomarker for exposure. The accumulation of these bioactive compounds, as well as the depletion of complex sphingolipids, may account for the toxicity, and perhaps the carcinogenicity, of fumonisins.

Antitumorigenic Effects of Limonene and Perillyl Alcohol Against Pancreatic and Breast Cancer

Abstract: Perillyl alcohol is a natural product from cherries and other edible plants. Perillyl alcohol and d-limonene, a closely related dietary monoterpene, have chemotherapeutic activity against pancreatic, mammary, and prostatic tumors. In addition, perillyl alcohol, limonene, and other dietary monoterpenes have chemopreventive activity. Several mechanisms may account for the antitumorigenic effects of monoterpenes. For example, many monoterpenes inhibit the post-translational isoprenylation of cell growth-regulatory proteins such as Ras. Perillyl alcohol induces apoptosis without affecting the rate of DNA synthesis in both liver and pancreatic tumor cells. In addition, monoterpene-treated, regressing rat mammary tumors exhibit increased expression of transforming growth factor β concomitant with tumor remodeling/redifferentiation to a more benign phenotype. Monoterpenes are effective, nontoxic dietary antitumor agents which act through a variety of mechanisms of action and hold promise as a novel class of antitumor drugs for human cancer.

Redox regulation of AP-1: a link between transcription factor signaling and DNA repair.

Abstract: The inducible transcription factor, AP-\, is a heterodimeric leucine zipper complex containing the protein products ofthejos andjun protooncogenes. The DNA binding activity of Fos and Jun is regulated in vitro by a posttranslational mechanism involving reduction­ oxidation. Redox regulation is mediated through a conserved cysteine residue located in the DNA binding domain of both proteins. Oxidation or chemical modification of the cysteine has an inhibitory effect on AP-l DNA binding activity. Conversely, reduction of this residue by chemical reducing agents or by a ubiquitous nuclear redox factor (Ref-I), purified and cloned from human cells, stimulates AP-l DNA binding activity. In addition, recombinant Ref-l stimulates the DNA binding activity of several other classes of redox regulated transcription factors. Immunodepletion studies indicate that Ref-l is the major AP-l redox activity in Hela cells. Interestingly, Ref-l is a bifunctional protein; it also possesses an apuriniclapyrimidinic (AlP) endonuclease DNA repair activity. However, the redox and DNA repair activities of Ref-l are physically and biochemically distinguishable. Ref-l may represent a novel component of the signal transduction processes that regulate eukaryotic gene expression in response to cellular stress.

Regulation of T Cell Activation by CD28 and CTLA4

Abstract: T cell activation is initiated by engagement of the T cell receptor (TCR)-CD3 complex by antigen displayed by major histocompatibility complex (MHC) molecules expressed on the surface of an antigen presenting cell (APC). However, under most circumstances, this initial signal is insufficient to induce a proliferative response. TCR engagement in the absence of a second or costimulatory signal can lead to a state of anergy or cell death1. CD28 is a T cell surface receptor capable of providing this critical second signal following ligation of the B7 family of counter-receptors that are expressed on APCs2,3. This interaction provides not only proliferative signals, but also crucial survival signals that are important for both initiation and maintenance of an immune response4,5.

Carbohydrate—Lectin Interactions in Infectious Disease

Abstract: For a long time, carbohydrates were believed to serve solely as a source of energy and as structural elements, and to lack biological activity. They were therefore considered as dull molecules lacking the glamor of the other major cell constituents, the proteins and nucleic acids. This attitude started to change, albeit slowly, around 1970. An early indication for the change can be found in the book written by me some 20 years ago, where I stated that “ carbohydrates are ideally suited for the formation of specificity determinants that may be recognized by complementary structures, presumably proteins, on other cells or macromolecules” (Sharon, 1975). This modern view of carbohydrates has now become widely accepted. Thus, the recently published 4th edition of the popular biochemistry textbook by Stryer, (1995) states that “ carbohydrate units on cell surfaces play key roles in cell-cell recognition processes” and that “ carbohydrate-binding proteins called lectins mediate many biological recognition processes”.

Studies on bioactive saponins from Chinese medicinal plants

Abstract: In our recent studies on bioactive saponins, two Chinese medicinal plants: Mussaenda pubescens Ait.f (Rubiaceae) and Clematis chinensis Osbeck (Ranunculaceae), were investigated. Of the two medicinal plants, M. pubescens is a Chinese folk medicine which has been used as a diuretic, antiphlogistic, diaphoretic and antipyretic agent, and has also been used to detoxify mushroom poisons and to terminate early pregnancy. Clematis chinensis is a Chinese traditional medicine which has been used as an analgesic, diuretic, antitumor, antiinflammatory and insecticidal agent for ages. As a result of our studies, eighteen saponins were identified from M. pubescens, among which seventeen were new compounds, while eleven saponins were isolated from C. chinensis, three of which were new compounds. In the course of our structural studies, mass fragment analysis in FAB-mass spectra and 1H- and 13C-NMR spectra were used to determine the structures of the sapogenin and oligosaccharide moieties. In those more complicated and minor saponins, various 2D-NMR experiments were carried out on 400-, 500- or 600-MHz NMR instruments, which permitted the identification of new sapogenins in glycoside form. In addition, it was also possible to assign all the proton and carbon signals of the sugar units on the basis of 1H-1H DQF COSY, TOCSY and HMQC spectra, which further permitted the establishment of linkage sites and sequences among the sugar units and aglycones by means of NOESY, ROESY and HMBC spectra. When the proton signals of sugar units are overlapped seriously in 1H-NMR spectra, peracetylation is a very helpful technique which can spread proton signals in wider range, thus simplifying their assignment by means of 2D-NMR spectra. When some of the sugar proton signals of a peracetylate are also overlapped coincidentally, alternation of deuterated solvents for measuring NMR spectra can circumvent the difficulties. Pharmacological tests indicated that mussaendoside O, the most abundant saponin from M. pubescens, can inhibit significantly the secretions of the lachrymal and salivary glands induced by galanthamine, and can also inhibit the contraction of the isolated longitudinal muscle strip from guinea pig ileum evoked by an M-Ach receptor agonist (carbachol, 10(-6) M) at concentrations of 10(-4) and 10(-5) M. From these results, the saponin should be an antagonist of the M-Ach receptor, and was presumed to be responsible for its antitoxicity activity toward some mushroom poisons of medicinal plant origin. In addition, mussaendoside O also showed immunopromotive and hemolytic activities.

Steroid Saponins from Fenugreek and some of their Biological Properties

Abstract: Fenugreek (Trigonella foenum-graecum L., Leguminosae, Trifoliae, Trigonellinae) is an annual plant found primarily in Mediterranean countries, the Middle East, and India. The seeds are most often used as a food spice (curry) and in traditional medicine. Fenugreek seeds are assumed to have restorative and nutritive properties (appetite stimulant) and hypocholesterolemic and antidiabetic effects.

Identification of Hydrogen Peroxide as the Relevant Messenger in the Activation Pathway of Transcription Factor NF-κB

Abstract: The inducible higher eukaryotic transcription factor NF-κB is activated by a large variety of distinct simuli [1–3]. In unstimulated cells, this factor resides in a latent form in the cytoplasm [4]. Latency is achieved by association of the DNA-binding NF-κB dimer with an inhibitory subunit, called IκB [5]. IΚB suppresses DNA-binding and nuclear transport of NF-κB. Upon stimulation of cells, IκB is phosphorylated and proteolytically degraded [6–9]. Both reactions are required for activation [10]. The released NF-κB is then translocated to the nucleus where it initiates transcription of target genes. Among the numerous proteins which are induced by a concerted action of NF-kB with other transcription factors are cytokines, chemokines, cell adhesion molecules, hematopoetic growth factors and receptors, histocompatibility antigens and acute phase proteins [1–3]. While NF-κB may be indispensable as inducer of many immediate-early inflammatory and immune reactions, the transcription factor is likely to play a fatal role in certain diseases and syndromes that involve an abberrant expression of inflammatory cytokines [22–24].

The Metzincin-Superfamily of Zinc-Peptidases

Abstract: Over the past three years, the three-dimensional structures of a number of zinc proteinases that share the zinc-binding motif HEXXHXXGXXH have been elucidated. These proteinases comprise astacin, a digestive enzyme from crayfish [1,2,3], adamalysin II [4,5] and atrolysin C [6] from snake venom, the Pseudomonas aeruginosa alkaline proteinase [7] and serralysin from Serratia marcescens proteinase [8], the collagenases from human neutrophils [9,10,11]) and fibroblasts [12,13,14,15], human stromelysin 1 [16; K. Appelt, personal communication] and matrilysin [M. Browner, Keystone Symposia, March 5–12, 1994]. These enzymes represent four different families of zinc peptidases: the astacins [3,17], the bacterial serralysins [18], the adamalysins/reprolysins [19,20], and the matrixins (matrix metalloproteinases, MMPs) [21,22].