Toxoplasma gondii induces changes in intracellular calcium in macrophages.

Toxoplasma gondii is an obligate intracellular parasite that interacts with calcium storage organelles and induces calcium-dependent signalling in macrophages. This study was performed to determine whether Toxoplasma induces changes in intracellular calcium in these cells. Ratiometric imaging of live, Fura-2 loaded macrophages challenged with T. gondii revealed robust elevations in intracellular calcium. These elevations were late in onset, beginning 15-20 min after addition of parasites and occurred in up to 20% of macrophages in an imaging field. Further characterization of these events revealed that they follow from challenge with live T. gondii, but not heat-killed parasites or soluble Toxoplasma antigen (STAg). Parasite-induced calcium elevations derived from extracellular sources, and were independent of host recognition factors MyD88 and CCR5. These findings indicate that Toxoplasma gondii alters calcium homeostasis in macrophages and this activity is independent of known pathways involved in the innate recognition of this organism.

Photoelectron spectroscopy characterization of diamond-like carbon films.

The electronic states of diamond-like hydrogenated carbon (DLC) films were studied by synchrotron radiation photoelectron spectroscopy. The valence band spectra measured at different excitation energies show the gradual emergence of the p-pi band in relation to the sample annealing and ion bombardment amorphization. The p-pi band of the annealed DLC was characterized by localized p(z) states, while the formation of the amorphous carbon surface was accompanied by appearance of the delocalized p(z) states, which reduce the optical gap. A simple approach permitting the extraction of the 2p band shape from the photoelectron spectra is proposed.

Comparison of FK 506, mycophenolate mofetil, and aminoguanidine effects on delay of corneal allograft rejection in an experimental model of low-risk and high-risk keratoplasty.

The purpose of our study was to compare the effectiveness of immunosuppressive drugs on the prevention of allograft rejection in a murine model of low-risk and high-risk keratoplasty. The therapy included FK 506 (tacrolimus; 0.2 mg/kg), mycophenolate mofetil (30 mg/kg), aminoguanidine (0.1 g/kg), and combination of FK506 + mycophenolate mofetil or FK506 + aminoguanidine. The results obtained from the Gray's survival model stratified according to the type of subjects suggest that a major rejection risk reduction was achieved using FK506; good results also were obtained for mycophenolate mofetil. Although the point estimates of both the survival and relative risk of rejection suggest a deferred effect of the combination FK506 + mycophenolate mofetil, this finding did not prove statistically significant.

Neuroendocrine immune interactions in health and disease.

The purpose of this paper is to review ways in which the neurohormonal system can interact with the immune system and to outline the main mechanisms which are involved in this interaction. Experimental as well as clinical evidence is presented to support the existence of a close interaction and bi-directional communication between the central nervous and immune systems. The role of major endocrine mechanisms and hormones is discussed. The evidences from experimental work to support the roles of the nervous system with neurotransmitters, the endocrine system with hormones, and the immune system with cytokines are presented. Aging, depression and cancer have a high degree of co-association and share mechanisms which result in cellular immune deficiency. Hormone therapy, zinc replenishment, antidepressants, immunomodulators like MDP act on these pathways to upregulate and improve cellular immunity. The authors believe that the central nervous system (CNS)-immune interaction is an important new frontier to be considered for new combination therapy in diseases with cellular immune deficiency such as cancer particularly in the aged with depression.

Structures and pathways of the central nervous system are potentially involved in the serotonergic modulation of gastrointestinal activity.

The supposed involvement of rat brain regions in the modulation of rat small intestine serotonergic activity was investigated. Small electrolytic lesions were placed in the areas of medulla oblongata and pons Varoli; one week later, changes in the serotonergic response of the intestine were detected. The contractions mediated by the activation of 5-HT2 receptors in the proximal ileum were investigated. The whole ileum segments were cut and placed into the bath. The preparations were contracted by adding increasing concentrations of 5-hydroxytryptamine (5-HT) (10 nM-1 microM) and noncumulative concentration-response curves (CRCs) were established. The differences between 5-HT responses of preparations from either sham-operated or experimental rats suggest the existence of brainstem regions (dorsal vagal and solitary nuclei, parvocellular reticular nuclei and serotonergic A1,2,5 groups) that either stimulate or inhibit 5-HT modulatory action in the rat gastrointestinal tract.

Stimulation of nonspecific immunity, haemopoiesis and protection of mice against radiation injury by 1-adamantylamide-L-alanyl-D-isoglutamine incorporated in liposomes.

1-Adamantylamide-L-alanyl-D-isoglutamine (adamantylamide dipeptide (AdDP)) belongs to a group of desmuramyl muramyl peptide derivatives which are able to protect an organism from some viral infections. Encapsulation of AdDP to egg phosphatidyl choline liposomes and the targeting of this drug to lymphatic node macrophages via subcutaneous (s.c.) administration proved to be the efficient way to protect mice against irradiation when administered s.c., 24 h prior to lethal gamma-irradiation (long-term survival rate in the range of 40% compared with 0% in saline or free drug control). Parameters characteristic for the recovery of haemopoiesis in the bone marrow (number of granulocyte-macrophage haemopoietic progenitor cells, granulocyte-macrophage colony forming cells (GM-CFC)) were significantly improved in comparison with the controls and free drug on day 10 after 6.5 Gy irradiation. The haemopoietic effect was observed in the broad application time window (72 h before and 48 h after irradiation). Very high radioprotective effect of s.c. administered liposomal AdDP (L-AdDP) can be explained (together with induction of haemopoiesis) by the effective and long-lasting activation of nonspecific immunity, which withholds the onset of septicemia in early days after irradiation. Induction of nonspecific immunity was proven in Candida albicans infectious model. L-AdDP significantly increased both the survival time and score (about 40% survival compared with 0% in controls and free drug). In conclusion, L-AdDP could be therapeutically beneficial to moderate the haemopoietic damage (undesirable effect of radiotherapy or chemotherapy) and induce the non-specific immunity to support the antimicrobial treatment of immunocompromised patients.

Modulatory effect of cyclosporin A on tert-butyl hydroperoxide-induced oxidative damage in hepatocytes.

In the present work, we followed an in vitro protective action of cyclosporin A (CsA) against tert-butyl hydroperoxide (t-BHP)-induced oxidative damage in hepatocytes. Various parameters (cell viability, cytosolic calcium level, rhodamine 123 accumulation as indicator of mitochondrial membrane potential and alanine-aminotransferase leakage from cells) were measured as an index of cytotoxicity. Tert-butyl hydroperoxide (1 mM) significantly increased cytosolic Ca2+ and affected mitochondrial membrane potential. Pretreatment with cyclosporin A (0.5 microM) reduced t-BHP-induced cytosolic Ca2+ increase and ALT (alanine-aminotransferase) leakage, but had no protective effect on t-BHP-induced changes of mitochondrial membrane potential. Our data thus suggest that the mechanism of cytoprotection of CsA on the cytosolic Ca2+ changes and ALT leakage induced by t-BHP, does not directly correlate with protection of t-BHP-induced changes of mitochondrial membrane potential.

Different mechanisms in inhibition of rat macrophage nitric oxide synthase expression by FK 506 and cyclosporin A.

The modulatory effect of FK 506 and cyclosporin A (CsA) on the expression of inducible nitric oxide synthase (iNOS) in macrophages and mechanisms of their action were analysed. Isolated rat peritoneal macrophages were cultured for 12 or 24 h with or without lipopolysaccharide (LPS) (5 microg/ml) and in the absence or presence of FK 506 or CsA (0.1 and 1 microg/ml). Total RNA from macrophages was isolated and the expression of the gene for iNOS was assessed by using RT-PCR. The concentration of NO2- in culture supernatants was taken as a measure of nitric oxide (NO) production. FK 506 (0.1 and 1 microg/ml) reduced the LPS-induced increase of NO2- levels by 68% and 81%, respectively. CsA (0.1 and 1 microg/ml) decreased levels of nitrites by 39% and 69%, respectively. The results obtained suggest that both immunosuppressive drugs exhibit dose-dependent inhibitory effect on NO production and that FK 506 is more potent agent than CsA, in this respect. FK 506 exhibits its inhibitory effect on a phosphatase at the transcriptional level in macrophages. iNOS expression down-regulation by CsA is occurred post-transcriptionally.

Hemopoiesis-stimulating action of adamantylamide dipeptide: kinetics of increase of GM-CFC in femur and co-stimulating activity of serum, role of bone marrow stromal cells.

Chief part of hemopoietic stromal cells in mediating hemopoiesis-stimulating effects of adamantylamide dipeptide (AdDP), a synthetic immunomodulatory compound, has been determined in a series of combined in vivo/in vitro studies. Indirect stimulatory effect of AdDP on proliferation of hemopoietic progenitor cells for granulocytes and macrophages (GM-CFC) was proved to be mediated by the cells of hemopoietic microenvironment growing as adherent stromal cell populations in vitro. These results supplement previously reported findings of a positive role which is played by AdDP at modulating the interplay among stimulatory cytokines and their cellular sources, and are in consent with the idea to introduce AdDP as a constituent of the hemopoiesis- and immunity-stimulating supportive medical care.

Two-dimensional model of thermal smoothing of laser imprint in a double-pulse plasma

The laser prepulse effect on the thermal smoothing of nonuniformities of target illumination is studied by means of a two-dimensional Lagrangian hydrodynamics simulation, based on the parameters of a real experiment. A substantial smoothing effect is demonstrated for the case of an optimum delay between the prepulse and the main heating laser pulse. The enhancement of the thermal smoothing effect by the laser prepulse is caused by the formation of a long hot layer between the region of laser absorption and the ablation surface. A comparison with experimental results is presented.

MDP and 5-HT receptors. Does MDP interact with 5-HT(7) receptors?

A possible interaction of immunomodulator muramyl dipeptide (MDP) with 5-HT(7) (5-hydroxytryptamine) receptors was investigated. The activation of 5-HT(7) receptors relaxes the guinea-pig distal ileum. The whole ileum segments were, therefore, cut and placed into the bath. The preparations were precontracted by substance P and potently relaxed by adding incremental concentrations of 5-carboxamidotryptamine (5-CT) (0.01-3.2 microM), less potently by 5-hydroxytryptamine (5-HT) (1-100 microM). The preparations most sensitive to 5-CT were also relaxed by MDP (1-100 microM). Noncumulative concentration-response curves (CRCs) for 5-HT or 5-CT were established in the absence or presence of 5-HT antagonist metergoline (320 nM). Metergoline inhibited the relaxations and shifted the CRCs to the right. In the preparations most sensitive to the effects of both 5-CT and metergoline, the latter substance also inhibited the effect of the highest concentration (100 microM) in CRCs for MDP. In another type of experiments, CRCs for 5-HT or 5-CT were constructed in the presence of low concentrations of MDP (5-500 nM). The relaxations evoked by either drug remained unchanged. These results suggest that low concentrations of MDP do not interact with activation of 5-HT(7) receptors. In higher concentrations MDP acts on this receptor type as a very weak partial agonist.

Potential role of cannabinoids in Parkinson's disease.

Parkinson's disease (PD) is a neurodegenerative disorder caused by a progressive loss of dopaminergic neurons of the substantia nigra, resulting from an oxidative stress. The lack of dopaminergic neurons is reflected by a disturbed balance of the neural circuitry in the basal ganglia. Cannabinoids might alleviate some parkinsonian symptoms by their remarkable receptor-mediated modulatory action in the basal ganglia output nuclei. Moreover, it was recently observed that some cannabinoids are potent antioxidants that can protect neurons from death even without cannabinoid receptor activation. It seems that cannabinoids could delay or even stop progressive degeneration of brain dopaminergic systems, a process for which there is presently no prevention. In combination with currently used drugs, cannabinoids might represent, qualitatively, a new approach to the treatment of PD, making it more effective.

Past, present and future of psychoneuroimmunology.

Psychoneuroimmunology was for the first time comprehensively described about 20 years ago. The influence of mental status on the course and outcome of a number of diseases, however, was suspected a long time before. Also the links between mental affective disorders and the immune status were repeatedly suggested. The authors in this paper shortly reviewed the most important clinical as well as experimental evidence which at present strongly supports the concept of a close and bidirectional communication between central nervous, neuroendocrine and immune systems. The most important anatomical, physiological as well as pharmacological experimental data, which were obtained by the authors during 20 years of research in this field, are presented. The data strongly suggest that in the very next future we will not only better understand a very complex communication between mind and body, but also completely new types of compounds might become available.

Adamantylamide dipeptide stimulates hematopoiesis and increases survival in irradiated mice.

It has been demonstrated that the synthetic immunostimulatory compound, adamantylamide dipeptide (AdDP) produces hematopoiesis-stimulating effects in mice exposed to sublethal doses of ionizing radiation and increases survival in experimental animals irradiated with a lethal dose. These findings might suggest contingent extension of clinical indications for the administration of AdDP for the conditions of hematopoietic suppression, especially in oncology.

[New, modern, centrally active antihypertensive agents in the treatment of essential hypertension]. / Nová moderní centrálne úcinkující antihypertenziva v lécení esenciální hypertenze.

It was recently found that the rise of blood pressure leads to the excitation of a vasomotor centre in the brain stem and that the accompanying decrease in brain cortex excitability results in the reduced sensitivity to various adverse stimuli. Centrally acting antihypertensives, moxonidine and rilmenidine, do not impair circulatory reflexes and therefore do not deprive the patient of a chance to resist the pressure; thus the compliance of the patient might be increased. Both drugs activate I1-imidazoline receptors on the neurons of the rostral ventrolateral medulla oblongata. The reduction of neuronal firing rate results in the decrease of sympathetic activity and arterial pressure. Beside other advantages, centrally acting antihypertensives might be more promising than peripherally acting drugs due to their possible more favourable psychopharmacological profile; this component of their action might be underestimated at present.

The amelioration of hepatocyte oxidative stress injury by nitric oxide released from S-nitroso-N-acetyl penicillamine: a study in immobilized perfused hepatocytes.

S-nitroso-N-acetyl penicillamine (SNAP, 0.1-0.5 mM) caused release of nitric oxide (NO) into the perfusion medium of immobilized hepatocytes. Oxidative injury of hepatocytes was evoked by tert-butyl hydroperoxide (TBH, 1 mM) and the functional and morphological ultrastructural integrity of the cells was monitored. At the end of a 270-min perfusion period, SNAP-induced NO reduced lactate dehydrogenase leakage in TBH-injured hepatocytes as compared to untreated TBH-injured cells (122% +/- 5 vs. 146% +/- 6 of control levels), lipid peroxides production (2.7 +/- 0.2 vs. 3.7 +/- 0.3 nmol/10(6) cells), increased O2 consumption (26 +/- 2 vs. 12 +/- 1 nmol/10(6) cells) although urea synthesis was reduced. SNAP improved the formation of granules in the Golgi complex as compared to untreated TBH-injured hepatocytes and preserved the ultrastructural architecture of mitochondria and the smooth endoplasmic reticulum. The present data support a possible protective role of NO in oxidative liver injury.

Prenatal dihydrotestosterone differentially masculinizes tonic and surge modes of luteinizing hormone secretion in sheep.

The control of LH secretion in sheep is sexually differentiated. Males begin to reduce their sensitivity to inhibitory steroid feedback, leading to a pubertal increase in tonic LH secretion by 10 weeks of age, but females remain hypersensitive until 30 weeks. Moreover, only females can respond to the positive feedback action of estradiol to produce a preovulatory LH surge. Prenatal exposure of the female lamb to testosterone masculinizes tonic LH and abolishes the LH surge postnatally. However, the type of steroid involved is not known because testosterone can be converted to estradiol or dihydrotestosterone (DHT). This study tested the hypothesis that DHT, which cannot be converted to an estrogen, masculinizes tonic LH without defeminizing the LH surge. Pregnant ewes were treated with DHT (800, 400, or 200 mg/week) during the critical period for sexual differentiation of gonadotropin secretion (days 30-90; 145 days is term). To evaluate the time of the decrease in responsiveness to steroid inhibition, a constant steroid feedback signal was produced. At 4 weeks of age, androgenized females (800 mg, n = 5; 400 mg, n = 4; 200 mg, n = 5) and control males (n = 7) and females (n = 9) were gonadectomized and implanted with a SILASTIC brand estradiol capsule. Tonic LH secretion in males began to increase at 6.7 +/- 0.5 weeks (mean +/- SEM). In DHT-treated females, the LH increase began at the same time (800 mg DHT, 10.7 +/- 3.9 weeks; 400 mg DHT, 9.9 +/- 5.9 weeks; 200 mg DHT, 7.1 +/- 4.9 weeks). This was several months earlier than in control females (29.1 +/- 0.8 weeks; P < 0.05). After puberty, estradiol induced LH surges in 8 of 9 control females and 11 of 12 DHT-treated females, but not in any control males. These results lead to the hypothesis that in the sheep, distinct requirements exist for differentiation of 2 types of reproductive hormone control systems, and that conversion of testosterone to an estrogen is not essential for both. Aromatization is necessary to prevent the surge control of GnRH from operating in the male, but nonaromatizable androgens differentiate the tonic control to permit high GnRH secretion earlier in life.

The interaction of immunomodulatory muramyl dipeptide with peripheral 5-HT receptors: overview of the current state.

Immunomodulator muramyl dipeptide (MDP) exerts also pronounced neuropharmacological activities which are probably mediated by an interaction with 5-HT receptors. Some of these effects are considered as undesirable by its clinical use. More precise information concerning MDP effects on 5-HT receptors with respect to their many subtypes could result from studies using isolated organs in vitro. Earlier conducted studies of this type provided data that are concisely overviewed and reinterpreted here from the view of current 5-HT receptor classification. Since new 5-HT receptor types have emerged recently, new studies are under way. The results might contribute to the development of novel immunomodulatory drugs devoid of adverse effects.

Effects of in vivo administration of adamantylamide dipeptide on bone marrow granulocyte-macrophage hemopoietic progenitor cells (GM-CFC) and on ability of serum of the treated mice to stimulate GM-CFC colony formation in vitro: comparison with muramyl dipeptide and glucan.

Adamantylamide dipeptide (AdDP), muramyl dipeptide (MDP), and glucan were shown to increase significantly the numbers of granulocyte-macrophage hemopoietic progenitor cells (GM-CFC) in the bone marrow of mice. However, whereas the sera of mice given MDP or glucan were found to stimulate the growth of colonies from GM-CFC in vitro, i.e. to produce a colony-stimulating activity (CSA), administration of AdDP did not lead to this effect. Nevertheless, when serum of mice given AdDP was added to the cultures concomitantly with a suboptimal concentration of mouse interleukin-3 (mIL-3), a broad spectrum hemopoietic stimulator, counts of colonies from GM-CFC were significantly increased, and accelerated growth of the colonies was found as well. This property of AdDP, i.e. its ability to exhibit co-stimulating activity (CoSA) without being able to exhibit CSA, suggests that AdDP acts in hemopoietic tissues differently as compared with the two other immunomodulators studied. It can be hypothesized that the action of AdDP is more specific when compared with its natural related compound, MDP, as well as with glucan. Our findings prove the possibility to stimulate by AdDP the granulopoietic compartment of hemopoiesis and are in agreement with previous observations concerning the absence of systemic side effects of AdDP. Both these qualities of AdDP may be advantageous when pondering over contingent clinical utilization of AdDP as hemopoietic stimulator.

Erratic behavior of nitric oxide within the immune system: illustrative review of conflicting data and their immunopharmacological aspects.

The literature data assembled in this article document the variation of immunobiological effects of nitric oxide (NO). A number of factors are obviously responsible for the diversity, ranging from inactivity, alleviation, but not rarely to exacerbation of certain pathogenetic processes. A better understanding of NO interactions with the immune system can only be reached if more complex experimental designs to study the effects of reactive nitrogen species are adopted in the future. They should integrate major participating variables and take into account pharmacodynamic/kinetic aspects of NO production in triggering the ultimate effects. If manipulation of NO in the organism by means of recently developed NO inhibitors and NO donors is to become a rational tool of immunopharmacological strategies, detailed knowledge of their pharmacologies and toxicologies is urgently needed in order to differentiate between the effects of NO and other side effects. Hopefully, this approach could improve the predictability of the clinical outcomes of NO manipulation.