Characterization of the various forms of the Reelin protein in the cerebrospinal fluid of normal subjects and in neurological diseases

Reelin is a large extracellular glycoprotein that is defective in reeler mutant mice and plays a well-established role during brain development in human as well as rodents. In the adult brain, Reelin is expressed in a subset of GABAergic interneurons. Its role in disease states is not clearly defined, although it is implicated in autism and psychoses such as schizophrenia. In this report, we show that Reelin immunoreactive proteins can be detected in the human cerebrospinal fluid (CSF) with monoclonal antibodies directed against the N- and C-terminal regions of the protein. In CSF, Reelin is present as different products due to processing at two main sites; preservation at -20 degrees C increases processing further. CSF Reelin originates from the brain tissue and not from plasma. The protein was detected in comparable concentrations in children and adults, and the signal varied largely from subject to subject with no obvious correlation with age or neurological disease state.     

Flow cytometry for pediatric platelets

The ability of platelets to carry out their hemostatic function can be impaired in a wide range of inherited and acquired conditions: trauma, surgery, inflammation, pre-term birth, sepsis, hematological malignancies, solid tumors, chemotherapy, autoimmune disorders, and many others. Evaluation of this impairment is vitally important for research and clinical purposes. This problem is particularly pronounced in pediatric patients, where these conditions occur frequently, while blood volume and the choice of blood collection methods could be limited. Here we describe a simple flow cytometry-based screening method of comprehensive whole blood platelet function testing that was validated for a range of pediatric and adult samples (n = 31) in the hematology hospital setting including but not limited to: classic inherited platelet function disorders (Glanzmann’s thrombasthenia; Bernard-Soulier, Wiscott-Aldrich, and Hermasky-Pudlak syndromes, MYH9-dependent thrombocytopenia), healthy and pre-term newborns, acute and chronic immune thrombocytopenia, chronic lympholeukemia, effects of therapy on platelet function, etc. The method output includes levels of forward and side scatter, levels of major adhesion and aggregation glycoproteins Ib and IIb-IIIa, active integrins’ level based on PAC-1 binding, major alpha-granule component P-selectin, dense granule function based on mepacrine uptake and release, and procoagulant activity quantified as a percentage of annexin V-positive platelets. This analysis is performed for both resting and dual-agonist-stimulated platelets. Preanalytical and analytical variables are provided and discussed. Parameter distribution within the healthy donor population for adults (n = 72) and children (n = 17) is analyzed.     

Effects of Electrical Stimulation of the Hunger Center in the Lateral Hypothalamus and Food Reinforcement on Impulse Activity of the Proper Masticatory Muscle in Rabbits under Conditions of Hunger and Satiation

Threshold stimulation of the lateral hypothalamus in starving and preliminary fed (satiated) rabbits in the absence of feed induces searching behavior associated with burst-like impulse activity of proper masticatory muscle with a bimodal distribution of interpulse intervals, what represents an anticipatory type of reaction. The increase in the level of food motivation during threshold stimulation of the lateral hypothalamus in starving and satiated rabbits with the food offered led to successful food-procuring behavior, during which the frequency and amplitude of spike bursts in the proper masticatory muscle become comparable with those under conditions of natural foraging behavior stimulated by the need in nutrients. Motivational excitation and backward afferentation from food reward are addressed to the same neurons of the masticatory center in the medulla oblongata.     

A rationally assembled microbial community for growing Tagetes patula L. in a lunar greenhouse

Well-defined plant-associated bacteria were used for growing French marigolds (Tagetes patula L.) in anorthosite, a substrate of low bioavailability, analogous to a lunar rock. The consortium was composed mainly of plant growth-promoting rhizobacteria and biocontrol agents that were used for seed inoculation. Simultaneously, the sterile substrate was inoculated with the siliceous bacterium Paenibacillus sp. IMBG156. The plant benefited from bacterial activity which resulted in stimulation of seed germination, better plant development, and finally in flowering of inoculated tagetes. In contrast, control plants grew poorly in sterile anorthosite and never flowered. Analysis of bacterial community composition showed that both species colonized plant roots, and there were no shifts in the consortium structure in the rhizosphere of French marigolds within 6 weeks. Paenibacillus sp. IMBG156 was able to release some elements (Ca, Fe, Si) from substrate anorthosite. It was assumed that a rationally assembled consortium of bacterial strains supported growth and development of the model plant under growth-limiting conditions, at least by means of bioleaching and delivering of essential nutritional elements to the plants, and by promoting plant growth.     

Angiogenic and anti-angiogenic therapy for gastrointestinal ulcers: new challenges for rational therapeutic predictions and drug design

Gastrointestinal (GI) ulcers are essentially internal wounds that resist normal healing processes. Since their pathogenesis is poorly understood, and the etiologic (e.g., gastric acid, aspirin-like drugs, stress) and aggravating factors (e.g., H. pylori) are not well characterized, the remaining therapeutic option is to accelerate healing. Superficial mucosal lesions, i.e., erosions usually heal by epithelial regeneration and restitution, but when ulcers involve the muscularis propria, smooth muscle cells do not divide/regenerate. These deep lesions are filled by granulation tissue, i.e., angiogenesis followed by proliferation of connective tissue fibroblasts that deposit collagen over which adjacent surviving and dividing epithelial cells migrate to complete the healing. Our laboratory was the first to postulate that stimulation of angiogenesis alone might be sufficient to accelerate ulcer healing in the GI tract. Indeed, daily treatment of rats with bFGF, PDGF or VEGF markedly improved the healing of cysteamine-induced chronic duodenal ulcers, without any reduction in gastric acid secretion. These results were reproduced by a single dose of gene therapy by adenoviral vectors encoding PDGF or VEGF genes. The molar potency of angiogenic growth factors was 2-7 million times better than the antiulcerogenic effect of antisecretory H2 antagonists. Since histologically & pathologically gastroduodenal ulcers look similar to ulcers in the lower GI tract, we also predicted that the healing of experimental ulcerative colitis might be also improved by these angiogenic growth factors. Rectal enemas containing bFGF or PDGF indeed accelerated the healing of chemically induced ulcerative colitis in rats. VEGF, also known as VPF (vascular permeability factor), however, had no effect or slightly aggravated the colonic lesions. Injection of anti-VEGF neutralizing antibodies, however, counteracted the increased vascular permeability in the early stages of experimental ulcerative colitis and subsequently decreased the number of inflammatory cells in colonic ulcers in rats, resulting in significantly improved healing in the lower GI tract lesions. Thus, the three angiogenic growth factors tested exerted beneficial effect on gastroduodenal ulcers, and rectal enemas with bFGF or PDGF also accelerated the healing of experimental ulcerative colitis. Surprisingly, we achieved the latter effect with anti-VEGF antibodies, most likely because of the pro-inflammatory actions of VEGF in the pathogenesis of ulcerative colitis.     

Effects of electrical stimulation of the hunger center in the lateral hypothalamus and food reinforcement on impulse activity of the mylohyoid muscle in rabbits under conditions of hunger and satiety

Effects of electrical stimulation of the hunger center in the lateral hypothalamus and food reinforcement on impulse activity of mylohyoid muscle were studied in chronic experiments under conditions of hunger and satiety. Threshold stimulation of the lateral hypothalamus in starving and satiated rabbits in the absence of food induced searching behavior associated with burst-like impulse activity with a bimodal distribution of interpulse intervals. Regular spike burst in the mylohyoid muscle during stimulation of the lateral hypothalamus in the absence of food serves as an example of the anticipatory type reaction. Increased food motivation during threshold stimulation of the lateral hypothalamus in starving and satiated rabbits with food offered led to successful food-procuring behavior, during which the frequency of spike bursts in the mylohyoid muscle became comparable with that under conditions of natural foraging behavior stimulated by the need in nutrients. Our results suggest that temporal structure of mylohyoid muscle impulse activity refl ects convergent interactions of food-motivation excitation with reinforcement excitation on neurons of the masticatory and deglutitive centers.