ABSTRACT
Metastatic chondrosarcoma is a bone malignancy not responsive to conventional therapies; new approaches and therapies are urgently needed. We have previously reported that mTORC1 inhibitor, antitumorigenic cytostatic proline rich polypeptide 1 (PRP-1), galarmin caused a significant upregulation of tumor suppressors including TET1/2 and SOCS3 (known to be involved in inflammatory processes), downregulation of oncoproteins and embryonic stem cell marker miR-302C and its targets Nanog, c-Myc and Bmi-1 in human chondrosarcoma. To understand better the mechanism of PRP-1 action it was very important to identify the receptor it binds to. Nuclear pathway receptor and GPCR assays indicated that PRP-1 receptors are not G protein coupled, neither do they belong to family of nuclear or orphan receptors. In the present study, we have demonstrated that PRP-1 binding interacting partners belong to innate immunity pattern recognition toll like receptors TLR1/2 and TLR6 and gel forming secreted mucin MUC5B. MUC5B was identified as PRP-1 receptor in human chondrosarcoma JJ012 cell line using Ligand-receptor capture technology. Toll like receptors TLR1/2 and TLR6 were identified as binding interaction partners with PRP-1 by western blot analysis in human chondrosarcoma JJ012 cell line lysates. Immunocytochemistry experiments confirmed the finding and indicated the localization of PRP-1 receptors in the tumor nucleus predominantly. TLR1/2, TLR6 and MUC5B were downregulated in human chondrosarcoma and upregulated in dose-response manner upon PRP-1 treatment. Experimental data indicated that in this cellular context the mentioned receptors had tumor suppressive function.
Subject(s)
Bone Neoplasms/metabolism , Chondrosarcoma/metabolism , Mucin-5B/metabolism , Peptides/metabolism , Toll-Like Receptors/metabolism , Antimicrobial Cationic Peptides , Bone Neoplasms/genetics , Bone Neoplasms/pathology , Cell Line, Tumor , Chondrosarcoma/genetics , Chondrosarcoma/pathology , Humans , Immunohistochemistry , Peptides/pharmacology , Protein Binding , Toll-Like Receptors/biosynthesis , Toll-Like Receptors/genetics , Up-Regulation/drug effectsABSTRACT
We investigated around 100 patients for one-year and then registered the information about the possible etiology of the stroke, type of disease, treatment, localization of the clot, age, as well as gender distribution of stroke patients' population. This general overview of risk factors, ways of treatment, diagnostics, care and monitoring of the stroke patients in Armenia might serve as a cornerstone work for further highlighting of new avenues for stroke treatment.
Subject(s)
Cost of Illness , Stroke/epidemiology , Stroke/therapy , Aged , Aged, 80 and over , Armenia/epidemiology , Atherosclerosis/complications , Atherosclerosis/epidemiology , Atherosclerosis/therapy , Female , Humans , Hypertension/complications , Hypertension/epidemiology , Hypertension/therapy , Male , Middle Aged , Risk Factors , Stroke/etiologyABSTRACT
Mammalian target of rapamycin (mTOR) serine threonine kinase is the enzyme that regulates cancer cell growth by altering nutrient supplies to cancer cells. The neuropeptide (proline-rich peptide 1 (PRP-1)), galarmin, produced by the brain neurosecretory cells is a mTOR kinase inhibitor with powerful 80% antiproliferative cytostatic effect in a high-grade chondosarcoma and other mesenchymal tumors. However, the negative feedback loop of phosphatidylinositol 3 kinase-Protein kinase B (PKB), PI3K-AKT and PI3K-rat sarcoma (RAS)-mitogen-activated protein kinase (MAPK) activation is well documented for mTOR inhibitors. This study explored the involvement of those loops in drug resistance after the treatment with mTOR complex 1 (mTORC1) inhibitor, PRP-1. Multidrug resistance assay (MDR) demonstrated that this cytokine did not inhibit permeability glycoprotein-mediated MDR in chondrosarcoma. Phospho-MAPK array in human chondrosarcoma cell line treated with galarmin (10 µg/ml,) showed a strong upregulation of phosphorylated glycogen synthase kinase 3ß (GSK3ß) via activation of PI3K-AKT and MAPK feedback loops. Such GSK3ß inactivation leads to ß-catenin accumulation that entails drug resistance. The ability of cells to metastasize is reflected in their capacity to adhere to extracellular matrix and endothelium. Laminin cell adhesion assay demonstrated that PRP-1 in the same concentrations that inhibit mTOR kinase inhibited JJ012 chondrosarcoma cell adhesion. The neuropeptide did not have any effect on the expression of total focal adhesion kinase and its phosphorylated form. Thus, it was not accompanied by total HAT downregulation and total HDAC upregulation. Combinatorial treatments of PRP-1 with MAPK and PI3K/AKT inhibitors most probably will lead to full cytotoxicity overcoming drug resistance.
Subject(s)
Extracellular Matrix/metabolism , Mitogen-Activated Protein Kinases/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Proteins/antagonists & inhibitors , Proto-Oncogene Proteins c-akt/metabolism , Signal Transduction/drug effects , ras Proteins/metabolism , Antimicrobial Cationic Peptides , Cell Adhesion/drug effects , Cell Line, Tumor , Drug Resistance, Neoplasm/drug effects , Histone Acetyltransferases/metabolism , Histone Deacetylases/metabolism , Humans , Mechanistic Target of Rapamycin Complex 1 , Multiprotein Complexes , Peptides/pharmacology , TOR Serine-Threonine KinasesABSTRACT
Crush-syndrom (CS) was characterized by Bywaters E.G.L. in 1941 after London blitz. The soft tissues is followed by acute hemodynamic shock, myoglobinuria, acute renal insufficiency, and lethal endotoxicity. Data of CS pathogenesis study has shown that the largest changes in Crush occur during decompression and are accompanied by acute alteration of brain protein synthesis and strong morphological changes of brain structures. The period of decompression might be characterized by the proteolytic breakdown of the myoglobine and formation of toxic peptides. In our current work we have identified four newly formed peptides in the brain of the animals subjected to the experimental muscle tissue injury. Our investigations related with the CS experimental model have demonstrated that during the 2-hours compression protein synthesis was decreased in cytosol (32,7%) and mitochondria (49%), after 5-h compression there were registered non-significant changes in the level of protein synthesis. Intraperitoneal administration of Proline-rich peptide, ((PRP), 1 mcg/100g weight of rats), originating from proteolysis of C-terminal glycoprotein a neurophysin II along with vasopressin and oxytocin and transferring from the hypothalamus to the neurohypophysis by axonal transport, initiates activation of the protein synthesis in all studied cellular subcomponents of brain cells. The positive effect of the peptide is conditioned, most probably, by activation of the immune system and adaptation mechanisms, including mobilization of endogen-protective mechanisms of the organism.
Subject(s)
Crush Syndrome/metabolism , Cytokines/pharmacology , Hypothalamus/metabolism , Peptides/pharmacology , Proteins/metabolism , Animals , Crush Syndrome/pathology , Crush Syndrome/physiopathology , Cytokines/chemistry , Cytokines/genetics , Humans , Male , Peptides/chemistry , Peptides/genetics , Proteins/chemistry , Proteins/genetics , Rats , Rats, WistarABSTRACT
It is recognized that the main trigger of Alzheimer disease related neurodegeneration is ß-amyloid peptide, which subsequently generates different metabolic disorders in neuron and finally leads to neuronal death. Several biologically active products were tested as neuroprotectors, but only few of them demonstrated any efficiency. Proline-rich polypeptide-1 was tested as a neuroprotective agent on Aß25-35 animal model of Alzheimer disease. Biochemical analysis (determination of spectrum of neuroactive amino acids, such as glutamate, gamma-aminobutyric acid, glycine, aspartate and taurine), as well as behavioral, electrophysiological and morphological studies were performed to reveal the neuroprotective potential of proline-rich polypeptide in rats. Based on the results of our study it can be concluded that proline-rich polypeptide-1 has a potential to be one of the effective preventive or therapeutic agents against neurodegenerative disorders, such as Alzheimer disease.
Subject(s)
Amyloid beta-Peptides/antagonists & inhibitors , Amyloid beta-Peptides/toxicity , Neurodegenerative Diseases/chemically induced , Neurodegenerative Diseases/prevention & control , Neuroprotective Agents , Peptide Fragments/antagonists & inhibitors , Peptide Fragments/toxicity , Peptides/pharmacology , Amino Acids/metabolism , Animals , Antimicrobial Cationic Peptides , Anxiety/psychology , Behavior, Animal/drug effects , Brain/pathology , Cerebral Cortex/metabolism , Cerebral Cortex/pathology , Chromatography, High Pressure Liquid , Corpus Striatum/metabolism , Corpus Striatum/pathology , Electroencephalography , Electrophysiological Phenomena , Hippocampus/metabolism , Hippocampus/pathology , Injections, Intraventricular , Male , Neurodegenerative Diseases/psychology , Rats , Rats, Sprague-DawleyABSTRACT
Activation of the PI3K-Akt-mTOR pathway is implicated both in the establishment of tumors and as well as a target for therapy in many types of solid malignancy, its blockade represents an opportunity to improve outcomes in patients with tumors that are associated with poor prognosis. Our experimental data indicates that proline-rich polypeptide-1 (PRP-1, galarmin) is immunomodulator cytokine, produced by hypothalamic neurosecretory cells and exerts its antiproliferative effect on the tumor cells of mesenchymal origin via inhibiting mTOR kinase activity and repressing cell cycle progression. The goal of these investigations was to elucidate the antiproliferative action of PRP-1 on the breast carcinoma cell line MDA 231 (ER-) and to compare PRP-1 action previously reported on other mesenchymal tumors. These experiments confirmed maximum inhibition of cell growth at 0.5 and 1 µg/ml PRP-1 (71% and 63%, respectively) and inhibition at 10 µg/ml of 44%. There was no inhibitory effect observed on luminal T47-D (ER+) cells. Videomicroscopy results demonstrated dividing cells in the cytokine-treated MDA 231 (ER-), suggesting that the cells were not in the state of dormancy. The flow cytometry experiments confirmed that PRP-1-treated cells were accumulated in S phase. No apoptosis, caspase activation, or senescence was detected after treatment with this cytokine. Experiments with mTOR with PRP-1 (10 µg/ml) indicated statistically significant 40% inhibition of mTOR kinase activity in immunoprecipitates of the MDA 231 (ER-) cell line. PRP-1 is a novel mTOR inhibitor with strong antiproliferative action in mesenchymal tumors mostly resistant to radiation and chemotherapy.
Subject(s)
Antineoplastic Agents/pharmacology , Breast Neoplasms/pathology , Peptides/pharmacology , Antimicrobial Cationic Peptides , Breast Neoplasms/genetics , Breast Neoplasms/metabolism , Cell Line, Tumor , Cell Proliferation/drug effects , Cytostatic Agents/pharmacology , Female , Flow Cytometry , Humans , Immunoprecipitation , Receptors, Estrogen/genetics , TOR Serine-Threonine Kinases/antagonists & inhibitorsABSTRACT
This study aimed to further elucidate the molecular mechanisms of antiproliferative action of proline rich polypeptide 1 (PRP-1) cytokine, produced by neurosecretory cells of the hypothalamus to be considered as alternative adjuvant therapy for metastatic chondrosarcoma, which does not respond to chemotherapy or radiation and currently without any effective treatment. Rapid cell proliferation assay of human primary cultures from high grade chondrosarcoma patients biopsies and human chondrosarcoma JJ012 cell line indicated 50 and 80% inhibition in PRP-1 treated samples correspondingly. Videomicroscopy detected that despite the treatment there are still dividing cells, meaning that cells are not in the state of dormancy, rather PRP-1 repressed the cell cycle progression, exhibited cytostatic effect. The mammalian target of rapamycin (mTOR) is an intracellular serine/threonine protein kinase that has a crucial role in a nutrient sensitive signaling pathway that regulates cell growth. Experiments with mTOR pathway after PRP-1 (10 µg/ml) treatment indicated statistically significant 30% inhibition of mTOR activity and its 56% inhibition in immunoprecipitates with PRP-1 concentrations effective for cell proliferation inhibition. Treatment with PRP- caused inhibition of mTOR and downstream target cMyc oncogenic transcription factor sufficient to trigger the cytostatic effect in high grade, but not in low grade chondrosarcomas. The fact that lower concentrations than 10 µg/ml peptide with cytostatic effect did not inhibit mTOR, but inhibited cMyc prompted us to assume that PRP-1 binds to two different receptors facilitating the antiproliferative effect.
Subject(s)
Chondrosarcoma/pathology , Cytokines/pharmacology , Hypothalamus/drug effects , Peptides/pharmacology , Proto-Oncogene Proteins c-myc/antagonists & inhibitors , TOR Serine-Threonine Kinases/physiology , Antimicrobial Cationic Peptides , Cell Line, Tumor , Enzyme-Linked Immunosorbent Assay , Humans , Hypothalamus/metabolism , Proto-Oncogene Proteins c-myc/physiologyABSTRACT
A proline-rich cytokine from neurosecretory granules of bovine neurohypophysis, 15 amino acids containing PRP-1 (Ala-Gly-Ala-Pro-Glu-Pro-Ala-Glu-Pro-Ala-Gln-Pro-Gly-Val-Tyr), had been demonstrated as a unique regulator of activity of neurons, strong antibacterial agent, and mediator of the hypothalamus-neurohypophysis-bone marrow-thymus axis, which participates in hematopoietic stem cells differentiation. In the present work it was shown that this neuropeptide represents a new natural substrate for Dipeptidyl Peptidase IV (DPPIV). The time-dependent increase of primary amines quantity in the assay mixture of PRP-1 and DPPIV has been observed allowing to conclude, that DPPIV catalyses the enzymatic reaction of PRP-1 cleavage. The amount of primary amines in the assay mixtures was evaluated using o-phtaldialdehyde dye. The gel-filtration and paper electrophoresis analyses proved this conclusion. The catalytic parameters of catalyzed by DPPIV enzymatic reaction of PRP-1 cleavage were determined as: V(max) = 1.27 ± 0.11 nmol/min and K(m) = 0.38 ± 0.1 mM.
Subject(s)
Cytokines/genetics , Cytokines/metabolism , Dipeptidyl Peptidase 4/metabolism , Proline/metabolism , Secretory Vesicles/chemistry , Amino Acid Sequence , Animals , Cattle , Chromatography, Gel , Dipeptidyl Peptidase 4/genetics , Molecular Sequence Data , Peptide Fragments/analysisABSTRACT
Discovery of neurosecretion of cardioactive neurohormones produced by hypothalamic nuclei (NSO and NPV), as well as the biosynthesis of several immunomodulators (signal molecules of the neuroendocrine immune system of brain), deciphering of their chemical structure and study of their biological properties led to the foundation of two important trends of neurobiology: neuroendocrine immunology and cardiology. Hormone formation by atrium ganglionary nerve cells and auriculum establishment of neurohumoral interactions between hypothalamic and atrium neurosecretion indicated the existence of the system neuroendocrine hypothalamus--endocrine heart. Study of their biological properties promoted creation of powerful neurohormonal preparations for the treatment of immune, cardio-vascular, neurodegenerative, infectious and tumor diseases. Concepts suggested by us on neuroendocrine cardiology and immunology, create large perspectives for development of the theory and its implementation in medicine.
Subject(s)
Allergy and Immunology , Heart/physiology , Neurosecretory Systems/metabolism , Signal Transduction , Animals , HumansABSTRACT
We tested the action of proline-rich peptide (PRP-1) and cobra venom Naja Naja Oxiana (NOX) on Deiters' nucleus neurons at 3rd, 15th and 35th days after unilateral labyrinthectomy (UL). Early and late tetanic, post-tetanic potentiation and depression of Deiters'neurons to bilateral high frequency stimulation of hypothalamic supraoptic and paraventricualar nuclei was studied. The analysis of spike activity was carried out by mean of on-line selection and special program. The complex averaged peri-event time and frequency histograms shows the increase of inhibitory and excitatory reactions of Deiters' neurons at early stage of vestibular compensation following PRP-1 and NOX injection, reaching the norm at the end of tests. In histochemical study the changes in Ca(2+)-dependent acidic phosphatase (AP) activity in neurons was discovered. It was shown that in UL animals the total disappearance or delay of decolorizing of Deiters' neurons lead to neurodegenerative pattern as cellular "shade". AP activity after UL and PRP-1 injection exerts more effective recovery of neurons in comparison with events, observed after the administration of NOX. The data of this study indicate that PRP-1 and NOX are protectors, which may successfully recover the disturbed vestibular functions.
Subject(s)
Elapid Venoms/pharmacology , Neurons/physiology , Peptides/pharmacology , Vestibular Nucleus, Lateral/physiology , Animals , Antimicrobial Cationic Peptides , Elapidae , Neurons/drug effects , Neurons/ultrastructure , Vestibular Nucleus, Lateral/drug effectsABSTRACT
In the present study we carried out experiments in vitro and in vivo and investigated the effect of proline-rich polypeptide (PRP) on the proliferation and effectiveness of colony formation of MMSCs in vitro. Various routes and doses of PRP administration to rats increased the number of MMSCs in bone marrow and spleen. Our research revealed opposite effects of PRP on the proliferation of bone marrow stromal cells obtained from normal humans and stromal cells isolated from a human giant-cell tumour.
Subject(s)
Bone Marrow Cells/drug effects , Giant Cell Tumors/pathology , Peptides/pharmacology , Animals , Bone Marrow Cells/cytology , Cell Proliferation , Cells, Cultured , Guinea Pigs , Humans , Male , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/drug effects , Multipotent Stem Cells/cytology , Multipotent Stem Cells/drug effects , Pituitary Gland/metabolism , Rats , Rats, Wistar , Stromal Cells/cytology , Stromal Cells/drug effects , Time FactorsABSTRACT
The AGAPEPAEPAQPGVY proline-rich polypeptide (PRP-1) was isolated from neurosecretory granules of the bovine neurohypophysis; it is produced by N. supraopticus and N. paraventricularis. It has been shown that PRP-1 has many potentially beneficial biological effects including immunoregulatory, hematopoietic, antimicrobial and anti-neurodegenerative properties. Here we demonstrated that PRP-1 administration influence on redistribution of monocytes, granulocytes and lymphocytes between bone marrow (BM) and peripheral blood and promotes the influx of granulocytes and monocytes/macrophages from BM into peripheral blood and accumulation of immature granulocyte and monocyte in BM and delayed the maturation of T cells in BM. PRP-1 increased colony-forming cell proliferation in rat cells in vivo. In PRP-treated rat BM, the CFU number at day 4, 7 and 14 was considerably increased in comparison with untreated rats BM and no difference was found at day 21 and day 28. We found that PRP-1 enhances erythroid and myeloid colonies formation in human CD34(+) progenitor cell culture in the presence of different growth factors and down-regulates T cells colony formation and specific surface markers expression during induction of human CD34(+) progenitor cells differentiation into T lymphocytes lineage. We suggested that the hypothalamic PRP-1 possibly represents an endogenous peptide whose primary functions are to regulate neuronal survival and differentiation and hematopoiesis within neurosecretory hypothalamus-bone marrow humoral axis.
Subject(s)
Hematopoiesis/drug effects , Peptides/pharmacology , Animals , Antigens, CD34/metabolism , Antimicrobial Cationic Peptides , Bone Marrow Cells/cytology , Bone Marrow Cells/drug effects , Cattle , Cell Differentiation , Cells, Cultured , Colony-Forming Units Assay , Dendritic Cells/cytology , Dendritic Cells/drug effects , Erythrocytes/cytology , Erythrocytes/drug effects , Humans , Lymphocytes/cytology , Lymphocytes/drug effects , Peptides/physiology , Pituitary Gland, Posterior/metabolism , Rats , Rats, Wistar , Stem Cells/cytology , Stem Cells/drug effects , Stem Cells/metabolismABSTRACT
Proline rich polypeptide (PRP-1) produced by neurosecretory cells of the hypothalamus is one of the fragments of neurophysin-vasopressin-associated glycoprotein. The primary structure of the neuropeptide PRP-1 isolated from neurosecretory granules of bovine neurohypophysis. We investigated PRP-1 action on chondrosarcoma, the second most common malignancy in bone, which primarily affects the cartilage cells. This deadly disease does not have any effective treatment. Earlier we demonstrated MYC oncogene inactivating effect by 1 lg/ml concentration brain PRP-1 In the present study we observed reduced viable sarcoma JJ012 cell numbers in comparison with control (89% growth inhibition) when treated with low concentrations of PRP-1 (0.51 lg/ml). Higher concentrations did not exhibit inhibitory effect. We assume that PRP-1 in low concentration impedes cell cycle progression. The fact that low concentrations of PRP-1 abolished Myc activity prompts to think that the antitumorigenic effect of PRP-1 in low concentrations is mediated through oncogene inactivation.
Subject(s)
Bone Neoplasms/drug therapy , Chondrosarcoma/drug therapy , Peptides/therapeutic use , Antimicrobial Cationic Peptides , Cell Line, Tumor , Genes, myc/drug effects , HumansABSTRACT
Proline rich polypeptide (PRP-1) produced by NPV and NSO cells is released into the general circulation and exerts its effect on the activity of immunocompetent and neuronal cells. PRP-1 is a unique regulator of hematopoiesis, stimulator of bone-marrow hematogenesis. Taking into consideration our preliminary data on antitumor and unique diverse biological properties of PRP-1 previously described by Galoyan et al., we proceeded with investigation of the PRP-1 effect on chondrosarcoma, the second most common malignancy in bone, which tends to be locally invasive and then metastatic. Currently it does not have any effective treatment and does not respond either to radiation or chemotherapy, leaving surgical resection as the only option. Our experimental results of PRP-1 action on human chondrosarcoma JJ012 cells demonstrated inactivation, abolishment of Myc oncogene activity usually upregulated in chondrosarcoma cells and other malignancies. The fact that addition of PRP-1 caused drastic inactivation of Myc-luc response element to the control level in human chondrosarcoma JJ012 cell line prompts to investigate further this neuropeptides powerful antioncogenic potential, opening up possibilities to consider PRP-1 as a potential therapeutic tool for chondrosarcoma treatment.
Subject(s)
Chondrosarcoma/metabolism , Gene Expression Regulation, Neoplastic/physiology , Genes, myc , Proline-Rich Protein Domains/physiology , Cell Line, Tumor , Chondrosarcoma/genetics , Chondrosarcoma/pathology , Enhancer Elements, Genetic , Genes, Reporter , Humans , Luciferases/geneticsABSTRACT
The effects of atrial polypeptide and neurohormone C upon the interaction of human factor Xa (FXa) and human antithrombin III (ATIII) were followed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. A pattern of bands consisting of a 1 degree duplex complex (FXaalpha-ATIII band of 109 kDa, FXabeta-ATIII band of 104 kDa), a 2 degree duplex complex (alpha band of 99 kDa, beta band of 95 kDa), a 3 degree duplex complex (alpha band of 66 kDa, beta band of 62 kDa), modified ATIII (ATIIIM, 58 kDa), native ATIII (55 kDa), FXaalpha (52 kDa), FXabeta (47 kDa), and a FXa degradation product (FXagamma, 35 kDa) was detected and quantitated. Preincubation of FXa, ATIII, or mixtures thereof with atrial polypeptide produced a shift from FXaalpha-ATIII to FXabeta-ATIII complexes and increases in both ATIIIM and FXagamma, reflecting degradation of the 1 degree and 2 degree complex to form the 3 degree complex. Atrial polypeptide appeared to promote FXa-ATIII complex formation when preincubated with ATIII, or when added within 1 min to FXa/ATIII mixtures. However, when atrial polypeptide was preincubated with FXa, inhibition of the 1 degree complex formation was suggested. Upon incubation of FXa, ATIII, or mixtures thereof with neurohormone C, there was an increase in total complex formation, a decrease in ATIIIM, a decrease in FXagamma, and little change in the ratio of free FXaalpha to FXabeta, or the ratio of FXaalpha-ATIII to FXabeta-ATIII complexes. Therefore, neurohormone C may act to suppress hydrolysis or proteolytic actions of excess FXa on FXa-ATIII complexes, or autolytic activity of FXa, to the level of FXagamma via an, as yet, unknown mechanism. Additionally, neurohormone C retards the hydrolysis of the FXa-ATIII complexes which form free FXa and ATIIIM. Hence, the role of atrial polypeptide in mixtures with ATIII and in mixtures with FXa is quite contrasting, and may reflect mechanistic effects of the atrial polypeptide molecule, as well as tissue-specific reactions.
Subject(s)
Antithrombin III/metabolism , Blood Coagulation/drug effects , Factor Xa/metabolism , Hypothalamic Hormones/pharmacology , Peptides/pharmacology , Animals , Blood Coagulation/physiology , Cattle , Electrophoresis, Polyacrylamide Gel , Heart Atria/chemistry , HumansABSTRACT
BACKGROUND: This work sought to determine the effects of hypothalamic proline-rich peptide (PRP)-1 in a rat model of Alzheimer's disease. METHODS: Complex histochemical, electrophysiologic, and behavioral analyses were performed on intact or diseased Wistar rats (n = 28). Pathologic conditions were induced by bilateral intracerebroventricular injection of amyloid peptide Abeta25-35. The diseased rats received systemic administration of PRP-1 or placebo control. RESULTS: Abeta25-35 caused cellular neurodegeneration with marked glial reaction in the hippocampal complex and almost full destruction of the dentate fascia, which was not observed in conditions of PRP-1 administration after Abeta25-35 injection. Hippocampal neurons of intact animals responded to high-frequency (tetanic) stimulation of entorhinal cortex of ipsilateral cerebral hemisphere by tetanic and posttetanic potentiation of a different intensity and duration, which was accompanied by posttetanic depression. Abeta25-35 led to significant changes in the level and pattern of hippocampal neuronal activity, indicating the absence of both tetanic and posttetanic activity. Poststimulus activity manifestations rarely occurred and rapidly decreased after repeated trials. This indicated the focal character of lesion. Regular administration of PRP-1 for 4 weeks resulted in optimal restoration of electrophysiologic parameters. PRP-1 maintained the initial learning level achieved in a behavioral study in a Morris water maze. CONCLUSIONS: Systemic administration of PRP-1 possesses neuroprotective effects and can prevent the neurodegeneration in hippocampus induced by Abeta25-35. This suggests that PRP-1 could be a potential therapeutic agent for specific neurodegenerative diseases.
Subject(s)
Alzheimer Disease/drug therapy , Amyloid beta-Peptides/toxicity , Hippocampus/drug effects , Neuroprotective Agents/therapeutic use , Proline-Rich Protein Domains , Alzheimer Disease/pathology , Amyloid beta-Peptides/administration & dosage , Animals , Disease Models, Animal , Electrophysiology , Female , Hippocampus/pathology , Hypothalamus/metabolism , Injections, Intraventricular , Long-Term Potentiation/drug effects , Long-Term Potentiation/physiology , Long-Term Synaptic Depression/drug effects , Long-Term Synaptic Depression/physiology , Male , Maze Learning/drug effects , Neurons/drug effects , Neurons/pathology , Rats , Rats, WistarABSTRACT
The AGAPEPAEPAQPGVY proline-rich peptide (PRP-1) was isolated from neurosecretory granules of the bovine neurohypophysis; it is produced by N. supraopticus and N. paraventricularis. It has been shown that PRP-1 has many potentially beneficial biological effects including immunoregulatory, hematopoietic, antimicrobial and anti-neurodegenerative properties. Here we investigated the influence of PRP-1 on staurosporine-induced apoptosis of postnatal hippocampal cells and on doxorubicin-induced bone marrow granulocyte- and monocyte apoptosis. The intention was to further characterize the effect of PRP-1 on the survival rate of neurons and in context with myelopoiesis. We demonstrate that PRP-1 significantly reduced apoptosis of postnatal hippocampal cells induced by staurosporine. The protective effect of PRP-1 against apoptotic cell death was shown to be both time- and dose-dependent. Neuroprotection was more pronounced after prolonged pretreatment of the cells with PRP-1 before the induction of apoptosis with staurosporine. The related peptide [arg(8)]vasopressin did not reveal neuroprotection. PRP-1 also significantly reduced apoptosis of bone marrow monocytes and granulocytes induced by doxorubicin. This protective effect lasted for 2-4 h and was not detectable anymore after 24 h when PRP-1 and doxorubicin were added simultaneously. Previously obtained data and results of the current studies suggested that the hypothalamic PRP-1 possibly represents an endogenous peptide whose primary functions are to regulate myelopoiesis and neuron survival as we provide evidence that PRP can differentially reduce both staurosporine- and doxorubicin-induced hippocampal and bone marrow cell apoptosis.
Subject(s)
Apoptosis/drug effects , Bone Marrow Cells/drug effects , Neurons/cytology , Peptides/pharmacology , Animals , Doxorubicin/pharmacology , Hippocampus/cytology , Hippocampus/drug effects , Neurons/drug effects , Proline-Rich Protein Domains , Rats , Staurosporine/antagonists & inhibitors , Staurosporine/pharmacologyABSTRACT
Proline-rich-polypeptides (PRPs) isolated from bovine hypothalamus have been shown to render protection against neuronal injury of the brain and spinal cord. We examined two PRPs containing 15 and 10 amino acid residues (PRP-1 and PRP-4 synthetic polypeptide) for their effect, if any, on dopaminergic neuronal damage caused by the parkinsonian neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Effects of these PRPs on hydroxyl radical ((*)OH) generation in a Fenton-like reaction as well as from isolated mitochondria were monitored, employing a sensitive salicylate hydroxylation procedure. Balb/c mice treated (i.p., twice, 16 h apart) with MPTP (30 mg/kg) or PRP-1 (1.6 mg/kg), but not PRP-4 (1.6 mg/kg) showed significant loss of striatal dopamine and norepinephrine as assayed by an HPLC-electrochemical procedure. Pretreatment with the PRPs, 30 min prior to the neurotoxin administration failed to attenuate MPTP-induced striatal dopamine or norepinephrine depletion, but significantly attenuated the MPTP-induced decrease in dopamine turnover. A significant increase in the generation of (*)OH by the PRPs in a Fenton-like reaction or from isolated mitochondria suggests their pro-oxidant action, and explains their failure to protect against MPTP-induced parkinsonism in mice.
Subject(s)
1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine/pharmacology , Dopamine/metabolism , Hydroxyl Radical/metabolism , MPTP Poisoning , Neurons/drug effects , Neurotoxins/pharmacology , Peptides/metabolism , Animals , Biogenic Amines/metabolism , Cattle , Cell-Free System , Corpus Striatum/cytology , Corpus Striatum/drug effects , Corpus Striatum/metabolism , Disease Models, Animal , Dopamine Agents/pharmacology , Dopamine Agents/toxicity , Mice , Mice, Inbred BALB C , Mitochondria/metabolism , Mitochondria/ultrastructure , Neurons/chemistry , Neurons/cytology , Neurons/metabolism , Neurotoxins/toxicity , Parkinson Disease/metabolism , Peptides/genetics , Proline-Rich Protein DomainsABSTRACT
OBJECTIVE: The effects of proline-rich polypeptide (PRP) isolated from neurosecretory granules of bovine neurohypophysis produced by nuclei supraopticus and paraventricularis on phagocytosis, bacterial intracellular killing and oxidative burst induction in normal human cells and inflammatory cells from patients with Behçet's disease (BD), i.e. peripheral blood neutrophils and monocytes, were investigated. METHODS: Intracellular killing of Staphylococcus aureus by neutrophils and monocytes of normal controls and BD patients, phagocytic activity as well as spontaneous and N-formyl-Met-Leu-Phe (fMLP)- or phorbol 12-myristate 13-acetate (PMA)-induced activation of their respiratory burst were determined by quantitative flow cytometry using highly specific fluorescence probes. RESULTS: PRP does not affect human peripheral blood neutrophil and monocyte phagocytosis but dramatically enhances spontaneous or fMLP- and PMA-induced oxidative burst as well as the intracellular killing of S. aureus. PRP induced the upregulation of the spontaneous or fMLP- and PMA-induced oxidative burst in normal PMNs and monocytes; the number of inflammatory BD cells did neither increase further nor undergo spontaneous or PMA-stimulated oxidative burst. In BD patients, increased spontaneous production of reactive oxygen intermediates (ROIs) by neutrophils and monocytes is characterized by impaired intracellular protein-kinase-C (PKC)-dependent oxidative burst regulation as well as over-regulation of chemotaxis/inflammation-mediated respiratory burst induction. PRP restores rather the impaired intracellular PKC-dependent regulation of ROI production in inflammatory diseased cells than the chemotaxis/induction of the inflammation-mediated respiratory burst. CONCLUSION: We demonstrated the regulatory role for PRP on oxidative burst in neutrophils and monocytes from normal controls and BD patients. Our results suggest that PRP differentially affects both chemotaxis- and PKC-dependent oxidative burst in normal and inflammatory cells from patients.