Proline-rich Polypeptide Research Articles

Proline-rich polypeptide complex (PRP) regulates secretion of inflammatory mediators by its effect on NF-κB activity

A proline-rich polypeptide complex (PRP) with immunoregulatory and procognitive activities shows beneficial effects in the Alzheimer's disease (AD). The mechanism of action of PRP is not yet fully clarified, we have shown that the PRP complex inhibits overproduction of reactive oxygen species, nitric oxide and proinflammatory cytokines induced by lipopolysaccharide (LPS). LPS stimulation exerts its inflammatory effects through the activation of the classical nuclear factor-κB (NF-κB) pathway. The results presented in this study showed the ability of PRP to inhibit the NF-κB activity induced by LPS while it increased activity of NF-κB in untreated cells. Examining the effect of PRP on IκB it was shown that relative level of IκB was lowered in the presence of PRP. It seems that in cells untreated with LPS, PRP can activate proteasome system and stimulate IκB degradation. Our results suggest that the regulatory effect of PRP on inflammatory processes may be associated with the influence of PRP on NF-κB translocation. Inhibitory effect of PRP on NF-κB activity might, at least in part, contribute to the beneficial therapeutic effects in the case of Alzheimer's disease.

Molecular and Biotechnological Advances in Milk Proteins in Relation to Human Health

Milk and colostrum is a rich source of proteins/peptides which have crucial roles in both neonates and adults. Milk bioactive proteins and peptides are potential health-enhancing nutraceuticals for food. Many bioactive peptides/proteins may be used as nutraceuticals, for example, in the treatment of cancer, asthma, diarrhea, hypertension, thrombosis, dental diseases, as well as mineral malabsorption, and immunodeficiency. The following components of milk are of particular interest in the recent years: 1) Lactoferrin [Lf] has antibacterial, antifungal, antiviral, antiparasite and antitumor activities and accelerates immunomodulatory properties. Lf is a potent inhibitor for several enveloped and naked viruses, such as rotavirus, enterovirus and adenovirus. Lf is resistant to tryptic digestion and breast-fed infants excrete high levels of faecal Lf, so that its effect on viruses replicating in the gastrointestinal tract is of great interest. 2) Casein has been protective in experimental bacteremia by eliciting myelopoiesis. Casein hydrolyzates were also protective in diabetic animals, reduced the tumor growth and diminished colicky symptoms in infants. 3) A Proline rich polypeptide [PRP] revealed variety of immunotropic functions, including promotion of T-cell activation and inhibition of autoimmune disorders such as multiple sclerosis. 4) alpha-Lactalbumin [LA] demonstrates antiviral, antitumor and anti-stress properties. 5) Lactoperoxidase shows antibacterial properties. 6) Lysozyme is effective in treatment of periodentitis and prevention of tooth decay. Taken together, milk-derived proteins and peptides are bio-available and safe for the prevention and treatment of various disorders in humans and may play a complementary [natural agents] rather than a substitutional role to the toxic synthetic pharmacological drugs.

The protective effects of the nutraceutical, colostrinin, against Alzheimer's disease, is mediated via prevention of apoptosis in human neurones induced by aggregated beta-amyloid.

It has previously been demonstrated that oral administration of ovine Colostrinin (CLN), a proline-rich polypeptide isolated from ovine colostrum, can effectively treat Alzheimer's disease patients. This study aims to determine whether CLN has effects on the aggregation and toxicity of synthetic beta-amyloid (Abeta), implicated as a causative agent of AD. Using cell assays, we examined if pre-treatment of neuronal cells with CLN confers protection. The data from cytotoxicity assays (using MTT and LDH) demonstrated that pre-treatment of human neuronal SHSY-5Y cells with 5 microg/ml CLN, for 24 hours, confers neuroprotection against Abeta-induced neurotoxicity. Twenty-four hour pre-treatment with 5 microg/ml CLN was also shown to reduce Abeta 1-40-induced apoptosis in human neuronal cells as determined via qualitative and quantitative apoptosis assays. The neuroprotection conferred with CLN pre-treatment was reduced with the Fas ligand (FasL) binding antibody Nok1, suggesting that the effects of CLN may involve a Fas:soluble FasL interaction. These findings indicate that CLN could possibly play a role in the prevention of AD pathogenesis, though the inhibition of Fas-mediated apoptosis.

Study of the genotoxic effects of a proline-rich polypeptide using the comet assay

The genotoxicity of a proline-rich polypeptide was investigated in vitro in the human myeloid leukemia KCL-22 cell line. A comet assay, based on single-cell gel electrophoresis, was used for the analysis of the DNA damage and reparation rate. After electrophoresis, images resembling comets were formed; the intensity of the “tail” relative to the “head” reflected the number of DNA breaks. We demonstrated that the level of DNA damage depended on the polypeptide concentration and incubation time. Estimation of DNA damage levels by the comet assay is recommended for the evaluation of the genotoxicity of new pharmaceuticals.

Antitumorigenic Effect of Brain Proline Rich Polypeptide-1 in Human Chondrosarcoma

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 (0.5–1 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.

Androgen-regulated proteins in the rat ventral prostate: Androgen-regulierte Proteine der ventralen Rattenprostata

The rat ventral prostate synthetizes and secretes a number of relatively abundant proteins, such as Prostatic Binding Protein (PBP), the Proline-Rich Polypeptides (PRP), the 20 or 22-kDa protein and a prostatic protease, which respectively constitute approx. 50, 5, 10 and 2% of the secreted protein. The synthesis of these proteins is androgen-regulated and this is also the case for the concentration of the corresponding mRNAs. The existence of a putative androgen response element in the first intron of the PBP-C3 gene supports the possibility of a direct control of its transcription by androgen receptor complexes, but there is also evidence for the requirement of protein synthesis for mRNA stimulation. Finally, changes of tissue composition and cell-cell interactions between epithelial and stromal cells most probably play an important role in the global effect of androgens on the rat ventral prostate.

The influence of hypothalamic proline-rich polypeptide on ATP-phosphohydrolase activity in rat brain mitochondria during epileptiform seizures induced by pentylenetetrazole

We have studied the shifts in the activity of Mg2+-, Ca2+-, and HCO3−-dependent ATPases in brain mitochondria of white rats during epileptiform seizures (ES) induced by pentylenetetrazole. Inhibition of the catalytic activity of Mg2+-, Ca2+-, and HCO3−-dependent ATPases was observed during ES. However, a preliminary intraperitoneal injection of proline-rich polypeptide (PRP) restored the activity of the investigated enzymes after ES to the control level. It is possible to assume that PRP has a protective effect on the body intoxication caused by pentylenetetrazole.

Ovine colostrum nanopeptide affects amyloid beta aggregation

A colostral proline-rich polypeptide complex (PRP) consisting of over 30 peptides shows beneficial effects in Alzheimer’s disease (AD) patients when administered in the form of sublinqual tablets called Colostrinin. The aim of the present studies was to investigate whether nanopeptide fragment of PRP (NP) – one of the PRP complex components can affect aggregation of amyloid β (Aβ1-42). The effect of NP on Aβ aggregation was studied using Thioflavin T (ThT) binding, atomic force microscopy, and analyzing circular dichroism spectra. Results presented suggest that NP can directly interact with amyloid beta, inhibit its aggregation and disrupt existing aggregates acting as a β sheet breaker and reduce toxicity induced by aggregated forms of Aβ.

Hypothalamic proline-rich polypeptide enhances human CD34+ progenitor cell differentiation into erythroid and granulomonocytic lineages

Hypothalamic proline-rich polypeptide enhances human CD34+ progenitor cell differentiation into erythroid and granulomonocytic lineages

Effects of Colostrinin™ on gene expression-transcriptomal network analysis

Colostrinin™ (CLN) is a uniform mixture of low-molecular weight proline-rich polypeptides isolated from the mother's first milk, colostrum. Exposure of cells to CLN decreases intracellular levels of reactive oxygen species by regulating glutathione metabolism and modulating activities of antioxidant enzymes and mitochondrial function. It also inhibits beta amyloid-induced apoptosis and induces neurite outgrowth of pheochromocytoma cells. Administration of CLN to Alzheimer's disease patients has resulted in a stabilizing effect on cognitive function. We analyzed CLN-induced gene expression changes using high-density oligonucleotide arrays and transcriptomal network analysis. We found that CLN elicited highly complex and multiphasic changes in the gene expression profile of treated cells. CLN treatment affected a total of 58 molecular networks, 27 of which contained at least 10 differentially expressed genes. Here we present CLN-modulated gene networks as potential underlying molecular mechanisms leading to the reported effects of CLN on cellular oxidative state, chemokine and cytokine production, and cell differentiation, as well as on pathological processes like allergy, asthma, Alzheimer's, and other neurological diseases. Based on our results, we also predict possible modulatory effects of CLN on adipocytokine gene networks that play a crucial role in the pathobiology of diabetes, cardiovascular disorders, obesity, and inflammation. Taken together, CLN-altered gene expression networks presented here provide the molecular basis for previously described biological phenomena and predict potential fields of application for CLN in the prevention and treatment of diseases.

Myc-oncogene Inactivating Effect by Proline Rich Polypeptide (PRP-1) in Chondrosarcoma JJ012 Cells

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.

Colostrinin™: An Oxidative Stress Modulator for Prevention and Treatment of Age-related Disorders

Colostrum-derived proline-rich polypeptide, also known as Colostrinin (CLN), has been shown to have a stabilizing effect on cognitive function in Alzheimer's disease patients. This complex action of CLN could be related to prevention of amyloid-beta peptide aggregation, as shown in in vitro studies, and its impact on delicate cassettes of signaling pathways common to cellular redox regulation, proliferation and differentiation. Studies on cultured cells showed that CLN modulates intracellular levels of reactive oxygen species (ROS), via regulation of glutathione metabolism, activity of antioxidant enzymes and mitochondria function. Due to an improvement in senescence-associated mitochondrial dysfunction and a decrease in ROS generation, CLN decelerates the aging processes of both cultured cells and experimental animals. When given orally to mice, CLN increased the lifespan and improved various motor and sensory activities. Although the molecular basis by which CLN exerts its diverse effects are still under investigation, the regulatory effect on the cellular redox state via maintenance of mitochondrial function and modification of ROS-induced cell signaling seem to be of great importance. In this article, we examine experimental data pertinent to the mechanism of action, including a review of CLN's utility in the maintenance of physiological processes in which oxidative stress has an etiological role.

Colostrinin Decreases Hypersensitivity and Allergic Responses to Common Allergens

Background: Colostrinin™ (CLN), isolated from mothers’ pre-milk fluid (colostrum), is a uniform mixture of low-molecular-weight, proline-rich polypeptides. CLN induces neurite outgrowth of pheochromocytoma cells, extends the lifespan of diploid fibroblast cells, inhibits β-amyloid-induced apoptosis and improves cognitive functions when administered to Alzheimer’s disease patients. Objective: The aim of this study was to investigate potential allergic responses to CLN and its impact on allergic sensitization and inflammation caused by common allergens. Methods: We used a well-characterized mouse model of allergic airway inflammation. Changes in IgE/IgG1 and mucin levels, airway eosinophilia and hyperreactivity to methacholine were determined by ELISA, differential cell counting and whole-body plethysmography, respectively. Results: CLN did not increase IgE/IgG1 levels or induce cutaneous hypersensitivity reaction, airway inflammation and mucin production. Importantly, CLN significantly (p < 0.001) decreased IgE/IgG1 production, airway eosinophilia, mucin production and hypersensitivity induced by allergenic extracts from ragweed pollen grains and house dust mites. Conclusion: CLN itself is non-allergenic; however, it is effective in preventing allergic responses to known indoor and outdoor allergens. These data support the safe application of CLN and its potential use in the prevention of allergic inflammation in humans.

Colostrum from different mammalian species—A rich source of colostrinin

Colostrinin™ (CLN), a proline-rich polypeptide complex, originally isolated from ovine colostrum, has been proven to be a promising and non-toxic natural preparation capable of preventing the development of Alzheimer's disease (AD). Utilizing a new purification protocol, colostrum from cows, goats or humans can also be used as alternative sources of this complex. Comparison of properties of preparation derived from these three species revealed no significant differences from that obtained from sheep. In amino-acid composition, as in ovine CLN, a high level of proline residues (over 20%) and acidic amino acids (approximately 18%) was found in CLN from cows, goats and humans. The comparison of the electrophoretic patterns and reversed-phase-high performance liquid chromatography (RP-HPLC) elution profiles showed the presence of additional proteins in the CLN preparations of different origin, consisting mainly of β-lactoglobulin and α-lactalbumin. These proteins could be removed by an ultrafiltration process. Colostrum, regardless of origin (ovine, bovine, caprine or human), appeared to be a rich source of CLN fractions that could be separated in gram quantities. Storage of colostrum in frozen or lyophilized form affected neither the quantity nor quality of the purified peptides compared with its preparation from fresh material.

The effect of proline-rich polypeptide on reduction of methemoglobin and formation of superoxide by the b558III cytochrome and suprol metalloproteins in vitro

A synthetic analogue of the proline-rich polypeptide (PRP-1) from neurosecretory granules of bovine neurohypophysis (1.5–7 μg/ml) was shown to stimulate reduction of methemoglobin (metHb) and NADPH-dependent formation of superoxide by the b558III cytochrome (cyt) from human erythrocyte membranes in the course of incubation under aerobic conditions at 4°C in vitro. However, increase in the PRP-1 concentration resulted in a decrease in the superoxide-producing activity of the b558III cyt. It was also demonstrated that PRP-1 inhibited superoxide formation in response to the action of the suprol lipoprotein of blood serum.

A proline-rich polypeptide complex (PRP) isolated from ovine colostrum. Modulation of H 2O 2 and cytokine induction in human leukocytes

A proline-rich polypeptide complex (PRP) has immunoregulatory properties and also shows beneficial effects in Alzheimer's disease (AD). It is known that the unregulated activation of microglial cells in AD may result in chronic inflammatory response. There is a link between the activation of immune cells on the periphery and in the central nervous system (CNS). Therefore, we studied the effect of the PRP on human peripheral blood mononuclear cells (PBMCs) stimulated by LPS with PHA (LP) or PMA as proinflammatory activators. PRP and its nonapeptide fragment (NP) inhibited by 40–60% production of H 2O 2 induced by PMA. The peptides also inhibited activity of superoxide dismutase. Both peptide preparations showed differential effects on the secretion of cytokines. NP induced TNF-α only while PRP induced IL-6, IL-10 and TNF-α. On the other hand, the release of TNF-α and IL-10 induced by LP in PBMCs was inhibited by PRP while NP inhibited the release of IFN-γ and IL-10. The results obtained showed that PRP may affect not only adaptive immunity but also innate immunity and thus may regulate secretions of mediators of inflammation. The regulatory effect of the PRP on the innate immunity may shed some light on understanding the beneficial effects of this polypeptide complex in AD patients.

Colostrinin delays the onset of proliferative senescence of diploid murine fibroblast cells

Colostrinin (CLN), a uniform mixture of low-molecular weight, proline-rich polypeptides, induces neurite outgrowth of pheochromocytoma cells and inhibits beta amyloid-induced apoptosis. Moreover, its administration to patients with Alzheimer's disease resulted in improved cognitive functions. In this study, we investigated the impact of CLN on the lifespan of murine diploid fibroblast cells (MDF), an in vitro model for cellular aging. Here, we show that CLN significantly decelerates the senescence of cultured MDF and increases their population doubling levels. This action of CLN is associated with a decrease in the intracellular levels of reactive oxygen species, which may be due to senescence-associated mitochondrial dysfunction. These data suggest that CLN may delay the development of cellular aging at the level of the organism. Thus, CLN may be used in the prevention and/or therapy of diseases associated with aging processes.

Whole Genome Phage Display Selects for Proline-rich Boi Polypeptides against Bem1p

Interaction selection by biopanning from a fragmented yeast proteome displayed on filamentous phage particles was successful in identifying proline-rich fragments of Boi1p and Boi2p. These proteins bind to the second "src homology region 3'' (SH3) domain of Bem1p, a protein of Saccharomyces cerevisiae involved in bud formation. Target Bem1p was a doubly-tagged recombinant, Bem1([Asn142-Ile551]), which strongly interacts in ELISA with a C-terminal 75 amino acids polypeptide from Cdc24p exposed on phage. The whole yeast genomic display library contained approximately 7.7 x 10(7) independent clones of sheared S. cerevisiae genomic DNA fused to a truncated M13 gene III. This study corroborates the value of fragmented-proteome display to identify strong and direct interacting protein modules.

Treatment and prophylaxis of anthrax by new neurosecretory cytokines.

In 1881, Louis Pasteur described the Bacillus anthracis vaccine, which plays an important role for the treatment and prophylaxis of anthrax. Currently, treatment for anthrax infection involves the use of several different antibiotics, used in combination with vaccines, which possess potential virulence in white mice and guinea pigs. We discovered several new immunomodulators cytokines (polypeptides) produced by the neurosecretory cells of hypothalamus, some of which can be used as drugs for the treatment and prophylaxis of the anthrax. The proline-rich polypeptides, which consist from 10 to 15 amino acids and four proline residues, are of the special interest; one of them (PRP-1), which consist of 15 amino acids and has the following primary structure ALa-GLy-ALa-Pro-GLu-Pro-Ala-GLu-Pro-Ala-GLn-Pro-GLy-Val-Tyr (AGAPEPAEPAQPGVY) possesses antibacterial activity, and a new proline-rich peptide described by Galoyan and called Gx-NH2. Both were tested for treatment against the anthrax bacillus or anthrax strain N55 vaccine in guinea pigs and mice in vivo, and in vitro preparations. The results of experiments show that these hypothalamic neurosecretory cytokines have a strong prophylaxis and therapeutic properties towards animals infected by episodic strain of anthrax and anthrax vaccine N55. The conventional concepts concerning the function of hypothalamic neurosecretion and hypothalamic mechanisms of adaptation have to be reconsidered.

Synthetic bovine proline-rich-polypeptides generate hydroxyl radicals and fail to protect dopaminergic neurons against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced dopaminergic neurotoxicity in mice

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.