RESUMEN
Activity of a peptide tuftsin analogue Selank was studied in outbred rats using the naloxone-precipitated morphine withdrawal model. Single intraperitoneal injection of Selank in an anxiolytic dose of 0.3 mg/kg reduced the total index of morphine withdrawal syndrome by 39.6%, significantly (Ñ<0.0001) attenuated convulsive reactions, ptosis, and posture disorders, and 9-fold increased the tactile sensitivity threshold in morphine-dependent rats in comparison with the group of active control; at the same time, Selank was slightly inferior to diazepam in a dose of 2 mg/kg by pharmacological activity (the decrease in total index of morphine withdrawal syndrome by 49.3% and 13-fold increase in sensitivity threshold). Thus, Selank, like diazepam, weakens the aversive signs of morphine withdrawal in rats with opiate dependence.
Asunto(s)
Dependencia de Morfina , Síndrome de Abstinencia a Sustancias , Tuftsina , Ratas , Animales , Morfina , Dependencia de Morfina/tratamiento farmacológico , Naloxona/farmacología , Síndrome de Abstinencia a Sustancias/tratamiento farmacológico , DiazepamRESUMEN
Tuftsin-PhosphorylCholine (TPC) is a novel bi-specific molecule which links tuftsin and phosphorylcholine. TPC has shown immunomodulatory activities in experimental mouse models of autoimmune diseases. We studied herein the effects of TPC ex vivo on both peripheral blood mononuclear cells (PBMCs) and temporal artery biopsies (TABs) obtained from patients with giant cell arteritis (GCA) and age-matched disease controls. GCA is an immune-mediated disease affecting large vessels. Levels of 18 cytokines in supernatants, PBMC viability, T helper (Th) cell differentiation of PBMCs and gene expression in TABs were analyzed. Treatment ex vivo with TPC decreased the production of IL-1ß, IL-2, IL-5, IL-6, IL-9, IL-12(p70), IL-13, IL-17A, IL-18, IL-21, IL-22, IL-23, IFNγ, TNFα, GM-CSF by CD3/CD28 activated PBMCs whereas it negligibly affected cell viability. It reduced Th1 and Th17 differentiation while did not impact Th22 differentiation in PBMCs stimulated by phorbol 12-myristate 13-acetate plus ionomycin. In inflamed TABs, treatment with TPC down-regulated the production of IL-1ß, IL-6, IL-13, IL-17A and CD68 gene expression. The effects of TPC were comparable to the effects of dexamethasone, included as the standard of care, with the exception of a greater reduction of IL-2, IL-18, IFNγ in CD3/CD28 activated PBMCs and CD68 gene in inflamed TABs. In conclusion our results warrant further investigations regarding TPC as an immunotherapeutic agent in GCA and potentially other autoimmune and inflammatory diseases.
Asunto(s)
Antiinflamatorios/uso terapéutico , Dexametasona/uso terapéutico , Arteritis de Células Gigantes/tratamiento farmacológico , Inmunoterapia/métodos , Fosforilcolina/análogos & derivados , Subgrupos de Linfocitos T/inmunología , Células TH1/inmunología , Tuftsina/uso terapéutico , Anciano , Anciano de 80 o más Años , Diferenciación Celular , Células Cultivadas , Citocinas/metabolismo , Combinación de Medicamentos , Femenino , Humanos , Activación de Linfocitos , Masculino , Fosforilcolina/uso terapéuticoRESUMEN
BACKGROUND The aim of this study was to investigate the effect of antigenic peptides on dendritic cell maturation and activation as well as the role of dendritic cell induced cell function. The tumor-specific cytotoxic T lymphocytes induced by activation of the dendritic cells were also evaluated. MATERIAL AND METHODS SW-480 cell lysate and peptide antigens were selected as adjuvants in dendritic cell sensitization, and tuftsin was used to induce the phagocytosis of dendritic cells. Immature dendritic cells were stimulated with the antigen and adjuvant as follows: group A was negative control; group B was SW-480 (20 µg/mL); group C was SW-480 (20 µg/mL)+tumor necrosis factor (TNF)-alpha (10 µg/mL); group D was SW-480 (20 µg/mL)+tuftsin (20 µg/mL); group E was antigen peptide (2 µg/mL); group F was antigen peptide (2 µg/mL)+TNF-alpha (10 µg/mL); group G was antigen peptide (2 µg/mL)+tuftsin (20 µg/mL). Cytotoxic T lymphocytes activation and in vitro anti-tumor effects were examined by detecting the maturation marks of dendritic cells as well as interleukin (IL)-10 and IL-12 levels secreted by dendritic cells. Cells with the strongest immunizing effects were injected into nude mice and tumor suppression status was evaluated. RESULTS Group D (SW-480+tuftsin), group E (antigen peptides), group F (antigen peptide+TNF-alpha), and group G (antigen peptides+tuftsin) displayed significant differences compared to the control group (P<0.05). Group G (antigen peptides+tuftsin) could also promote the secretion of cytokines IL-12, as well as inhibit cytokine IL-10 secretion, compared to the other experimental groups (P<0.05). In the in vivo experiments of tumor inhibitions, antigenic polypeptide+tuftsin was the most effective (P<0.05). CONCLUSIONS Combination of cytotoxic T lymphocytes and T peptide therapy in treating human colorectal cancer might be used as a new treatment strategy based on adoptive cellular immunotherapy.
Asunto(s)
Neoplasias Colorrectales/tratamiento farmacológico , Células Dendríticas/inmunología , Tuftsina/farmacología , Animales , Línea Celular Tumoral , Neoplasias del Colon/patología , Células Dendríticas/efectos de los fármacos , Células Dendríticas/metabolismo , Humanos , Inmunoterapia Adoptiva/métodos , Activación de Linfocitos/inmunología , Ratones , Ratones Desnudos , Péptidos/farmacología , Linfocitos T Citotóxicos , Tuftsina/metabolismo , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
BACKGROUND: The hygiene theory represents one of the environmental facets that modulate the risk for developing autoimmune diseases. There is a reverse correlation between the presence of helminthes and flares of autoimmune diseases, which explains the rise in incidence of certain autoimmune diseases in developed countries. The protective properties of certain helminthes are attributed to their secretory compounds which immunomodulate the host immune network in order to survive. Thus, the helminthes use an array of mechanisms. One of the major mechanisms enabling manipulation of the host-helminth interaction is by targeting the pattern recognition receptors (PRRs)-dependent and -independent mechanisms, which include toll-like receptors, C-type lectin receptors, and the inflammasome. The current review provides a glimpse of numerous helminth secreted products which have a role in the immunomodulation of the host immune network, focusing on bifunctional tuftsin-phosphorylcholine (TPC). TPC is a natural compound based on phosphorylcholine of helminth origin that was used in the past to cover stents and tuftsin, a self-peptide derived from the spleen. TPC was proven to be efficient in three murine experimental models (lupus, colitis, and arthritis) and ex vivo in giant cell arteritis.
Asunto(s)
Enfermedades Autoinmunes/prevención & control , Helmintiasis/inmunología , Helmintos/inmunología , Factores Inmunológicos/inmunología , Factores Inmunológicos/farmacología , Inmunomodulación/inmunología , Fosforilcolina/inmunología , Fosforilcolina/farmacología , Tuftsina/inmunología , Tuftsina/farmacología , Animales , Modelos Animales de Enfermedad , Humanos , RatonesRESUMEN
Peptide-based small molecule drug conjugates for targeted tumor therapy are currently in the focus of intensive research. Anthracyclines, like daunomycin, are commonly used anticancer drug molecules and are also often applied in peptide-drug conjugates. However, lability of the O-glycosidic bond during electrospray ionization mass spectrometric analysis hinders the analytical characterization of the constructs. "Overprotonation" can occur if daunomycin is linked to positively charged peptide carriers, like tuftsin derivatives. In these molecules, the high number of positive charges enhances the in-source fragmentation significantly, leading to complex mass spectra composed of mainly fragment ions. Therefore, we investigated different novel tuftsin-daunomycin conjugates to find an appropriate condition for mass spectrometric detection. Our results showed that shifting the charge states to lower charges helped to keep ions intact. In this way, a clear spectrum could be obtained containing intact protonated molecules only. Shifting of the protonation states to lower charges could be achieved with the use of appropriate neutral volatile buffers and with tuning the ion source parameters.
Asunto(s)
Antibióticos Antineoplásicos/análisis , Daunorrubicina/análisis , Glicoconjugados/análisis , Factores Inmunológicos/análisis , Tuftsina/análisis , Antibióticos Antineoplásicos/química , Daunorrubicina/química , Glicoconjugados/química , Humanos , Factores Inmunológicos/química , Estructura Molecular , Protones , Espectrometría de Masa por Ionización de Electrospray , Electricidad Estática , Tuftsina/químicaRESUMEN
The effects of a peptide anxiolytic Selank synthesized on the basis of the endogenous peptide tuftsin on memory impairment and content of brain-derived neurotrophic factor (BDNF) in brain structures were analyzed in outbred rats receiving 10% ethanol as the only source of fluid for 30 weeks. In the object recognition test, Selank (0.3 mg/kg a day, 7 days, intraperitoneally) produced a cognitive-stimulating effect in 9 months rats not exposed to ethanol (p<0.05) and prevented the formation of ethanol-induced memory and attention disturbances (p<0.01) developing during alcohol withdrawal. In ex vivo experiments, Selank prevented ethanol-induced increase in BDNF content in the hippocampus and frontal cortex (p<0.05). These results indicate positive effects of the tuftsin analogue on age-related memory disturbances associated with chronic alcohol intoxication and confirm the involvement of the neurotrophin mechanism related to BDNF production into the effect of Selank.
Asunto(s)
Ansiolíticos/farmacología , Factor Neurotrófico Derivado del Encéfalo/genética , Hipocampo/efectos de los fármacos , Trastornos de la Memoria/prevención & control , Nootrópicos/farmacología , Oligopéptidos/farmacología , Corteza Prefrontal/efectos de los fármacos , Alcoholismo/tratamiento farmacológico , Alcoholismo/etiología , Alcoholismo/metabolismo , Alcoholismo/fisiopatología , Animales , Animales no Consanguíneos , Ansiolíticos/síntesis química , Factor Neurotrófico Derivado del Encéfalo/agonistas , Factor Neurotrófico Derivado del Encéfalo/antagonistas & inhibidores , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Etanol/administración & dosificación , Expresión Génica/efectos de los fármacos , Hipocampo/metabolismo , Hipocampo/fisiopatología , Masculino , Trastornos de la Memoria/inducido químicamente , Trastornos de la Memoria/metabolismo , Trastornos de la Memoria/fisiopatología , Memoria a Corto Plazo/efectos de los fármacos , Nootrópicos/síntesis química , Oligopéptidos/síntesis química , Corteza Prefrontal/metabolismo , Corteza Prefrontal/fisiopatología , Ratas , Tuftsina/química , Tuftsina/metabolismoRESUMEN
The role of helminth treatment in autoimmune diseases is growing constantly. Systemic lupus erythematosus (SLE) is a multi-system autoimmune disease with challenging treatment options. Tuftsin-phosphorylcholine (TPC) is a novel helminth-based compound that modulates the host immune network. This study was conducted to evaluate the potential value of TPC in ameliorating lupus nephritis in a murine model and specifically to compare the efficacy of TPC to the existing first-line therapy for SLE: corticosteroids (methylprednisolone). Lupus-prone NZBxW/F1 mice were treated with TPC (5 µg/mouse), methylprednisolone (MP; 5 mg/body weight) or phosphate-buffered saline (PBS) (control) three times per week once glomerulonephritis, defined as proteinuria of grade > 100 mg/dl, was established. Levels of anti-dsDNA autoantibodies were evaluated by enzyme-linked immunosorbent assay (ELISA), splenic cytokines were measured in vitro and the kidney microscopy was analysed following staining. TPC and MP treatments improved lupus nephritis significantly and prolonged survival in NZBxW/F1 mice. TPC-treated mice showed a significantly decreased level of proteinuria (P < 0·001) and anti-dsDNA antibodies (P < 0·001) compared to PBS-treated mice. Moreover, TPC and MP inhibited the production of the proinflammatory cytokines interferon IFN-γ, interleukin IL-1ß and IL-6 (P < 0·001) and enhanced expression of the anti-inflammatory cytokine IL-10 (P < 0·001). Finally, microscopy analysis of the kidneys demonstrated that TPC-treated mice maintained normal structure equally to MP-treated mice. These data indicate that the small molecule named TPC hinders lupus development in genetically lupus-prone mice equally to methylprednisolone in most of the cases. Hence, TCP may be employed as a therapeutic potential for lupus nephritis.
Asunto(s)
Antiinflamatorios/uso terapéutico , Helmintos/inmunología , Riñón/patología , Lupus Eritematoso Sistémico/tratamiento farmacológico , Nefritis Lúpica/tratamiento farmacológico , Fosforilcolina/análogos & derivados , Fosforilcolina/uso terapéutico , Tuftsina/uso terapéutico , Animales , Anticuerpos Antinucleares/sangre , Citocinas/metabolismo , Modelos Animales de Enfermedad , Combinación de Medicamentos , Femenino , Humanos , Mediadores de Inflamación/metabolismo , Riñón/efectos de los fármacos , Metilprednisolona/uso terapéutico , Ratones , Ratones Endogámicos NZB , Fosforilcolina/química , Tuftsina/químicaRESUMEN
Nanoparticles consisting of biodegradable poly(d,l-lactic- co-glycolic acid) (PLGA) are promising carriers for drug molecules to improve the treatment of tuberculosis. Surface modifiers, such as Pluronic F127, are essential for biocompatibility and for the protection against particle aggregation. This study demonstrates a successful approach to conjugate Pluronic F127 coated PLGA nanoparticles with Tuftsin, which has been reported as a macrophage-targeting peptide. Transformation of Pluronic F127 hydroxyl groups-which have limited reactivity-into aldehyde groups provide a convenient way to bind aminooxy-peptide derivatives in a one-step reaction. We have also investigated that this change has no effect on the physicochemical properties of the nanoparticles. Our data showed that coating nanoparticles with Pluronic-Tuftsin conjugate markedly increased the internalization rate and the intracellular activity of the encapsulated drug candidate against Mycobacterium tuberculosis. By employing this approach, a large variety of peptide targeted PLGA nanoparticles can be designed for drug delivery.
Asunto(s)
Antituberculosos/administración & dosificación , Portadores de Fármacos/química , Sistemas de Liberación de Medicamentos , Nanopartículas/química , Poloxámero/química , Copolímero de Ácido Poliláctico-Ácido Poliglicólico/análogos & derivados , Tuftsina/química , Antituberculosos/farmacología , Línea Celular , Portadores de Fármacos/síntesis química , Humanos , Monocitos/microbiología , Mycobacterium tuberculosis/efectos de los fármacos , Poloxámero/síntesis química , Copolímero de Ácido Poliláctico-Ácido Poliglicólico/síntesis química , Propiedades de Superficie , Tuberculosis/tratamiento farmacológico , Tuftsina/síntesis químicaRESUMEN
Lymphatic filariasis is a "disease of poor people" due to a large section of affected people with economic backwardness. Therefore, successful elimination of this disease requires a cost-effective prophylactic agent such as vaccine along with conventional drugs. The Abundant Larval Transcript-2 (BmALT-2) protein of Brugia malayi has been recognized as the most potential vaccine candidate. Tuftsin, a tetra-peptide immunopotentiator has already shown the enhanced immunogenicity of various vaccine antigens in earlier studies. This study deals with the development of tuft-alt-2 fusion construct and a suitable culture condition for its large-scale production in Pichia pastoris. The recombinant P. pastoris/tuft-alt-2 with 9-11 copies of the gene construct exhibited the highest expression level. The molecular weight of P-TUFT-ALT-2 was determined as 28 kDa in SDS-PAGE including 3 kDa due to glycosylation. The dry cell biomass was 57.4 gL-1 in the bioreactor. The P-TUFT-ALT-2 expression was measured as about 35 mg L-1, which was 102% higher than flask culture. The P-TUFT-ALT-2 produced the highest 65,000 IgG peak titer in Balb/c mice. Moreover, P-TUFT-ALT-2 exhibited about 9.46% higher splenocyte proliferation than E. coli expressed E-ALT-2 alone. The enhanced secreted production of P-TUFT-ALT-2 in bioreactor would step up its commercialization as an inexpensive commercial vaccine for human lymphatic filariasis.
Asunto(s)
Antígenos Helmínticos/biosíntesis , Clonación Molecular/métodos , Factores Inmunológicos/biosíntesis , Pichia/metabolismo , Proteínas Recombinantes de Fusión/biosíntesis , Proteínas Recombinantes/biosíntesis , Tuftsina/biosíntesis , Animales , Antígenos Helmínticos/química , Antígenos Helmínticos/genética , Antígenos Helmínticos/inmunología , Secuencia de Bases , Brugia Malayi/química , Glicosilación , Factores Inmunológicos/química , Factores Inmunológicos/genética , Factores Inmunológicos/inmunología , Masculino , Ratones Endogámicos BALB C , Pichia/genética , Procesamiento Proteico-Postraduccional , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/inmunología , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/inmunología , Tuftsina/química , Tuftsina/genética , Tuftsina/inmunologíaRESUMEN
Experimental autoimmune encephalomyelitis (EAE) is an animal model of demyelinating autoimmune disease, such as multiple sclerosis (MS), which is characterized by central nervous system white matter lesions, microglial activation, and peripheral T-cell infiltration secondary to blood-brain barrier disruption. We have previously shown that treatment with tuftsin, a tetrapeptide generated from IgG proteolysis, dramatically improves disease symptoms in EAE. Here, we report that microglial expression of Neuropilin-1 (Nrp1) is required for tuftsin-driven amelioration of EAE symptoms. Nrp1 ablation in microglia blocks microglial signaling and polarization to the anti-inflammatory M2 phenotype, and ablation in either the microglia or immunosuppressive regulatory T cells (Tregs) reduces extended functional contacts between them and Treg activation, implicating a role for microglia in the activation process, and more generally, how immune surveillance is conducted in the CNS. Taken together, our findings delineate the mechanistic action of tuftsin as a candidate therapeutic against immune-mediated demyelinating lesions.
Asunto(s)
Encefalomielitis Autoinmune Experimental/metabolismo , Microglía/metabolismo , Neuropilina-1/metabolismo , Tuftsina/metabolismo , Animales , Modelos Animales de Enfermedad , Encefalomielitis Autoinmune Experimental/genética , Factores Inmunológicos/metabolismo , Activación de Macrófagos/genética , Activación de Macrófagos/fisiología , Ratones Transgénicos , Esclerosis Múltiple/genética , Esclerosis Múltiple/patología , Neuropilina-1/genética , Linfocitos T/metabolismoRESUMEN
Treatment with helminthes and helminthes ova improved the clinical symptoms of several autoimmune diseases in patients and in animal models. Phosphorylcholine (PC) proved to be the immunomodulatory molecule. We aimed to decipher the tolerogenic potential of tuftsin-PC (TPC), a novel helminth-based compound in collagen-induced arthritis (CIA) a mouse model of rheumatoid arthritis (RA). CIA DBA/1 mice were treated with TPC subcutaneously (5 µg/0.1 ml) or orally (250 µg/0.1 ml), starting prior to disease induction. The control groups were treated with PBS. Collagen antibodies were tested by enzyme-linked immunosorbent assay (ELISA), cytokine protein levels by ELISA kits and regulatory T (Treg ) and regulatory B (Breg ) cell phenotypes by fluorescence-activated cell sorter (FACS). TPC-treated mice had a significantly lower arthritis score of 1.5 in comparison with control mice 11.8 (P < 0.0001) in both subcutaneous and orally treated groups at day 31. Moreover, histology analysis demonstrated highly inflamed joints in control mice, whereas TPC-treated mice maintained normal joint structure. Furthermore, TPC decreased the titres of circulating collagen II antibodies in mice sera (P < 0.0001), enhanced expression of IL-10 (P < 0.0001) and inhibited production of tumour necrosis factor (TNF)-α, interleukin (IL)-17 and IL-1ß (P < 0.0001). TPC significantly expanded the CD4(+) CD25(+) forkhead box protein 3 (FoxP3(+) ) Treg cells and CD19(+) IL-10(+) CD5(high) CD1d(high) T cell immunoglobulin mucin-1 (TIM-1(+) ) Breg cell phenotypes (P < 0.0001) in treated mice. Our data indicate that treatment with TPC attenuates CIA in mice demonstrated by low arthritic score and normal joints histology. TPC treatment reduced proinflammatory cytokines and increased anti-inflammatory cytokine expression, as well as expansion of Treg and Breg cells. Our results may lead to a new approach for a natural therapy for early rheumatoid arthritis onset.
Asunto(s)
Antiinflamatorios no Esteroideos/farmacología , Anticuerpos/sangre , Artritis Experimental/tratamiento farmacológico , Fosforilcolina/farmacología , Tuftsina/farmacología , Administración Oral , Animales , Artritis Experimental/inmunología , Artritis Experimental/patología , Linfocitos B Reguladores/efectos de los fármacos , Linfocitos B Reguladores/inmunología , Linfocitos B Reguladores/patología , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/patología , Colágeno Tipo II/sangre , Colágeno Tipo II/inmunología , Factores de Transcripción Forkhead/genética , Factores de Transcripción Forkhead/inmunología , Expresión Génica , Receptor Celular 1 del Virus de la Hepatitis A , Inmunofenotipificación , Inyecciones Subcutáneas , Interleucina-10/genética , Interleucina-10/inmunología , Interleucina-17/genética , Interleucina-17/inmunología , Interleucina-1beta/genética , Interleucina-1beta/inmunología , Articulaciones/efectos de los fármacos , Articulaciones/inmunología , Articulaciones/patología , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/inmunología , Ratones , Ratones Endogámicos DBA , Linfocitos T Reguladores/efectos de los fármacos , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/patología , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/inmunologíaRESUMEN
Transmissible gastroenteritis coronavirus (TGEV) is a member of the genus Coronavirus, family Coronaviridae, order Nidovirales. TGEV is an enteropathogenic coronavirus that causes highly fatal acute diarrhoea in newborn pigs. An oral Lactobacillus casei (L. casei) vaccine against anti-transmissible gastroenteritis virus developed in our laboratory was used to study mucosal immune responses. In this L. casei vaccine, repetitive peptides expressed by L. casei (specifically the MDP and tuftsin fusion protein (MT)) were repeated 20 times and the D antigenic site of the TGEV spike (S) protein was repeated 6 times. Immunization with recombinant Lactobacillus is crucial for investigations of the effect of immunization, such as the first immunization time and dose. The first immunization is more important than the last immunization in the series. The recombinant Lactobacillus elicited specific systemic and mucosal immune responses. Recombinant L. casei had a strong potentiating effect on the cellular immunity induced by the oral L. casei vaccine. However, during TGEV infection, the systemic and local immune responses switched from Th1 to Th2-based immune responses. The systemic humoral immune response was stronger than the cellular immune response after TGEV infection. We found that the recombinant Lactobacillus stimulated IL-17 expression in both the systemic and mucosal immune responses against TGEV infection. Furthermore, the Lactobacillus vaccine stimulated an anti-TGEV infection Th17 pathway. The histopathological examination showed tremendous potential for recombinant Lactobacillus to enable rapid and effective treatment for TGEV with an intestinal tropism in piglets. The TGEV immune protection was primarily dependent on mucosal immunity.
Asunto(s)
Vacunas Bacterianas/inmunología , Gastroenteritis/prevención & control , Lacticaseibacillus casei/inmunología , Células TH1/inmunología , Células Th2/inmunología , Virus de la Gastroenteritis Transmisible/inmunología , Vacunas Virales/inmunología , Animales , Células Cultivadas , Gastroenteritis/inmunología , Inmunidad Mucosa/inmunología , Inmunización , Inmunoglobulina A/inmunología , Interleucina-17/biosíntesis , Interleucina-17/inmunología , Porcinos , Receptor Toll-Like 2/biosíntesis , Receptor Toll-Like 4/biosíntesis , Receptor Toll-Like 9/biosíntesis , Tuftsina/genéticaRESUMEN
Improved clinical findings of inflammatory bowel disease (IBD) upon treatment with helminthes and their ova were proven in animal models of IBD and in human clinical studies. The immunomodulatory properties of several helminthes were attributed to the phosphorylcholine (PC) molecule. We assessed the therapeutic potential of tuftsin-PC conjugate (TPC) to attenuate murine colitis. Colitis was induced by Dextransulfate-Sodium-Salt (DSS) in drinking water. TPC was given by daily oral ingestion (50 µg/0.1 ml/mouse or PBS) starting at day -2. Disease activity index (DAI) score was followed daily and histology of the colon was performed by H&E staining. Analysis of the cytokines profile in distal colon lysates was performed by immunoblot. Treatment of DSS induced colitis with TPC prevented the severity of colitis, including a reduction in the DAI score, less shortening of the colon and less inflammatory activity in histology. The immunoblot showed that the colitis preventive activity of TPC was associated with downregulation of colon pro-inflammatory IL-1ß, TNFα and IL-17 cytokines expression, and enhancement of anti-inflammatory IL-10 cytokine expression. In the current study, we demonstrated that TPC treatment can prevent significantly experimental colitis induction in naïve mice. We propose the TPC as a novel potential small synthetic molecule to treat colitis.
Asunto(s)
Colitis/patología , Factores Inmunológicos/farmacología , Fosforilcolina , Tuftsina/farmacología , Animales , Colitis/inducido químicamente , Colitis/tratamiento farmacológico , Colitis/inmunología , Citocinas/metabolismo , Sulfato de Dextran/efectos adversos , Modelos Animales de Enfermedad , Humanos , Factores Inmunológicos/administración & dosificación , Factores Inmunológicos/química , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Enfermedades Inflamatorias del Intestino/inmunología , Enfermedades Inflamatorias del Intestino/patología , Mucosa Intestinal/inmunología , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patología , Masculino , Ratones , Índice de Severidad de la Enfermedad , Tuftsina/administración & dosificación , Tuftsina/químicaRESUMEN
In areas where helminths infections are common, autoimmune diseases are rare. Treatment with helminths and ova from helminths, improved clinical findings of inflammatory bowel disease, multiple-sclerosis and rheumatoid-arthritis. The immunomodulatory functions of some helminths were attributed to the phosphorylcholine (PC) moiety. We aimed to decipher the tolerogenic potential of Tuftsin-PC (TPC) compound in mice genetically prone to develop lupus. Lupus prone NZBXW/F1 mice received subcutaneously TPC (5 µg/1 ml), 3 times a week starting at 14 weeks age. Autoantibodies were tested by ELISA, T-regulatory-cells by FACS, cytokines profile by RT-PCR and cytokines protein levels by DuoSet ELISA. Glomerulonephritis was addressed by detection of proteinuria, and immunoglobulin complex deposition in the mesangium of the kidneys of the mice by immunofluorescence. Our results show that TPC attenuated the development of glomerulonephritis in lupus prone mice, in particular, it ameliorated proteinuria (p < 0.02), and reduced immunoglobulin deposition in the kidney mesangium. TPC also enhanced the expression of TGFß and IL-10 (p < 0.001), and inhibited the production of IFNγ and IL-17 (p < 0.03). TPC Significantly enhanced the expansion of CD4+CD25+FOXP3+ T-regulatory cells (Tregs) phenotype in the treated mice. These data indicate that TPC hampered lupus development in genetically lupus prone mice which was exemplified by moderate glomerulonephritis, attenuation of pro-inflammatory cytokines and enhancement of anti-inflammatory cytokines expression, as well as Tregs expansion. Our results propose harnessing novel natural therapy for lupus patients.
Asunto(s)
Mesangio Glomerular/efectos de los fármacos , Glomerulonefritis/tratamiento farmacológico , Lupus Eritematoso Sistémico/tratamiento farmacológico , Nefritis Lúpica/tratamiento farmacológico , Fosforilcolina/administración & dosificación , Linfocitos T Reguladores/efectos de los fármacos , Tuftsina/administración & dosificación , Animales , Autoanticuerpos/metabolismo , Citocinas/metabolismo , Modelos Animales de Enfermedad , Factores de Transcripción Forkhead/metabolismo , Mesangio Glomerular/inmunología , Humanos , Inyecciones Subcutáneas , Subunidad alfa del Receptor de Interleucina-2/metabolismo , Activación de Linfocitos/efectos de los fármacos , Ratones , Ratones Endogámicos NZB , Fosforilcolina/análogos & derivados , Fosforilcolina/síntesis química , Linfocitos T Reguladores/inmunología , Tuftsina/síntesis químicaRESUMEN
Tuftsin (TF) is an immunomodulator tetrapeptide (Thr-Lys-Pro-Arg) that binds to the receptor neuropilin-1 (Nrp1) on the surface of cells. Many reports have described anti-tumor activity of tuftsin to relate with nonspecific activation of the host immune system. Lidamycin (LDM) that displays extremely potent cytotoxicity to cancer cells is composed of an apoprotein (LDP) and an enediyne chromophore (AE). In addition, Ec is an EGFR-targeting oligopeptide. In the present study, LDP was used as protein scaffold and the specific carrier for the highly potent AE. Genetically engineered fusion proteins LDP-TF and Ec-LDP-TF were prepared; then, the enediyne-energized fusion protein Ec-LDM-TF was generated by integration of AE into Ec-LDP-TF. The tuftsin-based fusion proteins LDP-TF and Ec-LDP-TF significantly enhanced the phagocytotic activity of macrophages as compared with LDP (P < 0.05). Ec-LDP-TF effectively bound to tumor cells and macrophages; furthermore, it markedly suppressed the growth of human epidermoid carcinoma A431 xenograft in athymic mice by 84.2 % (P < 0.05) with up-regulated expression of TNF-α and IFN-γ. Ec-LDM-TF further augmented the therapeutic efficacy, inhibiting the growth of A431 xenograft by 90.9 % (P < 0.05); notably, the Ec-LDM-TF caused marked down-regulation of CD47 in A431 cells. Moreover, the best therapeutic effect was recorded in the group of animals treated with the combination of Ec-LDP-TF with Ec-LDM-TF. The results suggest that tuftsin-based, enediyne-energized, and EGFR-targeting fusion proteins exert highly antitumor efficacy with CD47 modulation. Tuftsin-based fusion proteins are potentially useful for treatment of EGFR- and CD47-overexpressing cancers.
Asunto(s)
Antígeno CD47/inmunología , Enediinos/farmacología , Receptores ErbB/inmunología , Inmunotoxinas/farmacología , Proteínas Recombinantes de Fusión/farmacología , Tuftsina/farmacología , Animales , Carcinoma de Células Escamosas/inmunología , Carcinoma de Células Escamosas/terapia , Comunicación Celular , Línea Celular Tumoral , Regulación hacia Abajo , Enediinos/química , Femenino , Humanos , Inmunotoxinas/química , Inmunotoxinas/inmunología , Ratones , Ratones Endogámicos BALB C , Virus de la Neumonía Murina , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/inmunología , Tuftsina/química , Tuftsina/inmunología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Immune adjuvants have been used in cancer biotherapies to stimulate immune response to tumor cells. Despite their potential as anticancer reagents, there are several impediments to their use in clinical applications. In this study, we aim to modify the existing tuftsin structure and evaluate its antitumor activity in preclinical models. We synthesized a novel tuftsin derivative, namely, the T peptide (TP), by linking four tuftsin peptides, which showed enhanced stability in vivo. We then evaluated its anticancer activity in a postoperative residual tumor model in mice, where we surgically removed most of the primary tumor from the host, a procedure mimicking clinically postoperative patients. Despite the limited effect in intact solid tumors, TP strongly inhibited relapsed growth of residual tumors in postsurgical mice. Surgical resection of tumors accelerated residual tumor growth, but TP slowed down this process significantly. Interestingly, TP showed similar effects in human xenograft residual models. As an immunomodulator, TP could synergize the functions of macrophages, thus inhibiting the growth of cocultured tumor cells in vitro. Furthermore, TP could shift the macrophages to the tumor-suppressive M1 type and mobilize them to produce elevated cytotoxic TNF-α and NO. As a result, TP effectively prolonged the survival time of tumor-resected mice. Using the postoperative residual tumor models, we provide a body of evidence showing the antitumor activity of TP, which causes no obvious toxicity. Our study highlights the potential of TP as a postoperative adjuvant in cancer therapies.
Asunto(s)
Adyuvantes Inmunológicos/uso terapéutico , Antineoplásicos/uso terapéutico , Neoplasia Residual/tratamiento farmacológico , Tuftsina/análogos & derivados , Tuftsina/uso terapéutico , Adyuvantes Inmunológicos/farmacología , Animales , Antineoplásicos/química , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia sin Enfermedad , Humanos , Lisina/química , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Ratones , Neoplasia Residual/inmunología , Neoplasia Residual/patología , Tuftsina/química , Factor de Necrosis Tumoral alfa/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The role of muramyl dipeptide (MDP) and tuftsin in oral immune adjustment remains unclear, particularly in a Lactobacillus casei (L. casei) vaccine. To address this, we investigated the effects of different repetitive peptides expressed by L. casei, specifically the MDP and tuftsin fusion protein (MT) repeated 20 and 40 times (20MT and 40MT), in mice also expressing the D antigenic site of the spike (S) protein of transmissible gastroenteritis virus (TGEV) on intestinal and systemic immune responses and confirmed the immunoregulation of these peptides. Treatment of mice with a different vaccine consisting of L. casei expressing MDP and tuftsin stimulated humoral and cellular immune responses. Both 20MT and 40MT induced an increase in IgG and IgA levels against TGEV, as determined using enzyme-linked immunosorbent assay. Increased IgG and IgA resulted in the activation of TGEV-neutralising antibody activity in vitro. In addition, 20MT and 40MT stimulated the differentiation of innate immune cells, including T helper cell subclasses and regulatory T (Treg) cells, which induced robust T helper type 1 and T helper type 17 (Th17) responses and reduced Treg T cell immune responses in the 20MT and 40MT groups, respectively. Notably, treatment of mice with L. casei expressing 20MT and 40MT enhanced the anti-TGEV antibody immune responses of both the humoral and mucosal immune systems. These findings suggest that L. casei expressing MDP and tuftsin possesses substantial immunopotentiating properties, as it can induce humoral and T cell-mediated immune responses upon oral administration, and it may be useful in oral vaccines against TGEV challenge.
Asunto(s)
Acetilmuramil-Alanil-Isoglutamina/genética , Gastroenteritis Porcina Transmisible/inmunología , Lacticaseibacillus casei/genética , Células TH1/inmunología , Células Th17/inmunología , Virus de la Gastroenteritis Transmisible/inmunología , Tuftsina/genética , Vacunas Virales/inmunología , Acetilmuramil-Alanil-Isoglutamina/administración & dosificación , Acetilmuramil-Alanil-Isoglutamina/inmunología , Administración Oral , Animales , Femenino , Gastroenteritis Porcina Transmisible/prevención & control , Gastroenteritis Porcina Transmisible/virología , Lacticaseibacillus casei/inmunología , Masculino , Ratones , Glicoproteína de la Espiga del Coronavirus/administración & dosificación , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/inmunología , Porcinos , Virus de la Gastroenteritis Transmisible/genética , Tuftsina/administración & dosificación , Tuftsina/inmunología , Regulación hacia Arriba , Vacunas Virales/administración & dosificación , Vacunas Virales/genéticaRESUMEN
Antibodies (immunoglobulins, Ig) are used by the immune system to identify and neutralize foreign objects and are responsible for antigen-binding and effector functions. Immunoglobulin G (IgG) is the major serum immunoglobulin of a healthy human (~75% of the total Ig fraction). The discovery in 1970 of the endogenous tetrapeptide tuftsin (Thr-Lys-Pro-Arg, fragment 289-292 of the C(H2)-domain of the heavy (H) chain of IgG), possessing both immunostimulatory and neurotrophic activities, was an impetus for the search for new biologically active peptides of immunoglobulin origin. As a result, fragments of the H-chain of IgG produced as a result of enzymatic cleavage of IgG within the antigen-antibody complex were discovered, synthesized, and studied. These fragments include rigin (341-344), immunorphin (364-373), immunocortin (11-20), and peptide p24 (335-358) and its fragments. In this review the properties of these peptides and their role in regulating the immune response are analyzed.
Asunto(s)
Cadenas Pesadas de Inmunoglobulina/metabolismo , Complejo Antígeno-Anticuerpo/química , Complejo Antígeno-Anticuerpo/metabolismo , Humanos , Regiones Constantes de Inmunoglobulina/química , Regiones Constantes de Inmunoglobulina/metabolismo , Inmunoglobulina G/química , Inmunoglobulina G/metabolismo , Cadenas Pesadas de Inmunoglobulina/química , Cadenas gamma de Inmunoglobulina/química , Cadenas gamma de Inmunoglobulina/metabolismo , Oligopéptidos/química , Oligopéptidos/metabolismo , Fragmentos de Péptidos/química , Fragmentos de Péptidos/metabolismo , Transducción de Señal , Tuftsina/química , Tuftsina/metabolismo , betaendorfina/química , betaendorfina/metabolismoRESUMEN
Tuftsin (Thr-Lys-Pro-Arg) is a natural immunomodulating peptide found to stimulate phagocytosis in macrophages/microglia. Tuftsin binds to the receptor neuropilin-1 (Nrp1) on the surface of cells. Nrp1 is a single-pass transmembrane protein, but its intracellular C-terminal domain is too small to signal independently. Instead, it associates with a variety of coreceptors. Despite its long history, the pathway through which tuftsin signals has not been described. To investigate this question, we employed various inhibitors to Nrp1's coreceptors to determine which route is responsible for tuftsin signaling. We use the inhibitor EG00229, which prevents tuftsin binding to Nrp1 on the surface of microglia and reverses the anti-inflammatory M2 shift induced by tuftsin. Furthermore, we demonstrate that blockade of transforming growth factor beta (TGFß) signaling via TßR1 disrupts the M2 shift similar to EG00229. We report that tuftsin promotes Smad3 phosphorylation and reduces Akt phosphorylation. Taken together, our data show that tuftsin signals through Nrp1 and the canonical TGFß signaling pathway. Despite the 40-year history of the tetrapeptide tuftsin (TKPR), a macrophage and microglial activator, its mechanism of action has not been defined. Here, we report that the tuftsin-mediated anti-inflammatory M2 shift in microglia is caused specifically by tuftsin binding to the receptor neuropilin-1 (Nrp1) and signaling through TGFß receptor-1, a coreceptor of Nrp1. We further show that tuftsin signals via the canonical TGFß pathway and promotes TGFß release from target cells.
Asunto(s)
Factores Inmunológicos/fisiología , Neuropilina-1/fisiología , Transducción de Señal/fisiología , Factor de Crecimiento Transformador beta/fisiología , Tuftsina/fisiología , Animales , Western Blotting , Corteza Cerebral/citología , Citocinas/metabolismo , Técnica del Anticuerpo Fluorescente , Factores Inmunológicos/metabolismo , Metionina/fisiología , Ratones , Ratones Endogámicos C57BL , Microglía/metabolismo , Neuronas/fisiología , Neuropilina-1/antagonistas & inhibidores , Óxido Nítrico/metabolismo , Cultivo Primario de Células , ARN/biosíntesis , ARN/aislamiento & purificación , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptores de Superficie Celular/metabolismo , Tuftsina/metabolismoRESUMEN
The main objective of this study was to evaluate macrophage-targeted alginate nanoparticles as a noncondensing gene delivery system for potential anti-inflammatory therapy. An external gelation method was employed to form plasmid DNA-encapsulated alginate nanoparticles. The nanoparticle surface was modified with a peptide sequence containing tuftsin (TKPR), and transfection efficiency was determined in J774A.1 macrophages. The effect of transfected mIL-10 in blocking expression of tumor necrosis factor-alpha (TNF-α) was evaluated in lipopolysaccharide (LPS)-stimulated cells. Scrambled peptide- and tuftsin-modified cross-linked alginate nanoparticles efficiently encapsulated plasmid DNA and protected against DNase I degradation. The transgene expression efficiencies, measured using GFP and mIL-10 expressing plasmid DNA, were highest with tuftsin-modified nanoparticles. Levels of TNF-α were significantly lower (p < 0.0001) in LPS-stimulated cells that were transfected with mIL-10 using alginate nanoparticles. The results of the study show that noncondensing alginate nanoparticles can efficiently deliver plasmid DNA, leading to sustained in vitro gene expression in macrophages.