BACKGROUND: For clinical applications, dendritic cells (DCs) need to be generated using GMP-approved reagents. In this study, we tested the characteristics of DCs generated in two clinical grade culture media and activated by three maturation stimuli, Poly I: C, LPS and the mixture of proinflammatory cytokines in order to identify the optimal combination of culture media and activation stimulus for the clinical use. METHOD: We tested DCs generation using two GMP-certified culture media, CellGro and RPMI+5% human AB serum and evaluated DCs morphology, viability and capapability to mature. We tested three maturation stimuli, PolyI:C, LPS and the mixture of proinflammatory cytokines consisting of IL-1, IL-6, TNF and prostaglandin E2. We evaluated the capacity of activated DCs to induce antigen-specific T cells and regulatory T lymphocytes. RESULTS: Cell culture in CellGro resulted in a higher yield of immature DCs resulting from increased number of adherent monocytes. DCs that were generated in CellGro and activated using Poly I:C were the most efficient in expanding antigen-specific T cells compared to the DCs that were generated in other media and activated using LPS or the cocktail of proinflammatory cytokines. A comparison of all tested combinations revealed that DCs that were generated in CellGro and activated using Poly I:C induced low numbers of regulatory T cells. CONCLUSION: In this study, we identified monocyte-derived DCs that were generated in CellGro and activated using Poly I:C as the most potent clinical-grade DCs for the induction of antigen-specific T cells.
- MeSH
- Cell Differentiation drug effects MeSH
- Dendritic Cells cytology drug effects MeSH
- Epitopes immunology MeSH
- Phenotype MeSH
- Immunotherapy methods MeSH
- Clinical Trials as Topic MeSH
- Culture Media pharmacology MeSH
- Humans MeSH
- Neoplasms immunology therapy MeSH
- Poly I-C pharmacology MeSH
- Cell Proliferation drug effects MeSH
- T-Lymphocytes, Regulatory cytology drug effects immunology MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Parikalcitol (19-nor-1,25/OH 2 /D 2 ), aktivátor receptoru pro vitamín D (VDR) druhé generace, je syntetickým analogem vitamínu D3. Na rozdíl od kalcitriolu méně stimuluje resorpci kalcia ve střevě , a tak je jeho užití spojeno s menším rizikem rozvoje nežádoucí hyperkalcémie. Informace o imunomodulačních účincích parikalcitolu jsou omezené. V této studii jsme prokázali, že je ve všech námi sledovaných aspektech imunomodulační aktivita parikalcitolu srovnatelná s kalcitriolem. Oba VDR agonisté bránili diferenciaci nezralých dendritických buněk (DC) z monocyt ů . Po stimulaci prostřednictvím Toll-like receptorů v prostředí VDR agonistů z ůstávaly DC v nezralém stavu. Přítomnost VDR agonistů během diferenciace DC bránila jejich aktivaci – tyto DC neprodukovaly žádný bioaktivní IL-12 a ve srovnání s kontrolními DCs měly výrazně menší schopnost indukce antigen-specifických T-lymfocyt ů . Indukční kapacita funkčních T-reg se však nezměnila. DC a T-lymfocyty hrají důležitou roli v patogenezi aterosklerózy a end-stage onemocnění ledvin. Imunomodulace pomocí VDR agonistů je využitelná v praxi v terapii chronických imunitou ovlivňovaných zánětlivých onemocnění. Vzhledem k žádoucím imunomodulačním účinkům parikalcitolu a minimálnímu hyperkalcemickému efektu by měl být parikalcitol VDR agonistou první volby, především v rámci prevence aterosklerózy.
Paricalcitol (19-nor-1,25/OH 2 /D 2 ), a second generation vitamin D receptor (VDR) activator, is a synthetic analogue of vitamin D3. In contrast to calcitriol, paricalcitol has a reduced effect on intestinal calcium resorption thus avoiding undesirable hypercalcemia. Info rmation about immunomodulatory activity of paticalcitol is scarce. In this study we show that, in all investigated aspects, paricalcitol reta ins significant immunomodulatory activity, comparable to calcitriol. Both VDR agonists impaired differentiation of immature dendritic cells (DC s) from monocytes. Presence of VDR agonists during DCs differentiation abolished their capacity to be activated and, despite potent Tol l-like receptor mediated stimulation, VDR agonists-treated DCs remained in the immature state. In accordance with these findings, VDR-treated D Cs produ- ced no bioactive IL-12 and had significantly decreased capacity to induce antigen-specific T cells while the capacity to induce Tregs remained unchanched when compared to control DCs. As DCs and T cells play an important role in the pathogenesis of atherosclerosis, in e nd-stage renal disease patients, paricalcitol should be a VDR agonist of choice for the reduction of the risk of atherosclerosis due to its im munomodulatory effect proven in this study and known limited hypercalcemic effect. The immunomodulatory potency of paricalcitol make it a drug of interest in the therapy of chronic immune mediated inflammatory diseases other then atherosclerosis.
- Keywords
- VDR aktivátory, chronické onemocnění ledvin,
- MeSH
- Adjuvants, Immunologic therapeutic use MeSH
- Atherosclerosis drug therapy MeSH
- Autoimmune Diseases drug therapy immunology MeSH
- CD4-Positive T-Lymphocytes immunology metabolism MeSH
- Kidney Failure, Chronic immunology therapy MeSH
- Dendritic Cells immunology drug effects MeSH
- Ergocalciferols immunology metabolism therapeutic use MeSH
- Financing, Organized MeSH
- Humans MeSH
- Receptors, Calcitriol immunology metabolism therapeutic use MeSH
- Vitamin D MeSH
- Check Tag
- Humans MeSH
Paricalcitol (19-nor-1,25/OH(2)/D(2)), a second generation vitamin D receptor (VDR) activator, is a synthetic analogue of vitamin D3. In contrast to calcitriol, paricalcitol has a reduced effect on intestinal calcium resorption thus avoiding undesirable hypercalcemia. Information about immunomodulatory activity of paricalcitol is scarce. In this study we show that, in all investigated aspects, paricalcitol retains significant immunomodulatory activity, comparable to calcitriol. Both VDR agonists impaired differentiation of immature dendritic cells (DCs) from monocytes. The presence of VDR agonists during DC differentiation abolished their capacity to be activated and, despite potent Toll-like receptor mediated stimulation, VDR agonist-treated DCs remained in the immature state. In accordance with these findings, VDR-treated DCs produced no bioactive IL-12 and had a significantly decreased capacity to induce antigen-specific T cells while the capacity to induce functional Tregs remained unchanged when compared to control DCs. As DCs and T cells play an important role in the pathogenesis of atherosclerosis, in end-stage renal disease patients, paricalcitol should be a VDR agonist of choice for the reduction of the risk of atherosclerosis due to its immunomodulatory effect proven in this study and known limited hypercalcemic effect. The immunomodulatory potency of paricalcitol makes it a drug of interest in the therapy of chronic immune-mediated inflammatory diseases.
- MeSH
- Lymphocyte Activation drug effects MeSH
- Cell Differentiation immunology drug effects MeSH
- CD8-Positive T-Lymphocytes immunology drug effects MeSH
- Dendritic Cells immunology drug effects MeSH
- Ergocalciferols pharmacology MeSH
- Immunologic Factors pharmacology MeSH
- Interleukin-12 biosynthesis immunology MeSH
- Calcitriol pharmacology MeSH
- Cells, Cultured MeSH
- Humans MeSH
- Monocytes immunology drug effects MeSH
- Receptors, Calcitriol antagonists & inhibitors MeSH
- T-Lymphocytes, Regulatory immunology drug effects MeSH
- Check Tag
- Humans MeSH
- Publication type
- Research Support, Non-U.S. Gov't MeSH
- Comparative Study MeSH
Vitaminy jsou látky nezbytné pro normální fungování lidského organizmu. V posledních letech byla detailně popsána jejich role v jednotlivých metabolických i regulačních pochodech. Vitaminy také zasahují do mnoha procesů, které chrání tělo před narušením rovnováhy a rozvojem nemocí. V tomto článku jsou shrnuty dosud známé poznatky o možnosti použití vitaminů při řešení některých zdravotních problémů a o jejich užívání jako prevence před některými onemocněními.
- MeSH
- Child MeSH
- Adult MeSH
- Maternal Nutritional Physiological Phenomena physiology drug effects MeSH
- Breast Feeding MeSH
- Pharmaceutical Preparations administration & dosage metabolism MeSH
- Humans MeSH
- Drug-Related Side Effects and Adverse Reactions MeSH
- Nutrition Disorders drug therapy prevention & control MeSH
- Dietary Supplements utilization MeSH
- Self Medication MeSH
- Aged physiology MeSH
- Pregnancy physiology metabolism MeSH
- Vitamins pharmacology metabolism therapeutic use MeSH
- Check Tag
- Child MeSH
- Adult MeSH
- Humans MeSH
- Aged physiology MeSH
- Pregnancy physiology metabolism MeSH
Agamaglobulinemie vázaná na chromozom X (XLA) je způsobena mutacemi v genu, který kóduje Brutonovu tyrosin kinázu (Btk). Btk hr aje důležitou roli ve vývoji a diferenciaci B lymfocytů v kostní dřeni, a porucha funkce Btk vede tedy k deficitu B lymfocytů v per iferní krvi. Kromě klíčové role Btk pro vývoj B lymfocytů je tento enzym exprimován i v dalších buněčných populacích včetně myeloidních buněk. A p rávě funkce Btk v myeloidních buňkách není v současné době zcela objasněna. Na základě recentních prací, které naznačují, že Btk hraje roli v signalizaci přes Toll-like receptory (TLR), jsme v této práci studovali expresi a funkci TLR v dendritických buňkách pacientů s XLA. Prokáz ali jsme, že absence Btk neovlivňuje počet cirkulujících dendritických buněk. Dendritické buňky pacientů s XLA mají podobnou expresi TLR jak o DC zdravých dárců a stimulace specifickými agonisty TLR-1/2, TLR-2/6, TLR-3, TLR-4 a TLR-5 vede ke změnám fenotypu a funkčním změn ám srovnatelným se zdravými dárci. Důležitým zjištěním je, že jsm e u pacientů s XLA nalezli výrazný defekt v produkci IL-6 a TNF-α po stimulaci TLR-8 jednovláknovou RNA. Identifikace narušené signalizace přes TLR -8 je velmi zajímavá ve vztahu ke klinické prezentaci pacie ntů s XLA. Ačkoli pravidelná substituce imunoglobuliny zabezpečí dostateč né hladiny cirkulujících protilátek, pacienti s XLA často trpí tě žkými infekcemi enteroviry. Nedostatečná reaktivita na virovou jednovláknovou RNA zřejmě přispívá k vysoké náchylnosti pacientů s XLA k těmto v irům.
The critical role of Bruton's tyrosine kinase (Btk) in B-cells has been documented by the block of B-cell development in X-link ed agammaglobuli- nemia (XLA). Less is known about Btk function in myeloid cells. Several pieces of evidence indicate that Btk is a component of Toll-like receptors (TLRs) signaling. We analyzed whether Btk deficiency in XLA is associ ated with an impaired dendritic cells (DCs) compartment or defective TLR signaling. We analyzed the expression of TLRs 1-9 on myeloid DCs generated from XLA patients, and evaluated their response to activation by specific TLRs agonists. We show that XLA patients have normal numbers of circulating DCs. Btk deficient DCs have no defect i n response to stimulation of TLRs 1/2, 2/6, 3, 4 and 5, but display a profound impairment of IL-6 and TNF-α production in response to stimula tion by TLR-8 cognate agonist, ssRNA. These findings may provide an explanation for the susceptibility to enteroviral infections in XLA patie nts.
- MeSH
- Agammaglobulinemia etiology genetics immunology MeSH
- Dendritic Cells immunology pathology MeSH
- Enterovirus Infections genetics immunology blood MeSH
- Financing, Organized MeSH
- Genetic Diseases, X-Linked etiology genetics immunology MeSH
- Interleukin-6 genetics immunology blood MeSH
- Humans MeSH
- Mutation genetics immunology MeSH
- RNA genetics immunology isolation & purification MeSH
- Tumor Necrosis Factor-alpha genetics immunology blood MeSH
- Toll-Like Receptors genetics immunology MeSH
- Check Tag
- Humans MeSH
