dendritic cell

Dendritic cells (DCs), professional antigen-presenting cells that process and deliver antigens using MHC II/I molecules, can be enhanced in numerous ways.  Exosomes derived from heat‐shocked tumor cells (HS‐TEXs) contain high amounts of heat-shock proteins (HSPs). HSPs, as chaperons, can induce DC maturation. This study aimed to investigate whether HS‐TEXs can promote DC maturation. To generate DC, bone marrow-derived cells were treated with Interleukin-4 and GM-CSF. Exosomes were isolated from heat-treated CT-26 cells. The expression level of HSP in exosomes was checked by western blot and the increase in the expression of this protein was observed. Then, HS‐TEXs were co-cultured with iDCs to determine DC maturity, and then DCs were co-cultured with lymphocytes to determine DC activity. Our results showed that  DCs treated with HS‐TEXs express high levels of molecules involved in DC maturation and function including MHCII, CD40, CD83, and CD86. HS‐TEXs caused phenotypic and functional maturation of DCs. In addition, flow cytometric results reflected a higher proliferative response of lymphocytes in the iDC / Tex + HSP group. HS‐TEXs could be used as a strategy to improve DC maturation and activation.

Colorectal cancer remains a leading cause of cancer-related mortality worldwide. Cancer immunotherapy involves autologous tumor-derived antigen-loaded Dendritic Cells (DCs) that activate potent antitumor immunity. Cytokine-Induced Killer (CIK) cells are a heterogeneous population of anti-tumor cytotoxic CD8+ T-cells. Cancer immunotherapy using a combination of CIK cells with DCs vaccines is a promising strategy. We investigated some of the latest developments in the DCs vaccination field, with a special emphasis on strategies to prepare highly immunogenic tumor cell antigens to load and to activate DCs. In this context, we applied the effects of immunogenic treatmentmodalities (heat shock) and four potent inducers of immunogenic cell death and apoptosis (mitoxantrone, oxaliplatin, 5 fluorouracil, and staurosporine) on DCs biology and their employment in DC-based activation of CIK cells. DCs were generated from bone marrow cells using granulocyte-macrophage colony-stimulating factor and Interleukin (IL)-4. CIK cells were expanded by interferon-gamma (IFN-γ), anti-CD3 monoclonal antibody, and IL-2 stimulation. The cytotoxic activity of CIK cells against CMT-93 cancer cells was assessed by MTT assay. IFN-γ production of CIK cells was examined by ELISA. Coincubation of untreated DCs with CIK cells significantly induces antitumor immunity of CIK cells. Mature DCs loaded by polyethylene glycol with mitoxantrone-oxaliplatin-induced apoptotic tumor cells stimulate greater cytotoxicity of CIK cells compared to DCs loaded with staurosporine, oxaliplatin- and 5-fluorouracil-inactivated tumor cell antigens, whole tumor lysates, and total tumor RNA in CMT-93 cells. Heat shock and mitoxantrone-oxaliplatin-treatment are the best approaches for cancer cell antigens preparation for DC-induced CIK cells activation.

Background and purpose

One of the most effective methods for the development of dendritic cell (DC)- based cancer immunotherapy is ex vivo pulsing of DCs with tumor cell lysates (TCLs). However, antitumor immune responses of DCs are significantly influenced by how TCLs were prepared. Here, we compared four strategies of TCL preparation derived from colon cancer cells, HT-29, for ex vivo maturation of DCs.

Peripheral blood monocytes were isolated from healthy volunteers and incubated with granulocyte macrophage colony-stimulating factor and interleukin (IL)-4 to differentiate into DCs in 10 days. Morphological properties, phenotype characteristics (i.e. CD83 and CD86), and cytokine production (i.e. IL-10 and interferon gamma) of DCs loaded with four different TCLs (i.e. freeze-thaw, hypochlorous acid (HOCl), hyperthermia, and UV irradiation) were evaluated.

HOCl preparations led to the generation of DCs with higher surface expression of maturation biomarkers (particularly CD83), while UV preparations resulted in DCs with lower levels of surface biomarkers compared to freeze-thawed preparations. The supernatant of DCs pulsed with HOCl preparation showed significantly higher levels of interferon gamma and lower levels of IL-10 compared with the other groups.

Our results suggest that pulsing DCs with HOCl preparation may be superior to other TCLs preparation strategies, possibly due to induction of rapid necrotic cell death.

One of the most effective methods for the development of dendritic cell (DC)- based cancer immunotherapy is ex vivo pulsing of DCs with tumor cell lysates (TCLs). However, antitumor immune responses of DCs are significantly influenced by how TCLs were prepared. Here, we compared four strategies of TCL preparation derived from colon cancer cells, HT-29, for ex vivo maturation of DCs.

Peripheral blood monocytes were isolated from healthy volunteers and incubated with granulocyte macrophage colony-stimulating factor and interleukin (IL)-4 to differentiate into DCs in 10 days. Morphological properties, phenotype characteristics (i.e. CD83 and CD86), and cytokine production (i.e. IL-10 and interferon gamma) of DCs loaded with four different TCLs (i.e. freeze-thaw, hypochlorous acid (HOCl), hyperthermia, and UV irradiation) were evaluated.

HOCl preparations led to the generation of DCs with higher surface expression of maturation biomarkers (particularly CD83), while UV preparations resulted in DCs with lower levels of surface biomarkers compared to freeze-thawed preparations. The supernatant of DCs pulsed with HOCl preparation showed significantly higher levels of interferon gamma and lower levels of IL-10 compared with the other groups.

Our results suggest that pulsing DCs with HOCl preparation may be superior to other TCLs preparation strategies, possibly due to induction of rapid necrotic cell death.

Dendritic cells (DCs) are key arms of immune system, which act in antigen presenting processes, and are considered as a bridge between innate and adaptive immune responses. DCs are found in both lymphoid and non-lymphoid organs. They are called interdigitating dendritic cells (IDCs) in secondary lymphoid organs. IDCs lack lineage surface markers and are positive for S-100 and vimentin. IDC sarcoma (IDCS) is a very rare neoplasm, which mainly affects lymph nodes, though there are reports of extra-nodal involvement. IDCS is thought to have poor prognosis. Although there is no consensus on the treatment modalities, such options as radicalsurgery, chemotherapy, and radiotherapy are performed depending on severity and site of the lesion. In this study, we present a case of IDCS in a 53-year-old male with a history of several skin lesions and prior diagnoses of basal cell carcinoma (BCC), squamous cell carcinoma (SCC), and metatypical carcinoma (MTC).

Previously we reported functional leukocyte immunoglobulin-like receptor A3 (LILRA3) leads to susceptibility and sub-phenotypes of several autoimmune diseases. LILRA3 levels in blood serum and CD14+ monocytes enhanced in systemic lupus erythematosus and resulted in disease severity. However, the mechanism of LILRA3 in the pathogenesis of autoimmunity remains elusive. This study aims to explore the potential impact of LILRA3 on the differentiation, maturation, and function of monocyte-derived DCs (MoDCs). The human monocytic cell line (THP-1) was cultured to derive MoDCs in vitro. We performed plasmid transfection to examine the impact of LILRA3 on monocyte differentiation. Surface markers on MoDCs were measured using FACS. To assess the function of mature MoDCs, IL-12p70, IFN-γ and IL-4 levels were detected after the mixed leucocyte response by enzyme-linked immunosorbent assay. Western blot assay was employed in this study to determine the signaling pathways in MoDCs activation. LILRA3 promotes MoDCs maturation, our results showed significant up-regulation of CD40, CD80, CD86, CD209, and HLA-DR and increased production of pro-inflammatory cytokine IL-12. LILRA3-treated MoDCs exhibited a robust proliferation of allogeneic CD4+ T cells and induced naïve CD4+ T cell polarization into the Th1 phenotype. Furthermore, the preceding activation of MoDCs maturation and LILRA3 function might be attributed to p38 MAPK and STAT1 signaling pathway’s aberrant activation. This is the first study to report that LILRA3 played a critical role in promoting MoDCs maturation and directing MoDCs to modulate Th1 cell differentiation, which may have a role in the pathogenesis of autoimmune diseases.

 Dendritic cells (DCs) contribute essentially to the outset and course of immune responses. So in patients with malignancy, there have been considerable interests in use of these cells in different interventions.  To evaluate the impact of Leishmania major’s components on DC maturation and their use as a therapeutic agent against tumor cells.  The cancer model was induced by injection of WEHI-164 cells (BALB/c derived fibrosarcoma cell line) subcutaneously in the right flank of animals. Bone-marrow derived DCs (BMDCs) were cultured with granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-4. After 5 days, tumor lysate, Leishmania major’s lysate, and Lipopolysaccharide (LPS) were added to the culture and incubated for 2 days. IL-12 production in DCs was measured by ELISA. For Immunotherapy, Mice received DCs subcutaneously around the tumor site. Two weeks after DCs injection, cytotoxicity assay and infiltration of CD8+ lymphocytes were evaluated.  Our results showed that immunotherapy with dendritic cells exposed to Leishmania extract led to producing a higher amount of IL-12, compare to the control group. A considerable increment in specific cytotoxic T cells activity, diminished tumor growth rate and improved survival of immunized animals were seen.  This study indicates that the use of Leishmania major extract, as well as LPS, can enhance the efficiency of DC-based vaccines and provides a basis for the use of Leishmania major in DC-targeted clinical therapies.

The potential of Candida albicans to modulate antigen-presenting cells maturation has been documented in past studies. Dendritic cells are critical modulators in the orchestration of adaptive immune responses alongside myeloid subtypes, which play an important role in the presentation of antigens to T cells. The aim of this study was to evaluate the efficacy of splenocytes activated with the extract of heated 4T1 cells and the yeast form of C. albicans against breast cancer growth in vivo‏.

4T1 cells were subcutaneously injected into the left flanks of female BALB/c mice (n=40). At a time when palpable tumors had developed, experimental groups were immunized twice at one-week interim with either activated splenocytes with the extract of heated 4T1 or the killed preparation of yeast form of C. albicans or a combination of the two-One week after the second injection, one-half of animals (n=20) were euthanized to investigate the immune response profile.

Administration of activated splenocytes with the combination protocol caused a favorable survival curve and slower rates of tumor development compared to other tumor-bearing mice. Moreover, combination therapy significantly increased the secretion of IFN-γ, respiratory burst and nitric oxide production and conversely diminished the secretion of IL-4, IL-10 and TGF-β in the splenocyte population.

Since the murine 4T1 cell line is similar to the final stage of human breast carcinoma, we postulate that activated splenocytes with the extract of heated 4T1 cells and yeast form of C. albicans can reduce tumor development in tumor-bearing mice.

Stimulated dendritic cells (DCs) have been shown to be effective in the induction of specific immune cells. Also, the CMV pp65 plays an important role in CMV life cycle and immune recognition. To assess the effect of CMV pp65 on the maturity and function of dendritic cells. Splenic DCs were treated with non-cytotoxic concentrations of the pp65 and analyzed for MHC II, CD86, and CD40 expression by flow cytometry. Then, ROR-γ, GATA3, T-bet, and FOXP3 gene expression levels were evaluated in T cells co-cultured with DCs using Real time-PCR. Finally, the effects of pp65 on allogenic T-cell responses in mixed lymphocyte culture (MLR), and the release of cytokines were investigated by ELISA and flow cytometry. The phagocytosis rate was significantly lower in the pp65-treated DCs than the non-stimulated DCs. There were significant differences in the raised level of CD40, CD86, and CCR7 in DCs as maturation markers. Furthermore, ROR-γ, and T-bet overexpression in T cells of the pp65-treated group compared with the non-stimulated group was observed. Significant differences were observed in the levels of IL-2, IL-6, IL-17, IL-22, TNF-α, and IFN-γ in pp65-stimulated groups compared with the non-stimulated DCs. The pp65-treated DCs can induce differentiation and functional activity of the cellular immune system, including Th17, and Th1, but not other major T-cell subsets such as Tregs, and Th2 population.

Myeloid-derived suppressor cells (MDSCs) are capable of suppressing the immune response. 5-Fluorouracil (5-FU) compared to other chemotherapy drugs have shown considerable decreases in the number of MDSCs without visible effects on T, B and natural killer cells, as well as dendritic cells (DCs). DC-based vaccines considered to be appropriate candidates for cancer immunotherapy. However, due to the presence of various factors like MDSCs in tumor microenvironment, DC vaccine cannot effectively perform its function. The purpose of this study was to evaluate the effect of low doses of 5-FU on the efficacy of DC-based vaccines in preventing and treating of melanoma tumor model. This research was performed on 28 melanoma tumor bearing C57BL/6 female mice. The mice were randomly divided to 4 groups, group 1 is control population while group 2 and 3 were treated with DC vaccine and 5-FU respectively and group 4 was treated with both DC Vaccine and 5-FU. The mice survival, tumor growth rate, number of MDSC and CD8 CD107a T cells in mice spleen were evaluated in each group with maximum result in group 4. Our results revealed that combination of DC vaccine and 5-FU reduced number of MDSCs (3%) and also tumor growth rate(10%)(p

Natural killer (NK) cells are effector cells of the innate immune system that exert direct cytotoxic functions. Ubiquitously-expressed toll-like receptors (TLRs) have been recognized as one of the major components promoting dendritic cell (DC) maturation, which may induce polarized immune responses beneficial to cancer immunotherapy. TLR-activated NK cells and DCs are prerequisite for robust activation of the innate immune system against tumors. Recently, some medical research and clinical trials have proposed NK cells as a new therapy and potential strategy in both children and adults with those cancers which cannot be cured with the usual treatment modalities. As an example, the importance of DC/NK antitumor immunity in the outcome of breast and other cancers is recently recognized. Therefore, considering strategies which exploit TLR-mediated immunity in concordance with DC/NK system holds strong potential for cancer therapy. This review addresses the current knowledge about the potential role of TLR in tumor immunotherapy.

The expanse of dendritic cells (DC) differentiation plays an important role in determining immune response. DC-based immunosuppressive drugs have notable side effects in increasing the risk of infectious diseases and cancers. G2013, as a novel anti-inflammatory and immunosuppressive agent, has been tested in experimental model of multiple sclerosis. The aim of this study was to conduct the safety property of G2013 on dendritic cells biology.
The effect of G2013 on differentiation, maturation, and function of dendritic cells was examined at Tehran University in 2014. To investigate how G2013 affects human dendritic cells (DC) in a defined inflammatory environment, human peripheral blood mononuclear cells (PBMC) were isolated from healthy blood. Monocytes were then purified using anti-CD14 microbeads. Monocytes were treated with G2013 in two different doses (6 and 12 μg/well) along with adding granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 for inducing monocytes to immature DC and adding lipopolysaccharide for running DC maturation. Differentiation, maturation, and function of dendritic cells were examined with flow cytometry and ELISA.
G2013 therapy had no significant effect on CD83, CD86 and DR expression, as well as IL-10 and IL-12 cytokine levels and it, has no remarkable side on differentiation, maturation and function of dendritic cells in immature DC and mature DC process in vitro.
G2013 is a safe agent with no adverse effect on differentiation, maturation, and function of dendritic cells. It may be recommended as a novel immunosuppressive agent with no or little side effect in increasing the risk of infectious diseases and cancers.

Mesenchymal stem cells (MSCs) are multipotent cells with immunomodulatory effect on immune cells including dendritic cells (DCs). DCs are the most potent antigen-presenting cells (APC). MSCs have been found to modulate both differentiation and function of DCs. DCs express a broad range of Tolllike receptors (TLR), which play a critical role in DCs maturation and function.
To evaluate expression level of TLR3 and TLR9 transcripts in DCs following treatment with MSCs supernatant.
MSCs and DCs were derived from adult BALB/c mice bone marrow and spleen, respectively. MSCs supernatant was harvested following 24, 48, and 72 hours. Isolated DCs were treated with MSCs supernatant and incubated for 24 and 48 hours. TLR3 and TLR9 transcript levels were evaluated using real-time PCR.
The results showed that 48 and 72 hours MSCs supernatant significantly decreased the expression of TLR3 in DCs following 24 and 48 hours incubation in comparison with untreated cells (p TLR3 and TLR9 mRNA expression decreases in DCs after incubation with MSCs culture supernatant. This confirms the immunomodulatory role of MSCs in cell-base therapy.

The immunomagnetic separation technique is the basis of monocyte isolation and further generation of monocyte-derived dendritic cells.
To compare the efficiency of monocyte positive and negative separation, concentration of beads, and their impact on generated dendritic cells.
Monocytes were obtained using monoclonal antibody-coated magnetic beads followed the Ficoll-Paque gradient separation of mononuclear cell fraction from the peripheral blood of 6 healthy volunteers. CD14 expression was analyzed by flow cytometry.
Both types of magnetic separation including recommended and reduced concentrations of beads did not affect the yield and the purity of monocytes and their surface CD14 expression. However, DCs originated from the “positively” separated monocytes had noticeable higher expression of CD80 .

Fusion of dendritic cells (DCs) with melanoma cells could reinforce the antigenicity of tumors as a strategy for the treatment of malignant melanoma. However, the insufficient quantity of DCs and the low fusion efficiency limits the development of such approach. To define the dosage of the stimulating factors as well as the induction condition for the optimal DCs preparation and cell fusion. DCs were generated from murine bone marrow cells, and cultured with four different concentrations of the granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4). DCs were confirmed to be mature by detecting the expression of MHC-II, CD11c, CD80, and CD83 by flow cytometry. DCs-melanoma fusion cells were generated using polyethylene glycols (PEG) with different molecular weights and the fusion efficiency was detected by fluorescence-activated cell sorter (FACS). The largest quantity of DCs was found when cells were cultured with 1000 U/ ml of GM-CSF and 500 U/ml of IL-4 (1.69 ± 0.04 ×106 ml-1, p<0.001 when compared with the other three groups). The expression levels of MHC-II and CD83 on day 7 after incubation were significantly lower than those on day 3 (MHC-II: p<0.001; CD83: p<0.001). The efficiency of cell fusion under induction of PEG-3000 was significantly higher than that of PEG-4000 (15.4 ± 0.56% vs. 11.1 ± 0.45%, p<0.001). The largest quantity for mature DCs was stimulated with 1000 U/ml of GM-CSF and 500 U/ml of IL-4 and the highest fusion efficiency was under induction of PEG-3000.

Multiple sclerosis (MS) is an autoimmune disease with impairment in function of CNS, meanwhile macrophages and dendritic cells (DC) can cause inflammation and damage to the myelin of nerve cells by releasing Reactive oxygen species (ROS) and other harmful substances when these cells get matured. We investigated the effect of Alternaria alternata (A. alternata) extract on phagocytic T cell stimulation activity of DC pulsed with Myelin Basic Protein (MBP) as a laboratory model of MS. Plastic adherent monocytes were cultured with granulocyte-macrophage colony stimulating factor (GM-CSF), interleukin-4 for converting these cells to MoDc (Monocyte-Derived Dendritic Cell), pulsed with MBP, matured in the presence of monocyte-conditioned medium (MCM) in control group and MCM+Alternaria. alternata extract in treatment groups. Phagocytic activity of DC was evaluated and T cell responses were investigated by MTT test. Phagocytic activity in treatment groups decreased significantly in compare with control group. Meanwhile, DC couldn’t stimulate T cell proliferation. A. alternata extract decreased phagocytic activity of MoDc-pulsed with MBP and had no effect on T cell proliferation may provide a new strategy on immunotraphy of multiple sclerosis.

Chronicity of lupus nephritis (LN) should be considered for interaction of cell mediated immunity (CMI) and dendritic cells in glomeruli and tubulointerstitial areas. In this study establishment of immunohistopathological changes of dendritic cells and other immune effector cells in lupus nephritis comparing with non-lupus nephritis was performed. Renal needle biopsies of 35 cases of lupus nephritis and 35 cases of other causes of persistent proteinuria were compared for immunohistochemistry for plasmacytoid (CD123), myeloid (CD11c) dendritic cells, macrophages (CD68) and lymphocytes (CD4) markers. Statistical analysis of the data was performed using Spearman and Pearson correlation or ANOVA and t- student test (P < 0.05). Significant difference of glomerular and interstitial spaces for presence of myeloid-plasmacytoid dendritic cells and lymphocytes except macrophages between lupus nephritis and other causes of persistent proteinuria have found (P<0.001). Positive significant correlations were observed between glomerular presentation of myeloid dendritic cells and chronicity index but not with other markers in lupus nephritis (P <0.001). Statistically significant changes between presence of all markers and activity index were not observed (P >0.05). The myeloid dendritic cells might have synergistic role with other immune cells in pathogenesis and progression or chronicity of lupus nephritis.

Dendritic cells (DCs) are professional antigen presenting cells that have a potential role in the initiating of immune responses. The cell vaccination is a new strategy in treatment of infectious diseases and cancers. In this study, we have generated monocyte-derived dendritic cells of lymphoma patient''s peripheral blood mononuclear cells then; these cells were used as vaccine in lymphoma patients. We generated dendritic cell vaccine from lymphoma patient''s blood monocytes with human interleukin-4, granulocyte monocyte colony stimulating factor and then, antigen–primed Dcs were administrated subcutaneously close to the inguinal lymph nodes after maturation of dendritic cells. After 7 days, we analyzed immune response in lymphoma patients with determining of LDH, Beta 2 Microglobulin, CD4+T cell percent, CD8+ Tcell percent and Tumor size before and after vaccination. Furthermore, phenotypic and functional analysis of dendritic cells was performed using anti CD83-FITC monoclonal antibodies. Before vaccination, the mean ± SD of LDH was 530.62±140.65 but after vaccination it was 459±109.45 that significantly different between experimental groups (P=0.002). In addition, the CD8+ T cells percentage significantly different between two groups (P=0.002). We concluded that the use of dendritic cell probably is one of the suitable noninvasive treatments for lymphoma patients that they have not response to chemical drugs.