Giulia Orsini

Enumeration of human peripheral blood dendritic cells throughout the life

Human aging is associated with immunosenescence, a process characterized by alterations in numerical and functional features of immune system components. Dendritic cells (DCs) are the main antigen-presenting cells, playing a pivotal role in adaptive and innate immunity. Therefore, we investigated the distribution of human circulating DCs throughout the life, in order to contribute to the knowledge of the physiological background underlying the aging of immune system. Cytofluorimetric analysis of peripheral blood samples by all-aged healthy population showed a significant decrease of circulating DCs and of their two main subsets among age. This reduction was limited to the plasmacytoid cell subtype when young and old subjects were analyzed separately. The analysis of circulating Treg cell number in a cohort of the subjects showed a significant reduction with increasing age and a positive significant correlation to myeloid or plasmacytoid absolute numbers. In conclusion, this work provides a large set of data of normal reference values of peripheral blood dendritic cells in healthy population suitable for comparative clinical studies concerning pathological immune dysfunctions.

Defective Generation and Maturation of Dendritic Cells from Monocytes in Colorectal Cancer Patients during the Course of Disease

Colorectal cancer (CRC) is the second-leading cause of cancer-related deaths in Western countries. Today, the role of the host’s immune system in controlling the progression and spread of solid tumors is broadly established. Tumor immunosurveillance escape mechanisms, such as those involving dendritic cells (DCs), the most important antigen-presenting cells, are likewise recognized processes involved in cancer. The present study evaluates the ability of CRC patients to generate DCs in vitro from circulating monocytes at both pre- and post-operative timepoints; the results are correlated with the stage of disease to shed light on the systemic immune statuses of CRC patients. Our data showed that patients’ DCs had lower co-stimulatory molecule expression and were less able to present antigens to allogeneic T cells compared to healthy controls’ (HC) DCs. Furthermore altered cytokine secretion, such as increased IL-10 and reduced IL-12 and TNF-α, was observed. At the post-operative timepoints we observed a recovery of the patients’ ability to generate immature DCs, compared to HCs, but the maturational capacity remained affected. Our study conclusively highlights the persistently impaired in vitro generation of fully mature and functional DCs, which appears to be more altered during advanced stages. This work sheds light on a dendritic cell-based tumor immune escape mechanism that could be useful for the development of more effective immunotherapeutic strategies.

Dendritic cell defects in the colorectal cancer

Colorectal cancer (CRC) results from the accumulation of both genetic and epigenetic alterations of the genome. However, also the formation of an inflammatory milieu plays a pivotal role in tumor development and progression. Dendritic cells (DCs) play a relevant role in tumor by exerting differential pro-tumorigenic and anti-tumorigenic functions, depending on the local milieu. Quantitative and functional impairments of DCs have been widely observed in several types of cancer, including CRC, representing a tumor-escape mechanism employed by cancer cells to elude host immunosurveillance. Understanding the interactions between DCs and tumors is important for comprehending the mechanisms of tumor immune surveillance and escape, and provides novel approaches to therapy of cancer. This review summarizes updated information on the role of the DCs in colon cancer development and/or progression.

Numerical defect of circulating dendritic cell subsets and defective dendritic cell generation from monocytes of patients with advanced melanoma.

The behaviour of circulating dendritic cells (DCs) and DC generation from monocytes in melanoma patients during the progression of disease have not been described. We report a significant decrease in the absolute number of total DCs, which mainly affects plasmacytoid DCs in stage IV. Additionally, monocyte-DC generation is less efficient in advanced stages, resulting in DCs that exhibit increased phagocytic capacity, potentially indicating a less mature state. These findings elucidate aspects of basic tumour-mediated immunosuppression, which may have implications for immunotherapeutic approaches, suggesting that the selection of patients for immunotherapy should also be made on the basis of their immune status.

Zoledronic acid modulates maturation of human monocyte-derived dendritic cells:

Zoledronic acid (ZA) is a drug of the bisphosphonate class, which is widely used for the treatment of both osteoporosis and skeletal metastasis. Besides its main bone antiresorptive activity, ZA displays antitumor properties, by triggering the expansion and activation of γδ T-cells, which exert an antitumor effect through dendritic cells (DCs). Several studies have reported the interaction between ZA and γδ T-cells, but the potential immunoregulatory activity of this drug on DCs has scarcely been investigated. Therefore, in this paper, we evaluated the effects of a therapeutic dose of ZA on the in vitro generation and maturation of DCs derived from peripheral blood monocytes of healthy adult donors. We demonstrate that ZA treatment did not affect DC differentiation, but inhibited DC maturation on lipopolysaccharide activation, as shown by the impaired expression of maturation surface markers and reduced ability to induce allogeneic T-cell proliferation. Interestingly, IL-10 secretion by mature DCs was significantly lower in ZA-treated cells than in controls. We conclude that ZA exerts its immunological in vitro activity also by modulating the maturation of DCs.

Quantification of Blood Dendritic Cells in Colorectal Cancer Patients During the Course of Disease

Colorectal cancer is a malignancy with poor prognosis that might be associated with defective immune function. The aim of the present study was to investigate circulating dendritic cells in colorectal cancer patients, in order to contribute to elucidate tumor-escape mechanisms and to point out a possible correlation with the clinical condition of the disease. Therefore, we enumerated ex vivo myeloid and plasmacytoid dendritic cells, through multicolor flow cytometry, in 26 colorectal patients and 33 healthy controls. Furthermore we performed several analyses at determined time points in order to define the immunological trend of cancer patients after surgery and other conventional treatments. At the pre-operative time point the absolute number of plasmacytoid dendritic cells in cancer patients was significantly reduced in comparison to controls, this result being mainly referred to stage III-IV patients. The number of myeloid dendritic cells did not show any significant difference compared to healthy controls; interestingly the expression of the tolerogenic antigen CD85k was significantly higher on cancer patients’ myeloid dendritic cells than controls’. At the following samplings, circulating dendritic cell absolute number did not show any difference compared to controls. Conclusively the impairment of the number of circulating dendritic cells may represent one of the tumor escape mechanisms occurring in colorectal cancer. These alterations seem to be correlated to cancer progression. Our work sheds light on one of dendritic cell-based tumor immune escape mechanisms. This knowledge may be useful to the development of more effective immunotherapeutic strategies.

The effects of zoledronate on monocyte-derived dendritic cells from melanoma patients differ depending on the clinical stage of the disease

Zoledronic acid has shown indirect anticancer effects on angiogenesis, the tumor microenvironment and immune responses. Its immunological action is exerted, at least in part, via its modulating properties. The aim of this study was to investigate the in vitro effects of zoledronic acid on the dendritic cells of melanoma patients. Peripheral blood samples were collected from 26 patients with melanoma and 11 healthy donors. Dendritic cells were derived from purified monocytes, and zoledronic acid (ZA) was added on the first day of culture. The phenotype and function of the generated cells were evaluated by flow cytometry. The ZA-treated monocytes from patients with early-stage disease generated DCs characterized by reduced endocytic activity and increased allostimulatory capacity compared with the untreated samples, allowing restoration of the DC function observed in normal subjects. In contrast, the ZA-treated monocytes from patients at stage III generated cells with higher CD14 antigen expression and endocytosis than the untreated samples. Therefore, in melanoma patients, the in vitro ZA effects differ according to the progression of the disease. In addition, our preliminary results appear to suggest that ZA effects are also influenced by the expression of CD14 antigen, indicating that the DC phenotype together with clinical characteristics must be considered in the choice of patients to be treated with ZA. Our work focus on the effect of ZA on monocyte-derived DCs from melanoma patients, showing that the effects of therapeutic doses of this drug might be mediated at least in part by modulation of myeloid cell function.

Numerical alterations of dendritic cell subsets in the peripheral circulation of patients with partial 22q11.2 deletion syndrome

THE XXXI Italian Society of Cytometry (GIC) bi-annual meeting was held in Lucca (Tuscany, Italy) from October 8th to 11th 2013. All abstracts were carefully reviewed by the Scientific Program Committee and have been published here in full and categorized by scientific tracks such as cell cycle and apoptosis; environmental sciences and toxicology, hematology, immunology, methodology-technology, and oncology. The Abstracts accompany this report as online Supporting Information. To date there are over 600 members actively involved in educational programs, promotion of quality controls programs, drafting/validation of guidelines and accreditation, providing information for people involved that actively work in the field of basic and applied cytometry. This year, the Conference has been preceded by a half a day satellite event devoted to the establishment of the “Professional Certification GIC Register” which will be committed to test, train, certify, and monitor the professional levels of the “Italian Cytometrists” under the responsibility of GIC. At present more than 50 members have applied for enrollment in the formative training structure. After the series of “dedicated seminars” the attendees had the opportunity to discuss with the “Entry Committee” the consistency of the basic requirements they have declared in a preliminary “entry form.” Among all the CV items applicants should have a minimum of three years practical expertise in the field. Anyone passing this entry level will be examined by a formal “Examination Committee” that formally assigns the title of “Qualified Cytometrist.” This new GIC initiative had been very much appreciated by the members. Regarding the scientific program, this year the Scientific Committee selected 31 abstracts for oral presentation among those submitted (110). Plenary sessions included 9 invited lectures focused on the contribution of cytometry in different application fields, highlighting the interdisciplinary profile of GIC. The opening lecture was given by Prof. Dario Campana (University of Singapore) and was focused on how flow cytometry can guide treatment decisions in acute leukemia. This outstanding talk offered a rational and fascinating overview on the role of flow cytometry in the diagnosis and clinical management of acute leukemias including the future perspectives of monitoring the patient’s individual response to therapy in modern clinical trials. Each parallel session, offered along three days, was tailored on the traditional topics such as immunology, oncology, hematology, methodology, and environmental sciences with cytometry as the common denominator. Several invited and selected lectures were given to discuss more emerging and promising aspects of flow cytometry. Again in the “core” of the Society activities a special session had been programmed for reports of ongoing GIC Projects: