Number Of Peripheral Blood CD4 Research Articles

IMPACT OF BONE MARROW FIBROSIS IN MYELOMA PATIENTS UNDERGONE AUTOLOGOUS STEM CELL TRANSPLANTATION

Autologous hematopoetic stem cell transplantation (aHSCT) after high dose chemotherapy is a standard treatment for multiple myeloma (MM) patients. The successful aHSCT depends on collection of sufficient numbers of hematopoietic progenitor stem cells and sustained engraftment following infusion. The aim of the present study is to determine the the impact of bone marrow fibrosis (BMF) on the clinical outcomes of MM patients who underwent aHSCT. Retrospectively, bone marrow trephine biopsy analyzed in 73 MM patients who were treated with hematopoietic stem cell transplantation (aHSCT) following bortezomib based induction regimen. The BM biopsy samples of all patients were re-evaluated by a single pathologists The patients divided into 4 groups according to fibrosis degree and the correlations in initial characteristic features, therapeutic response, survival, mobilization and engraftment outcomes were reviewed between the groups. Comparative analyses revealed that the median apheresis number was found statistically different according to groups (p=0.04). No significance was detected between the fibrozis grade and the number of peripheral blood CD34+ cell collection results and recovery time of neutrophils and platelets. Overall survival and progression free survival were found similar in groups, however relapse of disease was statistically different in patients with fibrosis (p=0.01). After induction treatment, a regression was observed in fibrosis grade of patients who had fibrosis at the time of diagnosis. Therefore we suggest to evaluate fibrosis status in all MM patients during each histopathological examination. Difficulties may be experienced during stem cell collection in transplant eligible MM patients with fibrosis at diagnosis. Therefore, we recommend that clinicians should be more careful in these patients during the induction treatment and stem cell mobilization.

Pathologic tonsillar findings similar to IgA nephropathy and the role of tonsillectomy in a patient with nephrotic syndrome

Background: The pathological findings of tonsils in IgA nephropathy include the expansion of T-cell nodules around lymphoid follicles and abnormal reticulation of the crypt epithelium in contrast to chronic tonsillitis. Recently, several studies have reported that regulatory T cells play an important role in the maintenance of self-tolerance, an abnormality that is involved in the onset of nephrotic syndrome (NS). We encountered a patient of 28-year-old male with frequently relapsing nephrotic syndrome (FRNS) and chronic tonsillitis whose tonsils demonstrated pathological findings similar to those of IgA nephropathy.Case presentationA patient had developed NS at the age of 5 years, and was pathologically diagnosed with minimal change disease (MCD), for which he received various immunosuppressive agents as treatment for recurrence. Because tonsillitis often triggers the recurrence of NS, a tonsillectomy was performed for chronic tonsillitis at the age of 25 years. Immunohistochemical staining of his tonsils showed the expansion of CD4 positive lymphocytes around the lymphoid follicles and abnormal reticulation of the crypt epithelium. The number of peripheral blood CD4+CD25+ regulatory T cells increased, and the frequency of relapses decreased after tonsillectomy.ConclusionA similar self-tolerance abnormality exists in NS and IgA nephropathy; therefore, tonsillectomy might become a novel therapeutic approach for FRNS to redress the unbalanced self-tolerance and to remove the tonsillar focal infection. Further studies are necessary to verify the clinical efficiency of tonsillectomy for FRNS with recurrent episodes triggered by tonsillitis.

Rapid development of HIV elite control in a patient with acute infection

BackgroundElite controllers (EC), a small subset of the HIV-positive population (< 1%), suppress HIV viremia below the limit of quantification of clinical viral load assays in the absence of antiretroviral therapy (ART). However, there is a paucity of longitudinal data detailing the viral and immune dynamics or HIV reservoir seeding during acute infection in individuals that go on to become Elite Controllers.Case presentationIn this report, we describe a case of a 42 year old woman diagnosed during acute infection who rapidly and permanently suppressed her viremia in the absence of antiretroviral therapy (ART). Rapid antibody/antigen testing was either negative or equivocal during acute infection, despite subsequent viral load testing at that time point with 71,550 plasma HIV RNA copies/mL, making initial diagnosis challenging. The patient subsequently developed detectable anti-HIV antibodies and an increase in HIV-specific CD8+ T cell responses to overlapping subtype C HIV gag peptide; very low-level plasma viremia (0.84 RNA copies/mL) was detected by an ultrasensitive assay 2 years following infection. Subsequently, she was started on ART for multifocal furunculosis despite continued suppression of virus and stable CD4+ T cell counts. Following ART initiation, CD8+ T cell responses increased, but no HIV DNA or RNA was able to be isolated from large numbers of peripheral blood CD4+ T cells.ConclusionThis case provides important information regarding the establishment of elite HIV control during acute infection and also demonstrates an increase in HIV-specific immune responses following ART despite undetectable peripheral blood cellular measures of HIV persistence. This case also highlights the challenges in diagnosing acute HIV infection without the use of viral load testing in this rare elite controller phenotype.

Association of imbalance of effector T cells and regulatory cells with the severity of asthma and allergic rhinitis in children.

This study investigated the frequency and absolute numbers of different subsets of peripheral blood cells, including CD4+ T cells and CD19+ interleukin (IL) 10+ B regulatory cells (Breg) and their potential association with clinical laboratory measurements in children diagnosed with simple asthma or asthma plus allergic rhinitis (AR). The frequency and numbers of peripheral blood CD4+ interferon (IFN) gamma+ T-helper (Th) type 1, CD4+ IL-4+ Th2, CD4+ IL-17A+ Th17, CD4+CD25+ Forkhead Box P3+ regulatory T cells (Treg) and CD19+ IL-10+ Bregs in 22 children with asthma, 17 children with asthma and AR, and 25 healthy controls were determined by flow cytometry. The levels of serum cytokines were determined by enzyme-linked immunosorbent assay. In comparison with healthy controls, patients with asthma alone had significantly increased numbers of Th1, Th2, and Th17 cells, and their cytokines but decreased numbers of Tregs and Bregs, and the cytokines IL-10 and transforming growth factor beta 1. This imbalance between effector and regulatory cells and their cytokines further increased in patients with asthma and AR. The ratios of percentage effector T cells (Th1 + Th2 + Th17) to regulatory cells (Treg + Breg) were positively correlated with fractional exhaled nitric oxide but negatively correlated with forced expiratory volume in 1 second in patients with asthma or asthma plus AR. The imbalance of effector T cells and regulatory cells contributed to the pathogenesis of airway inflammation of asthma and AR in children.

Platelet Count before Peripheral Blood Stem Cell Mobilization Is Associated with the Need for Plerixafor But Not with the Collection Result

Background: A low platelet count before mobilization has recurrently been identified as risk factor for poor mobilization. Methods: To determine the relevance of this finding for peripheral blood stem cell (PBSC) mobilization, including pre-emptive or rescue plerixafor in the case of poor mobilization, we retrospectively analyzed all patients undergoing PBSC collection at our institution between January 2014 and December 2015 (n = 380). Results: In total, 99% of the patients (377/380) successfully collected a minimum of 2 × 10⁶ CD34+ cells/kg body weight sufficient for a single transplant. Rescue or pre-emptive plerixafor was administered to 11% of the patients (42/380). No correlations between the platelet count before mobilization and the number of peripheral blood CD34+ cells or the CD34+ cell collection result were detected in the entire population or the subgroups according to diagnosis (newly diagnosed multiple myeloma, relapsed multiple myeloma, lymphoma, amyloid light-chain amyloidosis, sarcoma, or germ cell tumor). However, patients requiring pre-emptive or rescue plerixafor had a significantly lower platelet count before mobilization (217/nl vs. 245/nl; p = 0.004). Conclusion: With the current state of the art PBSC mobilization strategies, the platelet count before mobilization was not associated with the CD34+ cell collection result but was associated with the need for pre-emptive or rescue application of plerixafor.

Guidelines for the use of flow cytometry and cell sorting in immunological studies.

The marriage between immunology and cytometry is one of the most stable and productive in the recent history of science. A rapid search in PubMed shows that, as of July 2017, using “flow cytometry immunology” as a search term yields more than 68 000 articles, the first of which, interestingly, is not about lymphocytes. It might be stated that, after a short engagement, the exchange of the wedding rings between immunology and cytometry officially occurred when the idea to link fluorochromes to monoclonal antibodies came about. After this, recognizing different types of cells became relatively easy and feasible not only by using a simple fluorescence microscope, but also by a complex and sometimes esoteric instrument, the flow cytometer that is able to count hundreds of cells in a single second, and can provide repetitive results in a tireless manner. Given this, the possibility to analyse immune phenotypes in a variety of clinical conditions has changed the use of the flow cytometer, which was incidentally invented in the late 1960s to measure cellular DNA by using intercalating dyes, such as ethidium bromide. The epidemics of HIV/AIDS in the 1980s then gave a dramatic impulse to the technology of counting specific cells, since it became clear that the quantification of the number of peripheral blood CD4+ T cells was crucial to follow the course of the infection, and eventually for monitoring the therapy. As a consequence, the development of flow cytometers that had to be easy-to-use in all clinical laboratories helped to widely disseminate this technology. Nowadays, it is rare to find an immunological paper or read a conference abstract in which the authors did not use flow cytometry as the main tool to dissect the immune system and identify its fine and complex functions. Of note, recent developments have created the sophisticated technology of mass cytometry, which is able to simultaneously identify dozens of molecules at the single cell level and allows us to better understand the complexity and beauty of the immune system.

Clinical and immunological parallels in periprosthetic infection after total large joints arthroplasty

Aim. To study peripheral blood lymphocytes and monocytes subpopulations and evaluate synthesis of serum autoantibodies in patients with periprosthetic infection after large joints arthroplasty.Methods. The study included 40 patients with infectious and inflammatory complications that developed at different times after knee or hip arthroplasty. All patients were divided into three groups according to the Coventry-Fitzgerald-Tsukayama classification. The number of cells expressing the CD3, CD4, CD8, CD19, CD14, CD16 and CD56 antigens, the expression of the activation markers HLA-DR, CD25 on lymphocytes and monocytes, the number of peripheral blood CD4+CD45RA+, CD4+CDRO+ cells, and the level of serum antibodies to double-stranded deoxyribonucleic acid (DNA) and anti-neutrophil antibodies were evaluated.Results. It is found that in acute development of infectious and inflammatory process in peripheral blood a significant increase in the number and functional activity of T-cells occur. For long-term low-grade infectious and inflammation process significant increase in the number of B-cells and memory T-cells, decrease in the number of naive T-cells are characteristic. Monocyte HLA-DR expression and the amount of B-lymphocytes correlate with the severity of inflammation. In seven patients elevated levels of antibodies to double-stranded DNA, in six - antineutrophil antibodies were revealed in blood serum.Conclusion. The assessment of lymphocytes subpopulations and the monocyte HLA-DR expression can be used as additional laboratory criteria for the diagnosis of infectious and inflammatory processes after large joints arthroplasty; the role of autoantibodies to double stranded DNA and anti-neutrophil antibodies, which were found in some patients, remains unclear.

CD4(+)CD25(-)Foxp3(+) T cells play a role in tuberculous hydrothorax rather than malignant hydrothorax.

BackgroundFoxp3+ T cells regulate inflammation and tumorigenesis. However, little is known about the role of different subsets of Foxp3+ T cells in malignant or tuberculous hydrothorax.MethodsThe numbers of CD4+CD25+Foxp3+, CD4+CD25−Foxp3+ T cells and the levels of some inflammatory cytokines in patients with tuberculous hydrothorax, malignant hydrothorax, and healthy controls (HCs) were examined by flow cytometry and ELISA. The potential association between the numbers of different subsets of Foxp3 + T cells and the values of clinical measures were analyzed.ResultsThe numbers of peripheral blood CD4+CD25+Foxp3+ T cells were greater in malignant hydrothorax patients than in HCs, but fewer than those of hydrothorax in patients. The percentages of circulating IL-10+ or LAP+ CD4+CD25+Foxp3+ T cells were higher than in the hydrothorax in patients with malignant hydrothorax. The numbers of circulating CD4+CD25−Foxp3+ T cells were significantly fewer in patients with tuberculous hydrothorax than in HCs, and both the numbers of circulating CD4+CD25+Foxp3+ and CD4+CD25−Foxp3+ T cells were significantly fewer than in the hydrothorax in patients. Significantly higher percentages of circulating IL-10+ or LAP+ CD4+CD25+Foxp3+ and CD4+CD25−Foxp3+ T cells were detected in tuberculous hydrothorax patients. The numbers of CD4+CD25+Foxp3+ and CD4+CD25−Foxp3+ T cells were associated with hydrothorax adenosine deaminase (ADA) levels in tuberculous hydrothorax patients, while CD4+CD25+Foxp3+ T cells were associated with carcino-embryonic antigen (CEA) in malignant hydrothorax patients. The concentrations of serum IL-6 and TGF-β in the patients were significantly higher than that in the HCs, but lower than that in the corresponding hydrothorax. A similar pattern of IL-10 was observed in different groups, except that there was no significant difference in the levels of serum IL-10 between the tuberculous hydrothorax patients and HCs.ConclusionsCD4+CD25−Foxp3+ T cells, which have lower inhibitory function than CD4+CD25+Foxp3+ T cells, may play a role in tuberculous hydrothorax.

HTLV-1-associated infective dermatitis demonstrates low frequency of FOXP3-positive T-regulatory lymphocytes

BackgroundHuman T-lymphotropic virus (HTLV)-1-associated infective dermatitis (ID) is a rare severe chronic eczema, considered as a harbinger for the development of cutaneous adult T-cell leukemia/lymphoma (ATLL) and/or HTLV-1-associated myelopathy (HAM)/tropical spastic paraparesis (TSP). The pathogenesis of ID remains unclear. High numbers of peripheral blood CD4+ CD25+ FoxP3+ regulatory T cells (Tregs) have been reported in ATLL and HAM/TSP. ObjectiveTo investigate the status of Tregs, unknown to date, and the histopathological features of ID. MethodsWe studied 16 skin biopsies from 15 Peruvian adults and children with ID by immunohistochemistry. ResultsHistopathological patterns were seborrheic dermatitis-like and lichenoid. Intraepidermal lymphocytes were conspicuous. The infiltrate was composed of a CD3+ T cell infiltrate with a predominance of CD8+ over CD4+ cells. CD4+ CD25+ FoxP3+ Tregs were rare and their numbers were significantly lower than those reported in other inflammatory dermatoses. ConclusionTregs have an essential role in maintaining immune homeostasis of skin. Treg dysregulation ends in severe clinical manifestations. The clinical presentation of ID, with lesions resembling those seen in patients with atopic dermatitis and with mutations in the FoxP3 gene, is in agreement with a common Treg-deficient skin environment in these disorders, possibly secondary to HTLV-1 infection.

Increased numbers of peripheral blood CD34+ cells in dogs with canine atopic dermatitis.

The bone marrow may be involved in human atopic diseases, as shown by the release of CD34+ cells into the peripheral blood. The aim was to determine the numbers of CD34+ cells in atopic dogs. The following three groups of dogs were studied: 27 dogs with nonfood-induced atopic dermatitis (NFICAD); 16 dogs with nonallergic inflammatory diseases; and 13 healthy control dogs. Dogs with NFICAD were selected after fulfilment of Favrot's criteria and exclusion of other pruritic dermatoses, including flea infestation and adverse reaction to foods. The Canine Atopic Dermatitis Extent and Severity Index (CADESI)-03 and a Visual Analog Scale (VAS) score for pruritus were used to quantify clinical signs. A phycoerythrin-conjugated anticanine CD34 antibody was used to stain peripheral blood CD34+ cells, and these were enumerated using a flow cytometer. The CD34+ cell counts were compared between groups and tested (in the NFICAD group) for correlation with the severity of clinical signs. The numbers of peripheral CD34+ cells in dogs with NFICAD (median 1.7) were statistically higher than in dogs with other nonallergic inflammatory diseases (median 1.0; P=0.01) and healthy control dogs (median 0.9; P=0.009). In dogs with NFICAD, there was no correlation between CD34+ cell numbers and CADESI-03 scores or owner-assessed pruritus (VAS score). The results of this study suggest the possible involvement of CD34+ cells in dogs with NFICAD. The role of CD34+ cells in the aetiopathogenesis of canine atopic dermatitis remains to be determined.

In vitro expansion of CD34(+)CD38(-) cells under stimulation with hematopoietic growth factors on AGM-S3 cells in juvenile myelomonocytic leukemia.

Using serum-containing culture, we examined whether AGM-S3 stromal cells, alone or in combination with hematopoietic growth factor(s), stimulated the proliferation of CD34(+) cells from patients with juvenile myelomonocytic leukemia (JMML). AGM-S3 cells in concert with stem cell factor plus thrombopoietin increased the numbers of peripheral blood CD34(+) cells to approximately 20-fold of the input value after 2 weeks in nine JMML patients with either PTPN11 mutations or RAS mutations, who received allogeneic hematopoietic transplantation. Granulocyte-macrophage colony-stimulating factor (GM-CSF) also augmented the proliferation of JMML CD34(+) cells on AGM-S3 cells. The expansion potential of CD34(+) cells was markedly low in four patients who achieved spontaneous hematological improvement. A large proportion of day-14-cultured CD34(+) cells were negative for CD38 and cryopreservable. Cultured JMML CD34(+)CD38(-) cells expressed CD117, CD116, c-mpl, CD123, CD90, but not CXCR4, and formed GM and erythroid colonies. Day-7-cultured CD34(+) cells from two of three JMML patients injected intrafemorally into immunodeficient mice stimulated with human GM-CSF after transplantation displayed significant hematopoietic reconstitution. The abilities of OP9 cells and MS-5 cells were one-third and one-tenth, respectively, of the value obtained with AGM-S3 cells. Our culture system may provide a useful tool for elucidating leukemogenesis and for therapeutic approaches in JMML.

FRI0377 Abatacept Reduces Circulating Effector Memory T-Helper Cells in Patients with Primary SjÖGren's Syndrome

BackgroundIn an open-label proof of concept study, Abatacept has been identified as an effective and safe treatment modality in primary Sjögren's syndrome (pSS).1 Abatacept is a fully human fusion molecule...

Autoimmune regulator expression in thymomas with or without autoimmune disease

BackgroundThe autoimmune regulator (AIRE) regulates autoimmunity and self-antigen expression, such as acetylcholine receptor (AchR), in the thymus. Regulatory T cells (Tregs) can down-regulate autoimmunity, but also promote tumor growth. The objective of this study was to examine the levels of AIRE, AchR, and Foxp3 expression in thymomas. MethodsThe relative levels of AIRE, AchR, and Foxp3 mRNA transcripts and the frequency of AIRE+, AchR+, and Foxp3+ cells were determined by quantitative RT-PCR and immunohistochemistry in 79 thymoma tissue samples from 21 patients with simple thymoma (the Tm group), 39 patients with myasthenia gravis (the MG group) and 19 patients with myasthenia gravis and one other autoimmune disease (the AD group). The numbers of peripheral blood CD4+CD25+Foxp3+ Tregs were determined by flow cytometry analysis. ResultsThe relative levels of AIRE and AchR mRNA transcripts in the MG group were significantly lower than that in the Tm group (p=0.04, p=0.03), but higher than that in the AD group (p=0.03, p=0.04). The relative levels of Foxp3 mRNA transcripts in the Tm group were significantly higher than that in the MG and AD groups (p=0.03 for both). A similar pattern of the percentages of AIRE+, AchR+, and Foxp3+ cells in the thymoma tissues and the numbers of peripheral blood Tregs were detected in these patients. The levels of AIRE mRNA transcripts were correlated positively with that of the AchR and Foxp3 in this population. The levels of AIRE and AchR mRNA transcripts in the A/AB/B1 types of thymomas were significantly higher than that in the B2/B3/C types of thymomas in this population. ConclusionSignificantly lower levels of AIRE, AchR, and Foxp3 expression are associated with the development of thymoma-related autoimmune diseases.

Decreased Peripheral Blood CD4+/CD25+ Regulatory T Cells in Patients with Alcoholic Hepatitis

Development of alcoholic hepatitis (AH) may be favored by the activation of the innate immune response. Recently, decreased numbers of circulating regulatory T cells (Tregs) have been reported in diseases associated with an immune activation status, but no studies have focused so far, in investigating the distribution of Tregs in chronic alcoholism and its potential association with liver disease. Here, we analyzed for the first time the frequency of peripheral blood (PB) Tregs and Treg subsets in AH and its relationship with the production of inflammatory cytokines by PB monocytes and dendritic cells (DCs). PB samples from 25 male patients with AH were studied; in parallel, 15 male chronic alcoholic patients without liver disease (AWLD) and 17 male healthy donors were also studied, as controls. The distribution of CD4⁺CD25hiCD127-/lo Tregs and their maturation subsets (naïve, central memory, and peripheral memory Tregs) was analyzed by flow cytometry. Spontaneous and in vitro-stimulated production of inflammatory cytokines by PB monocytes and DCs was analyzed by flow cytometry at the cytoplasmic level. Patients with AH showed decreased (p < 0.05) numbers of PB CD4⁺CD25hiCD127-/lo Tregs at the expense of all maturation-associated subsets, while AWLD and healthy subjects showed a similar (p > 0.05) distribution of PB CD4⁺CD25hiCD127-/lo Tregs. Interestingly, significantly increased amounts of spontaneously produced inflammatory cytokines were found among circulating monocyte-derived DCs and monocytes from AH (and AWLD) patients in comparison with healthy donors. Conversely, the ability of these cell subsets to produce cytokines after in vitro stimulation was lower (p < 0.05) in AH versus the 2 control groups. PB CD4⁺CD25hiCD127-/lo Tregs are significantly decreased in patients with AH when compared to both healthy and AWLD; this may contribute to explain the more pronounced activation of the innate immune response observed in AH, as reflected by an increased secretion of inflammatory cytokines by PB DCs and monocytes, and could facilitate the development of liver disease.

Prospective cohort study of the circadian rhythm pattern in allogeneic sibling donors undergoing standard granulocyte colony-stimulating factor mobilization

IntroductionPrior in vivo murine studies suggest circadian oscillations for hematopoietic stem cell release, which are maintained following administration of granulocyte colony-stimulating factor (G-CSF) or plerixafor. Furthermore, retrospective data analysis of healthy donors who underwent G-CSF-induced mobilization demonstrated significantly increased CD34+ cell yields when collected in the afternoon compared with the morning.MethodsA prospective study was conducted to directly examine the number of peripheral blood CD34+ and CD34+CD38– progenitor/stem cells at baseline and then every 6 hours for 24 hours on days 4 to 5 of G-CSF (10 μg/kg/day in the morning) mobilization in 11 allogeneic donors. Data were analyzed using mixed-model analysis of repeated measures.ResultsWhereas we observed a significant increase in CD34+ cell counts toward the evening, counts were then sustained on the morning of day 5. The correlation between CD34+CD38– cell counts and the less defined CD34+ populations was weak.ConclusionsOur results suggest that the pharmacodynamic activity and timing of G-CSF may alter endogenous progenitor rhythms. Donor age, medical history, and medications may also impact circadian rhythm. Further studies should examine the circadian rhythm at the peak of G-CSF mobilization and should consider potential confounders such as the time of G-CSF administration and the age of the subjects.

Impaired Thymic Export and Increased Apoptosis Account for Regulatory T Cell Defects in Patients with Non-ST Segment Elevation Acute Coronary Syndrome

Regulatory T (Treg) cells play a protective role against the development of atherosclerosis. Previous studies have revealed Treg cell defects in patients with non-ST elevation acute coronary syndrome (NSTACS), but the mechanisms underlying these defects remain unclear. In this study, we found that the numbers of peripheral blood CD4(+)CD25(+)CD127(low) Treg cells and CD4(+)CD25(+)CD127(low)CD45RA(+)CD45RO(-) naive Treg cells were lower in the NSTACS patients than in the chronic stable angina (CSA) and the chest pain syndrome (CPS) patients. However, the number of CD4(+)CD25(+)CD127(low)CD45RA(-)CD45RO(+) memory Treg cells was comparable in all of the groups. The frequency of CD4(+)CD25(+)CD127(low)CD45RO(-)CD45RA(+)CD31(+) recent thymic emigrant Treg cells and the T cell receptor excision circle content of purified Treg cells were lower in the NSTACS patients than in the CSA patients and the CPS controls. The spontaneous apoptosis of Treg cells (defined as CD4(+)CD25(+)CD127(low)annexin V(+)7-AAD(-)) was increased in the NSTACS patients compared with the CSA and CPS groups. Furthermore, oxidized LDL could induce Treg cell apoptosis, and the oxidized LDL levels were significantly higher in the NSTACS patients than in the CSA and CPS groups. In accordance with the altered Treg cell levels, the concentration of TNF-α was increased in the NSTACS patients, resulting in a decreased IL-10/TNF-α ratio. These findings indicate that the impaired thymic output of Treg cells and their enhanced susceptibility to apoptosis in the periphery were responsible for Treg cell defects observed in the NSTACS patients.

Very high efficacy of intermediate-dose cytarabine in combination with G-CSF as a second-line mobilization of hematopoietic stem cells

Autologous hematopoietic stem cell transplantation (autoHSCT) is used widely in the treatment of patients with lymphoid malignancies. Currently, 99 % of such procedures are performed using peripheral blood as a source of stem cells [1]. The generally accepted minimal level of CD34 cells required for rapid neutrophil and platelet recovery after autoHSCT is 2 9 10/kg. However, some data indicate that higher numbers are associated with less need for blood product transfusions and administration of antibiotics [2–4]. Mobilization regimens are based either on the use of granulocyte-colony stimulating factor (G-CSF) alone or G-CSF in combination with chemotherapy, most frequently cyclophosphamide 1.5–7 g/m or lymphoma-specific salvage regimens [5]. Unfortunately, a significant proportion of patients fail to mobilize sufficient number of CD34 cells, thus requiring additional attempts [6]. New mobilization strategies are being explored, including the use of plerixafor, CXCR4 inhibitor, in combination with G-CSF, with or without chemotherapy. This agent enabled sufficient CD34 cell harvest in 64.8–81.6 % of proven or predicted poor mobilizers [7, 8]. Unfortunately, such treatment is very expensive, which limits its worldwide application. Hence, the development of new strategies is still warranted. Although current studies focus on small molecules interfering with stem cell–stroma interactions, traditional chemotherapy-based salvage mobilization regimens have not been sufficiently explored. In our center, between November 2010 and September 2011, 14 patients who had failed chemotherapy-based mobilization were treated with salvage regimen including intermediate-dose cytarabine (ID-AraC) in combination with G-CSF. Most of the patients were referred for transplantation due to multiple myeloma (n = 8) or lymphoma (n = 5), while the remaining one had ovarian cancer. Patients had previously been treated with a median of 2 (range 1–5) lines of chemotherapy and most of them had received irradiation. Previous mobilization was usually based on cyclophosphamide plus G-CSF (Table 1). Six patients received AraC 400 mg/m (2 h infusions) every 12 h for three consecutive days (total dose, 2400 mg/m), while the following eight patients were treated with AraC for 2 days (total dose, 1600 mg/m). G-CSF 5–10 lg/kg was started on day 5 and continued daily until last leukapheresis. Leukaphereses were started once the peripheral blood CD34 cell count exceeded 15/lL and were performed using the Spectra-Optia Apheresis System (CaridianBCT Inc, Lakewood, CO, USA). The target number of CD34 cells collected was 5 9 10/kg for multiple myeloma (all planned for tandem autoHSCT) and 2 9 10/kg for the remaining patients. The maximum number of peripheral blood CD34 cells/lL after AraC ? G-CSF was 86 (17–312) and was significantly higher compared to the first-line mobilization: 8 (0–35) (Mann–Whitney U test, P = 0.002). No difference could be found according to the AraC dose: 80 (17–174) for 1.6 g/m versus 88 (24–312) for 2.4 g/m, P = 0.85. All patients collected required number of CD34 cells, which was achieved with a single apheresis in 10/14 (71.5 %) patients (Table 1). Although all patients experienced profound cytopenia and most required T. Kruzel M. Sadus-Wojciechowska J. Najda T. Czerw M. Glowala-Kosinska J. Holowiecki S. Giebel (&) Department of Bone Marrow Transplantation, Maria Sklodowska-Curie Memorial Cancer Center and Institute of Oncology, Gliwice Branch, Wybrzeze Armii Krajowej 15 Street, 44-101 Gliwice, Poland e-mail: sgiebel@io.gliwice.pl

Autologous Peripheral Blood-Derived Mononuclear Cells Induced by Erythropoietin Improve Critical Ischemic Limbs

Efficient and secure collection of CD34+ cells are crucial for the angiogenic therapies. We have developed autologous peripheral blood-mononuclear cell (MNC) transplantation induced by erythropoietin (rhEPO) for critical ischemic limbs. Seven patients, including five with arteriosclerosis obliterans, one with Buerger's disease and one with progressive systemic sclerosis, underwent ten cell therapies. The first administration of rhEPO was performed two weeks before apheresis, and the second administration and blood donation were performed one week before apheresis to activate bone marrow. MNCs including CD34+ cells, isolated from peripheral blood by apheresis, were immediately injected intramuscularly into ischemic limbs. The number of peripheral blood-CD34 + cells had significantly increased from 1.32 ± 0.83/microL, before the rhEPO induction, to 1.86 ± 0.94/microL, before the apheresis. The number of transplanted MNCs ranged between 0.5 × 10(9) and 16.5 × 10(9), and that of CD34+ cells, between 0.1 × 10(6) and 12.7 × 10(6), accounting for 0.02%-0.1% of MNCs. There were no serious complications. Finger ulcers with Buerger's disease were significantly improved one month after the transplantations, but the same or other ulcer(s) appeared 2-6 months later. Three patients had an improvement in rest pain, and one patient extended maximum pain-free walking distance. Erythropoietin-induced autologous peripheral blood-MNC transplantation is a useful and safe alternative for ischemic limbs.

Relationship of Th17 cells as well as interleukin (IL)-17A and IL-23R mRNA with psoriasis vulgaris

Objective To assess the number of peripheral blood Thl7 cells and mRNA expressions of IL-17A and IL-23R and their correlations with disease severity in patients with psoriasis vulgaris （PV）. Methods Tissue specimens were reseeted from the lesions of 25 patients with PV and normal skin of 10 human controls, and venous blood samples were obtained from 20 of the patients and all of the normal human controls. Reverse transcription PCR and flow cytometry were carried out to measure the mRNA levels of IL-17A and IL-23R in these tissue specimens and quantify the number of peripheral blood CD4＋IL-17＋ T lympbocytes. Psoriasis area and severity index （PASI） was calculated for these patients. Results A significant increase was observed in the IL-17A and IL-23R mRNA levels in the patients compared with the controls （0.996 ＋ 0.231 vs. 0.437 ~ 0.096, t-- 10.572, P 〈 0.05; 1.006 ~ 0.339 vs. 0.491 ~ 0.196, t = 6.015, P 〈 0.05）. The levels of both IL-17A mRNA and IL-23R mRNA were positively correlated with PASI （r~ = 0.67, 0.70, respectively, both P 〈 0.05）. The number of CD4＋IL-17＋ T lymphocytes in peripheral blood showed no significant differences between the patients and controls. No statistical correlation was observed between the counts of CD4＋IL-17＋ T lymphocytes and expression levels of IL-17A or 1L-23R mRNA in psoriatic lesions. Conclusions In patients with PV, there is an increase in the expressions of IL-17A and IL-23R mRNA in lesions, which are correlated with disease severity, while no significant change is observed in the number of peripheral blood CD4＊IL-17＋ T cells.

Regulat ory T-cells in periphera l blood of pat ientswith br onchial ast hma

Background. To investigate the expression of surface molecules CD4+CD25high and CD4+FoxP3+ on regulatory T-cells (Treg) in patients with different severity of bronchial asthma (BA). Methods. We included 19 patients with severe BA, 12 patients with mild BA and 17 healthy donors. The circulating percentage of CD4+CD25high and CD4+FoxP3+ T-reg in peripheral blood was estimated by the flow cytometry analysis (FACSCalibur, Becton Dickinson, USA). Results. Numbers of peripheral blood CD4+CD25high was significantly decreased in mild BA compared to healthy control. Patients with severe BA had increased levels of CD4+FoxP3+ and CD4+CD25high compared to mild BA. Conclusion. Our results demonstrate new scientific data on the phenotypic characteristics and the level of T-reg in peripheral blood at different clinical forms of asthma. This fact confirms the existence of immunoregulatory control mechanisms of allergic inflammation involving T-reg.