Mean Percentage Of CD4 Research Articles

The quality and quantity media-cultured mononuclear cell transplantation is safe and effective in ischemic hindlimb mouse model

ObjectiveThis study was conducted to investigate in vitro proangiogenic and anti-inflammatory phenotypes and functions and the in vivo efficacy and safety of quality and quantity (QQ) media-cultured mononuclear cells (MNCs) compared with standard cultured MNCs from the peripheral blood of patients with chronic limb-threatening ischemia (CLTI) with atherosclerotic risk factors. MethodsPeripheral blood MNCs (PBMNCs) from patients with CLTI were cultured in QQ culture media or standard culture media. Phenotypic analysis of progenitor cells (CD34+CD133+), M2 macrophages (CD206+), and inactivated T regulatory cells (CD4+CD25+CD127+), colony-forming assay, and tube formation assay of QQ media-cultured MNCs (QQMNCs) and PBMNCs, were conducted. Intramuscular transplantation of QQMNCs or PBMNCs was performed in the ischemic hindlimb model. The clinical appearance of ischemic limbs was observed, and blood flow in ischemic limbs was measured using a laser Doppler perfusion imager. Outcomes were compared between the QQMNC and PBMNC groups. ResultsTwenty patients with CLTI were included. The mean percentages of CD34+ cells, CD133+ cells, CD34+CD133+ progenitor cells, CD206+ cells, colony-forming cells, and tube formation were significantly higher in the QQMNCs. The mean percentage of CD4+CD25+CD127+ cells was significantly lower in QQMNC. The colony-forming unit count and Dil-acetylated low-density lipoprotein uptake were significantly greater in QQMNCs. The clinical appearance of post-QQMNC-injected limbs was less severe than the appearance of post-PBMNC-injected limbs. Limb perfusion was significantly better in the QQMNCs. ConclusionsProangiogenic and anti-inflammatory phenotypes of MNCs cultured in QQ culture media were reproducible. Intramuscular QQMNC transplantation was safe and resulted in better reperfusion of ischemic hindlimbs compared with PBMNCs.

Immunopathology of Cutaneous Leishmaniasis in a Cohort of Sri Lankan Patients.

Leishmania donovani is the causative organism of leishmaniasis in Sri Lanka. Studies on the immunopathology of leishmaniasis due to L. donovani are limited. The objective of this study was to describe the immunopathological characteristics of cutaneous leishmaniasis in a cohort of Sri Lankan patients. Fifty skin biopsies of cutaneous leishmaniasis confirmed by detection of organisms by histology, culture, slit-skin smear, and/or polymerase chain reaction were reviewed. The inflammatory infiltrate was characterized by immunohistochemical staining for CD4, CD8, CD20, and CD68. Associations and correlations between immunohistochemical staining pattern and the parasitic load, and patterns of inflammation were determined. The majority of biopsies showed a CD8+/CD4- T lymphocyte predominant infiltrate (84%, n = 42). A CD68 predominant infiltrate was seen in 16%(n = 8). The mean percentage of CD8+, CD4+, CD20+, and CD68+ inflammatory cells in the biopsies were 56.1% (SD = 16.5%), 2.6% (SD = 4.5%), 12.3% (SD = 10.9%), and 25.7% (SD = 15.8%) respectively. There was no association between the predominant inflammatory cell and the degree of inflammation (P = .173), presence of high RPI (P = .922), MRI(P = .367) or presence of granuloma (P = .247).The percentage of CD4+ cells showed a positive correlation with granuloma formation (Correlation coefficient = .411, P = .03). The percentage of CD20+ cells in the infiltrate showed a positive correlation with the degree of inflammation (Correlation coefficient = .491, P = .02) and the RPI (Correlation coefficient = .334, P = .018). Skin biopsies from cutaneous leishmaniasis due to L. donovani infection showed a CD8+/CD4- predominant infiltrate. This is similar to the findings of studies on cutaneous leishmaniasis due to some other species and suggests that the cytotoxic T cell response plays a role in infections due to L. donovani.

Abstract P041: Simultaneous checkpoint inhibition and immune cell activation that is safely localized to solid tumors

Abstract Unlike other checkpoint inhibitors, our targeted immunotherapeutic localizes to any solid tumor and simultaneously shields an agent of immunosuppression while presenting a signal for immunostimulation. Phosphatidylserine (PS) exposure on the extracellular surface of living tumor cells and their vasculatures provides one avenue by which the tumor microenvironment promotes immunosuppression. Extracellular surface PS is inherent to a tumor and its vasculature, even for inoperable tumors, and its expression cannot be mutated nor affected by acquired drug resistance. Annexin A5 (AnxA5) is a direct, high-affinity PS-binding protein that localizes to cells with PS exposed on the outer plasma membrane. In our studies, we conjugated a proprietary modified AnxA5, lacking cellular internalization, to TNFα (AnxA5MOD-TNFα) to convert the immunosuppresive environs of a murine 4T1 triple negative breast cancer (TNBC) into an immunostimulated one. This strategy localized the immune response to the tumor and minimized side effects, as evidenced by a lack of toxicity for up to 7 days in non-tumor bearing Balb/c female mice given up to 1 mg/kg. Proper assembly and functionality of AnxA5MOD-TNFα was verified simultaneously by ellipsometry, an optical technique similar to plasmon resonance. Fully assembled constructs were tested for binding to PS coated slides. The degree of light polarization is proportional to the amount of PS bound by the AnxA5 complex. Samples could be further incubated with TNF receptors to verify TNFα activity. Based on dose escalation studies in 4T1 tumor-bearing mice where the TNBC tumors were grown in the mammary fat pads, optimal dosages were determined for AnxA5MOD-TNFα (18 µg) and AnxA5MOD alone as a control (180 µg). These doses were further tested in a 4T1 growth inhibition study. Tumor size was tracked by caliper in two groups of mice (n=5/group) receiving drug treatment on days 12, 14 and 16 and a repeated measures ANOVA was conducted on measurements taken before, during and post-treatment. While median tumor size did not differ between control and drug treatment groups during the pre-treatment interval (p=0.84), there was a significant difference post-treatment (p<0.001) with mice receiving AnxA5MOD-TNFα having much smaller TNBC tumors. Tumors from the study were embedded in paraffin, sectioned (5 µm) and the overall immune cell content determined by H&E staining. Once it was evident there was a greater quantity of immune cells in AnxA5MOD-TNFα treated tumors vs. controls, sections were stained with validated antibodies to identify and count the immunoactivated T-cells, NK-cells and macrophages. There was a 3X greater mean percentage of CD8 and CD4 T-cells in mice receiving drug vs. control (p=0.03) along with 2.5X and 5X increases in NK-cells and M1 immunoactive macrophages, respectively. Conclusion: Our AnxA5MOD-TNFα inhibits the PS inhibitor while simultaneously activating TNF activators! Citation Format: Richard A. Richieri, Navneet Narula, Cynthia A. Loomis, Valeria Mezzano, John Billimek, Glenn T. Reynolds, Chris Reutelingsperger, Andries Zijlstra, Missag H. Parseghian. Simultaneous checkpoint inhibition and immune cell activation that is safely localized to solid tumors [abstract]. In: Abstracts: AACR Virtual Special Conference: Tumor Immunology and Immunotherapy; 2021 Oct 5-6. Philadelphia (PA): AACR; Cancer Immunol Res 2022;10(1 Suppl):Abstract nr P041.

Modulation of Tregs and iNKT by Fingolimod in Multiple Sclerosis Patients.

The altered numbers and functions of cells belonging to immunoregulatory cell networks such as T regulatory (Tregs) and invariant Natural Killer T (iNKT) cells have been reported in Multiple Sclerosis (MS), an immune-mediated disease. We aimed to assess the frequencies of Tregs and iNKT cells in MS patients throughout a one-year treatment with fingolimod (FTY) and to correlate immunological data with efficacy and safety data. The percentage of Tregs (defined as Live Dead-CD3 + CD4 + FoxP3 + CD25++/CD127− cells) increased steadily throughout the year, while there was no significant difference in the absolute number or percentage of iNKT cells (defined as CD3 + CD14−CD19− Vα24-Jα18 TCR+ cells). However, out of all the iNKT cells, the CD8+ iNKT and CD4−CD8− double-negative (DN) cell percentages steadily increased, while the CD4+ iNKT cell percentages decreased significantly. The mean percentage of CD8+ T cells at all time-points was lower in patients with infections throughout the study. The numbers and percentages of DN iNKT cells were more elevated, considering all time-points, in patients who presented a clinical relapse. FTY may, therefore, exert its beneficial effect in MS patients through various mechanisms, including the increase in Tregs and in iNKT subsets with immunomodulatory potential such as CD8+ iNKT cells. The occurrence of infections was associated with lower mean CD8+ cell counts during treatment with FTY.

Accumulation of Regulatory T Cells in Triple Negative Breast Cancer Can Boost Immune Disruption.

Background and AimThe present study was conducted to evaluate the number of Tregs in triple negative breast cancer (TNBC), in normal breast parenchyma and in the peripheral blood of these patients and controls, in addition to their correlations with the clinico-pathologic features and the outcomes of TNBC.MethodsThirty adult treatment-naïve women with non-metastatic TNBC were recruited. In addition, 20 ages matched healthy females participated as a control group. Peripheral blood samples were collected from all participants in tubes containing heparin, fresh tumor tissues were also obtained from all patients undergoing surgery, and 20 normal breast tissue samples were obtained from the same patients’ areas adjacent to the safety margins; all these samples were taken for flow cytometric detection of Tregs.ResultsThe mean percentages of CD4+CD25+highT cells and Tregs were higher in TNBC peripheral blood than healthy controls and in malignant tissue than normal tissue. Moreover, the frequencies of tumor-infiltrating CD4+T cells and Tregs were exceeding those in the peripheral blood of cancer patients. Only tumor-infiltrating Tregs have shown increasing levels with the increase in the tumor size and were significantly higher in patients with local recurrences than those without recurrence. In addition, Tregs showed significant inverse relation with DFS and direct relation with the level of the peripheral Tregs.ConclusionThe findings of the current study support the possibility that TNBC microenvironment conveys specific characteristics on Tregs distinguishing them from those in normal breast tissue or Tregs in peripheral blood, improving the capabilities of tumor-infiltrating Tregs to enhance tumor growth, local recurrence and reduce the DFS.

A clinical study of tremelimumab alone or in combination with olaparib in patients with advanced epithelial ovarian cancer.

6045 Background: Single agent immunotherapy (IO) has shown only modest clinical activity for the treatment of ovarian cancer. The combination of anti-programmed death-1 and PARP inhibitors showed promising activity in early trials. Here, we report the results of an open-label, parallel arm, dose escalation study of tremelimumab (T) alone or in combination with olaparib (O) in patients (pts) with advanced epithelial ovarian cancer (EOC). Methods: Pts with recurrent/persistent EOC who had progression < 12 months from last platinum exposure were enrolled. Prior therapy with IO (except anti-CTLA-4) or PARP inhibitor was allowed. Pts were randomized to either T 10mg/kg every 4 weeks (wks) x 7 then every 12 wks (Arm A) or T with O twice daily at three planned dose levels (Arm B). The primary objectives were safety, pharmacodynamic (PD) change in CD4+ICOShi peripheral T cells by flow cytometry, and identification of the optimal dose combination of T with O. Secondary objectives included 6-month progression-free survival (PFS6) and objective response rate (ORR). Results: A total of 24 pts were treated, 12 on Arm A, and 12 on two Arm B dose levels. Pts had a median age of 60 years (range 44-81). Histologic subtypes included high-grade serous EOC (20 pts, 83%), clear cell (3 pts, 13%), and moderately-differentiated adenocarcinoma (1 pt, 4%). BRCA1 mutation (mt) was present in 2 cases, BRCA2 mt in 1. Median number of prior regimens was 3.5 (range 1-9). Most adverse events (AEs) were attributable to T, the most common grade 3 toxicities were rash (13%), immune-mediated hepatitis (8%), and colitis (8%). No grade ≥4 toxicities were identified. Immune-mediated AEs also included acute kidney injury, hypophysitis, and hypothyroidism. No dose limiting toxicities were identified on Arm B. Two pts in Arm B had >PFS6. Of 20 pts evaluable for response, there was 1 partial response (Arm B), and 9 pts had stable disease (6 on Arm A, 3 on Arm B). Mean percentage of CD4+ICOShi T cells was significantly increased on Days 15 and 22 compared to Day 1 at both T dose levels (Table).T at 3 mg/kg with O at 150mg is the optimal dose of those tested. Conclusions: T and T with O was tolerable, with modest clinical activity in this pt population. AEs were as expected, and peripheral CD4+ICOShi T cells increased on therapy. Clinical trial information: 02485990. [Table: see text]

Toll-Like Receptors-2 and -4 in Graves' Disease-Key Players or Bystanders?

Graves’ disease (GD) is an autoimmune disease that affects the thyroid. The development of autoimmunity is associated with innate immune responses where the prominent role plays Toll-like receptors (TLRs). The aim of our study was to assess the relationship between the expression levels of TLR-2 and TLR-4 on CD4+ and CD8+ T as well as CD19+ B lymphocytes in patients with GD and selected clinical parameters. The study group consisted of 32 women with GD, the control group consisted of 20 healthy women. Immunophenotyping was performed using the flow cytometry and cytokines concentrations were assessed using ELISA assay. The mean percentage of CD4+/TLR-2+ and CD8+/TLR-2+ T cells in patients with GD was higher than in the control group (p < 0.0001). After obtaining euthyroidism, the mean percentage of CD4+/TLR-2+ T cells in patients with GD decreased (p < 0.0001). The expression level of TLR-2 on CD4+ T lymphocytes correlated with serum FT3 concentration in patients with GD (r = 0.47, p = 0.007). The mean percentage of CD8+/TLR-2+ T cells in patients with GD before treatment compared to patients with GD after obtaining euthyroidism was higher (p = 0.0163). Similar findings were found for TLR-4. Thus the TLR-2 and TLR-4 can be a prognostic marker for Graves’ disease.

CD4- and CD8-expressing cells found in the bovine and porcine anterior chamber of the eye.

The aim of the present study was to investigate whether the anterior chamber constitutes part of the normal migratory pathway of CD4+ and CD8+ lymphocytes in cattle and swine. The cells obtained from aqueous humor of cows and pigs were stained for CD4 and CD8 receptors, and subsequently analyzed with flow cytometry. The mean percentage of CD4+CD8-, CD4-CD8+ and CD4+CD8+ cells within the total lymphocyte population of the bovine anterior chamber was, respectively, 17.88, 12.64 and 27.26%. In turn, the mean values of these parameters in pigs were 1.77, 38.48 and 17.45, respectively. Among bovine and porcine CD4+CD8+ cells prevalent were those displaying CD4lowCD8low and CD4lowCD8high phenotypes, respectively. The results suggest that the anterior chamber in cattle and swine is an element in the normal migratory pathway of CD4+, CD8+ and CD4+CD8+ cells. Furthermore, the contribution of these subsets in the anterior chamber lymphocyte population can differ considerably between animal species.

CD25+CD127+Foxp3- Cells Represent a Major Subpopulation of CD8+ T Cells in the Eye Chambers of Normal Mice.

The aim of this study has been to determine whether eye chambers constitute part of the normal migratory pathway of naive CD4+ and CD8+ T cells in mouse and if natural CD4+CD25+Foxp3+ and CD8+CD25+Foxp3+ regulatory T cells are present within these eye compartments. To this aim, the cells obtained from aqueous humor (AH) of normal mice were phenotyped in terms of the expression CD4, CD8, CD25, CD127 and transcription factor Foxp3. The mean percentage of CD8+ T cells in the total AH lymphocyte population was as high as 28.69%; the mean percentage of CD8high and CD8low cells in this population was 34.09% and 65.91%, respectively. The presence of cells with the regulatory phenotype, i.e. CD25+Foxp3+ cells, constituted only 0.32% of CD8+ T cell subset. Regarding the expression of CD25, AH CD8+ T cells were an exceptional population in that nearly 85% of these cells expressed this molecule without concomitant Foxp3 expression. Despite having this phenotype, they should not be viewed as activated cells because most of them co-expressed CD127, which indicates that they are naive lymphocytes. With regard to the markers applied in the present research, CD8+CD25+CD127+Foxp3- T cells represent the most numerous subset of AH CD8+ cells. The results suggest that eye chambers in mice are an element in the normal migratory pathway of naive CD8+ T cells. The study presented herein demonstrated only trace presence of CD4+ cells in the eye chambers, as the mean percentage of these cells was just 0.56. Such selective and specific homing of CD8+ and CD4+ cells to the eye chambers is most clearly engaged in the induction and maintenance of ocular immune privilege.

Reactive T cells by flow cytometry distinguish Hodgkin lymphomas from T cell/histiocyte-rich large B cell lymphoma.

Classical Hodgkin lymphoma (CHL), nodular lymphocyte predominant Hodgkin lymphoma (NLPHL), and T-cell/histiocyte rich large B-cell lymphoma (T/HRLBCL) are B-cell lymphomas in which tumor cells are rare as compared with the background reactive infiltrate. We hypothesized that characterization of the reactive infiltrates can provide information to help diagnose these lymphomas. Lymphocyte subsets by flow cytometry were analyzed for 27 NLPHL, 20 T/HRLBCL, 34 CHL, and 49 reactive lymph nodes (RLN). CD4+ T cells with bright expression of CD7 and CD45 (CD3+CD4+CD7(bright) CD45(bright) ), CD4:CD8 ratio, and percentage of T cells, B cells, NK cells, CD4+ T cells, CD8+ T cells, T cells coexpressing CD4, and CD8 (CD4+CD8+), and plasma cells were measured. The CD3+CD4+CD7(bright) CD45(bright) T-cell population was present in the reactive infiltrate of CHL (76.5%) and T/HRLBCL (92.3%) but not in NLPHL (8.3%) or RLN (4.1%). In a separate analysis of 387 samples, the CD3+CD4+CD7(bright) CD45(bright) T-cell population was also observed in some CD10+ B-NHL. The mean percentage of CD4+CD8+ T cells was highest for NLPHL (11.7%) and differed significantly from T/HRLBCL, CHL, and RLN. Interestingly, the highest CD4:CD8 ratios were seen with T/HRLBCL. Finally, the percentage of B cells is decreased in T/HRLBCL relative to CHL and NLPHL. Differences in the reactive inflammatory infiltrate of CHL, NLPHL, and T/HRLBCL and can suggest these diagnoses. Additionally, in contrast to published studies, T/HRLBCL demonstrates both low and very high CD4 to CD8 T-cell ratios. © 2015 International Clinical Cytometry Society.

A role for helminth parasites in achieving immunological tolerance in transplantation

BackgroundHelminths infect more than a quarter of the world's population. Their success as parasites is the result of active immunomodulation of the host immune response, which can have benefits for the host, particularly in suppressing harmful allergic and autoimmune responses. Accordingly, we tested the hypothesis that helminth infection reduces the immune response to allograft transplantation. MethodsC57BL/6 mice were implanted with a subcutaneous minipump that delivered a continuous infusion of secreted products from the model mouse intestinal parasite, Heligmosomoides polygyrus (equivalent to 7 μg of protein per day). Simultaneously, fully allogeneic skin grafts from BALB/c donors were performed. 7 days later, lymphocytes were isolated from allograft draining lymph nodes and analysed by flow cytometry. FindingsFlow cytometric analysis showed a 41·7% increase in the mean percentage of CD4+CD25+Foxp3+ regulatory T cells (of total CD4 cells) from a baseline of 8·1% (95% CI 7·4–8·8) in untreated mice to 11·5% (8·8–14·2) in the treatment group (p=0·0085). Treatment with parasite products also increased mean expression of the regulatory cell surface receptor PD1 by 62·2% in the effector CD4 T-cell population from a baseline of 7·7% (5·7–9·6) to 12·5% (7·5–17·4) (p=0·03). InterpretationThe results show that helminth-derived products can powerfully induce regulatory immunological mechanisms in the presence of a fully allogeneic transplant. Identification of the specific mechanisms involved in suppression of allograft rejection by helminth parasites could lead towards development of safe and effective novel therapeutic strategies. FundingWellcome Trust.

High Percentage of Regulatory T Cells before and after Vitamin B12 Treatment in Patients with Pernicious Anemia

Introduction: In some previous studies, vitamin B₁₂ treatment showed immunomodulatory effects and restored the immunological abnormalities in patients with pernicious anemia (PA). In the present study, peripheral blood T cell subsets, including regulatory T cells (T_regs), were examined before and after vitamin B₁₂ treatment in PA patients. Patients and Methods: The percentages of CD4, CD8, Th1, Th2 and T_regs were examined in 23 PA patients before vitamin B₁₂ treatment, in 23 other PA patients after vitamin B₁₂ treatment and in 28 healthy controls. Results: The mean percentage of CD8+ T cells was significantly higher in the control group (23.0%; 95% CI, 20.4-25.6%) than in the pre- (16.0%; 95% CI, 12.1-20.0%) and posttreatment groups (15.2%; 95% CI, 11.8-18.6%; p < 0.05). The CD4/CD8 ratio was significantly lower in the control group (2.01; 95% CI, 1.66-2.34) than in the pre- (3.45; 95% CI, 2.55-7.80) and posttreatment groups (2.97; 95% CI, 2.22-3.72; p < 0.05). There was no significant difference in the mean Th1/Th2 ratio among these groups. There were significant increases in the mean percentage of T_regs in the pre- (6.29%; 95% CI, 5.04-7.54%) and posttreatment groups (7.77%; 95% CI, 6.34-9.20%) compared with the control group (4.18%; 95% CI, 3.92-4.47%; p < 0.05). Conclusions: The percentage of T_regs was significantly higher in PA patients than in normal subjects, and this high T_reg percentage was not different before and after vitamin B₁₂ treatment. Other immunological alterations also did not recover after vitamin B₁₂ treatment, so that these immunological changes appear to be the cause of PA and are not induced by vitamin B₁₂ deficiency.

Effect of hypotonic 0.18% sodium hyaluronate eyedrops on inflammation of the ocular surface in experimental dry eye.

To investigate the efficacy of hypotonic 0.18% sodium hyaluronate (SH) eyedrops in a mouse model of experimental dry eye (EDE). EDE was induced in C57BL/6 mice by a subcutaneous scopolamine injection and an air draft. The mice were divided into 4 groups according to topical treatment regimens: EDE control, isotonic 0.5% carboxymethycellulose (CMC), isotonic 0.1% SH, and hypotonic 0.18% SH. Tear volume, corneal smoothness, and corneal staining scores were measured at 5 and 10 days of EDE. Multiplex immunobead assay, immunohistochemistry, and flow cytometry for proinflammatory cytokines, chemokines, and inflammatory molecules were performed at 10 days of EDE. The 0.18% SH group had a significantly lower corneal smoothness and staining scores than the 0.5% CMC and 0.1% SH groups at 10 days of EDE (P<0.05). The 0.18% SH group showed significantly low levels of tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, monokine induced by interferon-γ, and interferon-γ-inducible protein 10 compared with the other groups (P<0.05). The mean percentages of CD4(+)CXCR3(+), CD40(+), and CD44(+) cells in the conjunctiva were significantly lower in the 0.18% SH group than in the other groups (P<0.05). In addition, the 0.1% SH group showed lower levels of TNF-α and IL-1β and percentages of CD40(+) and CD44(+) cells than the EDE and 0.5% CMC groups. Hypotonic 0.18% SH eyedrops are more effective in improving ocular surface irregularity and staining and decreasing inflammatory cytokines, chemokines, and cells on the ocular surface compared with isotonic 0.5% CMC or 0.1% SH eyedrops in the treatment of EDE.

Loss of CD28 Expression by Liver-Infiltrating T Cells Contributes to Pathogenesis of Primary Sclerosing Cholangitis

T-cell-mediated biliary injury is a feature of primary sclerosing cholangitis (PSC). We studied the roles of CD28(-) T cells in PSC and their regulation by vitamin D. Peripheral and liver-infiltrating mononuclear cells were isolated from blood or fresh liver tissue. We analyzed numbers, phenotypes, functions, and localization patterns of CD28(-) T cells, along with their ability to activate biliary epithelial cells. We measured levels of tumor necrosis factor (TNF)α in liver tissues from patients with PSC and the effects of exposure to active vitamin D (1,25[OH]2D3) on expression of CD28. A significantly greater proportion of CD4(+) and CD8(+) T cells that infiltrated liver tissues of patients with PSC were CD28(-), compared with control liver tissue (CD4(+): 30.3% vs 2.5%, P < .0001; and CD8(+): 68.5% vs 31.9%, P < .05). The mean percentage of CD4(+)CD28(-) T cells in liver tissues from patients with PSC was significantly higher than from patients with primary biliary cirrhosis or nonalcoholic steatohepatitis (P< .05). CD28(-) T cells were activated CD69(+)CD45RA(-) C-C chemokine receptor (CCR)7(-) effector memory and perforin(+) granzyme B(+) cytotoxic cells, which express CD11a, CX3CR1, C-X3-C motif receptor 6 (CXCR6), and CCR10-consistent with their infiltration of liver and localization around bile ducts. Compared with CD28(+) T cells, activated CD28(-) T cells produced significantly higher levels of interferon γ and TNFα (P < .05), and induced up-regulation of intercellular cell adhesion molecule-1, HLA-DR, and CD40 by primary epithelial cells (3.6-fold, 1.5-fold, and 1.2-fold, respectively). Liver tissue from patients with PSC contained high levels of TNFα; TNFα down-regulated the expression of CD28 by T cells invitro (P<.01); this effect was prevented by administration of 1,25(OH)2D3 (P < .05). Inflammatory CD28(-) Tcells accumulate in livers of patients with PSC and localize around bile ducts. The TNFα-rich microenvironment of this tissue promotes inflammation; these effects are reversed by vitamin D invitro.

Immunological Characterization of Compensatory Anti-Inflammatory Response Syndrome in Patients With Severe Sepsis

To perform a complete immunological characterization of compensatory anti-inflammatory response syndrome in patients with sepsis and to explore the relationship between these changes and clinical outcomes of 28-day mortality and secondary infections. Prospective single-center study conducted between April 2011 and December 2012. ICUs from Hospital Universitario San Vicente Fundación at Medellin, Colombia. One hundred forty-eight patients with severe sepsis. None. At days 0, 1, 3, 5, 10, and 28, we determined the expression of HLA-DR in monocytes and the apoptosis and the proliferation index in T lymphocytes, as well as the levels of tumor necrosis factor-α, interleukin-6, interleukin-1β, interleukin-10, and transforming growth factor-β in both plasma and cell culture supernatants of peripheral blood mononuclear cells. The mean percentage of HLA-DR was 60.7 at enrollment and increased by 0.9% (95% CI, 0.7-1.2%) per day. The mean percentage of CD4 T cells and CD8 T cells AV+/7-AAD- at enrollment was 37.2% and 20.4%, respectively, but it diminished at a rate of -0.5% (95% CI, -0.7% to -0.3%) and -0.3% (95% CI, -0.4% to -0.2%) per day, respectively. Plasma levels of interleukin-6 and interleukin-10 were 290 and 166 pg/mL and decreased at a rate of -7.8 pg/mL (95% CI, -9.5 to -6.1 pg/mL) and -4 pg/mL (95% CI, -5.1 to -2.8 pg/mL) per day, respectively. After controlling for confounders, only sustained plasma levels of interleukin-6 increase the risk of death (hazard ratio 1.003; 95% CI, 1.001-1.006). We found no evidence to support a two-phase model of sepsis pathophysiology. However, immunological variables did behave in a mixed and time-dependent manner. Further studies should evaluate changes over time of interleukin-6 plasma levels as a prognostic biomarker for critically ill patients.

Peripheral blood lymphocyte subsets (CD4+, CD8+ T cells), leptin level and weight loss after laparoscopic greater curvature plication in morbidly obese patients.

IntroductionThe aim of the study was to detect the effect of laparoscopic greater curvature plication (LGCP) on peripheral blood lymphocyte subsets (helper and cytotoxic T lymphocytes – CD4+ and CD8+ T cells respectively), leptin level and weight loss in morbidly obese patients.Material and methodsMorbidly obese patients (n = 20, age range: 25–50 years, body mass index (BMI) range: 37–45 kg/m2) who underwent LGCP were enrolled in a prospective study to determine the percentages of their peripheral blood T cells (CD4+ and CD8+) before and 4 months postoperatively using flow cytometry. Also, the level of their leptin before and 4 months postoperatively was established using enzyme-linked immunosorbent assay (ELISA). The data are expressed as the percentage of total lymphocytes ± the standard error of the mean.ResultsA decrease in the BMI and loss of weight (31.20 ±1.2%) were confirmed 4 months postoperatively since BMI was 44.71 ±4.3 (range: 37–45) kg/m2 preoperatively, and decreased to 31.80 ±1.1 (range: 24–33) kg/m2 after surgery. The mean percentage of CD4+ and CD8+ T lymphocytes significantly decreased postoperatively (38.2 ±1.5 before and 29.3 ±2.6 after operation for CD4+, 17.3 ±1.8 preoperatively and 9.5 ±1.7 postoperatively for CD8+, p < 0.05). The mean leptin level was 43.01 ±22.01 preoperatively while postoperatively it was 24.8 ±11.1 (p < 0.05), so the leptin level substantially decreased compared to its preoperative values.ConclusionsThis study found that weight loss after LGCP in morbidly obese patients led to decreases in levels of leptin and circulating immune cells compared to their preoperative values.

AB0046 Cytokine profile in CD4+ FOXP3+ lymphocytes of RA patients treated with tocilizumab and clinical correlation (DAS28-CPR, rheumatoid factor

BackgroundThe main group of suppressor cells/regulators of the immune response (Treg cells), usually co-expressed different combinations of antigens CD4, CD25, and FoxP3, among other markers (1).A decrease in the number-function...

The effect of laparoscopic greater curvature plication on peripheral blood lymphocyte subsets (CD4+, CDS+ T cells) in morbidly obese patients

Background: Bariatric surgery is the only and effective treatment for morbid obesity and it can also improve the obesity-related comorbidities. However, the effect ofbariatric surgery on immune status is still unclear. In our study we investigated the relationship between surgical weight loss and peripheral blood lymphocyte percentages. Methods: Morbidly obese patients (n=20, age range 25-50 years, body mass index [BMIJ range (37-45kgfm2) who had undergone laparoscopic greater curvature plication LGCP (were enrolled in a prospective study to determine the percentages of their peripheral blood T cells (CD4+, and CD8+ T cells) before and (4 months) postoperatively using flow cytometry. The data were expressed as the percentage oftotallymphocytes±the standard error ofthe mean. Results: A decrease in the BMI occurred at 4months postoperatively with loss of weight of (31.20±1.2%). PreoperativeBMJwas 44.71±4.3 (range 37-45kg fm2)and postoperative EM!was 31.80±1.1(range 24-33kg fm2).The mean percentage ofCD4+ T lymphocytes preoperatively was 38.2±1.5 and postoperatively was 29.3±2.6 p <0.05 (which is statistically significant) so there is postoperative decrease in the percentage ofCD4+T lymphocytes, the mean percentage ofCD8+ T lymphocytes preoperatively was 17.3±1.8 and postoperatively was 9.5±1.7p < 0.05 (which is statistically significant) so there is postoperative decrease in the percentage ofCD8+ T lymphocytes. Conclusion: This study found that weight loss after LGCP in morbidly obese patients showed attenuated activation of circulating immune cells (decrease in CD4+ T helper cells percentage and decrease in CD8+ killer T cells percentage) and more regulation of chronic inflammation in morbidly obese patients.

Evaluation of Natural Regulatory T Cells in Subjects with Selective IgA Deficiency: From Senior Idea to Novel Opportunities

Background: Selective IgA deficiency (SIgAD) is the most common primary immunodeficiency disorder, which is characterized by significantly decreased serum levels of IgA. Abnormalities of CD4+CD25^highforkhead box P3 (FoxP3)+ regulatory T cells (T_reg) have been shown in association with autoimmune and inflammatory disorders. Methods: In order to evaluate the relationship between autoimmunity and T_reg in SIgAD, we studied 26 IgA-deficient patients (aged 4–17 years) with serum IgA levels <7 mg/dl, 26 age- and sex-matched healthy controls and 26 age- and sex matched idiopathic thrombocytopenic purpura cases with normal immune system. T_reg were determined by flow cytometry using T_reg markers, including CD4, CD25 and FoxP3. Results: The mean percentage of CD4, CD25+FoxP3+ T_reg from all CD4+ cells was 4.08 ± 0.86 in healthy controls, which was significantly higher than in SIgAD patients (2.93 ± 1.3; p = 0.003). We set a cutoff point (2.36%) for T_reg, which was two standard deviations lower than the mean of normal controls. According to this cutoff point and in order to assess the role of T_reg in clinical SIgAD manifestation, we classified patients into two groups: 16 patients in G1 with T_reg <2.36% and 10 patients in G2 with T_reg >2.36%. Autoimmunity was recorded in 9 patients (53.3%) of G1 and only 1 patient of G2, respectively (p = 0.034). Although a defect in class switching recombination was observed in 40% of the patients in G1, none of the G2 patients had such a defect (p = 0.028). Conclusion: This study showed decreased proportions of T_reg in SIgAD patients, particularly in those with signs of chronic inflammation.

CD8+CD28− Lymphocytes in Peripheral Blood and Serum Concentrations of Soluble Interleukin 6 Receptor are Increased in Patients with Graves’ Orbitopathy and Correlate with Disease Activity

Background. The extrathyroid, orbital manifestation of Graves’ disease (GD)—Graves’ orbitopathy (GO)—presents a difficult clinical problem. The immunological status of GO patients is still under investigation. The aim of this study was to assess the serum concentration of interleukin 6 (IL-6), soluble interleukin 6 receptor (sIL-6R), and CD8+CD28− lymphocytes in GO patients and to evaluate if these parameters were associated with disease activity. Patients. Thirty-nine patients (29 women and 10 men, aged 24–71, mean 50.18) with newly diagnosed GD were enrolled in the study. Active GO was diagnosed in 20 patients. The control group included 12 healthy individuals. Methods. Serum concentrations of IL-6 and sIL-6R were estimated by ELISA. Percentages of CD8+CD28− lymphocytes in peripheral blood were assessed by flow cytometry. Results. Mean serum IL-6 and sIL-6R concentrations were significantly higher in all GD patients and in GO and non-GO patients than in normal controls. In all GD patients and the non-GO group, serum IL-6 and sIL-6R concentrations were significantly reduced after efficient treatment. In GO patients, only serum sIL-6R concentration was significantly lower after efficient treatment. In all GD patients, the mean percentage of CD8+CD28− lymphocytes was significantly lower after efficient treatment. In GO patients, the mean percentage of CD8+CD28− lymphocytes was significantly higher than in the non-GO group or in normals. Moreover, in the GO group, the mean percentage of CD8+CD28− lymphocytes was significantly lower after treatment. Conclusion. Our results have shown that CD8+CD28− lymphocyte percentage in peripheral blood and serum concentration of sIL-6R are increased in GO patients and correlate with disease activity.