Compact bone-derived mesenchymal stem cells attenuate nonalcoholic steatohepatitis in a mouse model by modulation of CD4 cells differentiation

Increasing evidence has accrued which indicates that mesenchymal stem cells (MSCs) have a potential clinical value in the treatment of certain diseases. Globally, nonalcoholic steatohepatitis (NASH) is a widespread disorder. In the present study, MSCs were isolated successfully from compact bone and a mouse model of NASH was established as achieved with use of a methionine-choline deficient (MCD) diet. Compact bone-derived MSCs transplantation reduced MCD diet-induced weight loss, hepatic lipid peroxidation, steatosis, ballooning, lobular inflammation and fibrogenesis. It was shown that MSCs treatment hampered MCD diet-induced proliferation of CD4⁺ IFN-γ⁺ and CD4⁺ IL-6⁺ T spleen cells. In addition, CD4⁺ IL-17⁺ lymphocytes that associated with anti-inflammation show little change in MCD as well as in MCD + MSCs splenocytes. We conclude that MSCs may have a potential clinical value upon NASH, through their capacity to suppress activation of CD4⁺ IFN-γ⁺ and CD4⁺ IL-6⁺ lymphocytes.     

Changes in regulatory T cells in patients with ovarian cancer undergoing surgery: Preliminary results

Regulatory T cells (Treg) suppress immune responses in patients with cancer. Surgery is the most effective therapeutic strategy for ovarian cancer (OC). However, the interplay between the Treg population and surgical resection remains unclear. 61 patients with OC who received no prior treatment were enrolled in the study. Treg percentages were characterized from peripheral blood mononuclear cells. We investigated CD4⁺ CD25⁺, CD4⁺ CD25⁺ Foxp3⁺, CD8⁺ CD28⁻, and CD8⁺ Foxp3⁺ Tregs in OC patients and their postoperative changes using flow cytometry. Treg percentages were significantly higher in OC patients than those in benign ovarian tumors (BOT) and healthy controls. Higher percentages of Tregs were found in patients with stage III/IV than stage I/II OC. Treg percentages were significantly decreased postoperatively. The postoperative Treg percentages in patients with stage I/II OC were similar to those in BOT patients, while postoperative Treg percentages in patients with stage III/IV OC remained higher. Tregs were markedly lower on postoperative day (POD) 3 than preoperatively. They increased slightly after 7 days, but remained lower than preoperative levels. These data suggested that Tregs continued to decline from POD 7 to POD 30. Treg percentages are correlated with the tumor burden and could be a key factor in monitoring the immunological status of patients with OC.     

Inosine Acedoben Dimepranol promotes an early and sustained increase in the natural killer cell component of circulating lymphocytes: A clinical trial supporting anti-viral indications

Inosine Acedoben Dimepranol (IAD), licensed for the treatment of cell-mediated immune deficiencies associated with viral infections, has been reported to impact a variety of immune parameters both in vitro and in vivo. Here we report the results from a clinical trial where multiple lymphocyte subsets – CD19 + B cells, CD3 + T cells, CD4 + T-helper cells, FoxP3^hi/CD25^hi/CD127^lo regulatory T cells (Tregs), CD3 −/CD56 + NK cells, and CD3 +/CD56 + NKT cells – were, together with serum immunoglobulins and IgG subclasses, followed during 14 days of IAD administration to ten healthy volunteers; these selected from 27 individuals pre-screened in vitro for their capacity to respond to IAD as gauged by increases in the percentage of Treg and/or NKT cells arising in PHA-stimulated cultures. While a transient spike and dip in Treg and T-helper fractions, respectively, was noted, the outstanding consequence of IAD administration (1 g po, qds) was an early and durable rise in NK cells. For half the cohort, NK cells increased as a percentage of total peripheral blood lymphocytes within 1.5 h of receiving drug. By Day 5, all but one of the volunteers displayed higher NK cell percentages, such elevation – effectively a doubling or greater – being maintained at termination of study. The IAD-induced populations were as replete in Granzyme A and Perforin as basal NK cells. The novel finding of IAD boosting phenotypically competent NK numbers in healthy individuals supports the drug's indicated benefit in conditions associated with viral infection and reinforces the potential for uplift where immune performance may be compromised.     

Soluble serum HLA-G and HLA-A, -B, -C molecules in patients with seasonal allergic rhinitis exposed to pollens

BackgroundAllergic rhinitis (AR) is characterized by a Th2 polarized immune response and soluble HLA (sHLA) molecules play an immunomodulatory role in this response. Previously, it has been reported that these molecules are increased in sera of patients with pollen-induced allergic rhinitis studied outside the pollen season. To date, however, no study has investigated there in AR patients during the pollen season.ObjectiveThe aim of this study was to evaluate serum sHLA-G and sHLA-A, -B, -C levels in both AR patients and healthy controls.Methods60 symptomatic allergic patients were enrolled. A group of 50 healthy subjects was included as a control. Serum sHLA-G and sHLA-A, -B, -C levels were determined by an immunoenzymatic method. Allergy severity was assessed by VAS for symptoms and drug use.ResultsAllergic patients had significantly higher levels of both sHLA-G (p < 0.001) and sHLA-A, -B, -C (p = 0.001) than normal controls. In addition, there was a very strong correlation between sHLA-G levels and clinical severity.ConclusionThe present study confirms evidence that serum sHLA-G and sHLA-A, -B, -C molecules are significantly increased in patients with pollen-induced AR also during the pollen season. Moreover, sHLA-G might be considered as a biomarker for assessing clinical severity.     

Modulation of regulatory T cells by intranasal allergen immunotherapy in an experimental rat model of airway allergy

Allergic airway diseases such as asthma and allergic rhinitis are increasing in prevalence worldwide. The theory of an altered Th1/Th2 balance in allergic diathesis has recently been termed a “procrustean paradigm” as it failed to explain many preclinical findings. Regulatory T cells (Treg) have now been shown to be critical in T-cell homeostasis and in the maintenance of peripheral tolerance to allergens. Allergen specific immunotherapy (SIT) has been shown to induce regulatory T cells in allergic patients. Among various types of SIT, intranasal immunotherapy had not been studied in detail for the treatment of allergic airway diseases. So, there was a need to study the contribution of regulatory T cells and their mechanistic pathways following intranasal immunotherapy in-vivo. It had been previously shown that intranasal allergen immunotherapy using Alstonia scholaris pollen extract abrogates allergic airway inflammation with decline in IgE and Th2 cytokine levels. The present study for the first time offers a multi-targeted approach towards attenuation of airway allergy by the generation of CD4 + CD25 + Foxp3 + T cells and other subsets of Treg cells like Tr1 cells, Th3 cells, CTLA4 + Treg cells, and also modulation of various Treg cell surface molecules like GITR, OX40, CD39 and CD73 by intranasal immunotherapy in the same animal model. This animal experiment will thus help to chart out newer molecular targets for treating allergic asthma or rhinitis.     

CXCR5+ CD8+ T cells could induce the death of tumor cells in HBV-related hepatocellular carcinoma

The follicular CXCR5⁺ CD8⁺ T cells have recently emerged as a critical cell type in mediating peripheral tolerance as well as antiviral immune responses during chronic infections. In this study, we investigated the function of CXCR5⁺ CD8⁺ T cells in HBV-related hepatocellular carcinoma patients. Compared to CXCR5⁻ CD8⁺ T cells, CXCR5⁺ CD8⁺ T cells presented elevated PD-1 expression but reduced Tim-3 and CTLA-4 expression. Upon anti-CD3/CD28 stimulation, CXCR5⁺ CD8⁺ T cells demonstrated higher proliferation potency than CXCR5⁻ CD8⁺ T cells, especially after PD-1 blockade. CXCR5⁺ CD8⁺ T cells also demonstrated significantly higher granzyme B synthesis and release, as well as higher level of degranulation. Tumor cells were more readily eliminated by CXCR5⁺ CD8⁺ T cells than by CXCR5⁻ CD8⁺ T cells. Interestingly, we found that B cells were more resistant to CXCR5⁺ CD8⁺ T cell-mediated killing than tumor cells, possibly through IL-10-mediated protection. In addition, the CXCR5⁺ CD8⁺ T cell-mediated cytotoxic effects on tumor cells could be significantly enhanced by PD-L1 blockade. Together, we presented that in patients with in HBV-related hepatocellular carcinoma, CXCR5⁺ CD8⁺ T cells could mediate tumor cell death more potently than the CXCR5⁻ CD8⁺ T cells in vitro while the autologous B cells were protected.     

A higher frequency of CD4+ CXCR5+ T follicular helper cells in patients with newly diagnosed Henoch–Schönlein purpura nephritis

T follicular helper (TFH) cells play an important role in the humoral immune responses. The aim of this study was to examine the frequency of different subsets of CD4⁺ CXCR5⁺ TFH cells and B cells in patients with new-onset Henoch–Schönlein purpura nephritis (HSPN). The numbers of different subsets of CD4⁺ CXCR5⁺ TFH cells, B cells and the constituents of serum cytokines were detected in a total of 25 patients with newly diagnosed HSPN before and after treatment, and in 14 healthy controls (HC). The potential connection of these cells with the clinical characteristics in HSPN patients was analyzed. The numbers of circulating CD4⁺ CXCR5⁺, CD4⁺ CXCR5⁺ ICOS⁺ and CD4⁺ CXCR5⁺ PD-1⁺ TFH cells, CD86⁺ CD19⁺, CD38⁺ CD19⁺ B cells and serum IL-2, IL-4, IL-17A, IL-21 and IFN-γ were significantly higher in HSPN patients (p < 0.05) than in HC. Before and after treatment the numbers of CD4⁺ CXCR5⁺ TFH cells were negatively correlated with the values of eGFR (r = − 0.7162, p < 0.05; r = − 0.732, p < 0.05, respectively). Similarly the numbers of CD4⁺ CXCR5⁺ PD-1⁺ TFH cells were negatively correlated with 24-h urinary proteins (r = − 0.4013, p < 0.05; r = − 0.7857, p < 0.05, respectively), and the numbers of CD4⁺ CXCR5⁺ ICOS⁺ TFH cells were positively correlated with the levels of serum IL-21 (r = 0.5186, p < 0.05; r = 0.8503, p < 0.05, respectively) and 24-h urinary protein (r = 0.6045, p < 0.05; r = 0.833, p < 0.05, respectively) in these patients, regardless of treatment. Following treatment the numbers of CD4⁺ CXCR5⁺, CD4⁺ CXCR5⁺ PD-1⁺, and CD4⁺ CXCR5⁺ ICOS⁺ TFH cells, as well as serum levels of IL-21 were significantly reduced, however IL-4 levels were noticeably increased (p < 0.05). A higher frequency of circulating CD4⁺ CXCR5⁺ TFH cells existed in patients with HSPN and may be a viable therapeutic target.     

Gabapentin-induced changes of plasma cortisol level and immune status in hysterectomized women

AimWe have examined the effects of gabapentin (GBP) on stress-related changes of cortisol and catecholamines in patients who underwent hysterectomy because of uterine fibrinoids. Additionally, we have observed the effect of GBP on the immune status in the acute stress response to surgery.MethodsSixty patients scheduled for an abdominal hysterectomy were randomly assigned to the GBP administration 1 h before surgery (n = 30 pts), or to the placebo group (n = 30 pts). Blood samples were collected before and 24 h after the surgery. The intensity of pain was assessed by a visual analogue scale (VAS) every 8 h at rest. Immunomodulatory effects of GBP were determined by flow cytometry. We followed the total proportion of CD3⁺ lymphocytes, CD3⁺CD4⁺, CD3⁺CD8⁺, CD19⁺ B lymphocytes, CD16⁺CD56⁺CD3⁻NK cells and CD16⁺CD56⁺CD3⁺ NKT cells before and 24 h after hysterectomy. The plasma cortisol and catecholamines concentration was used to estimate the level of the stress response.ResultsVAS pain score at rest was significantly lower in the GBP group than in the placebo group (P = 0.003). Application of GBP significantly decreased the plasma cortisol level 24 h after the operation in comparison to the placebo group (P < 0,001). We found significant positive correlation between the VAS pain score and concentration of cortisol in all patients (P = 0.025). GBP reduced the concentration of catecholamines (p < 0.05). The proportion of CD3⁺ (P = 0.027) and CD3⁺CD4⁺cells (P = 0.006) was significantly lower in the GBP group 24 h after operation, while the contribution of CD19⁺ (P = 0.033) was significantly higher.ConclusionPreoperative administration of GBP reduced the pain scores at rest in patients at 0, 16 and 24 h after abdominal hysterectomy. Additionally, GBP reduced the stress response and changed immune parameters in the reaction to surgery.     

Neutrophil-to-lymphocyte ratio (NLR) and platelet-to-lymphocyte ratio (PLR) were associated with disease activity in patients with systemic lupus erythematosus

ObjectiveNeutrophil-to-lymphocyte ratio (NLR) and platelet-to-lymphocyte ratio (PLR) have recently been investigated as two new inflammatory markers used in the assessment of systemic inflammation in many diseases. The purpose of the study was to investigate their relation with disease activity in newly diagnosed SLE patients.MethodsThe study population consisted of 116 SLE patients who did not receive any treatment and 136 healthy controls. We divided the patients into two groups according to the SLE Disease Activity Index 2000 (SLEDAI-2K) system. Group 1 included patients with a score of 9 and lower (patients with mild disease activity), and Group 2 included patients with a score of > 9 (patients with severe disease activity). Correlations between NLR, PLR and disease activity were analyzed.ResultsThe NLR and PLR of SLE patients were significantly higher compared to those of the controls (both P < 0.001). There was a statistically significant difference in NLR and PLR between Group 1 and Group 2 (both P < 0.05). SLEDAI scores positively correlated with NLR (r = 0.312, P < 0.001) and PLR (r = 0.298, P < 0.001). Furthermore, SLE patients with nephritis had higher NLR levels than those without nephritis (P = 0.027). Based on the ROC curve, the best NLR cut-off value to predict SLE patients with severe disease activity was 2.26, with 75% sensitivity and 50% specificity, whereas the best PLR cut-off value was 203.85, with 42.3% sensitivity and 83.9% specificity.ConclusionNLR and PLR were two useful inflammatory markers for assessment of disease activity in patients with SLE.     

Sublingual immunotherapy reduces soluble HLA-G and HLA-A,-B,-C serum levels in patients with allergic rhinitis

Allergic rhinitis (AR) is characterized by Th2 polarized immune response. Soluble HLA (sHLA) molecules play an immunomodulatory activity. Specific immunotherapy is the only causal treatment for AR. So far no study investigated the effect of sublingual immunotherapy (SLIT) on sHLA molecules.The aim of the study was to evaluate sHLA-G and sHLA-A,-B,-C serum levels in AR patients with pollen allergy before and after a pre-seasonal course of SLIT. Forty AR patients with pollen allergy were enrolled and they assumed a pre-seasonal SLIT course for 3 months. Serum sHLA-G and sHLA-A,-B,-C and IFN-γ and IL-4 levels were determined by ELISA method at baseline and 3 months after the end of the SLIT course. Symptoms severity was assessed by a Visual Analogue Scale.Both sHLA-G and sHLA-A,-B,-C levels significantly diminished (p < 0.0001 for both) after SLIT. Moreover, there was a highly significant relationship between the serum levels of these two soluble molecules (r = 0.84). Significant relationship between symptoms evaluated by VAS and change of sHLA molecules was also evidenced (r = 0.60 and 0.63). Serum cytokines were not affected by SLIT.Therefore, this preliminary study provides the first evidence that both sHLA-G and sHLA-A,-B,-C levels are significantly reduced by SLIT in AR patients with pollen allergy. Therefore, the clinical implication of this study is that these soluble molecules might be interpreted as biomarker of response to SLIT.     

P-glycoprotein function in peripheral T lymphocyte subsets of myasthenia gravis patients: Clinical implications and influence of glucocorticoid administration

Myasthenia gravis (MG) is an autoimmune neuromuscular disorder with a chronic clinical course that requires long-term glucocorticoid (GC) therapy. A drug efflux pump, P-glycoprotein (P-gp), actively transports GC out of target cells, thereby reducing its efficacy. We evaluated the P-gp function of peripheral-blood mononuclear cells in 59 MG patients. P-gp function was estimated from a decrease in fluorescent P-gp substrate Rhodamine 123 and its inhibition by the conformation-sensitive UIC2 monoclonal antibody. P-gp function on CD8⁺ T cells in 21 MG patients having experienced GC therapy was higher than that in 19 MG patients having no history of GC therapy (p = 0.026). There was a significant correlation between P-gp function in CD3⁺ (r = 0.55, p = 0.014) or CD4⁺ (r = 0.48, p = 0.034) T cells and the total dose of prednisolone for treatment. P-gp function on CD4⁺ T cells in MG patients who showed low responses to prednisolone therapy (n = 8) was higher than that in patients who showed relatively high responses to prednisolone therapy (n = 10) (p = 0.045). These results suggest that higher P-glycoprotein activity on CD3⁺ or CD4⁺ cells necessitated treatment with higher steroid doses in order to achieve a clinical response. The measurement of P-gp function on CD4⁺ T cells is useful in the assessment of clinical response to GC therapy.     

Changes of CD4+CD25+FOXP3 + and CD8+CD28 − regulatory T cells in non-small cell lung cancer patients undergoing surgery

Little is known about the regulatory T cells (Tregs) in the peripheral blood after surgery of non-small cell lung cancer (NSCLC) patients. In this study, we investigated whether CD4+CD25+FOXP3 + and CD8+CD28 − regulatory T cells are decreased in the peripheral blood of NSCLC patients undergoing surgery. The study group (n = 49) comprised NSCLC, and the control group (n = 24) consisted of age- and sex-matched nonmalignant diseases. The prevalence of CD4+CD25+FOXP3 + and CD8+CD28 − Tregs was analyzed using flow cytometry. The study group showed significantly higher percentage of CD4+CD25+FOXP3 + and CD8+CD28 − Tregs than control. The percentage of CD4+CD25+FOXP3 + and CD8+CD28 − Tregs increased with tumor stage. One way ANOVA test shows the significant differences between all subgroups. LSD test shows that there was a statistical significance between each of the two subgroups except stage II in CD4+CD25+FOXP3 + Tregs and control vs. each stage, stage I vs. stage III, and stage IV in CD8+CD28 − Tregs. There is no significant difference among stages II, III, and IV in CD8+CD28 − Tregs. No differences were found between squamous carcinoma and adenocarcinoma. These levels were dropped significantly after operation. Furthermore postoperative Treg percentage in the early stages (stage I and stage II) was not statistically different from that of controls. Postoperative Treg percentage in advanced stage (III + IV) remained above the values shown by controls. Our findings indicate that the percentage of CD4+CD25+FOXP3 + and CD8+CD28 − Tregs correlated with the pathological stage in NSCLC and tumor burden.     

Reversing the polarization of tumor-associated macrophages inhibits tumor metastasis

ObjectiveThe M2 phenotype is dominant in tumor associated macrophages (TAM), and plays a key role in promoting tumor growth, invasion and metastasis. Converting TAM polarization from M2 to M1 may contribute to eliciting anti-tumor-specific immune responses and inhibiting tumor metastasis. In this study, the effect of reversing the polarization of TAM on tumor metastasis was investigated.MethodsPeritoneal macrophages were obtained from BABL/c mice, and M2 polarization was induced by IL-4. In an in vivo experiment, BABL/c mice were transplanted with 4 T1 tumor cells. In vitro and in vivo experimental studies, M2 macrophage polarization was reversed with CpG-DNA or CpG-DNA combined with anti-IL-10R Ab. CD68, MHCII and FRβ molecular expression in macrophages were examined with immunofluorescence staining. The mRNA expression of IL-2, IL-6, IL-13, VEGF and MMP-9 were detected with RT-PCR. VEGF and MMP-9 protein expression of tumors in situ was measured by western blot assay. Lung-metastasis of the tumor was observed and assessed by micro-CT.ResultsCpG-DNA and CpG-DNA combined with anti-IL-10R Ab could promote MHCII, IL-2, IL-6 and IL-13 molecular expression, and suppress the expression of FRβ, MMP-9 and VEGF, in both freshly isolated peritoneal macrophages and M2 macrophages. In the CpG-DNA combined with anti-IL-10R Ab injecting group, the percentage of CD68⁺ MHCII⁺ cells were significantly higher than that of CD68⁺ FRβ⁺ cells (P < 0.05). This was distinct from the result of the control group, which CD68⁺ FRβ⁺ was higher than CD68⁺ MHCII⁺ cells (P < 0.01). Furthermore, VEGF-A and MMP-9 level in primary tumor tissues in the experimental group was significantly lower (P < 0.01), compared to the control group. Moreover, the number of detectable lung-metastasis foci was significantly lower in the experimental group than in the control group (P < 0.05).ConclusionReversing the polarization of TAM from M2 to M1 phenotype can inhibit tumor metastasis.     

Influence of anticancer agents on cell survival,proliferation, and CD4+CD25+Foxp3+ regulatory T cell-frequency in human peripheral-blood mononuclear cells activated by T cell-mitogen

Many anticancer agents currently used are considered to be cytotoxic not only to cancer cells but also to functional immune cells. To learn more about the immunosuppressive adverse influence of chemotherapeutic drugs in cancer chemotherapy, we examined the effects of arsenic trioxide, dacarbazine, 5-fluorouracil, and methotrexate on the survival, proliferation, cytokine production, and CD4⁺CD25⁺Foxp3⁺ regulatory T (Treg) cell-frequency in human peripheral blood mononuclear cells (PBMCs) activated by T cell mitogen in vitro. Arsenic trioxide, dacarbazine, and 5-fluorouracil increased trypan-blue stained (dead) cell rates and suppressed the mitogen-activated proliferation of PBMCs significantly at 1–100 μM (p < 0.05). Methotrexate also significantly increased the percentages of dead cells and suppressed the mitogen-activated PBMC-proliferation at concentrations of more than 0.05 μM (p < 0.01). Arsenic trioxide significantly inhibited the production of interferon γ, interleukin (IL)-4, -6, and -10 from the activated PBMCs at 5 μM (p < 0.05). In contrast, the anticancer agents significantly increased Treg cell-frequency in the activated PBMCs at concentrations of more than 0.1 μM for methotrexate, 5 μM for arsenic trioxide and 5-fluorouracil, and 50 μM for dacarbazine, respectively (p < 0.05). These agents did not significantly influence the production of transforming growth factor (TGF) β from the activated PBMCs at a concentration range of 0.05–50 μM. Our data suggest that the anticancer agents: arsenic trioxide, dacarbazine, 5-fluorouracil, and methotrexate attenuate T cell mediated immunity by not only inhibiting the proliferative response of T cells but by also increasing the frequency of Treg cells, which may result in the suppression of the effector T cell function.     

Circulating CD4+ CXCR5+ T cells contribute to proinflammatory responses in multiple ways in coronary artery disease

Coronary artery disease (CAD) is a common subtype of cardiovascular disease. The major contributing event is atherosclerosis, which is a progressive inflammatory condition resulting in the thickening of the arterial wall and the formation of atheromatous plaques. Recent evidence suggests that circulating CD4⁺ CXCR5⁺ T cells can contribute to inflammatory reactions. In this study, the frequency, phenotype, and function of circulating CD4⁺ CXCR5⁺ T cells in CAD patients were examined. Data showed that circulating CD4⁺ CXCR5⁺ T cells in CAD patients were enriched with a PD-1⁺ CCR7⁻ subset, which was previously identified as the most potent in B cell help. The CD4⁺ CXCR5⁺ T cells in CAD patients also secreted significantly higher levels of IFN-γ, IL-17A, and IL-21 than those from healthy controls. Depleting the PD-1⁺ population significantly reduced the cytokine secretion. Interestingly, the CD4⁺ CXCR5⁺ PD-1⁻ T cells significantly upregulated PD-1 following anti-CD3/CD28 or SEB stimulation. CD4⁺ CXCR5⁺ T cells from CAD patients also demonstrated more potent capacity to stimulate B cell inflammation than those from healthy individuals. The phosphorylation of STAT1 and STAT3 were significantly higher in B cells incubated with CD4⁺ CXCR5⁺ T cells from CAD than controls. The IL-6 and IFN-γ expression were also significantly higher in B cells incubated with CD4⁺ CXCR5⁺ T cells from CAD. Together, this study demonstrated that CAD patients presented a highly activated CD4⁺ CXCR5⁺ T cell subset that could contribute to proinflammatory responses in multiple ways. The possibility of using CD4⁺ CXCR5⁺ T cells as a therapeutic target should therefore be examined in CAD patients.     

CXCR5+ CD8+ T cells present elevated capacity in mediating cytotoxicity toward autologous tumor cells through interleukin 10 in diffuse large B-cell lymphoma

Diffuse large B-cell lymphoma (DLBCL) is a common and aggressive subtype of non-Hodgkin's lymphomas, with limited treatment options in refractory and relapsed patients. Growing evidence supports the notion that CD8⁺ T cell immunity could be utilized to eliminate B cell lymphomas. CXCR5⁺ CD8⁺ T cell is a novel cell subtype and share CXCR5 expression with CD19⁺ tumor cells. In this study, we investigated the frequency and function of existing CXCR5⁺ CD8⁺ T cells in DLBCL patients. We found that DLBCL patients as a group demonstrated significantly higher level of CXCR5⁺ CD8⁺ T cells than healthy individuals, with huge variability in each patient. Using anti-CD3/CD28-stimulated CD8⁺ T cells as effector (E) cells and autologous CD19⁺ tumor cells as target (T) cells, at high E:T ratio, no difference between the intensities of CXCR5⁺ CD8⁺ T cell- and CXCR5⁻ CD8⁺ T cell-mediated cytotoxicity were observed. However, at intermediate and low E:T ratios, the CXCR5⁺ CD8⁺ T cells presented stronger cytotoxicity than CXCR5⁻ CD8⁺ T cells. The expressions of granzyme A, granzyme B, and perforin were significantly higher in CXCR5⁺ CD8⁺ T cells than in CXCR5⁻ CD8⁺ T cells, with no significant difference in the level of degranulation. Tumor cells in DLBCL were known to secrete high level of interleukin 10 (IL-10). We therefore blocked the IL-10/IL-10R pathway, and found that the expressions of granzyme A, granzyme B, and perforin by CXCR5⁺ CD8⁺ T cells were significantly elevated. Together, these results suggest that CXCR5⁺ CD8⁺ T cells are potential candidates of CD8⁺ T cell-based immunotherapies, could mediate elimination of autologous tumor cells in DLBCL patients, but are also susceptible to IL-10-mediated suppression.     

Suppressive effect of β,β-dimethylacryloyl alkannin on activated dendritic cells in psoriasis by the TLR7/8 pathway

β,β-dimethylacryloyl alkannin (DMA) is a key component of Lithospermum and possesses good efficacy for treating psoriasis. DMA inhibits activated dendritic cells (DCs), but the mechanism is unknown. Therefore, this study aimed to explore the modulation of the TLR7/8 pathway by DMA in psoriasis-activated DCs. Models of psoriasis-like skin lesions were established using BALB/c mice; 8 mice were treated with DMA (2.5 mg/kg). Bone marrow cells were isolated and induced into DCs using R848, a TLR7/8 agonist. Splenic CD11c + cells were detected by flow cytometry. Skin CD11c + cells were detected by immunofluorescence. TLR7, TLR8, MYD88, and IRAKM proteins were detected by Western blot. The effects of DMA on surface molecules of DCs were observed by flow cytometry. mRNA expression of inflammatory factors was detected by qRT-PCR. Secreted cytokines were detected by cytometric bead array. Compared with the model group, psoriasis-like skin lesions were alleviated by DMA, the splenic CD11c + cells were significantly decreased (P < 0.01), and CD11c + cell numbers in skin lesions were decreased (P < 0.01). Expression levels of TLR7, MYD88, and IRAKM were significantly decreased (P < 0.05). R848-stimulated DCs showed increased expression of I-A/I-E, CD80, and CD86 (P < 0.01), increased IL-23 and IL-1β mRNA and secretion (P < 0.05), and increased TLR7, TLR8, MYD88, and IRAKM expression (P < 0.01); DMA inhibited all of these effects of the TLR7/8 pathway activation by R848 (P < 0.05). In conclusion, DMA could inhibit psoriasis-activated DCs via the TLR7/8 pathway.     

APL-1, an altered peptide ligand derived from human heat-shock protein 60, selectively induces apoptosis in activated CD4 + CD25 + T cells from peripheral blood of rheumatoid arthritis patients

Rheumatoid arthritis (RA) is a chronic T-cell mediated autoimmune disease that affects primarily the joints. The induction of immune tolerance through antigen-specific therapies for the blockade of pathogenic CD4 + T cells constitutes a current focus of research. In this focus it is attempted to simultaneously activate multiple regulatory mechanisms, such as: apoptosis and regulatory T cells (Tregs). APL-1 is an altered peptide ligand derived from a novel CD4 + T-cell epitope of human heat-shock protein of 60 kDa, an autoantigen involved in the pathogenesis of RA. Previously, we have reported that APL-1 induces CD4 + CD25^highFoxp3 + Tregs in several systems. Here, we investigated the ability of APL-1 in inducing apoptosis in PBMCs from RA patients, who were classified as active or inactive according to their DAS28 score. APL-1 decreased the viability of PBMCs from active but not from inactive patients. DNA fragmentation assays and typical morphological features clearly demonstrated that APL-1 induced apoptosis in these cells. Activated CD4 + CD25 + T cells but not resting CD4 + CD25 − T cells were identified as targets of APL-1. Furthermore, CD4 + T-cell responses to APL-1 were found to be dependent on antigen presentation via the HLA-DR molecule. Thus, APL-1 is a regulatory CD4 + T cell epitope which might modulate inflammatory immune responses in PBMCs from RA patients by inducing CD4 + CD25^highFoxp3 + Tregs and apoptosis in activated CD4 + T cells. These results support further investigation of this candidate drug for the treatment of RA.     

Circulating follicular helper T cells presented distinctively different responses toward bacterial antigens in primary biliary cholangitis

Primary biliary cholangitis (PBC) is a chronic and progressive cholestatic liver disease with unknown causes. The initiation of PBC is associated with bacterial infections and abnormal immune correlates, such as the presence of self-reactive anti-mitochondrial antibodies and shifted balance of T cell subsets. In particular, the CD4⁺ CXCR5⁺ follicular helper T (Tfh) cells are highly activated in PBC patients and are significantly associated with PBC severity, but the underlying reasons are unknown. In this study, we found that the circulating CD4⁺ CXCR5⁺ T cells were enriched with the interferon (IFN)-γ-secreting Th1-subtype and the interleukin (IL)-17-secreting Th17-subtype, but not the IL-4-secreting Th2 subtype. We further demonstrated that a host of microbial motifs, including Pam3CSK4, poly(I:C), LPS, imiquimod, and CpG, could significantly stimulate IFN-γ, IL-17, and/or IL-21 from circulating CD4⁺ CXCR5⁺ T cells in PBC patients, especially in the presence of monocytes and B cells. Whole bacterial cells of Escherichia coli, Novosphingobium aromaticivorans, and Mycobacterium gordonae, could also potently stimulate IFN-γ, IL-17, and/or IL-21 production from circulating CD4⁺ CXCR5⁺ T cells. But interestingly, while the whole cell could potently stimulate circulating CD4⁺ CXCR5⁺ T cells from both healthy controls and PBC patients, the cell protein lysate could only potently stimulate circulating CD4⁺ CXCR5⁺ T cells from PBC patients, but not those from healthy controls, suggesting that circulating CD4⁺ CXCR5⁺ T cells in PBC patients had distinctive antigen-specificity from those in healthy individuals. Together, these data demonstrated that bacterial antigen stimulation is a potential source of aberrant Tfh cell activation in PBC patients.     

Evaluación retrospectiva de la efectividad y seguridad de cinacalcet para el tratamiento de hiperparatiroidismo secundario dependiendo del valor basal de paratohormona intacta

ObjectiveCinacalcet is a calcimimetic agent, recommended for treating refractory secondary hyperparathyroidism in patients undergoing dialysis.The aim of this study was to evaluate the efficacy and safety of cinacalcet, comparing patients with baseline iPTH > 300 pg/ml with those with iPTH < 300 pg/ml.MethodObservational retrospective study of patients being treated with cinacalcet 30 mg/day from January 2008 to January 2009.We studied 26 patients, 15 with iPTH > 300 pg/ml and 11 with iPTH < 300pg/ml.The primary efficacy outcome was that there was a reduction between baseline and final iPTH (4 month). The secondary efficacy outcome were the reduction between basal and final calcium, phosphorus, Ca x P, and the percentage of patients with an iPTH decrease > 30%. The safety was evaluated based on the most frequent adverse effects and the levels of serum calcium < 8.6 mg/dl.ResultsPatients with initial iPTH > 300 had significant differences before and after cinacalcet treatment in iPTH (563.49 + 286.88 pg/ml vs 315.15 + 201.948 pg/ml; P = .017) and serum calcium (9.1 + 1.77 mg/dl vs 8.15 + 1.2 mg/dl; P = .02). There were no significant differences in patients with initial iPTH < 300 pg/ml. A decrease greater than 30% from baseline iPTH was observed in 60% of patients with baseline iPTH > 300pg/ml, and only in 27.3% of those with basal iPTH < 300 pg/ml (P = .098). Patients did not show gastric intolerance.ConclusionsCinacalcet is an effective and safe drug for controlling secondary hyperparathyroidism in dialysis, mainly when it is used in patients with baseline iPTH > 300 pg/ml.