Intratumoral NKT cell accumulation promotes antitumor immunity in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDA) is a potentially lethal disease lacking effective treatments. Its immunosuppressive tumor microenvironment (TME) allows it to evade host immunosurveillance and limits response to immunotherapy. Here, using the mouse KRT19-deficient (sgKRT19-edited) PDA model, we find that intratumoral accumulation of natural killer T (NKT) cells is required to establish an immunologically active TME. Mechanistically, intratumoral NKT cells facilitate type I interferon (IFN) production to initiate an antitumor adaptive immune response, and orchestrate the intratumoral infiltration of T cells, dendritic cells, natural killer cells, and myeloid-derived suppressor cells. At the molecular level, NKT cells promote the production of type I IFN through the interaction of their CD40L with CD40 on myeloid cells. To evaluate the therapeutic potential of these observations, we find that administration of folinic acid to mice bearing PDA increases NKT cells in the TME and improves their response to anti-PD-1 antibody treatment. In conclusion, NKT cells have an essential role in the immune response to mouse PDA and are potential targets for immunotherapy.

Development of an immunoassay for quantification of soluble human CD40L (CD154) in plasma and serum samples.

When the membrane protein CD40 ligand (CD40L) on activated T cells binds the receptor CD40 on B-cells, it provides a co-stimulatory signal for B cell activation. Dysregulation of the CD40L:CD40 axis is associated with inflammatory and autoimmune diseases. The presence of soluble CD40L (sCD40L) in plasma is implicated in several diseases, from cardiovascular and autoimmune diseases to different types of cancer, and sCD40L has been suggested as a valuable marker of disease. If sCD40L is to be used as a biomarker, being able to precisely measure and quantify the levels of sCD40L in human blood samples is of utmost importance. We demonstrate the development of a sandwich-type time-resolved immunofluorometric assay for quantification of sCD40L in plasma or serum samples. For this, we generate 29 monoclonal anti-CD40L antibodies, and from these, we select the optimal combination of capture antibody and detection antibody. A number of variables were tested: the influence of the type of sample (comparing 3 different blood collection tubes for serum sampling and 4 different types of tubes for plasma sampling), the influence of freeze-thaw cycles, the influence of sampling time during night and day, and the influence of centrifugation of the samples. We found a very similar level of sCD40L in paired EDTA plasma and serum samples. Out of 100 healthy blood donor samples 61 had a level of sCD40L below the detection level of the assay, whereas the remaining 39 samples had ranging levels of sCD40L from 1.14 to 33.14 ng/mL. In summary, we present a time-resolved immunofluorometric assay based on paired monoclonal antibodies, ensuring high specificity, sensitivity, and homogeneity. The Eu3+-based assay additionally provides consistent assay readouts due to the extended decay time not seen in standard enzyme-linked immunosorbent assays. The assay paves the way for specific and consistent quantification of sCD40L in human plasma and serum samples.

The CD40/CD40 ligand dyad and its downstream effector molecule ISG54 in relating acute neuroinflammation with persistent, progressive demyelination.

Although Multiple Sclerosis (MS) is primarily thought to be an autoimmune condition, its possible viral etiology must be taken into consideration. When mice are administered neurotropic viruses like mouse hepatitis virus MHV-A59, a murine coronavirus, or its isogenic recombinant strain RSA59, neuroinflammation along with demyelination are observed, which are some of the significant manifestations of MS. MHV-A59/RSA59 induced neuroinflammation is one of the best-studied experimental animal models to understand the viral-induced demyelination concurrent with axonal loss. In this experimental animal model, one of the major immune checkpoint regulators is the CD40-CD40L dyad, which helps in mediating both acute-innate, innate-adaptive, and chronic-adaptive immune responses. Hence, they are essential in reducing acute neuroinflammation and chronic progressive adaptive demyelination. While CD40 is expressed on antigen-presenting cells and endothelial cells, CD40L is expressed primarily on activated T cells and during severe inflammation on NK cells and mast cells. Experimental evidences revealed that genetic deficiency of both these proteins can lead to deleterious effects in an individual. On the other hand, interferon-stimulated genes (ISGs) possess potent antiviral properties and directly or indirectly alter acute neuroinflammation. In this review, we will discuss the role of an ISG, ISG54, and its tetratricopeptide repeat protein Ifit2; the genetic and experimental studies on the role of CD40 and CD40L in a virus-induced neuroinflammatory demyelination model.

The mechanism of cilostazole alleviating intestinal mucosal barrier dysfunction in type 2 diabetic mice / 西安交通大学学报(医学版)

【Objective】 To explore the effect of cilostazol on intestinal barrier function in type 2 diabetes (T2DM). 【Methods】 The GSE142153 dataset was downloaded from GEO database to analyze gene changes in diabetic patients. Eight-week-old male db/db mice and control m/m mice were randomly divided into m/m+cmc, m/m+cilo, db/db+cmc, and db/db+cilo groups. Mice in different groups were given cilostazol and corresponding solvents for 4 weeks. We detected the levels of serum sCD40L and the expression of CD40 in intestinal tissue, and evaluated the mice’s intestinal barrier function by examining intestinal permeability, water content, bacterial number, and tight junction protein expression in different groups. 【Results】 Differential expressed genes were enriched in platelet activation and endothelial barrier function pathways in diabetic patients. Compared with those in the control group, the levels of serum sCD40L in db/db diabetic mice elevated significantly, and the CD40 expression, permeability, water content and bacterial number in intestinal tissue increased obviously, while the expression of tight junction protein decreased. Cilostazol treatment in diabetic mice decreased the levels of serum sCD40L and CD40, and alleviated significantly the intestinal barrier dysfunction. 【Conclusion】 Cilostazol attenuated the damage of intestinal barrier function in T2DM, and its protective effect may be related to the inhibition of platelet activation in diabetic mice.

Impact of immune inflammation on the proliferation and apoptosis of benign prostatic hyperplasia cells / 中华男科学杂志

Objective@#To investigate the impact of macrophage-induced immune inflammation on the proliferation and apoptosis of BPH cells and its possible mechanism.@*METHODS@#Macrophages were stimulated with phorbol myristate acetate, co-cultured with BPH-1 cells, and then treated with the androgen receptor (AR) inhibitor or anti-CD40L antibody. The immunohistochemical biomarkers of the T lymphocytes (CD4 and CD8), B lymphocyte (CD20) and macrophages (CD68), AR, CD40/CD40L, and inflammatory factors IL-1, IL-6 and TNF-α were measured before and after treatment. The proliferation and apoptosis of the cells were observed by MTT assay, colony-forming assay and flow cytometry, and the expressions of cell apoptosis- and MAPK signaling pathway-related proteins were determined by qRT-PCR and Western blot.@*RESULTS@#Significantly increased proliferation and decreased apoptosis of the cells, up-regulated expressions of Bcl-2, IL-1, IL-6, TNF-α, AR, CD40 and CD40L, and down-regulated expression of Bax were observed in the BPH-1 cells co-cultured with macrophages (the M-BPH-1 group) compared with those in the blank control (B-BPH-1) group (P < 0.01). In comparison with the BPH-1 cells treated with normal saline, those treated with either low-dose CD40L (L-CD40L) or high-dose CD40L (H-CD40L) showed markedly inhibited proliferation, increased apoptosis, up-regulated expression of Bax, and down-regulated expressions of Bcl-2, IL-1, IL-6 and TNF-α (P < 0.01), and those in the low- and high-dose AR (L-AR and H-AR) inhibitor groups exhibited remarkably reduced proliferation, increased apoptosis, down-regulated expressions of Bcl-2, IL-1, IL-6 and TNF-α, and up-regulated expression of Bax (P < 0.01). The phosphorylation levels of JNK, ERK and P38 were significantly elevated in the M-BPH-1 group, but declined in the H-CD40L and the H-AR inhibitor groups compared with those in the B-BPH-1 group, all in a concentration-dependent manner (P < 0.01).@*CONCLUSIONS@#Macrophage-induced immune inflammation regulates AR and CD40/CD40L expressions and promotes the proliferation and inhibits the apoptosis of BPH-1 cells by activating the MAPK signaling pathway. /.

Clinical significance of soluble CD30/CD30L and CD40/CD40L system imbalance in ovarian serous tumors / 中华内分泌外科杂志

Objective To investigate clinical significance of soluble CD30/CD30L and CD40/CD40L system imbalance in ovarian serous tumors.Methods 40 patients of serous cystadenoma and 30 patients of serous cystadenocarcinoma were selected,and 40 age-and weight-matched healthy women were also recruited as the control group.Peripheral venous blood (3 ml) of the healthy control and patients with ovarian serous tumors before surgery and 7 days after surgery were collected.After separation of serum,ELISA was used to detect levels of sCD30,sCD30L,sCD40 and sCD40L.Results Compared to the control group,levels of sCD30,sCD30L,sCD40 and sCD40L in both serous cystadenoma and serous cystadenocarcinoma groups were significantly in creased (P＜0.05).And in those serous cystadenocarcinoma group,levels of such soluble proteins were much higher than in serous cystadenoma group (P＜0.05).7 days after surgery,levels of such soluble proteins were significantly decreased in both serous cystadenoma and serous cystadenocarcinoma groups (P＜0.05).Conclusion Detection of serum sCD30/sCD30L and sCD40/sCD40L is possible to have a certain guiding significance to early diagnosis of ovarian tumors and the prognosis of patients.

Effects of immunosuppressants on co-stimulated lymphocyte's Th1/Th2 cytokine production / 中华泌尿外科杂志

Objective To analyze the effect of cyclosporin A(CsA), rapamycin(RPM) and macophenolic acid(MPA) on the co-stimulated lymphocytes, CD28 and CD40, and their production of Th1/Th2 cytokine, IL-2, IFN-γ, IL-4, IL-10 and IL-12. Methods The experimental groups were divided into ①mono-stimulating and co-stimulating groups: CD3 mAb mono-stimulating (group a),CD3 mAb+CD28 mAb co-stimulating (group b), CD3 mAb+CD28 mAb+CD40 L mAb co-stimulating(group c), CD3 mAb+CD28 mAb+CTLA4 mAb co-stimulating (group d). ②CsA groups: 300 ng/ml of CsA was added to group a, b, c and d. ③RPM groups: 300 ng/ml of RPM was added to group a,b, c and d. ④MPA groups:300 ng/ml of MPA was added to group a, b, c and d. The cytokine production was measured by ELISA.Results The co-stimulated CD28 and CD40 Th1/Th2 cytokines production of IFN-γ, IL-2, IL-4 and IL-10 were significantly increased. Compared with group a, IFN-γ increased from (248.91±11.20)ng/ml to (555.08±24.42)ng/ml and (548.19±33.06)ng/ml, IL-2 increased from (29.48±8.61)ng/ml to (1100.82±99.29)ng/ml and (842.23±29.31)ng/ml, IL-4 increased from (32.29±6.76)ng/ml to (116.02±15.03)ng/ml and (147.28±18.07)ng/ml, IL-10 increased from (147.01±10.47)ng/ml to (291.79±12.47)ng/ml and (302.52±35.18)ng/ml,respectively, P＜0. 01. Compared group b with group c, the Th1 cytokines production was decreased.IL-2 and IL-12 decreased (P＜0.05). The Th2 cytokine IL-4 production was increased (P＜0. 05).CTLA4 mAb and three other immunosuppressants, CsA, RPM and MPA, inhibited co-stimulated lymphocyte's both cytokines Th1 and Th2 production. The inhibitory effect of CsA on Th1/Th2 cytokine production was more significant than RPM and MPA did. The co-inhibitory effect of CTLA4 mAb and CsA was observed as well. The increased co-stimulated CD28 and CD40 IL-12 production could be suppressed by MPA. CsA and RPM had no inhibitory effect on the IL-12 production.Conclusions CD28/CD40 co-stimulatory pathway plays the key role in lymphocyte activation and Th1/Th2 cytokine production. CsA, RPM and MPA can inhibit co-stimulated lymphocyte's Th1 and Th2 cytokine production. CsA and CTLA4 mAb have co-inhibitory effect on co-stimulated lymphocyte's Th1/Th2 cytokines production. CD40 L mAb decreases the Th1 cytokines production(including IL-12) and increases the Th2 (mainly IL-4) production, which may be the mechanism of its longevity effect on allograft.

Correlation investigation of signal molecule CD_(40)/CD_(40L) and children asthma / 中国医师进修杂志

0.05) among three groups in expression rate of CD_(40L) on B lymphocyte and CD_8~+T cells.Conclusions Abnormal level of CD_(40)/CD_(40L) what attack asthma and stable asthma showed on their relevant immune cells would have an important significance in development of allergic asthma.