GPRC5D CAR T cells (OriCAR-017) in patients with relapsed or refractory multiple myeloma (POLARIS): a first-in-human, single-centre, single-arm, phase 1 trial.

BACKGROUND: Chimeric antigen receptor (CAR) T-cell therapy targeting B-cell maturation antigen (BCMA) has shown activity in treating relapsed or refractory multiple myeloma; however, relapse is still common, and new targets are needed. We aimed to assess the activity and safety profile of G protein-coupled receptor class C group 5 member D (GPRC5D)-targeted CAR T cells (OriCAR-017) in patients with relapsed or refractory multiple myeloma. METHODS: POLARIS was a first-in-human, single-centre, single-arm, phase 1 trial of GPRC5D-targeted CAR T cells (OriCAR-017) done at the First Affiliated Hospital of Zhejiang University School of Medicine, Hangzhou, China. Eligible patients were adults aged 18-75 years with a diagnosis of relapsed or refractory multiple myeloma and an ECOG performance status of 0-2, had GPRC5D expression in bone marrow plasma cells greater than 20% or were positive for GPRC5D by immunohistochemistry, and had received at least three previous lines of treatment including proteasome inhibitors, immunomodulatory drugs, and chemotherapy. Patients were consecutively assigned to receive a single dose of intravenous OriCAR-017 at 1 × 106 CAR T cells per kg, 3 × 106 CAR T cells per kg, or 6 × 106 CAR T cells per kg in the dose-escalation phase. In the expansion phase, patients received the recommended phase 2 dose. Recruitment to the expansion phase terminated early due to the COVID-19 pandemic on May 1, 2022. The primary endpoints were safety, the maximum tolerated dose and the recommended phase 2 dose. Safety and activity analyses included all patients who received OriCAR-017. This trial is registered with ClinicalTrials.gov, NCT05016778. This trial has been completed and is entering long-term follow-up. FINDINGS: Between June 9, 2021, and Feb 28, 2022, we recruited 13 patients for inclusion into the study. One patient was excluded because of GPRC5D negativity and two patients discontinued after apheresis because of rapid progression. Nine patients were assigned to the dose escalation phase (three received 1 × 106 CAR T cells per kg, three received 3 × 106 CAR T cells per kg, and three received 6 × 106 CAR T cells per kg). The maximum tolerated dose was not identified, because no dose-limiting toxic effects were observed. On the basis of safety and preliminary activity, the recommended phase 2 dose was set at 3 × 106 CAR T cells per kg, which was received by one additional patient in the dose expansion phase. Five patients (50%) were female, five (50%) were male, and all were Chinese. Five patients (50%) were previously treated with BCMA-targeted CAR T-cell therapy. Median follow-up was 238 days (IQR 182-307). There were no serious adverse events and no treatment-related deaths. The most common grade 3 or worse adverse events were haematological, including neutropenia (ten [100%] of ten patients), thrombocytopenia (nine [90%]), leukopenia (nine [90%]), and anaemia (seven [70%]). All patients had cytokine release syndrome (nine [90%] grade 1 and one [10%] grade 2). No neurological toxic effects were reported. Ten (100%) of ten patients had an overall response, of whom six (60%) had a stringent complete response and four (40%) had very good partial response. Two patients discontinued due to disease progression (one GPRC5D-positive patient in the middle-dose group and one GPRC5D-negative patient in the low-dose group). INTERPRETATION: The results of this study suggest that GPRC5D is an active target for immunotherapy in multiple myeloma. GPRC5D-targeted CAR T-cell therapy is a promising treatment modality for patients with relapsed or refractory multiple myeloma and deserves further testing. FUNDING: OriCell Therapeutics.

Treatment of Severe Destructive Foot and Ankle Injury in a Young Child with a Skin Regeneration Technique.

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Effect of acute ammonia toxicity on inflammation, oxidative stress and apoptosis in head kidney macrophage of Pelteobagrus fulvidraco and the alleviation of curcumin.

Ammonia is one of the most major pollutant and stress factors of aquaculture systems, and has seriously endangered fish health. However, few studies have been performed on mechanisms of the detrimental impact of ammonia stress and mitigation in fish. A study was carried out to investigate the response of genes involved in inflammation, antioxidation, polarization and apoptosis in head kidney macrophages to acute ammonia toxicity, and the alleviation effect of curcumin. The cells were divided into six groups, as follows: The control group composed of untreated macrophages (CON), the experimental groups, consisting of macrophages treated with 0.23 mg L-1 ammonia (AM), 45 µmol L-1 curcumin (CUR), 0.23 mg L-1 ammonia and 5 µmol L-1 curcumin (5A), 0.23 mg L-1 ammonia and 25 µmol L-1 curcumin (25A), 0.23 mg L-1 ammonia and 45 µmol L-1 curcumin (45A). The cells were pretreated with different concentrations of curcumin for 1 h and then incubated with ammonia for 24 h. The results showed that ammonia poisoning could increase ROS levels, up-regulate the expression of antioxidant enzymes (SOD and GPx), inflammatory cytokines (IL-1, IL-6 and TNF-α) and inflammatory mediators (NF-κB p65 and COX-2), decrease cell viability, down-regulate the expression of M2 marker (Arg-1) and anti-apoptosis (Bcl-2), but curcumin could alleviate the adverse effect of ammonia toxicity. Overall, these results have important implications for understanding of the mechanism of ammonia toxicity and the mitigating effect of curcumin in fish.

Naringenin inhibits migration, invasion, induces apoptosis in human lung cancer cells and arrests tumour progression in vitro.

Lung cancer is one of the major cause for high-death rate all over the world, due to increased metastasize and difficulties in diagnosis. Naringenin is naturally occurring flavonoid found in various fruits including tomatoes, citrus fruit and figs. Naringenin is known to have several therapeutic effects including anti-atherogenic, antimicrobial, anti-inflammatory, hepatoprotective, anticancer and anti-mutagenic. The present study was aimed to analyse the naringenin induced anti-proliferative and apoptosis effects in human lung cancer cells. Cells were treated with various concentrations of naringenin (10, 100 & 200 µmol/L) for 48 hours. Cisplatin (20 µg/mL) was used as positive control. Cell viability, apoptosis, migration and mRNA, and protein expression of caspase-3, matrixmetallo proteinases-2 (MMP-2) and MMP-9 were determined. The cell viability was 93.7 ± 7.5, 51.4 ± 4.4 and 32.1 ± 2.1 at 10, 100 and 200 µmol/L of naringenin respectively. Naringenin significantly increased apoptotic cells. The 100 and 200 µmol/L of naringenin significantly suppressed the larger wounds of cultured human cancer cells compared with the untreated lung cancer cells. Naringenin increased d the expression of caspase-3 and reduced the expression of MMP-2 and MMP-9. Taking all these data together, it is suggested that the naringenin was effective against human lung cancer proliferation, migration and metastasis.

Different screening frequencies of carbapenem-resistant Enterobacteriaceae in patients undergoing hematopoietic stem cell transplantation: which one is better?

BACKGROUND: A consensus has been reached that carbapenem-resistant Enterobacteriaceae (CRE) screening in immunosuppressed individuals can reduce the incidence of CRE bloodstream infection (BSI). METHODS: We retrospectively studied the clinical data of 395 consecutive HSCT patients from September 2017 to April 2019. From September 2017 to June 2018 (period 1), 200 patients received single CRE screening before transplantation. From July 2018 to April 2019 (period 2), 195 patients received continuous weekly CRE screening after admission. For patients colonized with CRE, targeted managements were received: (1) contact precautions and (2) preemptive CRE-targeted treatment if necessary. RESULTS: During period 1, 3 patients with CRE colonization were detected (1.5%). The CRE BSI rate was 2.0% (4 patients), and the related 30-day mortality was 50.0% (2 out of 4 patients). During period 2, 21 patients with CRE colonization were detected, and the detection rate was significantly higher than that in period 1 (P < 0.001). Of the 21 colonized patients, 4 (19.0%) patients were identified as positive for CRE at the first screening, 5 (23.8%) were identified at the second screening, and the remaining 12 (57.1%) were identified at the third or later screening. The CRE BSI rate decreased to 0.5% (1/195), and there were no CRE-related death. Fifteen colonized patients developed neutropenic fever. Thirteen colonizers were preemptively treated with tigecycline within 24 h of fever onset, and they achieved rapid temperature control. One colonizer received tigecycline later than 48 h after fever onset and ultimately survived due to the addition of polymyxin. The other received tigecycline later than 72 h after fever onset and died of septic shock. CONCLUSION: The increase in screening frequency contributed to the detection of patients with CRE colonization. Targeted managements for these colonized patients may contribute to reducing the incidence and mortality of CRE BSI, therefore improving the prognosis of patients.

Preparation of acrylamide and carboxymethyl cellulose graft copolymers and the effect of molecular weight on the flocculation properties in simulated dyeing wastewater under different pH conditions.

In this study, graft copolymers with different graft ratios (GRs) and molecular weights (MWs) were prepared by water bath polymerization using acrylamide (AM) and carboxymethyl cellulose (CMC) as substrates; additionally, the best preparation parameters were provided. The MW of the copolymer was inversely correlated with the GR (R = -0.82). The obtained graft copolymer was characterized by FTIR, 1H NMR, XPS and DSC. The results showed that CMC and AM were successfully copolymerized and the thermal stability of the product is improved. The graft copolymers with different MWs were used to flocculate a simulated dyeing wastewater at pH values of 3, 5, 7, 9 and 11. The results showed that as the molecular weight of the graft copolymers increased, the average flocculation settling ratio decreased drastically from 61.2% to 19.4% at pH 3, and the higher the MW, the better the flocculation settling performance. But the average settling ratio increased sharply at pH 11, and the increase grew from 21.0% to 50.3%. The larger the MW, the more obvious the decrease in flocculation sedimentation performance. The supernatant turbidity was reduced from 13324 NTU to less than 150 NTU, and the turbidity removal ratio exceeded 99%. The flocculation mechanism was discussed.

Elimination of Autofluorescence in Archival Formaldehyde-Fixed, Paraffin-Embedded Bone Marrow Biopsies.

CONTEXT.­: High levels of autofluorescence in bone marrow tissue constitute a major obstacle to immunofluorescence analysis of bone marrow biopsies. OBJECTIVE.­: To present a simple, efficient method to eliminate autofluorescence in bone marrow biopsies. DESIGN.­: Autofluorescence of paraffin bone marrow tissues was examined in different hematologic disorders with confocal laser scanning microscopy. Strong autofluorescence was observed in primary myelofibrosis and acute leukemia with reticulin myelofibrosis in 488-nm and 561-nm channels. To eliminate autofluorescence, AutoFluo Quencher was used on bone marrow sections with different incubation times. The effects of AutoFluo Quencher on immunofluorescence analysis of bone marrow biopsies was tested using antibodies tagged with different fluorophores. RESULTS.­: AutoFluo Quencher thoroughly eliminated the strong autofluorescence of bone marrow but did not decrease the intensity of fluorophores, leaving the specific signals of target proteins clearly visible. CONCLUSIONS.­: This study presents a simple, efficient method to eliminate autofluorescence in bone marrow paraffin tissue, and it opens the way to better results in the immunofluorescence analysis of bone marrow biopsies.

Thalidomide plus prednisone with or without danazol therapy in myelofibrosis: a retrospective analysis of incidence and durability of anemia response.

Low-dose thalidomide and prednisone alone or combined are effective therapies in some persons with primary myelofibrosis (PMF) and anemia with or with RBC transfusion dependence. Danazol is also effective in some persons with PMF and anemia. Responses to these drugs are typically incomplete and not sustained. It is unclear whether adding danazol to thalidomide and prednisone would improve efficacy. We retrospectively compared the outcomes of 88 subjects with PMF and anemia receiving thalidomide and prednisone without (n = 46) or with danazol (n = 42). The primary end point was anemia response, which was 71% (95% confidence interval (CI), 57, 85%) in subjects receiving thalidomide/prednisone/danazol compared with 46% (32, 60%; P = 0.014) in those receiving thalidomide/prednisone. Response rates in subjects who were RBC transfusion dependent was also higher in the danazol cohort (61% (38, 84%)) vs. 25% (6, 44%); P = 0.024). Time to response was rapid (median, 2 months (range, 1-11 months)) and similar between the cohorts. Response duration was longer in the thalidomide/prednisone/danazol cohort (HR 2.18 (1.18-5.42); P = 0.019). Adverse effects were mild and similar between the cohorts. In conclusion, thalidomide/prednisone/danazol seems superior to thalidomide/prednisone in persons with PMF and anemia. Our conclusion requires confirmation in a randomized trial.

Candida glabrata induced infection of rat tracheal epithelial cells is mediated by TLR-2 induced activation of NF-κB.

An increasing number of reports identified Candida glabrata (C. glabrata) as the important causative agent of invasive pulmonary fungal infection. However, little is known about immune responses to C. glabrata in rat tracheal epithelial cell (RTEC). Here, the effect of C. glabrata on RTEC and the role of TLR-2 and NF-κB in the immune response were investigated by treatment with TLR-2 siRNA and NF-κB inhibitor pyrrolidine dithiocarbamate (PDTC), respectively. Our results showed that the knockdown of TLR-2 and pretreatment of PDTC led to inhibition of cell proliferation by C. glabrata, further enhanced cells in G0/G1 phases, and promoted C. glabrata -induced apoptosis. C. glabrata infection induced the expression or secretion of TLR-2, NF-κB, TNF-α, and IL-6, and its effect was inhibited by knockdown of TLR-2. Pretreatment with PDTC inhibited the C. glabrata -induced expression of TLR2, and also inhibited the expression of p65 subunit of NF-κB in the first 4 h. Although the expression of p65 subunit at 6 h was elevated compared to baseline, the C. glabrata -induced expression of TNF-α and IL-6 remained attenuated by PDTC pretreatment. Therefore, C. glabrata recognized the TLR-2 in rat tracheal epithelial cell (RTEC), and then activated the transcription factor NF-κB and further promoted the secretion of TNF-α and IL-6 to contribute to the immune response and inflammation.

Characteristics of γδ T cells in Schistosoma japonicum-infected mouse mesenteric lymph nodes.

Gamma delta (γδ) T cells are mainly present in mucosa-associated lymphoid tissues, which play an important role in mucosal immunity. In this study, C57BL/6 mice were infected by Schistosoma japonicum and lymphocytes were isolated from the mesenteric lymph node (MLN) to identify changes in the phenotype and function of γδ T cells using flow cytometry. Our results indicated that the absolute number of γδ T cells from the MLNs of infected mice was significantly higher compared with normal mice (P < 0.05). In addition, the infected γδ T cells expressed a high level of the activated molecule CD69 (P < 0.01) and demonstrated an increasing population of CD4(+) γδ T cells (P < 0.05). MLN γδ T cells secrete interferon-γ (IFN-γ), interleukin (IL)-4, IL-9, and IL-17 in response to propylene glycol monomethyl acetate (PMA) plus ionomycin simulation, and the levels of IL-4, IL-9, and IL-17 increased significantly after S. japonicum infection (P < 0.05). Taken together, these findings indicated that S. japonicum infection could induce γδ T cell activation, proliferation, and differentiation in the MLN. Moreover, our results indicated that the expression of NKG2D (CD314) was not increased in γδ T cells after infection, suggesting that other mechanisms are involved in activating γδ T cells. Furthermore, higher expression of programmed death-1 (CD279) but not IL-10 was detected in the γδ T cells isolated from infected mice (P < 0.05), suggesting that the function of γδ T cells is inhibited gradually over the course of S. japonicum infection.

Phospholipase C ε-1 inhibits p53 expression in lung cancer.

The pathogenesis of lung cancer is to be further investigated. Recent reports indicate that phospholipase C ε-1 (PLCE1) is a critical molecule involved in tumour growth. This study aims to investigate the role of PLCE1 in the regulation of apoptosis in lung cancer cells. In this study, the surgically removed non-small-cell lung cancer (NSCLC) tissue was collected from 36 patients. Single NSCLC cells were prepared from the tissue, in which immune cells of CD3(+) , CD11c(+) , CD19(+) , CD68(+) and CD14(+) were eliminated by magnetic cell sorting. The expression of PLCE1 and p53 was assessed by quantitative real-time polymerase chain reaction and Western blotting. Apoptosis of NSCLC cells was analysed by flow cytometry. The results showed that, in cultured NSCLC cells, high levels of PLCE1 and low levels p53 were detected; the two molecules showed a negative correlation (p < 0.01). The addition of anti-PLCE1 antibody increased the expression of p53 in NSCLC cells, which increased the frequency of apoptotic NSCLC cells. We conclude that NSCLC cells express high levels of PLCE1, which suppresses the expression of p53 in NSCLC cells. PLCE1 can be a therapeutic target of NSCLC.

Changes in NK and NKT cells in mesenteric lymph nodes after a Schistosoma japonicum infection.

The mesenteric lymph node (MLN) is the main draining lymph node in mouse enterocoelia, which contains many types of immune cells. Among these cells, natural killer (NK) and natural killer T (NKT) cells belong to innate lymphoid cells (ILCs), which have potent activities for controlling a variety of pathogenic infections. In this study, C57BL/6 mice were infected with Schistosoma japonicum for 5-7 weeks. Lymphocytes were isolated from the MLN to detect changes in the phenotype and function of NK and NKT cells using a fluorescence activating cell sorter (FACS). These results demonstrated that a S. japonicum infection could significantly increase the percentage of NK cells in the mouse MLN, (P < 0.05). We found an increase in the cell number of both NK and NKT cells. In addition, we found that NK and NKT cells from infected mice expressed higher levels of CD69 compared to normal mice (P < 0.05). These results demonstrated that a S. japonicum infection could induce MLN NK and NKT cell activation. Moreover, we found that the expression of CD4 was increased in infected MLN NK cells (P < 0.05). Furthermore, intracellular cytokine staining revealed that expression of IL-4 and IL-17 were significantly enhanced in both the NK and NKT cells of infected mice after phorbol 12-myristate 13-acetate (PMA) and ionomycin stimulation (P < 0.05). Taken together, these results indicated that infection-induced MLN NK and NKT cells might play roles in modulating the classical T cell response. Finally, our results indicated that the expression of CD94 was decreased in NK cells, suggesting that the downregulation of CD94 expression might served as a mechanism in NK cell activation.

Roles of Th17 cells in pulmonary granulomas induced by Schistosoma japonicum in C57BL/6 mice.

In schistosomiasis, limited information is available about the role of interleukin-17 (IL-17) in lung, despite the fact that this cytokine plays a crucial role during pro-inflammatory immune responses. In our study, we observed CD4(+)T cells changed after the infection. Furthermore, ELISA and FACS results revealed that Schistosomajaponicum infection could induce a large amount of IL-17 in mouse pulmonary lymphocytes. IL-17-producing cells, including Th17 cells, CD8(+)T (Tc) cells, γδT cells and natural killer T cells, was also associated with the development of lung inflammatory diseases. FACS results indicated that Th17 cell was the main source of IL-17 in the infected pulmonary lymphocytes after phorbol-12-myristate-13-acetate (PMA) and Ionomycin stimulation. Moreover, FACS results revealed that the percentage of Th17 cells continued to increase as over the course of S. japonicum infection. Additionally, cytokines co-expression results demonstrated that Th17 cells could express more IL-4 and IL-5 than IFN-γ. Reducing IL-17 activity by using anti-IL-17 ameliorated the damage and decreased infiltration of inflammatory cells in infected C57BL/6 mouse lungs. Collectively, these results suggest Th17 cells is the major IL-17-producing cells population and IL-17 contributes to pulmonary granulomatous inflammatory during the S. japonicum infection.

[Proportion and characteristics of γδT cells in different tissues and organs of C57BL/6 mice].

OBJECTIVE: To compare the percentages of γδT cells in the lymphocytes and CD3⁺; T cells isolated from the livers, lungs, spleen and mesenteric lymph nodes of C57BL/6 mice and the phonotype and function of γδT cells in different tissues or organs. METHODS: Lymphocytes were isolated from livers, lungs, spleen and mesenteric lymph nodes of normal C57BL/6 mice, respectively. Then, percentages of γδT cells in lymphocytes and CD3⁺; T cells, and phenotypic characteristics of γδT cells were examined by flow cytometry. Moreover, IFN-γ, IL-4, IL-9 and IL-17 secreted by γδT cells were detected by means of intracellular cytokine staining after stimulation with PMA plus ionomycin. RESULTS: The percentage of γδT in lymphocyte cells of the livers was significantly higher than that of the lungs, spleen and mesenteric lymph nodes (P<0.05), and the percentage in CD3⁺; T cells of the mesenteric lymph nodes was the lowest (P<0.05). CD4⁻; CD8⁻; γδT cells were the main subpopulation in these tissues and organs and there was also a small proportion of CD8⁺; γδT cells. The proportion of CD4⁺; γδT cells in the mesenteric lymph nodes was significantly higher than that in the others (P<0.05), and a group of CD4⁺; CD8⁺; γδT cells existed obviously. The percentage of IL-17⁺; cells in γδT cells was significantly higher than IFN-γ⁺; and IL-4⁺; cells, and almost no IL-9 γδT cells were found. The ability of secreting cytokines of γδT cells in the lungs was the strongest and the percentage of IL-17⁺; γδT was (26.6 ± 12.1) %. In addition, the proportion of IFN-γ⁺⁻; γδT cells in the livers and lungs were (1.36 ± 0.37)% and (1.6 ± 0.7)%, respectively. CONCLUSION: The proportion, phenotype and function of γδT cells had significant difference in the livers, lungs, spleen, mesenteric lymph nodes of C57BL/6 mice.

Characteristics of IL-17 induction by Schistosoma japonicum infection in C57BL/6 mouse liver.

Schistosomiasis japonica is a severe tropical disease caused by the parasitic worm Schistosoma japonicum. Among the most serious pathological effects of S. japonicum infection are hepatic lesions (cirrhosis and fibrosis) and portal hypertension. Interleukin-17 (IL-17) is a pro-inflammatory cytokine involved in the pathogenesis of many inflammatory and infectious conditions, including schistosomiasis. We infected C57BL/6 mice with S. japonicum and isolated lymphocytes from the liver to identify cell subsets with high IL-17 expression and release using flow cytometry and ELISA. Expression and release of IL-17 was significantly higher in hepatic lymphocytes from infected mice compared with control mice in response to both non-specific stimulation with anti-CD3 monoclonal antibody plus/anti-CD28 monoclonal antibody and PMA plus ionomycin. We then compared IL-17 expression in three hepatic T-cell subsets, T helper, natural killer T and γδT cells, to determine the major source of IL-17 during infection. Interleukin-17 was induced in all three subsets by PMA + ionomycin, but γδT lymphocytes exhibited the largest increase in expression. We then established a mouse model to further investigate the role of IL-17 in granulomatous and fibrosing inflammation against parasite eggs. Reducing IL-17 activity using anti-IL-17A antibodies decreased infiltration of inflammatory cells and collagen deposition in the livers of infected C57BL/6 mice. The serum levels of soluble egg antigen (IL)-specific IgGs were enhanced by anti-IL-17A monoclonal antibody blockade, suggesting that IL-17 normally serves to suppress this humoral response. These findings suggest that γδT cells are the most IL-17-producing cells and that IL-17 contributes to granulomatous inflammatory and fibrosing reactions in S. japonicum-infected C57BL/6 mouse liver.

Some characteristics of IL-5-producing T cells in mouse liver induced by Schistosoma japonicum infection.

Schistosome infection could cause significant liver damage in animal; Th2 cells play an important role in the progress of this disease. In our study, C57BL/6 mice were infected by Schistosoma japonicum and lymphocytes were isolated from the liver to detect some characteristics of interleukin-5 (IL-5)-producing T cells by different methods. The results revealed that S. japonicum infection could induce a large amount of IL-5 in mouse liver T cells by the means of fluorescent bead immunoassay and RT-PCR. Although, mouse liver contained many T cell subsets, such as Th cells, Tc cells, NKT cells, and γδ T cells. Fluorescence activated cell sorting results indicated that Th cells were the main source of IL-5 in the T cell population after phorbol 12-myristate 13-acetate and ionomycin stimulation. Moreover, the percentage of IL-5-producing Th cells continued to increase from 4 to 8 weeks after S. japonicum infection, which differed from the changes of IFN-γ(+) Th1 cells, IL-4(+) Th2 cells, and IL-17A(+) Th17 cells during S. japonicum infection. Additionally, cytokines co-expression results demonstrated that 36.2 % of IL-5(+) Th cells could express IL-4, and 10 % of it could produce IFN-γ or IL-17A. Collectively, these findings implied that IL-5-producing Th cells posses some properties which differ from other cytokines secreting Th cells.

[Immune response of Th17 cells in mesenteric lymph node of mice infected by Schistosoma japonicum].

OBJECTIVE: To observe the immune response of Th17 cells in mesenteric lymph node (MLN) of C57BL/6 mice infected by Schistosoma japonicum. METHODS: Twenty C57BL/6 mice were randomly divided into infected group and control group each with ten mice. The mice in infected group were infected each with 40 +/- 5 S. japonicum cercariae. Five to six weeks later, MLN lymphocytes were separated and stimulated for 4 h by anti-CD3 (1 microg/ml) and anti-CD28 (1 microg/ml) before examination of IL-17 and retinoic acid receptor-related orphan receptor gammat (ROR-gammat) mRNA by reverse transcription PCR. The level of IL-17 and IFN-gamma was detected by ELISA after culturing with supernatant for 72h. MLN lymphocytes were stimulated for 5h by 10 ng/ml phorbol myristoyl acetate (PMA) and 1 microg/ml ionomycin. The intracellular cytokines were stained and the content of Th17 and other cytokines was examined by flow cytometry. RESULTS: The level of IFN-gamma [(214.3 +/- 62.6) pg/ml] and IL-17 [(176.8 +/- 62.1) pg/ml] in the supernatant of cultured MLN cells from the infected mice was significantly higher than that of normal mice [(467 +/- 13.9) and 0 pg/ml) (P < 0.05). The expression level of IL-17 and ROR-gammat mRNA was also considerably higher than that of normal mice. IL-17+ IL-4+, IL-17+ IFN-gamma+, IL-17+ IL-5+ and IL-17+ IL-9 cells accounted for 0.06%, 0.02%, 0.02%, and 0.01% of the mesenteric lymph node CD4+ T cells of the infected mice, respectively. However, IL-17+ IL-10+ and IL-17+ Foxp3+ cells were undetected. CONCLUSION: The MLN of S. japonicum-infected C57BL/6 mice can induce the production of Th17 cells, and these cells can secrete IL-4, less IFN-gamma, IL-5 and IL-9, but not IL-10, and can not express Foxp3 in the infected mice.