The novel α-glucan YCP improves the survival rates and symptoms in septic mice by regulating myeloid-derived suppressor cells.

Sepsis is a life-threatening health condition that is initially characterized by uncontrolled inflammation, followed by the development of persistent immunosuppression. YCP is a novel α-glucan purified from the mycelium of the marine fungus Phoma herbarum YS4108, which has displayed strong antitumor activity via enhancing host immune responses. In this study, we investigated whether YCP could influence the development of sepsis in a mouse model. Caecal ligation and puncture (CLP)-induced sepsis was established in mice that were treated with YCP (20 mg/kg, ip or iv) 2 h before, 4 and 24 h after the CLP procedure, and then every other day. YCP administration greatly improved the survival rate (from 39% to 72% on d 10 post-CLP) and ameliorated disease symptoms in the septic mice. Furthermore, YCP administration significantly decreased the percentage of myeloid-derived suppressor cells (MDSCs) in the lungs and livers, which were dramatically elevated during sepsis. In cultured BM-derived cells, addition of YCP (30, 100 µg/mL) significantly decreased the expansion of MDSCs; YCP dose-dependently decreased the phosphorylation of STAT3 and increased the expression of interferon regulatory factor-8 (IRF-8). When BM-derived MDSCs were co-cultured with T cells, YCP dose-dependently increased the production of arginase-1 (Arg-1) and inducible nitric oxide synthase (iNOS), and activated the NF-κB pathway. In addition, the effects of YCP on MDSCs appeared to be dependent on toll-like receptor (TLR) 4. These results reveal that YCP inhibits the expansion of MDSCs via STAT3 while enhancing their immunosuppressive function, partially through NF-κB. Our findings suggest that YCP protects mice against sepsis by regulating MDSCs. Thus, YCP may be a potential therapeutic agent for sepsis.

Distinct expression patterns of Toll-like receptor 7 in tumour cells and fibroblast-like cells in oral squamous cell carcinoma.

AIMS: Toll-like receptor (TLR)-7 agonists have been used in cancer immunotherapy, but tumour heterogeneity means that TLR-7 activity is variable in different components of the tumour microenvironment and the characteristics of TLR-7 in oral squamous cell carcinoma (OSCC) are unclear. METHODS AND RESULTS: Twenty healthy oral tissues, 50 oral leukoplakia tissues and 166 retrospective primary OSCC samples were collected for immunohistochemical staining of TLR-7 and showed up-regulated expression during carcinogenesis. Moreover, patients with high expression of TLR-7 in tumour cells (TCs) had poor differentiation and prognosis. Interestingly, patients with high expression of TLR-7 in stroma fibroblast-like cells (FLCs) had low tumour stage, no lymph node metastasis (LNM) and better prognosis. Furthermore, Ki-67, CD3, CD4, CD8 and forkhead box protein 3 (FoxP3)(+) tumour-infiltrated lymphocytes were assessed and we found that TLR-7(high) TCs were infiltrated by fewer CD3(+) CD4(+) but more FoxP3(+) lymphocytes. Importantly, patients with TLR-7(low) TCs and TLR-7(high) FLCs had less FoxP3(+) lymphocyte infiltration and longer survival time than those with TLR-7(high) TCs/TLR-7(low) FLCs, although TLR-7 was not an independent prognostic factor for OSCC. CONCLUSIONS: The low expression of TLR-7 in tumour and high expression of TLR-7 in stroma predict a good clinical outcome for OSCC patients, and stroma FLCs might be amenable to immunotherapy by a TLR-7 agonist.

Microlocalization of CD68+ tumor-associated macrophages in tumor stroma correlated with poor clinical outcomes in oral squamous cell carcinoma patients.

CD68 has been widely used as a pan-macrophage marker for tumor-associated macrophages (TAM) which always involve in carcinogenesis. But the correlations between CD68(+) TAMs and prognosis of patients show to be inconsistent, which might due to lack of specific markers of TAMs. We here found that the microlocalization of CD68(+) TAMs also played a unique role in prognosis of patients with oral squamous cell carcinoma (OSCC). CD68(+) TAMs were identified in paraffin-embedded OSCC specimens (n = 91) by using immunohistochemistry. The number of CD68(+) TAMs was remarkably increased from adjacent none-neoplasia tissues (NT) to tumor nest (TN), but tumor stroma (TS) was infiltrated with highest frequency of CD68(+) TAMs (P < 0.0001). Unexpectedly, more CD68(+) TAMs in TS, but not NT or TN, were associated with high tumor grade (P = 0.033), lymph node metastasis (P = 0.034), and shorter 10-year overall survival time, disease free survival. Considering TAMs was derived from monocytes in peripheral blood, we assessed the relationship between leukocytes in peripheral blood and CD68(+) TAMs in OSCC and found that more CD68(+) TAMs in TS were accompanied with decreased monocytes and lymphocytes in peripheral blood (P < 0.05). Although Cox regression analysis revealed that CD68(+) TAMs in TS were not an independent prognostic factor for OSCC patients, we raised a possibility that the microlocalization of CD68(+) TAMs was an indispensable factor for the advance of OSCC.

Serum IL-17F combined with VEGF as potential diagnostic biomarkers for oral squamous cell carcinoma.

Although interleukin (IL) 17A can promote angiogenesis in several tumors, there are limited clinical evidences on cancer about the correlation between serum vascular endothelial growth factor (VEGF) and IL-17F, which is the most homologous to IL-17A. In this study, serum concentration of IL-17F and VEGF from healthy (n = 28), leukoplakia (n = 15), and oral squamous cell carcinoma (OSCC) groups (n = 85) were assessed and showed that IL-17F level was remarkably downregulated from healthy group (394.3 pg/ml) to OSCC group (82.96 pg/ml). Conversely, the OSCC group had a highest level of VEGF (P < 0.05) in whole groups, and there was a negative correlation between IL-17F and VEGF in serum or in the peripheral blood mononuclear cells (PBMCs) at mRNA level. Moreover, the lowest ratio of IL-17F/VEGF was found in OSCC patients (P < 0.05) and lower ratio of IL-17F/VEGF correlated to higher tumor stage and lymph node metastasis. Furthermore, the serum level of IL-17F and the ratio of IL-17F/VEGF were positively associated with the numbers of CD3(+) CD4(+) T cells, which indicated that serum IL-17F could originate from PBMCs during the development of OSCC, and could be used for the diagnosis by effectively distinguishing OSCC patients from healthy individuals.

Meroterpenes with toll-like receptor 3 regulating activity from the endophytic fungus Guignardia mangiferae.

The endophytic fungus Guignardia mangiferae isolated from Ilex cornuta leaves was shown to produce a family of meroterpenes with toll-like receptor 3 regulating activity (1-9), of which 1-3 possessed new structures. The absolute stereochemistry of 1-3 was assigned through a combination of nuclear magnetic resonance experiments, chemical derivation, CD spectra, and single-crystal X-ray diffraction analyses (CuK α ). The precursor labeled cultivation suggests that these meroterpenes are most likely assembled through terpenoid-shikimate pathways. Moreover, meroterpenes 1-3, 5-7, and 9 selectively upregulate, but 4 and 8 downregulate the toll-like receptor 3 expression in mouse dendritic cells at 10.0 µM.

Serum CCL2 and CCL3 as potential biomarkers for the diagnosis of oral squamous cell carcinoma.

Monocyte chemotactic protein-1 (MCP-1/CCL2) and macrophage inflammatory protein-1α (MIP-1α/CCL3) are small chemotactic proteins that have been found in several kinds of tumor tissue samples and function as key regulators of cancer progression. However, the expression of CCL2 and CCL3 in serum samples of oral squamous cell carcinoma (OSCC) patients remains unknown. This study aimed to investigate the prognostic meaning of serum CCL2 and CCL3 in OSCC. The concentration of CCL2 and CCL3 was assessed by ELISA in serum of OSCC patients (n = 98), leukoplakia patients (n = 14), and healthy donors (n = 27). The results showed that the concentration of CCL2 in the OSCC group was significantly lower compared to that in the healthy controls (67.81 vs. 108.1 pg/ml, P < 0.0001). The CCL3 concentration was higher in leukoplakia patients than in OSCC patients and healthy donors (201.9 vs. 153.9 or 118.3 pg/ml, P < 0.05). No significant difference in CCL3 concentration was observed between OSCC patients and healthy donors. However, the OSCC group clearly presented two subclusters, i.e., CCL3 (LOW) and CCL3 (HIGH) OSCC subclusters, in which the serum level of CCL3 was positively related to the tumor size. Interestingly, the ratio of CCL2/CCL3 in OSCC patients was correlated to TNM (tumor, node, metastasis), smoking habits, and differentiation. The receiver operating characteristic (ROC) curve suggests that serum CCL2 is a good diagnostic marker to discriminate OSCC patients from healthy people (cutoff value, 101.1 pg/ml) and the ratio of CCL2/CCL3 also is a good diagnostic marker to discriminate leukoplakia patients and CCL3 (HIGH) OSCC patients from healthy people (cutoff values, 1.080 and 0.424, respectively). These results indicate that CCL2 and CCL3 are associated with progression of OSCC and may be potential biomarkers.

Accumulation of CD208⁺ mature dendritic cells does not correlate with survival time in oral squamous cell carcinoma patients.

PURPOSE: To investigate the distribution and clinical outcomes of tumor-infiltrating CD208(+) mature dendritic cells (mDCs) in patients with oral squamous cell carcinoma (OSCC). MATERIALS AND METHODS: Using immunohistochemical analysis, the distribution of CD208(+) mDCs in adjacent non-neoplastic tissues (NTs) and tumor tissue, including the tumor stroma (TS) and tumor nest (TN), of 79 patients with OSCC was evaluated. The analysis was quantitative, and the number of positive cells was counted in 5 microscopic high-power fields (×400). At the gene expression level, CD208 expression also was assessed by quantitative polymerase chain reaction. Kaplan Meier analyses were used to analyze the prognostic value of tumor-infiltrating CD208(+) mDCs. RESULTS: The number of tumor-infiltrating CD208(+) mDCs was larger in the TS and TN than in the NTs (P < .0001) and the number of CD208(+) mDCs in the TS was larger in patients with OSCC and lymph node positivity (P < .05). At the gene expression level, CD208 also was upregulated in patients with lymph node positivity (P < .05). The number of infiltrating CD208(+) mDCs was not associated with a patient's survival time. CONCLUSIONS: The number of tumor-infiltrating CD208(+) mDCs in the TS was larger in patients with OSCC and lymph node positivity, but the accumulation of CD208(+) mDCs did not correlate with survival time in patients with OSCC.

Emergence of peripheral CD3+CD56+ cytokine-induced killer cell in HIV-1-infected Chinese children.

Cytokine-induced killer (CIK) cells are immune effector cells characterized by co-expression of CD3 and CD56 molecules. We examined the quantities of CIK cells and the changes of these cell expressing NK cell receptors in HIV-1-positive children infected via mother-to-child transmission. The percentage of CIK cells was quantified and the changes in the surface cell receptor profiles in 18 HIV-1-infected children were examined. We found that CIK cell percentages were dramatically increased in HIV-1-infected children. Furthermore, the expressions of CD16, NKp30, NKp44, NKp46, NKp80 and CD244 on CIK cells were decreased, while the expressions of KIR3DL1 and NKG2D on CIK cells were increased in HIV-1-infected children. However, the expressions of KIR2D and NTB-A on CIK cells did not change in the HIV-1-infected children. CIK cells possessed the characteristics of promoting the maturation of dendritic cells and killing functions in HIV-1-infected children. Moreover, serum concentrations of IL-4 and IFN-γ were significantly increased in HIV-1-infected children compared with the HIV-negative controls. These changes likely occurred as a protective mechanism against transmission of maternal HIV-1 virus and thereby helped to limit viral spread, eliminate infected cells and help HIV-1-infected patients to slow the progression to AIDS.

Long non-coding RNA MEG3 inhibits NSCLC cells proliferation and induces apoptosis by affecting p53 expression.

BACKGROUND: Long non-coding RNAs play an important role in tumorigenesis, hence, identification of cancer-associated lncRNAs and investigation of their biological functions and molecular mechanisms are important for understanding the development and progression of cancer. Recently, the downregulation of lncRNA MEG3 has been observed in various human cancers. However, its role in non-small cell lung cancer (NSCLC) is unknown. The aim of this study was to examine the expression pattern of MEG3 in NSCLC and to evaluate its biological role and clinical significance in tumor progression. METHODS: Expression of MEG3 was analyzed in 44 NSCLC tissues and 7 NSCLC cell lines by qRT-PCR. Over-expression approaches were used to investigate the biological functions of MEG3 in NSCLC cells. Bisulfite sequencing was used to investigate DNA methylation on MEG3 expression. The effect of MEG3 on proliferation was evaluated by MTT and colony formation assays, and cell apoptosis was evaluated by Hoechst staining and Flow-cytometric analysis. NSCLC cells transfected with pCDNA-MEG3 were injection into nude mice to study the effect of MEG3 on tumorigenesis in vivo . Protein levels of MEG3 targets were determined by western blot analysis. Differences between groups were tested for significance using Student's t-test (two-tailed). RESULTS: MEG3 expression was decreased in non-small cell lung cancer (NSCLC) tumor tissues compared with normal tissues, and associated with advanced pathologic stage, and tumor size. Moreover, patients with lower levels of MEG3 expression had a relatively poor prognosis. Overexpression of MEG3 decreased NSCLC cells proliferation and induced apoptosis in vitro and impeded tumorigenesis in vivo. MDM2 and p53 protein levels were affected by MEG3 over-expression in vitro. CONCLUSIONS: Our findings indicate that MEG3 is significantly down-regulated in NSCLC tissues that could be affected by DNA methylation, and regulates NSCLC cell proliferation and apoptosis, partially via the activition of p53. Thus, MEG3 may represent a new marker of poor prognosis and is a potential therapeutic target for NSCLC intervention.

Relative and absolute configuration of vatiparol (1 mg): a novel anti-inflammatory polyphenol.

Bioactive natural products offer multiple opportunities for the discovery of novel chemical entities with potential pharmaceutical, nutraceutical and agrochemical applications. Many new organic compounds with novel scaffolds are isolated in small quantities and established methods often fail to determine the structure and bioactivity of such novel natural products. To meet this challenge, we present here a new methodology combining RDC (residual dipolar coupling)-based NMR spectroscopy in microtubes, with a motif-inspired biological assessment strategy. Using only one milligram (ca. 1.5 µmol) of sample, the new protocol established the bioactivity as well as the relative and absolute configuration of vatiparol obtained from Vatica parvifolia. Vatiparol is unique in its unprecedented carbon skeleton and selective inhibitory effect on the expression of monocyte chemo-attractant protein-1 (MCP-1, also known as CCL2). The plausible biosynthetic pathway of vatiparol is briefly discussed. The approach introduced here promises to be widely applicable to the determination of the structure and bioactivity of structurally unknown organic samples available in very limited amounts.

Stanniocalcin-1 promotes tumor angiogenesis through up-regulation of VEGF in gastric cancer cells.

BACKGROUND: Stanniocalcin-1(STC-1) is up-regulated in several cancers including gastric cancer. Evidences suggest that STC-1 is associated with carcinogenesis and angiogenic process. However, it is unclear on the exact role for STC-1 in inducing angiogenesis and tumorigeneisis. METHOD: BGC/STC cells (high-expression of STC-1) and BGC/shSTC cells (low- expression of STC-1) were constructed to investigate the effect of STC-1 on the xenograft tumor growth and angiogenesis in vitro and in vivo. ELISA assay was used to detect the expression of vascular endothelial growth factor (VEGF) in the supernatants. Neutralizing antibody was used to inhibit VEGF expression in supernatants. The expression of phosphorylated -PKCßII, phosphorylated -ERK1/2 and phosphorylated -P38 in the BGC treated with STC-1protein was detected by western blot. RESULTS: STC-1 could promote angiogenesis in vitro and in vivo, and the angiogenesis was consistent with VEGF expression in vitro. Inhibition of VEGF expression in supernatants with neutralizing antibody markedly abolished angiogenesis induced by STC-1 in vitro. The process of STC-1-regulated VEGF expression was mediated via PKCßII and ERK1/2. CONCLUSIONS: STC-1 promotes the expression of VEGF depended on the activation of PKCßII and ERK1/2 pathways. VEGF subsequently enhances tumor angiogenesis which in turn promotes the gastric tumor growth.

Differences in natural killer cell quantification and receptor profile expression in HIV-1 infected Chinese children.

Natural killer (NK) cells are believed to play a role in the progression of human immunodeficiency virus 1 (HIV-1) disease, and NK cell levels are reduced in individuals with chronic HIV-1 infection. To assess the effects on quantity of NK cells and the changes of NK cell receptors in HIV-1 infected children via mother-to-child transmission, the percentage of NK cells is quantified and the changes in the NK cell receptor profiles in 20 HIV-1 infected children who are not progressing into AIDS were examined. The results showed that NK cell percentage was decreased in the HIV-1 infected children. The expression of NKp30 on NK cells was increased, while the expressions of CD16, NKp44, NKp46, NKp80, NTB-A, CD244, KIR2D, KIR3DL1 and NKG2D on NK cells were decreased in the HIV-1 infected children. NK cell cytolytic activity was elevated in HIV-1 infected children. These results indicate that the acute changes in NK cell percentage and NK cell receptors in HIV-1 infected children are different from the HIV-1 infected adult individuals. Moreover, serum concentrations of IL-18 were elevated in HIV-infected children compared to HIV-uninfected controls. These differences probably play a role in protecting against transmission of maternal HIV-1 virus and guiding the therapeutic strategies for HIV-1 infected children.

Tumor-Infiltrating CD1a+ DCs and CD8+/FoxP3+ Ratios Served as Predictors for Clinical Outcomes in Tongue Squamous Cell Carcinoma Patients.

Tumor-infiltrating immune cells engage in an extensive crosstalk with tumors and act as two-edged swords by inhibiting or promoting cancer growth. Therefore, identifying the density and prognostic values of tumor-infiltrating immune cells will provide valuable tips for cancer treatments. In this study, we identified the density of tumor inflammatory infiltrates and the number of tumor-infiltrating immune cells, including CD3+, CD4+, CD8+, FoxP3+ T cells and CD1a+ dendritic cells (DCs) in 153 tongue squamous cell carcinomas (TSCC). High inflammatory cell infiltration was associated with better overall survival (OS) and disease free survival (DFS). Moreover, the number of CD3+, CD4+, FoxP3+ and CD1a+ cells were associated with tumor differentiation (P<0.001) and the number of FoxP3+, CD1a+ cells and CD8+/FoxP3+ ratios were also associated with tumor stage (P<0.01, P<0.01, P<0.05). In addition, patients with higher CD1a+ DCs had better OS and increased CD8+/FoxP3+ ratios were associated with improved OS and DFS (P = 0.037; P = 0.047; P = 0.033). In conclusion, our results indicated that tumor-infiltrating CD1a+ DCs and CD8+/FoxP3+ ratios were associated with favorable clinical outcomes but not independent prognostic factors for TSCC patients.

C-type lectin receptor-mediated immune recognition and response of the microbiota in the gut.

C-type lectin receptors (CLRs) are powerful pattern-recognition receptors that discern 'self' and 'non-self' in our body and protect us from invasive pathogens by mediating immune recognition and response. The gastrointestinal tract is very important for the maintenance of homeostasis; it is the largest shelter for the billions of microorganisms in the body and CLRs play a crucial regulatory role in this system. This study focuses on several CLRs, including Dectin-1, Dectin-2, Dectin-3 and Mincle. We summarize the roles of CLRs in maintaining gastrointestinal immune-system homeostasis, especially their functions in mediating immune recognition and responses in the gut, discuss their relationships to some diseases, highlight the significance of CLR-mediated sensing of microbial and non-microbial compounds in the gut immune system and identify new therapeutic targets.

Over-expressed and truncated midkines promote proliferation of BGC823 cells in vitro and tumor growth in vivo.

AIM: To determine whether midkine (MK) and its truncated form (tMK) contribute to gastric tumorigenesis using in vitro and in vivo models. METHODS: Human MK and tMK plasmids were constructed and expressed in BGC823 (a gastric adenocarcinoma cell line) to investigate the effect of over-expressed MK or tMK on cell growth and turmorigenesis in nude mice. RESULTS: The growth of MK-transfected or tMK-transfected cells was significantly increased compared with that of the control cells, and tMK-transfected cells grew more rapidly than MK-transfected cells. The number of colony formation of the cells transfected with MK or tMK gene was larger than the control cells. In nude mice injected with MK-transfected or tMK-transfected cells, visible tumor was observed earlier and the tumor tissues were larger in size and weight than in control animals that were injected with cells without the transfection of either genes. CONCLUSION: Over-expressed MK or tMK can promote human gastric cancer cell growth in vitro and in vivo, and tMK has greater effect than MK. tMK may be a more promising gene therapeutic target compared with MK for treatment of malignant tumors.

[Aberrant expression of sperm protein 17 enhances migration of ovarian cancer cell line HO-8910].

OBJECTIVE: To explore the effects of aberrant expression of sperm protein 17 (Sp17) on the migration of the ovarian cancer cell line HO-8910. METHODS: The recombinant plasmid pEGFP-Sp17 containing Sp17 and enhanced green fluorescent protein gene was transfected into the human ovarian cancer cell line HO-8910 with Lipofectamine 2000. The expression of Sp17 was examined by RT-PCR and Western blot, and the cell migratory capability detected by Transwell chamber assays. RESULTS: Sp17 was expressed as a fusion protein with EGFP after transfected. There was a significant difference in the migratory cell number of the transfected and the control cells (156.6 +/- 14.9/HP vs 39.3 +/- 8.53/HP, P < 0.05). CONCLUSION: The aberrant expression of Sp17 greatly enhances the migration of ovarian cancer cells.

Abnormal surface markers expression on bone marrow CD34+ cells and correlation with disease activity in patients with systemic lupus erythematosus.

Defects of hematopoietic stem cells (HSCs) have been suggested to contribute to the development of systemic lupus erythematosus (SLE). The aim of this study was to investigate the phenotypic characteristics of bone marrow (BM) CD34(+) cells in patients with SLE and its relationship with SLE disease activity. Ten SLE patients and 10 healthy subjects were recruited and their BM CD34(+) cells were analyzed by flow cytometric analysis with CD45/SSC gating for the expression of CD90, CD95, CD117, CD123, CD164, CD166, FAS-L, and HLA-DR. The percentage of BM CD34(+) cells was significantly decreased in active SLE patients (1.48 +/- 0.41%, n = 7) compared to the healthy controls (2.31 +/- 0.75%, n = 10, p < 0.01), but no significant difference was found between the inactive patients (2.04 +/- 0.44%, n = 3) and the controls. The expression of CD95, CD123, and CD166 on BM CD34(+) cells were significantly increased in SLE patients (48.31 +/- 10.59%, 44.9 +/- 21.5%, 30.9 +/- 19.54%, respectively, n = 10) when compared with the control subjects (24.33 +/- 11.1%, 19.5 +/- 4.4%, 10.7 +/- 5.5%, respectively, n = 10, p < 0.05). The increased CD123 expression was negatively correlated with the number of peripheral white blood cells (r = -0.700, p < 0.05, n = 10). The percentage of CD166 expression was found significantly correlated with the index of SLE disease activity (r = 0.472, p < 0.05, n = 10) and 24 h proteinuria (r = 0.558, p < 0.05, n = 10), but negatively correlated with serum C3 level (r = -0.712, p < 0.01, n = 10). Our study found that the surface marker expression of CD95, CD123, and CD166 on BM CD34(+) cells were significantly increased in patients. This supports the hypothesis that there are abnormalities of the HSC in SLE. Since CD166 and CD123 correlated with the overall lupus activity, their role as a biomarker of inflammatory disease activity also requires further study.

[Immunophenotypic characteristics of uterine natural killer cells and helper T cell 1/helper T cell 2 immunity in the third trimester decidua of preeclampsia patients].

OBJECTIVE: To investigate immunophenotypic characteristics of uterine natural killer (uNK) cells and helper T cell 1/helper T cell 2 (Th1/Th2) immunity in third trimester decidua in preeclampsia. METHODS: The proportions of uNK cell subsets, expression of CD(69) and CD(94) on uNK cells and Th1/Th2 immunity in decidua were determined in 20 cases of preeclampsia patients and 11 cases of normal term pregnancies by flow cytometric analysis. RESULTS: The percentage of CD(56)(bright)CD(16)(-) uNK cell subset in preeclampsia patients and the controls was (17.3 +/- 11.1)% vs (17.9 +/- 16.8)%, that of CD(56)(dim)CD(16)(+) uNK cell subset was (16.3 +/- 8.7)% vs (16.2 +/- 8.8)%; that of CD(56)(+)CD(69)(+) uNK cells was (37.9 +/- 18.9)% vs (36.8 +/- 19.7)%, that of CD(56)(+)CD(94)(+) uNK cells was (34.9 +/- 15.2)% vs (32.7 +/- 16.2)% and the ratio of CD(56)(+)CD(69)(+)/CD(56)(+)CD(94)(+) was 1.1 +/- 0.2, 1.2 +/- 0.6. No statistical difference was shown in the above values between the preeclampsia patients and controls. The percentage of cytotoxic T cell (Tc) 2 cells was significantly lower in the decidua of preeclampsia patients [(3.0 +/- 1.0)% vs (4.3 +/- 0.9)%, P=0.001], and the ratio of Tc1/Tc2 in preeclampsia patients was significantly higher than that of normal term pregnancies (17.8 +/- 3.4 vs 11.8 +/- 4.6; P=0.001); the ratio of Th1/Th2 was increased (15.1 +/- 2.4 vs 13.2 +/- 3.1; P=0.06). CONCLUSIONS: The immunophenotypic characteristics of uNK cells do not present any significant change in preeclampsia patients. Owing to Tc2 cell decrease, the Th1/Th2 immunity shifts to Th1 type immunity in the decidua, which might contribute to the pathogenesis of preeclampsia.

[Recent advances for research on anti-tumor effects of TNF-related apoptosis-inducing ligand].

Tumor necrosis factor (TNF)-related apoptosis--inducing ligand (TRAIL) is a member of TNF family. TRAIL selectively induces apoptosis in a wide variety of tumor cells, but not most normal cells through binding to its receptors. The ability of TRAIL to induce apoptosis in a large number of tumors has stimulated interest in TRAIL as a tumor therapeutic agent.

[Cloning, expression and purification of human TNF-related apoptosis-inducing ligand in Escherichia coli].

OBJECTIVE: To clone human tumor necrosis factor-related apoptosis-inducing ligand (TRAIL, 114-281) cDNA and develop an inducible system for expression in E. coli. METHODS: The human TRAIL (114-281) cDNA was amplified with the total RNA from the human peripheral blood monocytes by RT-PCR. The cDNA was inserted into pMD T vector. The selected integrants were confirmed by PCR screen, digestion with restriction enzymes and DNA sequencing. Then, the insert of human TRAIL cDNA was subcloned into prokaryotic expression vector pET28a. The construction of prokaryotic expression vector was proofed correct by RT-PCR and digestive identification. The recombinant protein was induced by IPTG (isopropyl-beta-D-thiogalactopyranoside) . The sTRAIL inclusion bodies were refolded by dilution method. Refolded protein was purified with column chromatography. RESULTS: Agarose gel electrophoresis of product of RT-PCR revealed a band around 500bp, which was expected. The positive integrants were confirmed by PCR screen and digestion with restriction enzymes. The same band as RT-PCR product was showed by PCR screen and digestion with restriction enzymes. Then the selected integrants were confirmed by DNA sequencing. The sequence was checked in GenBank. The construction of prokaryotic expression vector pET 28a was proofed correct by RT-PCR and digestive identification. TRAIL protein was successfully induced by IPTG in E. coli BL21. The results also showed that the protein was expressed as inclusion bodies. After the sTRAIL inclusion bodies were solubilized and refolded, the protein expressed was purified with one band, which was about 20kD, analyzed by SDS-PAGE. CONCLUSION: These results suggested that the hsTRAIL was cloned, expressed and purified in the present study. Significant quantities of TRAIL produced by this method should allow further studies in determining the physiological significance and function of TRAIL in the future.