Structure and function of Gab2 and its role in cancer (Review).

The docking proteins of the Grb-associated binder (Gab) family transduce cellular signals between receptors and intracellular downstream effectors, and provide a platform for protein­protein interactions. Gab2, a key member of the Gab family of proteins, is involved in the amplification and integration of signal transduction, evoked by a variety of extracellular stimuli, including growth factors, cytokines and antigen receptors. Gab2 protein lacks intrinsic catalytic activity; however, when phosphorylated by protein­tyrosine kinases (PTKs), Gab2 recruits several Src homology­2 (SH2) domain­containing proteins, including the SH2­containing protein tyrosine phosphatase 2 (SHP2), the p85 subunit of phosphoinositide­3 kinase (PI3K), phospholipase C­Î³ (PLCγ)1, Crk, and GC­GAP. Through these interactions, the Gab2 protein triggers various downstream signal effectors, including SHP2/rat sarcoma viral oncogene/RAF/mitogen­activated protein kinase kinase/extracellular signal­regulated kinase and PI3K/AKT, involved in cell growth, differentiation, migration and apoptosis. It has been previously reported that aberrant Gab2 and/or Gab2 signaling is closely associated with human tumorigenesis, particularly in breast cancer, leukemia and melanoma. The present review aimed to focus on the structure and effector function of Gab2, its role in cancer and its potential for use as an effective therapeutic target.

Effects of recombinant human interleukin-10 on Treg cells, IL-10 and TGF-ß in transplantation of rabbit skin.

The current study aimed to investigate the rejection and survival time of grafted skin, and the changes of Treg cells, interleukin 10 (IL-10) and transforming growth factor-ß (TGF-ß) in peripheral blood following skin transplantation with recombinant human interleukin-10 (rhIL-10) or cyclosporin A (CsA), as well as the role of IL-10 in immunological rejection mechanisms. A total of 36 rabbits were divided into two groups. The skin of a donor rabbit was transplanted onto the back of one receptor rabbit. Receptors were randomly divided into six groups, including rhIL-10 low-dose (5 µg/kg/d), rhIL-10 high-dose (10 µg/kg/d), CsA low-dose (5 mg/kg/d), CsA high-dose (10 mg/kg/d), rhIL-10 (5 µg/kg/d) and CsA (5 mg/kg/d) and negative control normal saline (NS; 1 ml/d). All groups received intramuscular drug injection for ten days, beginning one day prior to skin transplantation surgery. Following transplantation, each rabbit's peripheral blood was collected at different times. The changes of CD4+CD25+ regulatory T cells, IL-10 and TGF-ß were determined by flow cytometry and enzyme-linked immunosorbent assay. When compared with the control group, the rejection and survival times of the experimental groups were longer following skin graft. Compared with the two CsA groups and the control group, the proportion of CD4+CD25+ regulatory T cells of rhIL-10 groups was significantly upregulated on the 4th and 7th days following surgery. However, TGF-ß levels were not significantly different. Data suggested that the concentration of IL-10 was positively correlated with the proportion of CD4+CD25+ regulatory T cells. In addition, IL-10 may delay the rejection time of rabbit skin transplantation and prolong the survival time. Thus, the role of IL-10 in inhibited allograft rejection may be associated with CD4+CD25+ regulatory T cells and IL-10, and may be independent of TGF-ß.

TLR9 signaling repressed tumor suppressor miR-7 expression through up-regulation of HuR in human lung cancer cells.

BACKGROUND: Our recent evidence showed that Toll like receptor 9 (TLR9) signaling could enhance the growth and metastatic potential of human lung cancer cells through repressing microRNA-7 (miR-7) expression. Human antigen R (HuR) has been involved in stabilizing multiple mRNAs in cellular biology. However, whether HuR also contributed to the altered expression of miR-7 in TLR9 signaling stimulated human lung cancer cells remains to be elucidated. METHODS: The expression of HuR in human lung cancer 95D cells treated with TLR9 agonist CpG Oligonucleotides (ODNs) was detected by Real-time PCR and Western blot assay. To explore the possible role of HuR on miR-7 expression, eukaryotic expression vector encoding HuR was transiently transfected into 95D cells and then the expression of miR-7 was detected by Real-time PCR assay. Moreover, RNA interference, western blot, Real-time PCR, MTT assay, BrdU labeling, invasion assay and scratch assay were employed to examine the disrupt effect of HuR on miR-7 expression in human lung cancer cells treated with CpG ODNs. Finally, inhibitors for PI3K, Akt or Erk respectively, and western blot were performed to explore the possible signaling pathway related to HuR expression in CpG ODNs treated human lung cancer cells. RESULTS: Our data showed that TLR9 agonist CpG ODNs could induce the expression of HuR in human lung cancer cells. Moreover, overexpression of HuR could reduce the expression of miR-7 in lung cancer cells. Notably, down-regulation of HuR using RNA interference restored miR-7 expression in CpG ODNs treated lung cancer cells, accompanied by enhanced growth and metastatic potential. Finally, CpG ODNs could induce HuR expression through Akt pathway. CONCLUSION: Our findings indicated that HuR could act as regulator in regulating TLR9 signaling associated biological effect in human lung cancer cells, which might be helpful for the understanding of the potential role of HuR in tumor biology.

Profiling of human leukocyte antigens in Eales disease and tuberculosis.

To analyze the association between Eales disease, histocompatibility leukocyte antigen alleles (HLA-A/B, HLA-DRB/DQB) and tuberculosis infection, and to explore susceptible genes and protective genes associated with Eales disease and tuberculosis infection in a population of Han nationals from Zunyi City in Guizhou Province, China. The subjects were analyzed by a case-control study consisting of three groups--Eales disease group (47 cases), pulmonary tuberculosis group (36 cases) and normal control group (100 cases). The Eales disease group was divided into four parts. Part one consisted of 12 patients who had suffered from pulmonary tuberculosis. Part two consisted of 27 patients who had not suffered from pulmonary tuberculosis. Parts three and four consisted of 27 patients with a positive purified protein derivative test and 12 patients with a negative test, respectively. DNA samples from 47 patients with Eales disease, 36 patients with pulmonary tuberculosis and 100 healthy people were detected by polymerase chain reaction with sequence-specific primers. The 59 HLA-A/B and HLA-DRB/DQB alleles from Eales disease were compared with those from tuberculosis and normal control, and a correlativity test of common susceptible genes was performed to analyze the potential relationship between Eales disease and pulmonary tuberculosis. The frequency distribution of HLA-A*02 alleles (OR 9.719, OR 95 % CI 4.377-21.580, P = 0.000) and HLA-B*07 (OR 11.605, OR 95 % CI 2.397-56.191, P = 0.001) in the Eales disease group was higher than in the normal control group, but the HLA-A*11 alleles (OR 0.495, OR 95 % CI 0.245-1.000, P = 0.048) were lower than in the normal control group, showing a significant difference. Compared with parts two and four, the frequency distribution of HLA-A*02, HLA-A*11 and HLA-B*07 alleles in parts one and three showed no significant difference (P > 0.05). HLA-A*A02, HLA-A*24, HLA-B*07 and HLA-DRB*16 alleles between the Eales disease, pulmonary tuberculosis and normal control group showed statistical significance (P < 0.05). HLA-A*24 alleles in the pulmonary tuberculosis group were lower than the Eales disease group (χ(2) 7.289, P = 0.007), but HLA-A*02 alleles showed no significant difference (P > 0.05) between the two groups. The results show that HLA-A*02 and HLA-B*07 may be genetic predisposing genes, but HLA-A*11 alleles may be protective genes of Eales disease, the HLA-A*02 allele may be a common susceptible gene of Eales disease and pulmonary tuberculosis. HLA-A*11 and HLA-A24 alleles are protective genes of Eales disease and pulmonary tuberculosis, respectively. In summary, the frequency distribution of susceptible genes of Eales disease and pulmonary tuberculosis in a population of Han nationals from Zunyi City in Guizhou Province, China showed no significant difference.

[The change of CD4+ CD25high CCR6+ regulatory T cells in breast cancer patients].

OBJECTIVE: To detect the proportion of CD4+ CD25high CCR6+ regulatory T cells in peripheral blood mononuclear cells and tumor infiltration lymphocytes from breast cancer patients and explore its significance. METHODS: Lymphocytes isolated from blood and tumor mass of breast cancer patients were analyzed for the proportion of CD4+ CD25high CCR6+ regulatory T cells by flow cytometry. The expression of Foxp3 on CD4+ CD25high CCR6+ regulatory T cells, as well as CD45RO, CD44 and CD62L, were also analyzed. The effects of CD4+ CD25high CCR6+ regulatory T cells on the proliferation of CD4+ CD25- T cells also were detected by 3H-incorporation assay. RESULTS: Compared to the control, increased proportion of CD4+ CD25high CCR6+ regulatory T cells was observed in PBMCs and TILs from breast cancer patients. Moreover, CD4+ CD25high CCR6+ regulatory T cells, expressing high level of Foxp3, displayed effector memory phenotype determined by high level expression of CD45RO, CD44 and low level of CD62L. In addition, CD4+ CD25high CCR6+ regulatory T cells could inhibit the proliferation of CD4+ CD25- T cells in vitro. CONCLUSION: The enrichment of CD4+ CD25high CCR6+ regulatory T cells in tumor mass in breast cancer patients, which might be close related to long term immunoescape of tumor.

CXCR4/SDF-1 pathway is crucial for TLR9 agonist enhanced metastasis of human lung cancer cell.

Accumulating data suggested that CXCR4/SDF-1 pathway may play an important role in the metastasis of tumor. We previously demonstrated that CpG ODN could enhance the metastasis of human lung cancer cell via TLR9. Here we further evaluated the possible role of CXCR4/SDF-1 pathway in the enhanced metastasis of human lung cancer 95D cells induced by CpG ODN. Our data showed down-regulation of CXCR4 expression using siRNA against CXCR4 could significantly reduce the enhanced metastasis of 95D cells induced by CpG ODN both in vitro and in vivo. These results suggested that TLR9 agonist might promote the metastasis of human lung cancer cells via CXCR4/SDF-1 pathway.

Analysis of the CDR3 length repertoire and the diversity of TCR alpha chain in human peripheral blood T lymphocytes.

Analysis of complementarity determining region 3 (CDR3) length of T lymphocyte receptors (TCRs) by immunoscope spectratyping technique has been used successfully to investigate the diversity of TCR in autoimmune diseases and infection diseases. In this study, we investigated the patterns of CDR3 length distribution for all 32 TCR AV gene families in human peripheral blood lymphocytes of four normal volunteers by the immunoscope spectratyping technique. It was found that PCR products exhibited an obscure band on 1.5% agarose gel electrophoresis. Each TCR AV family exhibited more than 8 bands on 6% sequencing gel electrophoresis. The CDR3 spectratyping of all TCR AV families showed a standard Gaussian distribution with different CDR3 length, and the expression frequency of CDR3 was similar among the gene families. Most of CDR3 in TCR AV family recombine in frame. However, some of the CDR3 showed out-of frame gene rearrangement. Additionally, we found that in some of TCR AV families there were 18 amino acid discrepancies between the longest CDR3 and shortest CDR3. These results may be helpful to further study the recombination mechanism of human TCR genes, the TCR CDR3 gene repertoire, and the repertoire drift in health people and disease state.