OBJECTIVE: To prepare anti-LRRC4 polyclonal antibody and analyze the correlation between the expression of LRRC4 and pathological grades of gliomas in rabbits. METHODS: Appropriate protein sequence with good hydrophilicity and antigenicity was chosen by analyzing with DS Gene 1.1 software. The corresponding nucleic acid sequence amplified by PCR was used to construct a recombinant pGEX-4T-2/276 bp. E.coli JM109 transformed with the recombinant was induced by IPTG to express GST-fusion protein, and the fusion protein expressed as insoluble inclusion bodies. Then the purified inclusion body was used to immunize rabbits. Once the titer of antiserum reached 1:10(8) by indirect ELISA, the serum was collected and purified. The expression-profile of LRRC4 in embryonic tissues and gliomas with various pathological grades were obtained by western blot and immunohistochemistry with the anti-LRRC4 polyclonal antibody. RESULTS: The highly specific anti-LRRC4 polyclonal antibody whose titer reached 1:10(8) was prepared. The relatively specific expression of LRRC4 was detected in the normal brain, but reduced expression or loss of expression in gliomas was also noticed by immunohistochemistry, and there was a correlation between the expression level of lrrc4 and the pathological grade of gliomas. CONCLUSION The anti-LRRC4 polyclonal antibody with high titer and specificity has been obtained. A correlation between the expression level of LRRC4 and the pathological grade of gliomas is detected, which lays the foundation for advanced research of LRRC4.
Animals ; Antibodies, Monoclonal ; immunology ; Antibody Specificity ; immunology ; Blotting, Western ; Brain ; metabolism ; pathology ; Brain Neoplasms ; immunology ; metabolism ; pathology ; Glioma ; immunology ; metabolism ; pathology ; Immunohistochemistry ; Male ; Nerve Tissue Proteins ; biosynthesis ; genetics ; immunology ; Rabbits ; Recombinant Fusion Proteins ; biosynthesis ; immunology

Adult ; Antigens, CD20 ; metabolism ; Brain ; pathology ; Brain Neoplasms ; immunology ; pathology ; CD3 Complex ; metabolism ; Diagnosis, Differential ; Humans ; Leukocyte Common Antigens ; metabolism ; Lymphomatoid Granulomatosis ; immunology ; pathology ; Male

U251 cell is a sensitive cell line to serum, which stops at G0 phase of cell cycle in no-serum medium, and recovers growth when the serum is added into no-serum medium. The cell can express corresponding proteins in different phase of cell cycle. Therefore it is very signification for the study of cell cycle regulation mechanism that explores these proteins. In this paper, the mouse antibody phage display library was added into the bottle in which the serum starvation U251 cells had been cultured, and the special antibody phages were absorbed. Then the absorbed antibody phages were amplified by adding E. coli TG1 and helper phage M13K07. Amplified antibody phages were added into bottle in which the serum cultured cell after serum starvation (follow named as serum recovered cells) were incubated, so that the cell absorbed the no-special antibody phages for the serum starvation cell and the special antibody phages were in supernatant. The remaining no-special antibody phages in the supernatant were discarded by repeating above program 3-4 times. The pure special antibody phages were gotten, and amplified by adding the host cell E. coli TG1 and helper phage M13K07. Then the host bacterium infected special antibody phage was spread on the plate medium with ampicillin, and the monoclonal antibody phages were gotten. Using same as above program, the monoclonal antibody phages absorbed specially for serum recovered U251 cells were obtained when the serum recovered cells instead of serum starvation cells and serum starvation cells instead of serum recovered cells. In this study, ninety-six positive monoclonal antibody phages that absorbed specially the serum starvation cells and eighty-two positive monoclonal antibody phages that absorbed specially the serum recovered cells were obtained. By using cell immunochemistry assay, two special signification antibodies were obtained. one (No.11) was the strong response in serum starvation cells, the other (No.2) was the strong response in serum recovered cells. The antibody No.2 had the distinctive response to the serum recovered cells in different incubation time (15min, 30min, 1h, 2h, 4h, 8h, 12h and 48h) after serum starvation. The results showed that No.2 antibody would be useful to research the factors of cell cycle regulation and apply to tumor diagnosis.
Antibodies, Neoplasm ; genetics ; immunology ; isolation & purification ; Bacteriophages ; genetics ; Brain Neoplasms ; immunology ; pathology ; Cell Line, Tumor ; Escherichia coli ; genetics ; metabolism ; Glioma ; immunology ; pathology ; Humans ; Peptide Library

Secretory meningioma have been described as a distinct variant of meningioma based on their histologic, immunohistochemical and ultrastructural features of epithelial and secretory differentiation of meningothelial cells with accumulation of secretory material in the form of hyaline inclusion. Secretory meningioma is also a benign tumor having similar biological behaviour to that of typical meningiomas: hence, it is important for it to be recognized and diagnosed correctly to avoid unnecessary radiation and chemotherapy. Here we present a case of secretory meningioma with typical morphologic features. The patient was a 56-year-old woman with bilateral visual disturbance. A well-circumscribed mass was present in the left frontal lobe of cerebrum with surrounding edema. The tumor was composed of whorls of meningothelial cells and abundant intra- and extracellular eosinophilic hyaline inclusions which showed immunoreactivity for epithelial membrane antigen(EMA) and carcinoembryonic antigen(CEA). Ultrastructural features also supported epithelial and secretory differentiation of tumor cells.
Brain Neoplasms/immunology/*pathology ; CA-15-3 Antigen/analysis ; Carcinoembryonic Antigen/analysis ; Case Report ; Female ; Human ; Meningioma/immunology/*pathology ; Middle Age ; Tomography Scanners, X-Ray Computed

Brain metastasis was a common metastasis site and leading cause of death in non-small cell lung cancer (NSCLC). Tyrosine kinase inhibitors had improved survival of NSCLC patients with positive drive gene. It also brings good news to NSCLC patients with positive drive gene and brain metastases. However, there is still no effective treatment for NSCLC patients with drive gene-negative and brain metastases. In recent years, immunotherapy has made breakthrough progress and become important first and second line treatment options of NSCLC especially in patients with drive gene-negative. The role of immunotherapy in specific populations of NSCLC-brain metastasis patients, especially drive gene-negative patients has become the focus of attention. In this report, we review the research progress of immunotherapy in NSCLC with brain metastases, especially in driver-negative patients, analyze the limitations of existing research and future challenge.
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Brain Neoplasms ; immunology ; secondary ; therapy ; Carcinoma, Non-Small-Cell Lung ; genetics ; pathology ; Humans ; Immunotherapy ; methods ; Lung Neoplasms ; genetics ; pathology ; Patient Selection

Dendritic cells (DCs) derived from bone marrow cells are specialized cells for the uptake, processing, and presentation of foreign and self-antigens. The study indicated that re-transfusion of DCs pulsed with tumor-associated antigen can induce an vigorous specific anti-tumor response in clinic. The present study was aimed to investigate the enhancing effect of DCs derived from human cord blood on T cells in killing tumor cells. Human cord blood mononuclear cells were isolated from human cord blood by density gradient centrifugation using lymphocyte separating medium, and cord blood mononuclear cells were obtained by adherence and cultured in a liquid culture system with GM-CSF and IL-4 for 15 days. Then the cells were analyzed for phenotypes of CD1a by indirect immunofluorescence. The capacity of DCs to initiate T cell-dependent anti-tumor immune responses was assayed by MTT kit. The ratios of DCs to tumor cells in experimental groups were 20:1, 50:1 and 100:1 respectively. The DCs were not added in control group. The results indicated that in the presence of GM-CSF and IL-4, the DCs with typical morphological features at days 15 were observed. At that time, (43.12 +/- 5.83)% CD1a(+) cells were obtained. In addition to these phenotypic properties, the DC of experimental groups could remarkably initiate T cell-dependent anti-tumor immune responses with different ratios compared with control group (P < 0.01), there were no significant difference of killing effects between 100:1 and 50:1 groups (P > 0.05), and killing effect of DC in 20:1 group was higher than that in 100:1 or 50:1 groups (P < 0.05). It is concluded that human cord blood mononuclear cells can serve as a better source of DC, which can promote the capacity to initiate T cell-dependent anti-tumor immune responses.
Apoptosis ; immunology ; Brain Neoplasms ; pathology ; Cell Differentiation ; Cells, Cultured ; Dendritic Cells ; cytology ; immunology ; Fetal Blood ; cytology ; Granulocyte-Macrophage Colony-Stimulating Factor ; pharmacology ; Humans ; Interleukin-4 ; pharmacology ; Leukocytes, Mononuclear ; cytology ; Neuroblastoma ; pathology ; Recombinant Proteins ; T-Lymphocytes ; immunology ; Tumor Cells, Cultured

Gliomas are extremely aggressive brain tumors with a very poor prognosis. One of the more promising strategies for the treatment of human gliomas is targeted immunotherapy where antigens that are unique to the tumors are exploited to generate vaccines. The approach, however, is complicated by the fact that human gliomas escape immune surveillance by creating an immune suppressed microenvironment. In order to oppose the glioma imposed immune suppression, molecules and pathways involved in immune cell maturation, expansion, and migration are under intensive clinical investigation as adjuvant therapy. Toll-like receptors (TLRs) mediate many of these functions in immune cell types, and TLR agonists, thus, are currently primary candidate molecules to be used as important adjuvants in a variety of cancers. In animal models for glioma, TLR agonists have exhibited antitumor properties by facilitating antigen presentation and stimulating innate and adaptive immunity. In clinical trials, several TLR agonists have achieved survival benefit, and many more trials are recruiting or ongoing. However, a second complicating factor is that TLRs are also expressed on cancer cells where they can participate instead in a variety of tumor promoting activities including cell growth, proliferation, invasion, migration, and even stem cell maintenance. TLR agonists can, therefore, possibly play dual roles in tumor biology. Here, how TLRs and TLR agonists function in glioma biology and in anti-glioma therapies is summarized in an effort to provide a current picture of the sophisticated relationship of glioma with the immune system and the implications for immunotherapy.
Animals ; Antigens, Neoplasm ; chemistry ; immunology ; Antineoplastic Agents ; chemistry ; immunology ; therapeutic use ; Brain Neoplasms ; genetics ; immunology ; pathology ; therapy ; Chemotherapy, Adjuvant ; Clinical Trials as Topic ; Disease Models, Animal ; Gene Expression Regulation, Neoplastic ; drug effects ; immunology ; Glioma ; genetics ; immunology ; pathology ; therapy ; Humans ; Immunotherapy ; methods ; Signal Transduction ; Toll-Like Receptors ; agonists ; genetics ; immunology

This study was aimed to further explore whether brain derived neurotrophic factor (BDNF) pathway is a potential therapeutic target in multiple myeloma (MM) and whether anti-BDNF monoclonal antibody can prevent the development of this disease. The in vivo antitumor effect of anti-BDNF monoclonal antibody (McAb) on a human myeloma xenograft animal model was evaluated. The model of xenograft tumors was established in the nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice by subcutaneous injection of human myeloma cell line RPMI8226. The antibodies were injected intraperitoneally at a dose of 20 microg/mouse at day 1, 2, 3 after inoculation or at a dose of 100 microg/mouse once a week after tumors were detected. The microvascular densities in tumors were analyzed by immunohistochemistry study. The effect of anti-BDNF McAb on the proliferation of RPMI8226 cells in vitro and on endothelial cells network formation in the co-culture system were determined by using a (3)H-thymidine incorporation assay and a Matrigel network formation assay, respectively. The results showed that multiple injections of anti-BDNF McAb reduced the tumor size, decreased the microvascular density and significantly prolonged tumor-free time and survival time. Moreover, the proliferation of RPMI8226 cells was inhibited in vitro by anti-BDNF McAb, but not by the control IgG. Anti-BDNF McAb also inhibited RPMI8226-induced network formation in endothelial cells in vitro. It is concluded that anti-BDNF monoclonal antibody can inhibit cell growth and angiogenesis in subcutaneous plasmacytoma.
Animals ; Antibodies, Monoclonal ; therapeutic use ; Brain-Derived Neurotrophic Factor ; immunology ; Cell Line, Tumor ; Humans ; Male ; Mice ; Mice, SCID ; Multiple Myeloma ; drug therapy ; metabolism ; pathology ; Neoplasms, Plasma Cell ; drug therapy ; Xenograft Model Antitumor Assays

Animals ; Antigens, CD20 ; metabolism ; Brain Neoplasms ; metabolism ; pathology ; Cell Line, Tumor ; Disease Models, Animal ; Female ; Humans ; Ki-67 Antigen ; metabolism ; Leukocyte Common Antigens ; metabolism ; Lymphoma, Large B-Cell, Diffuse ; immunology ; pathology ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Neoplasm Transplantation