Efficacy of inverso isomer of CendR peptide on tumor tissue penetration

The dense extracellular matrix and high interstitial fluid pressure of tumor tissues prevent the ability of anti-tumor agents to penetrate deep into the tumor parenchyma for treatment effects. C-end rule (CendR) peptides can enhance the permeability of tumor blood vessels and tumor tissues binding to neuropilin-1 (NRP-1), thus aiding in drug delivery. In this study, we selected one of the CendR peptides (sequence RGERPPR) as the parent l-peptide and substituted d-amino acids for the l-amino acids to synthesize its inverso peptide (RGERPPR). We investigated the NRP-1 binding activity and tumor-penetrating ability of (RGERPPR). We found that the binding affinity of (RGERPPR) with NRP-1 and the cellular uptake was significantly higher than that of RGERPPR. Evans Blue tests revealed that (RGERPPR) exhibited improved tumor-penetrating ability in C6, U87 and A549 tumor-bearing nude mice. Using nude mice bearing A549 xenograft tumors as a model, we found that the rate of tumor growth in the group co-administered with (RGERPPR) and gemcitabine (Gem) was significantly lower than the gemcitabine-treated group with a tumor suppression rate (TSR%) of 55.4%. Together, our results demonstrate that (RGERPPR) is a potential tumor-penetrating peptide.

Effects on proliferation and apoptosis of T-cell leukemia cells by silencing NRP-1 / 中国免疫学杂志

Objective:To investigate the effect on proliferation and apoptosis of T-cell leukemia cells by silencing NRP-1 ( Jurkat cells).Methods:The lentivirus plasmid which expresses NRP1 gene specific shRNA was constructed in our preliminary ex-perimental.We transfected the lentivirus plasmid to human T-cell Lymphoma cells.The proliferation of Jurkat cells different groups and effect on cell proliferation after chemotherapy drug EPI-treated were found by CCK-8 kit.The proliferation level and apoptosis rate of the cells were detected by flow cytometry and Annexin-V-FITC/PI method.Results:The proliferation level of NRP-1 /shRNA interference group was decreased significantly in 48 h,72 h,96 h,which was compared with the control groups.The apoptosis rate of the NRP-1/shRNA interference group was increased compared with control groups.The chemotherapy drug sensitivity of epirubicin ( EPI ) test results showed that EPI concentration was 0.025,0.05,0.1,0.2,0.4 μg/ml,the NRP-1/shRNA interference group of cell growth inhibition rate was increased,the corresponding control group difference had statistical significance(P<0.05).We choose the drug con-centration of the EPI IC50 for next experiments.NRP-1/shRNA interference group cell apoptosis rate increased significantly after induction,compared with the control groups difference was statistically significant ( P<0.05 ).Compared with control group, the expression level of Bcl-2 protein was decreased and the expression level of bax protein was increased significantly after EPI induction.The percentage of cells at G0/G1 phase increased significantly,while those at S phase decreased significantly.Conclusion:Plasmid shRNA-NRP1 inhibited the expression of NRP1 in Jurkat cells and decreased the proliferation level of Jurkat cells and promote their apoptosis and enhance their drug sensitivity;the molecular mechanism may relate to down-regulation of Bcl-2 and up-regulation of Bax.and arrested the cell cycle at G0/G1 phase.

Role of PLC-PIP2 and cAMP-PKA signal pathways in radiation-induced immune-suppressing effect / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>The purpose of the present study was to observe the changes in CD4+CD25+Nrp1+Treg cells after irradiation with different doses and explore the possible molecular mechanisms involved.</p><p><b>METHODS</b>ICR mice and mouse lymphoma cell line (EL-4 cells) was used. The expressions of CD4, CD25, Nrp1, calcineurin and PKC-α were detected by flow cytometry. The expressions of TGF-β1, IL-10, PKA and cAMP were estimated with ELISA.</p><p><b>RESULTS</b>At 12 h after irradiation, the expression of Nrp1 increased significantly in 4.0 Gy group, compared with sham-irradiation group (P<0.05) in the spleen and thymus, respectively, when ICR mice received whole-body irradiation (WBI). Meanwhile the synthesis of Interleukin 10 (IL-10) and transforming growth factor-β1 (TGF-β1) increased significantly after high dose irradiation (HDR) (> or = 1.0 Gy). In addition, the expression of cAMP and PKA protein increased, while PKC-α, calcineurin decreased at 12h in thymus cells after 4.0 Gy X-irradiation. While TGF-β1 was clearly inhibited when the PLC-PIP2 signal pathway was stimulated or the cAMP-PKA signal pathway was blocked after 4.0 Gy X-irradiation, this did not limit the up-regulation of CD4+CD25+Nrp1+Treg cells after ionizing radiation.</p><p><b>CONCLUSION</b>These results indicated that HDR might induce CD4+CD25+Nrp1+Treg cells production and stimulate TGF-β1 secretion by regulating signal molecules in mice.</p>

Construction and identification of recombinant adenovirus vector of human neuropilin-1 gene / 西安交通大学学报(医学版)

Objective To construct a adenovirus vector containing human NRP-1 gene and 3Flag gene to interaction between tumor and interstitial cell.Methods Plasmid containing NRP-1 gene was digested by AgeⅠand NheⅠ restriction endonuclease.Then the DNA was ligated into linearized pDC315-3Flag vector.After having been constructed,the pDC3 1 5-NRP-1-3 Flag plasmid was co-transfected with framework plasmid pBHGlox△E1 , 3Cre into HEK 293 cells to obtain the homologous recombinant adenovirus,which was then amplified and purified its titer tested.Expression of NRP-1 protein was detected using Western blot.Results Polymerase chain reaction and sequencing analysis confirmed that the shuttle plasmid pDC3 1 5-NRP-1-3 Flag and design were consistent. Cytopathic effect was observed by inverted phase contrast after transfecting HEK2 9 3 cells with shuttle plasmid pDC315-NRP-1-3Flag.95-130 ku was detected by Western lot after transfecting HEK293 cells with shuttle plasmid pDC315-NRP-1-3Flag and recombinant adenovirus,the size being consistent with the NRP-1-3Flag fusion protein (104 ku),with a titer of 2.00E+11PFU/mL.Conclusion The recombinant adenovirus vector for human NRP-1 gene was successfully constructed expressed in HEK 2 9 3 cells.

Establishment and characterization of a targeting tumor vesseles system for inducing thrombosis / 中国药学杂志

METHODS: Chemical cross-linking technique was used to prepare a cross-linking agent of the A6-Streptavidin (A6-SA), MF-A6-SA and Biotein-tTF (B-tTF). FX coagulation assay was used to test MF-A6-SA:B-tTF system's FX activity. Fluorescence microscopy and prussian blue staining were used to simultaneously observe the targeting activity of MF-A6-SA:B-tTF with an external magnetic field. Hemagglutination was directly used to study the system's biological amplification by SA/B. Biodistribution experiment was used to observe the toxicity of MF-A6-SA:B-tTF.