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Mesenchymal stem cells (MSC) are important cellular component of the bone marrow microenvironment in supporting hemopoiesis. Li J et al reported previously that MSCs are resistant to chemotherapy commonly used in hematologic malignancies but are relatively sensitive to anti-microtubule agents. However, the response of MSCs to other chemotherapeutic agents commonly used in solid tumour settings remains unknown. This study was purposed to evaluate the acute direct effects of 4 individual chemotherapeutic agents on human MSCs (hMSC), including cisplatin, topotecan, daunorubicin and hydroxyurea. Using an in vitro culture system, the chemosensitivity of hMSC was determined by XTT assay and compared with NB-4 cells and normal peripheral blood mononuclear cells (PBMNC). The recovery of cell numbers following exposure to chemotherapeutic agents and apoptosis induced by chemotherapy in hMSC were evaluated. The results showed that although hMSCs were more resistant to the 4 agents above mentioned than NB-4 cells, they were sensitive to topotecan, cisplatin and daunorubicin than PBMNCs. The IC₅₀ values of hMSCs for topotecan, cisplatin, hydroxyurea and daunorubicin were 636, 24.8, > 20 and 2.4 times of those of NB-4 cells respectively. The IC₅₀ values of human PBMNCs for topotecan, cisplatin and daunorubicin were > 27, 1.9 and 1.4 times of those of hMSCs respectively. Reduction of cell number was observed in hMSCs treated with the 4 drugs in clinically relative concentrations. Sustained suppression in hMSCs was observed following 3 days exposure to the 4 agents. It is concluded that the cisplatin, topotecan, daunorubicin and hydroxyurea alone can induce apoptosis of hMSCs and exert persistent suppressive effect on the proliferation of hMSCs even with short term exposure.

Objective: To observe the epithelial Toll like receptor (TLR)4 expression changes and the signaling pathway function after respiratory syncytial virus (RSV) infection and to explore the mechanisms of RSV-induced airway inflammation., Methods: 9HTEo-human tracheal epithelial cell line was infected by RSV (MOI = 10), and TLR1-10 mRNA were detected by RT-PCR assay at 3 h post RSV infection. TLR4 mRNA was detected by real time Q-PCR assay at 3 h, 6 h and 9 h post RSV infection, and TLR4 protein expression and cell apoptosis were determined by flow cytometry at 24 h post RSV infection. IL-8 in supernatant was detected by ELISA after RSV-infected cells exposed to lipopolysaccharide (LPS). A normal control group and a RSV infection group were set up for the RT-PCR and flow cytometry experiments, and the data were analyzed by paired t test using GraphPad 4.0 software. A normal group, a RSV group and a UV-inactivated RSV group were set up for the real time Q-PCR, experiments, and the data were analyzed by Kruskal-Wallis test. The ELISA experiments were divided into 4 groups including a normal control, a RSV, a LPS stimulation, and a RSV plus LPS co-stimulation groups, and the data were analyzed by One-way ANOVA test., Results: (1) TLR2-10 mRNA level was significantly up-regulated (t value of TLR2-10: 3.49 -14.47, P < 0.05), especially TLR-2, 6 enhanced expression, compared with the normal epithelial cells. Real time Q-PCR assay showed that TLR4 mRNA started to increase at 3hr (Kruskal-Wallis test value = 8.82, P < 0.05, n = 6) and significantly elevated at 9 hour (Kruskal-Wallis test value = 6.62, P < 0.05, n = 6). UV inactivated-RSV had no effect on the TLR4 mRNA level. (2) Flow cytometry showed that membrane TLR4 mean fluorescence intensity (MFI) increased (RSV: 1.27 +/- 0.48, normal: 0.97 +/- 0.25; t = 2.39, P > 0.05, n = 10) while cytoplasmic TLR4 MFI simultaneously decreased (RSV: 3.08 +/- 1.38, normal: 3.36 +/- 1.31, t = 2.92, P = 0.225, n = 10). Percentage of membrane TLR4-positive cells was higher in RSV infected population [RSV: (11.99 +/- 7.74)%, normal: (1.16 +/- 0.47)%, Mann-Whitney t value = 0.001, P < 0.01, n = 8], most (93.32 +/- 1.7)% of which were Annexin V positive. IL-8 was significantly induced in the RSV plus LPS costimulation group compared with RSV group (F = 59.29, P < 0.01, n = 3)., Conclusions: RSV induced epithelial TLR4 up-regulation, localization changes from cytoplasm to membrane, IL-8 secretion through TLR4 signaling pathway and epithelial cell apoptosis in membrane TLR4 positive population. These results indicate TLR4 is involved in RSV-induced acute or chronic epithelial-dependent inflammation, which might contribute to acute or chronic airway inflammation.