Gut commensal bacteria, Paneth cells and their relations to radiation enteropathy.

In steady state, the intestinal epithelium forms an important part of the gut barrier to defend against luminal bacterial attack. However, the intestinal epithelium is compromised by ionizing irradiation due to its inherent self-renewing capacity. In this process, small intestinal bacterial overgrowth is a critical event that reciprocally alters the immune milieu. In other words, intestinal bacterial dysbiosis induces inflammation in response to intestinal injuries, thus influencing the repair process of irradiated lesions. In fact, it is accepted that commensal bacteria can generally enhance the host radiation sensitivity. To address the determination of radiation sensitivity, we hypothesize that Paneth cells press a critical "button" because these cells are central to intestinal health and disease by using their peptides, which are responsible for controlling stem cell development in the small intestine and luminal bacterial diversity. Herein, the most important question is whether Paneth cells alter their secretion profiles in the situation of ionizing irradiation. On this basis, the tolerance of Paneth cells to ionizing radiation and related mechanisms by which radiation affects Paneth cell survival and death will be discussed in this review. We hope that the relevant results will be helpful in developing new approaches against radiation enteropathy.

MicroRNA-9 enhanced radiosensitivity and its mechanism of DNA methylation in non-small cell lung cancer.

In order to improve the therapeutic effect of non-small cell lung cancer (NSCLC), it is critical to combine radiation and gene therapy. Our study found that the activation of microRNA-9 (miR-9) conferred ionizing radiation (IR) sensitivity in cancer cells. Furthermore, increased microRNA-9 promoter methylation level was observed after IR. Our study combined the IR and microRNA-9 overexpression treatment which leads to a significant enhancement in the therapeutic efficiency in lung cancer both in vitro and in vivo. Therefore, it is plausible that microRNA-9 can be used as a novel therapeutic strategy of NSCLC. MTT assay was performed to detect the effect of microRNA-9 on the survival and growth of NSCLC cells. Flow cytometry results showed that microRNA-9 enhanced the apoptosis of NSCLC cells. Wound healing assay found that microRNA-9 can inhibit the migration of NSCLC cells and enhance the effect of radiation on the migration of NSCLC cells. In addition, bisulfate sequencing PCR was performed to analyze the methylation status of the microRNA-9 promoter. In order to determine the effect of microRNA-9 and its promoter methylation status on proliferation and radio-sensitivity in vivo, a subcutaneous tumor formation assay in nude mice was performed. Results have shown that microRNA-9 overexpression increased the radiosensitivity of A549 cells by inhibiting cell activity and migration, and by increasing apoptosis. In addition, the promoter methylation status of the microRNA-9 gene increased in response to ionizing radiation. Our study demonstrated that microRNA-9 enhanced radiosensitivity in NSCLC and this effect is highly regulated by its promoter methylation status. These results will help to clarify regulatory mechanisms of radiation resistance thus stimulate new methods for improving radiotherapy for NSCLC.

MicroRNA-9 functions as a tumor suppressor and enhances radio-sensitivity in radio-resistant A549 cells by targeting neuropilin 1.

Radiotherapy is commonly used to treat lung cancer but may not kill all cancer cells, which may be attributed to the radiotherapy resistance that often occurs in non-small cell lung cancer (NSCLC). At present, the molecular mechanism of radio-resistance remains unclear. Neuropilin 1 (NRP1), a co-receptor for vascular endothelial growth factor (VEGF), was demonstrated to be associated with radio-resistance of NSCLC cells via the VEGF-phosphoinositide 3-kinase-nuclear factor-κB pathway in our previous study. It was hypothesized that certain microRNAs (miRs) may serve crucial functions in radio-sensitivity by regulating NRP1. Bioinformatics predicted that NRP1 was a potential target of miR-9, and this was validated by luciferase reporter assays. Functionally, miR-9-transfected A549 cells exhibited a decreased proliferation rate, increased apoptosis rate and attenuated migratory and invasive abilities. Additionally, a high expression of miR-9 also significantly enhanced the radio-sensitivity of A549 cells in vitro and in vivo. These data improve understanding of the mechanisms of cell radio-resistance, and suggest that miR-9 may be a molecular target for the prediction of radio-sensitivity in NSCLC.

MSC-derived cytokines repair radiation-induced intra-villi microvascular injury.

Microvascular injury initiates the pathogenesis of radiation enteropathy. As previously demonstrated, the secretome from mesenchymal stem cells contains various angiogenic cytokines that exhibited therapeutic potential for ischemic lesions. As such, the present study aimed to investigate whether cytokines derived from mesenchymal stem cells can repair endothelial injuries from irradiated intestine. Here, serum-free medium was conditioned by human adipose-derived mesenchymal stem cells, and we found that there were several angiogenic cytokines in the medium, including IL-8, angiogenin, HGF and VEGF. This medium promoted the formation of tubules between human umbilical cord vein endothelial cells and protected these cells against radiation-induced apoptosis in vitro. Likewise, our in vivo results revealed that repeated injections of mesenchymal stem cell-conditioned medium could accelerate the recovery of irradiated mice by reducing the serum levels of pro-inflammatory cytokines, including IL-1α, IL-6 and TNF-α, and promoting intra-villi angiogenesis. Herein, intervention by conditioned medium could increase the number of circulating endothelial progenitors, whereas neutralizing SDF-1α and/or inhibiting PI3K would hamper the recruitment of endothelial progenitors to the injured sites. Such results suggested that SDF-1α and PI3K-mediated phosphorylation were required for intra-villi angiogenesis. To illustrate this, we found that conditioned medium enabled endothelial cells to increase intracellular levels of phosphorylated Akt Ser473, both under irradiated and steady state conditions, and to up-regulate the expression of the CXCR4 and CXCR7 genes. Collectively, the present results revealed the therapeutic effects of mesenchymal stem cell-derived cytokines on microvascular injury of irradiated intestine.

Neuropilin 1 expression correlates with the Radio-resistance of human non-small-cell lung cancer cells.

The purpose of this study was to determine the correlation between over-expression of the neuropilin 1 (NRP1) gene and growth, survival, and radio-sensitivity of non-small cell lung carcinoma (NSCLC) cells. 3-[4,5-dimethylthylthiazol-2-yl]-2,5 diphenyltetrazolium broide (MTT) and colony assays were then performed to determine the effect of NRP1 inhibition on the in vitro growth of NSCLC cells. The Annexin V-Fluorescein Isothiocyanate (FITC) apoptosis detection assay was performed to analyse the effect of NRP1 enhancement on apoptosis of NSCLC cells. Transwell invasion and migration assays were employed to examine the metastatic ability of A549 cells post X-ray irradiation. In addition, Western blot assays were carried out to detect the protein level of VEGFR2, PI3K and NF-κB. Finally, to examine the effect of shNRP1 on proliferation and radio-sensitivity in vivo, a subcutaneous tumour formation assay in nude mice was performed. Microvessel density in tumour tissues was assessed by immunohistochemistry. The stable transfected cell line (shNRP1-A549) showed a significant reduction in colony-forming ability and proliferation not only in vitro, but also in vivo. Moreover, shRNA-mediated NRP1 inhibition also significantly enhanced the radio-sensitivity of NSCLC cells both in vitro and in vivo. The over-expression of NRP1 was correlated with growth, survival and radio-resistance of NSCLC cells via the VEGF-PI3K- NF-κB pathway, and NRP1 may be a molecular therapeutic target for gene therapy or radio-sensitization of NSCLC.