Irradiated lung cancer cell-derived exosomes modulate macrophage polarization by inhibiting MID1 via miR-4655-5p.

Radiation Pneumonitis (RP) is one of the most common and severe complication in patients receiving thoracic radiotherapy. The release of cytokines contribute to activating the RP process. Macrophages also play an important role in the pathogenesis of RP. The differential activation of macrophages is regulated by microRNA (miRNA). Exosomes containing miRNAs are one of the important ways to mediate cellular communication. However, the exosomes mediate communication between tumor cells and macrophages during the pathogenesis of RP remains understudied. In this study, we isolated and characterized the exosomes secreted by lung cancer cells after irradiation. Co-culture of exosomes with macrophages revealed that exosomes could induce macrophage proliferation activation and M2 polarization. miRNA array was used to analyze the differential expression of miRNAs in exosomes, and it was found that miR-4655-5p was stably and highly expressed in exosomes. The function of miR-4655-5p in macrophages was confirmed by overexpression/inhibition of miR-4655-5p expression in macrophages. The targeting association between miR-4655-5p and MID1 was determined by bioinformatics prediction followed by a confirmatory dual luciferase reporter assay. We showed that miR-4655-5p regulate the macrophage proliferation and inflammatory response by forming a negative regulatory loop that alters MID1 activity and its downstream PP2Ac. Overall, our results indicated that exosomal miR-4655-5p secreted by lung cancer cells after irradiation promoted the proliferation and M2 polarization of macrophages. It can be speculated that exosomes play an immunomodulatory role in the pathogenesis of RP and provided a new target for the prevention and treatment of RP.

BPA modulates the WDR5/TET2 complex to regulate ERß expression in eutopic endometrium and drives the development of endometriosis.

Overexpression of estrogen receptor ß (ERß) in endometrium contributes to endometriosis (EM) pathogenesis. Trimethylation of the H3 lysine (K) 4 (H3K4me3) in promoters is strongly correlated with gene expression. This study aimed to explore the effects of bisphenol A (BPA) exposure on EM development from the perspective of the regulation of ERß expression in eutopic endometrium via the H3K4me3-related epigenetic pathway. A mouse EM model was established to investigate the effects of BPA. Immortalized human normal endometrial stromal cells (iESCs) were cultured and treated with BPA to explore the underlying mechanism. Eutopic endometria from patients with or without EM were collected and analyzed. Results showed that BPA elevated ERß expression in mouse eutopic endometrium and promoted lesion growth. BPA also promoted WD repeat domain 5 (WDR5) expression and upregulated H3K4me3 levels in the ERß promoter and Exon 1. Further research indicated that WDR5 interacted with tet methylcytosine dioxygenase 2 (TET2), while BPA exposure enhanced the interaction between these two proteins, promoted the recruitment of the WDR5/TET2 complex to the ERß promoter and Exon 1, and inhibited DNA methylation of CpG islands. The WDR5/TET2 interaction was essential for BPA-induced ERß overexpression. Enhanced WDR5/TET2 interaction was also observed in eutopic endometria from EM patients. Further results showed that BPA upregulated WDR5 expression through the G protein-coupled estrogen receptor (GPER)-mediated PI3K/mTOR signaling pathway. In conclusion, our study suggests that BPA exposure promotes EM development by upregulating ERß expression in eutopic endometrium via the WDR5/TET2-mediated epigenetic pathway.

Structural basis for Epstein-Barr virus host cell tropism mediated by gp42 and gHgL entry glycoproteins.

Herpesvirus entry into host cells is mediated by multiple virally encoded receptor binding and membrane fusion glycoproteins. Despite their importance in host cell tropism and associated disease pathology, the underlying and essential interactions between these viral glycoproteins remain poorly understood. For Epstein-Barr virus (EBV), gHgL/gp42 complexes bind HLA class II to activate membrane fusion with B cells, but gp42 inhibits fusion and entry into epithelial cells. To clarify the mechanism by which gp42 controls the cell specificity of EBV infection, here we determined the structure of gHgL/gp42 complex bound to an anti-gHgL antibody (E1D1). The critical regulator of EBV tropism is the gp42 N-terminal domain, which tethers the HLA-binding domain to gHgL by wrapping around the exterior of three gH domains. Both the gp42 N-terminal domain and E1D1 selectively inhibit epithelial-cell fusion; however, they engage distinct surfaces of gHgL. These observations clarify key determinants of EBV host cell tropism.

Radiotherapy concurrent versus sequential with endocrine therapy in breast cancer: A meta-analysis.

INTRODUCTION: This study compared treatment outcomes of radiotherapy concurrent with endocrine therapy and radiotherapy sequential with endocrine therapy in breast cancer. MATERIALS AND METHODS: Eligible studies of radiotherapy concurrent and sequential with endocrine therapy in breast cancer were retrieved through extensive searches of the PubMed, Medline, Embase, Cochrane library, FEBM, FMJS, Web of science, Wiley, CBM, CNKI, Wang fang, Cqvip databases from 2000 to 2014. Original English and Chinese publications of radiotherapy concurrent and sequential with endocrine therapy in breast cancer were included. The primary endpoint was radiation-induced toxicity including upper than grade 2 skin related toxicity, radiation pneumonia and pulmonary fibrosis; the second endpoint was survival date, including local recurrence, distant metastasis, 5-year OS, 10-year OS. RESULTS: Eleven eligible trials were identified, six in English and five in Chinese. Totally, there were 1291 women in concurrent groups, and 1179 in sequential groups. Statistical analysis showed that there was no statistical difference between concurrent and sequential groups in skin related toxicity (RR 1.20, 95% CI 0.92-1.56, P = 0.17), radiation pneumonia (RR 1.11, 95% CI 0.46-2.70, P = 0.81) and pulmonary fibrosis (RR 1.35, 95% CI 0.75-2.41, P = 0.32). Meanwhile, no statistical difference was found in survival data, (RR 0.97, 95% CI 0.79-1.28, P = 0.26), (RR 0.86, 95% CI 0.66-1.12, P = 0.27) in local recurrence and distant metastasis respectively, (RR 1.01, 95% CI 0.96-1.06, P = 0.65), (RR 0.98, 95% CI 0.93-1.02, P = 0.32) in 5-year and 10-year overall survival respectively. Stratification analysis was proceeded, grouped by tamoxifen and AI in different treatment timing, however, no statistical difference was found in radiation-induced toxicity and survival outcomes. CONCLUSION: Radiotherapy concurrent with endocrine therapy didn't increase or decrease neither the incidence of radiation-induced toxicity nor the survival rate compared with that of sequential group; Endocrine therapy drugs didn't influence outcomes in different treatment timing.

MicroRNAs level as an initial screening method for early-stage lung cancer: a bivariate diagnostic random-effects meta-analysis.

Accumulating studies suggested that microRNAs (miRNAs) can have high diagnostic value as a non-invasive and cost-effective procedure with high sensitivity and specificity in the detection of early-stage lung cancer. However, there is inconsistency observed in the results of relevant studies. Therefore, we performed this meta-analysis to evaluate diagnostic value of miRNAs based on all related studies. A total of 38 studies from 13 included articles were used for the analysis, consisting of 510 patients and 465 healthy controls. All analyses were performed on the R 3.2.0 software. The bivariate random-effects meta-analysis model was applied to obtain the following pooled parameters: sensitivity, 0.797 (95% CI: 0.756-0.832); false positive rate, 0.296 (95% CI: 0.250-0.346); and AUC, 0.818. In addition, subgroup analyses were conducted, showing not only that a combination of multiple miRNAs as biomarkers have greater diagnostic value for early-stage lung cancer (sensitivity, false positive rate and AUC of 83%, 25.2% and 0.858, respectively) had a higher diagnostic accuracy than single miRNA (sensitivity, false positive rate and AUC of 78.3%, 31.6% and 0.799, respectively), but also that specimen from circulating system (sensitivity, false positive rate and AUC of 82.5%, 30.5% and 0.836, respectively) provide better biomarkers than specimen from non-circulating system (sensitivity, false positive rate and AUC of 73.8%, 26.5% and 0.796, respectively). In summary, the current meta-analysis suggests that miRNAs as biomarkers, particularly a combination of multiple tumor-specific miRNAs from circulating system, have moderately high clinical diagnostic value in the detection of early-stage lung cancer. However, the clinical diagnostic utilization and additional improvements of miRNAs as biomarkers for early-stage lung cancer detection still remain to be further validated by more future studies.

Microarray Analysis of Long Non-coding RNA Expression Profile Associated with 5-Fluorouracil-Based Chemoradiation Resistance in Colorectal Cancer Cells.

BACKGROUND: Preoperative 5-fluorouracil (5-FU)-based chemoradiotherapy is a standard treatment for locally advanced colorectal cancer (CRC). However, CRC cells often develop chemoradiation resistance (CRR). Recent studies have shown that long non-coding RNA (lncRNA) plays critical roles in a myriad of biological processes and human diseases, as well as chemotherapy resistance. Since the roles of lncRNAs in 5-FU-based CRR in human CRC cells remain unknown, they were investigated in this study. MATERIALS AND METHODS: A 5-FU-based concurrent CRR cell model was established using human CRC cell line HCT116. Microarray expression profiling of lncRNAs and mRNAs was undertaken in parental HCT116 and 5-FU-based CRR cell lines. RESULTS: In total, 2,662 differentially expressed lncRNAs and 2,398 mRNAs were identified in 5-FU-based CRR HCT116 cells when compared with those in parental HCT116. Moreover, 6 lncRNAs and 6 mRNAs found to be differentially expressed were validated by quantitative real time PCR (qRT-PCR). Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis for the differentially expressed mRNAs indicated involvement of many, such as Jak- STAT, PI3K-Akt and NF-kappa B signaling pathways. To better understand the molecular basis of 5-FU-based CRR in CRC cells, correlated expression networks were constructed based on 8 intergenic lncRNAs and their nearby coding genes. CONCLUSIONS: Changes in lncRNA expression are involved in 5-FU-based CRR in CRC cells. These findings may provide novel insight for the prognosis and prediction of response to therapy in CRC patients.

Assembly and architecture of the EBV B cell entry triggering complex.

Epstein-Barr Virus (EBV) is an enveloped double-stranded DNA virus of the gammaherpesvirinae sub-family that predominantly infects humans through epithelial cells and B cells. Three EBV glycoproteins, gH, gL and gp42, form a complex that targets EBV infection of B cells. Human leukocyte antigen (HLA) class II molecules expressed on B cells serve as the receptor for gp42, triggering membrane fusion and virus entry. The mechanistic role of gHgL in herpesvirus entry has been largely unresolved, but it is thought to regulate the activation of the virally-encoded gB protein, which acts as the primary fusogen. Here we study the assembly and function of the reconstituted B cell entry complex comprised of gHgL, gp42 and HLA class II. The structure from negative-stain electron microscopy provides a detailed snapshot of an intermediate state in EBV entry and highlights the potential for the triggering complex to bring the two membrane bilayers into proximity. Furthermore, gHgL interacts with a previously identified, functionally important hydrophobic pocket on gp42, defining the overall architecture of the complex and playing a critical role in membrane fusion activation. We propose a macroscopic model of the initiating events in EBV B cell fusion centered on the formation of the triggering complex in the context of both viral and host membranes. This model suggests how the triggering complex may bridge the two membrane bilayers, orienting critical regions of the N- and C- terminal ends of gHgL to promote the activation of gB and efficient membrane fusion.

[Surface-enhanced Raman spectra analysis of trace degradation products from goat horn].

Nano-silver colloid was synthesized by using microwave method on the mixtures of sodium citrate solution and silver nitrate solution. The method has advantages of fast heating speed, uniform temperature distribution and easily controlled reaction conditions. The sizes and size distributions of the silver particles were characterized by means of quasi-elastic laser scattering (QLS). The average particles size was (53.27 +/- 2.65) nm and the size of the particles was mainly distributed around 56 nm. Surface-enhanced Raman spectra of the degradation products from goat horn were obtained with silver colloid as active substrate. It was observed that the Raman signal of SERS was enhanced significantly compared with that of regular Raman spectrum, especially at the Raman bands of 659, 830, 850, 929, 999, 1 028, 1 280, 1 439 and 1 599 cm(-1) which reflect the biochemical components in degradation products. The characteristic Raman bands of degradation products from goat horn were preliminary assigned. The assignments showed that the main constituents of the degradation products from goat horn were amino acids and polypeptides. It was for the first time that Surface-enhanced Raman spectroscopy was used to detect trace degradation products from the horns. Raman signal enhancement can be obtained with high sensitivity for the trace concentrations as low as ppm level. It is concluded that surface-enhanced Raman spectroscopy can provide a fast, direct and precise detecting method for the detection of trace degradation solution from horns.

Characteristic variation of α-fetoprotein DNA nanometer-range irradiated by iodine-125.

To obtain the characteristic variation of structure and functional groups of α-fetoprotein (AFP) DNA irradiated by iodine-125((125)I), the AFP antisense oligonucleotide labeled with various radioactivity dose (125)I was mixed with the AFP DNA in a simulated polymerase chain reaction temperature condition. After the mixtures were irradiated by the (125)I from 2 to 72 hours, the mutation of the biogenic conformation and functional groups of the irradiated DNA were investigated using laser Raman spectroscopy. The shifted peak and the decreased intensity of the characteristic Raman spectra were found, which demonstrated that the structure of the phosphodiester linkage was broke, the pyridine and purine bases in DNA emerged and damaged. The model of gene conformation changed from form B to form C spectrum after the nanometer-range irradiation with (125)I from 2 to 24 hours. The damage of local pyridine and purine bases gradually increased along with the accumulation of irradiation, and the bases and ribosome were finally dissociated and stacked.

Reactive oxygen species formation and apoptosis in human peripheral blood mononuclear cell induced by 900 MHz mobile phone radiation.

We demonstrate that reactive oxygen species (ROS) plays an important role in the process of apoptosis in human peripheral blood mononuclear cell (PBMC) which is induced by the radiation of 900 MHz radiofrequency electromagnetic field (RFEMF) at a specific absorption rate (SAR) of ~0.4 W/kg when the exposure lasts longer than two hours. The apoptosis is induced through the mitochondrial pathway and mediated by activating ROS and caspase-3, and decreasing the mitochondrial potential. The activation of ROS is triggered by the conformation disturbance of lipids, protein, and DNA induced by the exposure of GSM RFEMF. Although human PBMC was found to have a self-protection mechanism of releasing carotenoid in response to oxidative stress to lessen the further increase of ROS, the imbalance between the antioxidant defenses and ROS formation still results in an increase of cell death with the exposure time and can cause about 37% human PBMC death in eight hours.

ß,ß-Dimethylacrylshikonin exerts antitumor activity via Notch-1 signaling pathway in vitro and in vivo.

ß,ß-Dimethylacrylshikonin (DA) is a major component of Radix Lithospermum erythrorhizon and has various biological activities. We have investigated the inhibitory effect of DA on the growth of hepatocellular carcinoma in vitro and in vivo. Notch signaling plays a critical role in maintaining the balance between cell proliferation, differentiation and apoptosis. Hence, perturbed Notch signaling may contribute to tumorigenesis. In the present study, we evaluated whether DA could be an effective inhibitor on cell growth in human gastric cancer cell line, and also the molecular mechanisms. Using multiple cellular and molecular approaches such as MTT assay, colony formation assay, DAPI staining, flow cytometry, real-time PCR and Western blot analysis, we found that DA inhibited cell growth in a dose- and time-dependent manner. Biochemical analysis revealed the involvement of cell cycle regulated proteins in DA-mediated of G0-G1 arrest of SGC-7901 cells. Furthermore, DA treatment led to reduced Notch-1 activation, expression of Jagged-1 and its downstream target Hes-1 in vitro and in vivo. Our data demonstrated that DA is a potent inhibitor of progression of gastric cancer cells, which could be due to attenuation of Notch-1. We also suggest that DA could be further developed as a potential therapeutic agent for the treatment of gastric cancer.

Over-expression of extracellular matrix metalloproteinase inducer in prostate cancer is associated with high risk of prostate-specific antigen relapse after radical prostatectomy.

PURPOSE: The prognostic efficiency of clinical grading and staging in patients with confined or moderately differentiated prostate cancer (PCa) has been markedly improved, which underscores the importance of new prognostic markers. Extracellular matrix metalloproteinase inducer (EMMPRIN) has been demonstrated to be involved in cancerangiogenesis, metastasis and invasion. EMMPRIN expression was evaluated by measuring mRNA and protein levels in a large cohort of patients with PCa following prostatectomy and the findings were compared with clinico-pathological parameters, including prostate-specific antigen (PSA) relapse time. METHODS: EMMPRIN mRNA levels in 20 pairs of normal and cancerous prostate tissues were determined by quantitative real-time PCR. Protein expression in paraffin-embedded specimens of prostates gathered from 300 patients with PCa was detected by immunohistochemistry using a monoclonal antibody against EMMPRIN. The associations of EMMPRIN protein expression with the clinico-pathological parameters and PSA relapse-free time after radical prostatectomy were subsequently assessed. RESULTS: Both EMMPRIN mRNA and protein levels were higher in PCa tissue, compared with adjacent normal tissue. In addition, the positive expression rates of EMMPRIN in PCa tissues were significantly associated with preoperative PSA levels (p=0.008), AJCC stage (p=0.006) and Gleason Score (p < 0.001), Risk classification (p < 0.001), lymph node status post-surgery (p < 0.001) and surgical margin status (p < 0.001) were also determined. Multivariate analysis, using the Cox proportional hazards model, revealed that positive EMMPRIN expression was an independent prognostic factor for an increased risk of PSA relapse. CONCLUSION: Over-expression of EMMPRIN correlated with the aggressiveness of PCa, and the PSA relapse-free time, and may be a novel and useful biomarker for follow-up and treatment decisions for PCa.

Quantum dots as a sensor for quantitative visualization of surface charges on single living cells with nano-scale resolution.

We developed a technique using quantum dot (QD) as a sensor for quantitative visualization of the surface charge on biological cells with nano-scale resolution. The QD system was designed and synthesized using amino modified CdSe/ZnS nanoparticles. In a specially designed buffer solution, they are positively charged and can homogeneously disperse in the aqueous environment to label all the negative charges on the surfaces of living cells. Using a wide-field optical sectioning microscopy to achieve 2D/3D imaging of the QD-labeled cells, we determined the charge densities of different kinds of cells from normal to mutant ones. The information about the surface charge distribution is significant in evaluating the structure, function, biological behavior and even malignant transformation of cells.

The periplasmic bacterial molecular chaperone SurA adapts its structure to bind peptides in different conformations to assert a sequence preference for aromatic residues.

The periplasmic molecular chaperone protein SurA facilitates correct folding and maturation of outer membrane proteins in Gram-negative bacteria. It preferentially binds peptides that have a high fraction of aromatic amino acids. Phage display selections, isothermal titration calorimetry and crystallographic structure determination have been used to elucidate the basis of the binding specificity. The peptide recognition is imparted by the first peptidyl-prolyl isomerase (PPIase) domain of SurA. Crystal structures of complexes between peptides of sequence WEYIPNV and NFTLKFWDIFRK with the first PPIase domain of the Escherichia coli SurA protein at 1.3 A resolution, and of a complex between the dodecapeptide and a SurA fragment lacking the second PPIase domain at 3.4 A resolution, have been solved. SurA binds as a monomer to the heptapeptide in an extended conformation. It binds as a dimer to the dodecapeptide in an alpha-helical conformation, predicated on a substantial structural rearrangement of the SurA protein. In both cases, side-chains of aromatic residues of the peptides contribute a large fraction of the binding interactions. SurA therefore asserts a recognition preference for aromatic amino acids in a variety of sequence configurations by adopting alternative tertiary and quaternary structures to bind peptides in different conformations.

Improvement in erectile dysfunction after insulin-like growth factor-1 gene therapy in diabetic rats.

AIM: To determine whether adenoviral gene transfer of insulin like growth factor-1 (IGF-1) to the penis of streptozotocin (STZ)-induced diabetic rats could improve erectile capacity. METHODS: THE STZ diabetic rats were transfected with AdCMV-betagal or AdCMV-IGF-1. These rats underwent cavernous nerve stimulation to assess erectile function and their responses were compared with those of age-matched control rats 1 to 2 days after transfection. In control and transfected STZ diabetic rats, IGF-1 expression were examined by reverse transcription polymerase chain reaction (RT-PCR), Western blot and histology. The penis beta-galactosidase activity and localization of the STZ diabetic rats were also determined. RESULTS: One to two days after transfection, the beta-galactosidase was found in the smooth muscle cells of the diabetic rat penis transfected with AdCMV-betagal. One to 2 days after administration of AdCMV-IGF-1, the cavernosal pressure, as determined by the ratio of maximal intracavernous pressure-to-mean arterial pressure (ICP/MAP) and total intracavernous pressure (ICP), was increased in response to cavernous nerve stimulation. Transgene expression was confirmed by RT-PCR, Western blot and histology. CONCLUSION: Gene transfer of IGF-1 significantly increased erectile function in the STZ diabetic rats. These results suggest that in vivo gene transfer of IGF-1 might be a new therapeutic intervention for the treatment of erectile dysfunction (ED) in the STZ diabetic rats.

An active artificial cornea with the function of inducing new corneal tissue generation in vivo-a new approach to corneal tissue engineering.

An active artificial cornea which can perform the function of inducing new cornea generation in vivo but does not need culture cells in vitro and which has similar optical and mechanical properties to those of the human cornea was constructed. An animal keratoplasty experiment using the artificial cornea as the implant showed that the animals' corneas could keep smooth surface and clear stroma postoperatively, and that the repopulation of the host's keratocytes, the degradation of the implant and new corneal tissue generation were completed at 5-6 months after surgery. Such an artificial cornea has several advantages over other corneal equivalents constructed in the typical way of tissue engineering: in having similar mechanical and optical properties to those of the human cornea and with no exogenetic cells, it can be used universally in different implantation surgeries without immunoreaction; it is easy to prepare and process into different shapes and sizes on a large scale, and suitable for long-distance transportation and long-term storage. All these characteristics make it a new approach to cornea tissue engineering having potential in many clinical applications.

[The methods of shorting of proximal femoral and total hip arthroplasty for old femoral neck fracture with severe hip joint dislocation].

OBJECTIVE: To discuss the effective method and outcome to old femoral neck fracture with severe hip joint dislocation. METHODS: From April 1996, 7 cases of old femoral neck fracture with severe hip joint dislocation were treated by the shorting of posterior femoral and total hip arthroplasty. RESULTS: The average age of 7 patients was 51 years old and the mean follow-up was 27.3 months. The mean Harris score was improved to 84.3 from 36.7 before operation in the latest follow-up. The prosthesis position of acetabular and femoral was excellent, there was no sinking or softening of artificial joint, nonunion of femoral osteotomy. CONCLUSION: The preliminary clinical results are quite satisfactory of the shorting of posterior femoral and total hip arthroplasty to old femoral neck fracture with severe hip joint dislocation, the follow-up is necessary for further long-term outcome.