Nancy Lan Guo

Perfluorooctane sulfonate (PFOS) induces reactive oxygen species (ROS) production in human microvascular endothelial cells: role in endothelial permeability

Perfluorooctane sulfonate (PFOS) is a member of the perfluoroalkyl acids (PFAA) containing an eight-carbon backbone. PFOS is a man-made chemical with carbon–fluorine bonds that are among the strongest in organic chemistry, and PFOS is widely used in industry. Human occupational and environmental exposure to PFOS occurs globally. PFOS is non-biodegradable and is persistent in the human body and environment. In this study, data demonstrated that exposure of human microvascular endothelial cells (HMVEC) to PFOS induced the production of reactive oxygen species (ROS) at both high and low concentrations. Morphologically, it was found that exposure to PFOS induced actin filament remodeling and endothelial permeability changes in HMVEC. Furthermore, data demonstrated that the production of ROS plays a regulatory role in PFOS-induced actin filament remodeling and the increase in endothelial permeability. Our results indicate that the generation of ROS may play a role in PFOS-induced aberrations of the endothelial permeability barrier. The results generated from this study may provide a new insight into the potential adverse effects of PFOS exposure on humans at the cellular level.

Iron oxide nanoparticles induce human microvascular endothelial cell permeability through reactive oxygen species production and microtubule remodeling

BackgroundEngineered iron nanoparticles are being explored for the development of biomedical applications and many other industry purposes. However, to date little is known concerning the precise mechanisms of translocation of iron nanoparticles into targeted tissues and organs from blood circulation, as well as the underlying implications of potential harmful health effects in human.ResultsThe confocal microscopy imaging analysis demonstrates that exposure to engineered iron nanoparticles induces an increase in cell permeability in human microvascular endothelial cells. Our studies further reveal iron nanoparticles enhance the permeability through the production of reactive oxygen species (ROS) and the stabilization of microtubules. We also showed Akt/GSK-3β signaling pathways are involved in iron nanoparticle-induced cell permeability. The inhibition of ROS demonstrate ROS play a major role in regulating Akt/GSK-3β – mediated cell permeability upon iron nanoparticle exposure. These results provide new insights into the bioreactivity of engineered iron nanoparticles which can inform potential applications in medical imaging or drug delivery.ConclusionOur results indicate that exposure to iron nanoparticles induces an increase in endothelial cell permeability through ROS oxidative stress-modulated microtubule remodeling. The findings from this study provide new understandings on the effects of nanoparticles on vascular transport of macromolecules and drugs.

Cadmium Increases HIF-1 and VEGF Expression through ROS, ERK, and AKT Signaling Pathways and Induces Malignant Transformation of Human Bronchial Epithelial Cells

Cadmium is categorized as a human carcinogen especially involved in lung cancers. Angiogenesis is considered a fundamental requirement for tumorigenesis, but the mechanisms underlying the tumor angiogenesis induced by cadmium are poorly understood. Using in vitro and in vivo models, we investigated the angiogenic mechanisms of cadmium in human bronchial epithelial cells and tumor formation. Our results demonstrated that cadmium (CdCl(2)) activated extracellular signal-regulated kinases (ERK) and AKT signaling and elevated the expression of a key downstream proangiogenic molecule hypoxia-inducible factor-1 (HIF-1) in immortalized human lung epithelial BEAS-2B cells. Cadmium also induced reactive oxygen species (ROS) production, which could be inhibited by ROS scavengers, catalase and diphenyleneiodonium chloride. Inhibition of ROS generation also attenuated ERK, AKT, p70S6K1 activation, and HIF-1α expression. Similar results were obtained in normal human bronchial epithelial (NHBE) cells, showing that cadmium induced HIF-1 expression via ROS/ERK/AKT signaling pathway. Furthermore, cadmium induced vascular endothelial growth factor expression and transcriptional activation through ROS, ERK, and AKT pathways. Finally, cadmium transformed human bronchial epithelial cells in culture; the transformed cells induced tube formation in vitro, angiogenesis on chicken chorioallantoic membrane, and formed tumors in nude mice. Taken together, the results of this study provide explanation for the role and molecular mechanisms of cadmium in promoting angiogenesis in lung epithelial cells and malignant transformation and will be helpful for improved occupational protection, prevention, as well as chemotherapy of human lung cancers caused by heavy metal cadmium.

Confirmation of Gene Expression–Based Prediction of Survival in Non–Small Cell Lung Cancer

Purpose: It is a critical challenge to determine the risk of recurrence in early stage non–small cell lung cancer (NSCLC) patients. Accurate gene expression signatures are needed to classify patients into high- and low-risk groups to improve the selection of patients for adjuvant therapy. Experimental Design: Multiple published microarray data sets were used to evaluate our previously identified lung cancer prognostic gene signature. Expression of the signature genes was further validated with real-time reverse transcription-PCR and Western blot assays of snap-frozen lung cancer tumor tissues. Results: Our previously identified 35-gene signature stratified 264 patients with NSCLC into high- and low-risk groups with distinct overall survival rates (P < 0.05, Kaplan-Meier analysis, log-rank tests). The 35-gene signature further stratified patients with clinical stage 1A diseases into poor prognostic and good prognostic subgroups (P = 0.0007, Kaplan-Meier analysis, log-rank tests). This signature is independent of other prognostic factors for NSCLC, including age, sex, tumor differentiation, tumor grade, and tumor stage. The expression of the signature genes was validated with real-time reverse transcription-PCR analysis of lung cancer tumor specimens. Protein expression of two signature genes, TAL2 and ILF3, was confirmed in lung adenocarcinoma tumors by using Western blot analysis. These two biomarkers showed correlated mRNA and protein overexpression in lung cancer development and progression. Conclusions: The results indicate that the identified 35-gene signature is an accurate predictor of survival in NSCLC. It provides independent prognostic information in addition to traditional clinicopathologic criteria.

Cell Permeability, Migration, and Reactive Oxygen Species Induced by Multiwalled Carbon Nanotubes in Human Microvascular Endothelial Cells

Multiwalled carbon nanotubes (MWCNT) have elicited great interest in biomedical applications due to their extraordinary physical, chemical, and optical properties. Intravenous administration of MWCNT-based medical imaging agents and drugs in animal models was utilized. However, the potential harmful health effects of MWCNT administration in humans have not yet been elucidated. Furthermore, to date, there are no apparent reports regarding the precise mechanisms of translocation of MWCNT into target tissues and organs from blood circulation. This study demonstrates that exposure to MWCNT leads to an increase in cell permeability in human microvascular endothelial cells (HMVEC). The results obtained from this study also showed that the MWCNT-induced rise in endothelial permeability is mediated by reactive oxygen species (ROS) production and actin filament remodeling. In addition, it was found that MWCNT promoted cell migration in HMVEC. Mechanistically, MWCNT exposure elevated the levels of monocyte chemoattractant protein-1 (MCP-1) and intercellular adhesion molecule 1 (ICAM-1) in HMVEC. Taken together, these results provide new insights into the bioreactivity of MWCNT, which may have implications in the biomedical application of MWCNT in vascular targeting, imaging, and drug delivery. The results generated from this study also elucidate the potential adverse effects of MWCNT exposure on humans at the cellular level.

The gene expression signature of genomic instability in breast cancer is an independent predictor of clinical outcome.

Recently, expression profiling of breast carcinomas has revealed gene signatures that predict clinical outcome, and discerned prognostically relevant breast cancer subtypes. Measurement of the degree of genomic instability provides a very similar stratification of prognostic groups. We therefore hypothesized that these features are linked. We used gene expression profiling of 48 breast cancer specimens that profoundly differed in their degree of genomic instability and identified a set of 12 genes that defines the 2 groups. The biological and prognostic significance of this gene set was established through survival prediction in published datasets from patients with breast cancer. Of note, the gene expression signatures that define specific prognostic subtypes in other breast cancer datasets, such as luminal A and B, basal, normal‐like, and ERBB2+, and prognostic signatures including MammaPrint® and Oncotype DX, predicted genomic instability in our samples. This remarkable congruence suggests a biological interdependence of poor‐prognosis gene signatures, breast cancer subtypes, genomic instability, and clinical outcome.

The microRNA-200 family targets multiple non-small cell lung cancer prognostic markers in H1299 cells and BEAS-2B cells

Lung cancer remains the leading cause of cancer-related mortality for both men and women. Tumor recurrence and metastasis is the major cause of lung cancer treatment failure and death. The microRNA-200 (miR-200) family is a powerful regulator of the epithelial-mesenchymal transition (EMT) process, which is essential in tumor metastasis. Nevertheless, miR-200 family target genes that promote metastasis in non-small cell lung cancer (NSCLC) remain largely unknown. Here, we sought to investigate whether the microRNA-200 family regulates our previously identified NSCLC prognostic marker genes associated with metastasis, as potential molecular targets. Novel miRNA targets were predicted using bioinformatics tools based on correlation analyses of miRNA and mRNA expression in 57 squamous cell lung cancer tumor samples. The predicted target genes were validated with quantitative RT-PCR assays and western blot analysis following re-expression of miR-200a, -200b and -200c in the metastatic NSCLC H1299 cell line. The results show that restoring miR-200a or miR-200c in H1299 cells induces downregulation of DLC1, ATRX and HFE. Reinforced miR-200b expression results in downregulation of DLC1, HNRNPA3 and HFE. Additionally, miR-200 family downregulates HNRNPR3, HFE and ATRX in BEAS-2B immortalized lung epithelial cells in quantitative RT-PCR and western blot assays. The miR-200 family and these potential targets are functionally involved in canonical pathways of immune response, molecular mechanisms of cancer, metastasis signaling, cell-cell communication, proliferation and DNA repair in Ingenuity pathway analysis (IPA). These results indicate that re-expression of miR-200 downregulates our previously identified NSCLC prognostic biomarkers in metastatic NSCLC cells. These results provide new insights into miR-200 regulation in lung cancer metastasis and consequent clinical outcome, and may provide a potential basis for innovative therapeutic approaches for the treatment of this deadly disease.

Association of arsenic exposure with lung cancer incidence rates in the United States.

Background Although strong exposure to arsenic has been shown to be carcinogenic, its contribution to lung cancer incidence in the United States is not well characterized. We sought to determine if the low-level exposures to arsenic seen in the U.S. are associated with lung cancer incidence after controlling for possible confounders, and to assess the interaction with smoking behavior. Methodology Measurements of arsenic stream sediment and soil concentration obtained from the USGS National Geochemical Survey were combined, respectively, with 2008 BRFSS estimates on smoking prevalence and 2000 U.S. Census county level income to determine the effects of these factors on lung cancer incidence, as estimated from respective state-wide cancer registries and the SEER database. Poisson regression was used to determine the association between each variable and age-adjusted county-level lung cancer incidence. ANOVA was used to assess interaction effects between covariates. Principal Findings Sediment levels of arsenic were significantly associated with an increase in incident cases of lung cancer (P<0.0001). These effects persisted after controlling for smoking and income (P<0.0001). Across the U.S., exposure to arsenic may contribute to up to 5,297 lung cancer cases per year. There was also a significant interaction between arsenic exposure levels and smoking prevalence (P<0.05). Conclusions/Significance Arsenic was significantly associated with lung cancer incidence rates in the U.S. after controlling for smoking and income, indicating that low-level exposure to arsenic is responsible for excess cancer cases in many parts of the U.S. Elevated county smoking prevalence strengthened the association between arsenic exposure and lung cancer incidence rate, an effect previously unseen on a population level.

Multi-walled carbon nanotubes induce human microvascular endothelial cellular effects in an alveolar-capillary co-culture with small airway epithelial cells

BackgroundNanotechnology, particularly the use of multi-walled carbon nanotubes (MWCNT), is a rapidly growing discipline with implications for advancement in a variety of fields. A major route of exposure to MWCNT during both occupational and environmental contact is inhalation. While many studies showed adverse effects to the vascular endothelium upon MWCNT exposure, in vitro results often do not correlate with in vivo effects. This study aimed to determine if an alveolar-capillary co-culture model could determine changes in the vascular endothelium after epithelial exposure to MWCNT.MethodsA co-culture system in which both human small airway epithelial cells and human microvascular endothelial cells were separated by a Transwell membrane so as to resemble an alveolar-capillary interaction was used. Following exposure of the epithelial layer to MWCNT, the effects to the endothelial barrier were determined.ResultsExposure of the epithelial layer to MWCNT induced multiple changes in the endothelial cell barrier, including an increase in reactive oxygen species, actin rearrangement, loss of VE-cadherin at the cell surface, and an increase in endothelial angiogenic ability. Overall increases in secreted VEGFA, sICAM-1, and sVCAM-1 protein levels, as well as increases in intracellular phospho-NF-κB, phospho-Stat3, and phospho-p38 MAPK, were also noted in HMVEC after epithelial exposure.ConclusionThe co-culture system identified that alveolar-capillary exposure to MWCNT induced multiple changes to the underlying endothelium, potentially through cell signaling mediators derived from MWCNT-exposed epithelial cells. Therefore, the co-culture system appears to be a relevant in vitro method to study the pulmonary toxicity of MWCNT.

Multiwalled Carbon Nanotube-Induced Gene Signatures in the Mouse Lung: Potential Predictive Value for Human Lung Cancer Risk and Prognosis

Concerns over the potential for multiwalled carbon nanotubes (MWCNT) to induce lung carcinogenesis have emerged. This study sought to (1) identify gene expression signatures in the mouse lungs following pharyngeal aspiration of well-dispersed MWCNT and (2) determine if these genes were associated with human lung cancer risk and progression. Genome-wide mRNA expression profiles were analyzed in mouse lungs (n = 160) exposed to 0, 10, 20, 40, or 80 μg of MWCNT by pharyngeal aspiration at 1, 7, 28, and 56 d postexposure. By using pairwise statistical analysis of microarray (SAM) and linear modeling, 24 genes were selected, which have significant changes in at least two time points, have a more than 1.5-fold change at all doses, and are significant in the linear model for the dose or the interaction of time and dose. Additionally, a 38-gene set was identified as related to cancer from 330 genes differentially expressed at d 56 postexposure in functional pathway analysis. Using the expression profiles of the cancer-related gene set in 8 mice at d 56 postexposure to 10 μg of MWCNT, a nearest centroid classification accurately predicts human lung cancer survival with a significant hazard ratio in training set (n = 256) and test set (n = 186). Furthermore, both gene signatures were associated with human lung cancer risk (n = 164) with significant odds ratios. These results may lead to development of a surveillance approach for early detection of lung cancer and prognosis associated with MWCNT in the workplace.