Loss Of Cell Function Research Articles

Inherited Retinal Disease Therapies Targeting Precursor Messenger Ribonucleic Acid.

Inherited retinal diseases are an extremely diverse group of genetically and phenotypically heterogeneous conditions characterized by variable maturation of retinal development, impairment of photoreceptor cell function and gradual loss of photoreceptor cells and vision. Significant progress has been made over the last two decades in identifying the many genes implicated in inherited retinal diseases and developing novel therapies to address the underlying genetic defects. Approximately one-quarter of exonic mutations related to human inherited diseases are likely to induce aberrant splicing products, providing opportunities for the development of novel therapeutics that target splicing processes. The feasibility of antisense oligomer mediated splice intervention to treat inherited diseases has been demonstrated in vitro, in vivo and in clinical trials. In this review, we will discuss therapeutic approaches to treat inherited retinal disease, including strategies to correct splicing and modify exon selection at the level of pre-mRNA. The challenges of clinical translation of this class of emerging therapeutics will also be discussed.

Chronic viral infections induce major disruption of bone marrow stromal cell networks and persistent loss of hematopoietic stem cell function

Hematopoiesis is a highly demand-adapted and tightly regulated process that is sustained by a rare population of self-renewing, multipotent hematopoietic stem and progenitor cells (HSPCs) residing in specialized microenvironments within bone marrow (BM) cavities. The basic tissue infrastructure of the BM is provided by stromal cellular networks of mesenchymal, neural and vascular origin, which are critically involved in the fine regulation of different stages of hematopoiesis. Viral infections act as major stressors to the hematopoietic system, inducing massive and adaptive responses in cellular output. Albeit the effects of viral challenge and ensuing inflammatory responses on hematopoietic cells have been studied, whether viral infections alter BM stromal scaffolds and thus shape hematopoietic responses remains poorly defined. By combining conventional in vitro and in vivo assays with 3D confocal imaging, we herein investigated the structural and functional alterations imposed on the BM after a chronic infection with Lymphocytic Choriomeningitis Virus (LCMV). Our data shows that chronic LCMV infections trigger a substantial loss of BM endothelial and mesenchymal CXCL12-abundant reticular cells. Upon viral challenge, conspicuous vasodilation of BM sinusoidal vessels is induced which was followed by intense vascular remodeling and substantial disruption of extracellular matrix networks throughout the BM cavity. Major damage to BM stromal integrity is accompanied by a profound and sustained reduction in the number of both HSPCs as well as hematopoietic stem cells by phenotype. Competitive repopulation assays revealed that the remaining HSCs are additionally impaired in their repopulation capacity for prolonged times after LCMV infection. Finally, our results indicate that the observed alterations in the composition and functionality of cells in the BM are mediated by virus-specific CD8 T cells and partly dependent on the production of specific cytokines. Our observations thus indicate that chronic infections result in persistent damage to HSPC function, which might be partially explained by an impaired regulatory function of the stromal compartment of the BM.

Design and Acquisition of EOG Based Interactive Communications for ALS Patients

Amyotrophic lateral sclerosis (ALS) is a motor neuron disease caused by loss of function of spinal cord and brain stem nerve cells. Loss of function in nerve cells leads to weakness and explosion (atrophy) in the muscles. The outcome of the weakness in the muscles needs the help of someone. Despite the limitations of the movement, studies on methods of increasing the daily quality of life ALS patients are continuing. Electrooculogram (EOG) signals were taken from an instrumentation amplifier with 48dB gain and 107dB CMMR ratio. A 16Hz Low Pass Filter and a 50Hz Notch Filter were used to increase the signal to noise ratio. EOG signals are digitized with 10bit resolution ADC and applied to ATmega328 microcontroller. So, the software of microcontroller determines the horizontal and vertical movements of the eye. In this way, the interactive PC software was controlled by the EOG signals. EOG-based interactive software which was developed using the C # programming language has provided to patients' daily requirements, social media accounts etc. The system has been tested on healthy subjects and it has been seen that people can control the software by eye movements. As a result, system will be useful not only for ALS patients, but also for permanent or partially bedridden patient (MS, Hemiplegia, etc.) groups. And also, while the quality of life in the patients' own habitats is being raised, they will also be able to benefit from health services within the scope of home health services.

Up-regulation of NADPH oxidase-mediated redox signaling contributes to the loss of barrier function in KRIT1 deficient endothelium

The intracellular scaffold KRIT1/CCM1 is an established regulator of vascular barrier function. Loss of KRIT1 leads to decreased microvessel barrier function and to the development of the vascular disorder Cerebral Cavernous Malformation (CCM). However, how loss of KRIT1 causes the subsequent deficit in barrier function remains undefined. Previous studies have shown that loss of KRIT1 increases the production of reactive oxygen species (ROS) and exacerbates vascular permeability triggered by several inflammatory stimuli, but not TNF−α. We now show that endothelial ROS production directly contributes to the loss of barrier function in KRIT1 deficient animals and cells, as targeted antioxidant enzymes reversed the increase in permeability in KRIT1 heterozygous mice as shown by intravital microscopy. Rescue of the redox state restored responsiveness to TNF-α in KRIT1 deficient arterioles, but not venules. In vitro, KRIT1 depletion increased endothelial ROS production via NADPH oxidase signaling, up-regulated Nox4 expression, and promoted NF-κB dependent promoter activity. Recombinant yeast avenanthramide I, an antioxidant and inhibitor of NF-κB signaling, rescued barrier function in KRIT1 deficient cells. However, KRIT1 depletion blunted ROS production in response to TNF-α. Together, our data indicate that ROS signaling is critical for the loss of barrier function following genetic deletion of KRIT1.

Molecular regulative mechanisms of aging and interventional effects of Chinese herbal medicine

Aging is a gradual process during the loss of functions in cells,organs and tissues by time. The molecular mechanisms of aging-related theories include the classical ones such as telomere,oxygen radical and nonenzymatic glycosylation,as well as the newly proposed ones such as DNA methylation,mitochondrial DNA （mtDNA）and autophagy. The latest study showed the anti-aging effect of autophagy in hematopoietic stem cells. In recent years,based on the molecular regulative mechanisms of aging,a number of the promising anti-aging drugs have been found,including nicotinamide mononucleotide（NMN）and FOXO4-DRI,a peptide of anti-aging. In addition,there are many new discoveries in the field of plant extracts,in which,the extracts from Chinese herbal medicine（CHM）,some single CHMs and the classical prescriptions of CHM,represented by curcumin and resveratrol,have the partial anti-aging effects by regulating the molecular mechanisms of aging both in vivo and in vitro. In brief,developing or exploring anti-aging drugs,especially the natural drugs,is one of the main development directions in the field of anti-aging research in the basis of the molecular regulative mechanisms of aging.

Normal tissue tolerance

Effects of radiation exposure are observed in virtually all normal tissues. Early reactions occur primarily in turnover tissues (e.g., bone marrow, epidermis, mucosae of the gastrointestinal tract), where proliferative impairment results in progressive hypoplasia and eventually complete loss of functional cells, after a tissue dependent but dose independent latent time. These early radiation reponses are regularly preceded and accompanied by vascular and inflammatory reactions. In contrast, late reactions are based on combined parenchymal, vascular, and connective tissue changes; very late effects are dominated by vascular sequelae. In most instances, a significant involvement of the immune system can also be demonstrated for chronic radiation sequelae, and a contribution of neural changes is discussed. The orchestrated response of all tissue components results in loss of function within the exposed volume. Importantly, latent times of late effects are inversely dependent on dose. Hence modern, highly conformal treatment techniques with relatively low and inhomogeneous doses in the organs at risk (OAR) require very long follow-up intervals with a precise assessment and documentation of the complication endpoints for characterisation of the treatment-induced morbidity profile. Consequential late effects (CLEs) develop through interactions between early and late effects in the same organ; they follow the radiobiological principles of the early reactions. The clinical manifestation of radiation responses is defined by several parameters, summarized as the “R’s of radiobiology”. First, each individual symptom or endpoint of radiation-induced morbidity follows an individual dose-effect relationship (intrinsic ‘R’adiosensitivity), in many instances related to the dose within specific subvolumes of the individual OAR, rather than e.g., the mean organ dose. The biological effectiveness of a certain (total) dose is modulated by exposure conditions: Changes in dose fractionation protocols (‘R’recovery) predominantly impact on late responding tissues, while overall treatment time (‘R’epopulation) predominantly affects early (and consequential late) reactions. Consequences of partial organ exposure (i’R’adiated volume”) are related to tissue architecture. In mainly ‘tubular’ or ‘serial’ organs (e.g., gastrointestinal tract, but also vasculature), local exposure affects function in downstream compartments. In contrast, in predominantly ‘parallel’ organs, such as liver or lungs, only exposure of a significant (organ-dependent) fraction of the total volume results in clinical consequences. However, all organs in fact are composed of tubular and serial components. Translational studies into damage processing (molecular ‘R’adiopathology), starting immediately after the onset of radiation exposure, but proceeding for long and very long time intervals even at subclinical levels, intra- and intercellular signals and signalling pathways may be identified that are relevant or even specific for the clinical manifestation of morbidity endpoints. These can serve as a basis to identify (early) biomarkers of the individual risk for specific tissue reactions and endpoints, and also for establishment of strategies to prevent/mitigate tissue effects after exposure.

Modeling the effect of blunt impact on mitochondrial function in cartilage: implications for development of osteoarthritis.

ObjectiveOsteoarthritis (OA) is a disease characterized by degeneration of joint cartilage. It is associated with pain and disability and is the result of either age and activity related joint wear or an injury. Non-invasive treatment options are scarce and prevention and early intervention methods are practically non-existent. The modeling effort presented in this article is constructed based on an emerging biological hypothesis—post-impact oxidative stress leads to cartilage cell apoptosis and hence the degeneration observed with the disease. The objective is to quantitatively describe the loss of cell viability and function in cartilage after an injurious impact and identify the key parameters and variables that contribute to this phenomenon.MethodsWe constructed a system of differential equations that tracks cell viability, mitochondrial function, and concentrations of reactive oxygen species (ROS), adenosine triphosphate (ATP), and glycosaminoglycans (GAG). The system was solved using MATLAB and the equations’ parameters were fit to existing data using a particle swarm algorithm.ResultsThe model fits well the available data for cell viability, ATP production, and GAG content. Local sensitivity analysis shows that the initial amount of ROS is the most important parameter.DiscussionThe model we constructed is a viable method for producing in silico studies and with a few modifications, and data calibration and validation, may be a powerful predictive tool in the search for a non-invasive treatment for post-traumatic osteoarthritis.

A review of the toxicity of nanoparticles to <italic>Daphnia magna</italic>

With the wide production and usage of nanoparticles, nanotoxicity has received worldwide researches. As a model organism, Daphnia magna has been extensively used in the nanotoxicity studies. In this review, we overviewed research advance in the toxicity of nanoparticles to Daphnia magna , including acute toxicity, chronic toxicity, bioaccumulation, and the underlying mechanisms. Nanoparticles have dose-responsive acute toxicity to Daphnia manga and their toxicity is related to types of nanoparticles and culture media. Smaller nanoparticles generally have higher toxicity to Daphnia manga , presumably due to their easier transport through cell membranes. Due to polar functional groups and organic macromolecules are able to change surface properties of nanoparticles, most studies suggest that the acute toxicity of nanoparticles might be weakened by surface modification. High concentrations of nanoparticles can suppress the growth and reproduction of Daphnia manga . Bioaccumulation studies showed that nanoparticles could accumulate in Daphnia magna via water, feeding food, and absorbing through the skin. Since the concentration of nanoparticles, exposure time and cultivation media adopted in the studies varied greatly, the body burden and bioconcentration factor of nanoparticles were largely different, which makes it hard to compare accumulation capacity of Daphnia manga to different nanoparticles. At high exposure concentrations, accumulation amount of nanoparticles by Daphnia manga could reach µg/mg level. Although the toxicity mechanisms of nanoparticles to Daphnia manga have not been fully understood, below points are generally used to explain the toxicity: nanoparticle specific toxic, the release of toxic metal ions, and forming toxic complexes with other compounds. Nanoparticles and the release of metal ions can adhere and aggregate onto surfaces of Daphnia manga , causing the physical damage, membrane permeability changes, and cell function loss. They can also promote the production of the reactive oxygen species (ROS) and free radical, leading to lipid peroxidation, protein carbonylation and DNA damage. In addition, some nanoparticles themselves are not toxic, but can form complexes with toxic substance. At the end of text, we proposed several future research needs. First, compared with the experimental cultivation media, the physical and chemical properties of natural waters are obviously different, likely to cause big difference in the toxicity results. Therefore, realistic toxicity in natural waters needs to be studied. Second, the exposure concentrations of nanoparticles in chronic experiments are much higher than that in the real water environment, and the exposure time is also not long enough compared with practical condition. Lower concentrations and longer exposure time of nanoparticles are of great significance. Third, there is no unified approach in bioaccumulation experiments, making it difficult to compare and analyze the results from different labs. A standard test method established for the bioaccumulation test is thus required. Moreover, nanoparticles can remain in Daphnia magna and transfer to their offspring and other organisms though food chains. Hence, it is necessary to study the transformation, chronic toxicity, intergenerational transfer, and migration though food chains of nanoparticles. Lastly, up to date, toxicity mechanisms of nanoparticles to Daphnia manga have yet to reach a consensus. Nanoparticle characteristics, the growth status of Daphnia magna , culture medium compositions, toxicity experimental method, and many other factors might influence the judgment of the toxicity mechanisms. In order to better understand the toxic effects caused by nanoparticles, more researches are supposed to be done to find the impacts of these factors on the toxicity mechanisms.

Suppression of HSP27 Restores Retinal Function and Protects Photoreceptors From Apoptosis in a Light-Induced Retinal Degeneration Animal Model.

We used a light-induced retinal degeneration animal model to investigate possible roles of heat shock protein 27 (HSP27) in retinal/photoreceptor protection. Sprague-Dawley rats were used for the light-induced retinal degeneration animal model. The histology of eye sections was observed for morphologic changes in the retina. Cell apoptosis was examined in each group using the terminal deoxynucleotidyl transferase dUTP nick-end labeling assay, and electroretinography was used to evaluate retinal function. Protein and mRNA expression levels of different retinal cell markers were also detected through immunofluorescence staining, Western blotting, and real-time PCR. The thickness of the outer nuclear layer significantly decreased after 7-day light exposure. Moreover, we injected a viral vector for silencing HSP27 expression into the eyes and observed that photoreceptors were better preserved in the HSP27-suppressed (sHSP27) retina 2 weeks after injection. HSP27 suppression also reduced retinal cell apoptosis caused by light exposure. In addition, the loss of retinal function caused by light exposure was reversed on suppressing HSP27 expression. We subsequently found that the expression of the Rho gene and immunofluorescence staining of rhodopsin and arrestin (cell markers for photoreceptors) increased in sHSP27-treated retinas. HSP27 suppression did not affect the survival of ganglion and amacrine cells. Retinal cell apoptosis and functional loss were observed after 7-day light exposure. However, in the following 2 weeks after light exposure, HSP27 suppression may initiate a protective effect for retinal cells, particularly photoreceptors, from light-induced retinal degeneration.

Proline-Rich Homeodomain protein (PRH/HHEX) is a suppressor of breast tumour growth

Breast tumours progress from hyperplasia to ductal carcinoma in situ (DCIS) and invasive breast carcinoma (IBC). PRH/HHEX (proline-rich homeodomain/haematopoietically expressed homeobox) is a transcription factor that displays both tumour suppressor and oncogenic activity in different disease contexts; however, the role of PRH in breast cancer is poorly understood. Here we show that nuclear localization of the PRH protein is decreased in DCIS and IBC compared with normal breast. Our previous work has shown that PRH phosphorylation by protein kinase CK2 prevents PRH from binding to DNA and regulating the transcription of multiple genes encoding growth factors and growth factor receptors. Here we show that transcriptionally inactive phosphorylated PRH is elevated in DCIS and IBC compared with normal breast. To determine the consequences of PRH loss of function in breast cancer cells, we generated inducible PRH depletion in MCF-7 cells. We show that PRH depletion results in increased MCF-7 cell proliferation in part at least due to increased vascular endothelial growth factor signalling. Moreover, we demonstrate that PRH depletion increases the formation of breast cancer cells with cancer stem cell-like properties. Finally, and in keeping with these findings, we show that PRH overexpression inhibits the growth of mammary tumours in mice. Collectively, these data indicate that PRH plays a tumour suppressive role in the breast and they provide an explanation for the finding that low PRH mRNA levels are associated with a poor prognosis in breast cancer.

DYSGLYCEMIA TO TYPE 1 DIABETES: EVALUATING THE ASSOCIATION OF RISK FACTORS WITH DISEASE PROGRESSION

BACKGROUND: Type 1 diabetes (T1D) has long been recognized as an autoimmune disorder, likely stimulated by environmental factors and immune dysregulation in individuals with an increased genetic susceptibility. Accelerated beta cell function loss precedes disease detection, indicating a need to identify high-risk individuals prior to diagnosis. While parameters such as number of autoantibodies against beta cells, age, body-mass index, and measures of insulin and glucose response have been shown to be predictive of T1D, prediction of the timing of diagnosis remains challenging. OBJECTIVES: To evaluate the association between risk factors for T1D and the rate of progression from an abnormal oral glucose tolerance test (OGTT) to the development of T1D. DESIGN/METHODS: Data from a prospective cohort study (n = 1127), Type 1 Diabetes TrialNet, was employed to perform a multivariate Cox regression using a forward stepwise approach to identify variables that increased risk of T1D. Cut points that defined high and low risk sub-populations based on identified variables of interest were selected through recursive partitioning analysis. C-statistics and life tables were then used to explore model discrimination and time from first abnormal OGTT to T1D. RESULTS: While there was some variability between study sub-groups (all ages, < 18 years, and > 18 years), ICA or ICA 512 positivity, DPTRS (Diabetes Prevention Trial-Type 1 Risk Score), number of antibodies, Index60 (metabolic index), and HbA1c were identified as being significant T1D risk factors. In the all ages group, DPTRS < 7 and <= 2 antibodies identified a lower risk population with slower progression to T1D compared to those with DPTRS > 7 and >= 3 antibodies (model C-statistic = 0.63). In the < 18 years group, DPTRS > 8 and HbA1c > 5.4 defined a faster progressing population (model C-statistic = 0.69). In the > 18 years group, ICA positivity and Index60 >= 1 identified a population at higher risk of progression to T1D (model C-statistic = 0.66). CONCLUSION: Several T1D risk factors predict the rate of progression from dysglycemia to T1D onset. These may be used to identify individuals at a higher risk for disease development and may help inform efforts aimed at T1D prevention.

The peroxisomal AAA ATPase complex prevents pexophagy and development of peroxisome biogenesis disorders

ABSTRACTPeroxisome biogenesis disorders (PBDs) are metabolic disorders caused by the loss of peroxisomes. The majority of PBDs result from mutation in one of 3 genes that encode for the peroxisomal AAA ATPase complex (AAA-complex) required for cycling PEX5 for peroxisomal matrix protein import. Mutations in these genes are thought to result in a defect in peroxisome assembly by preventing the import of matrix proteins. However, we show here that loss of the AAA-complex does not prevent matrix protein import, but instead causes an upregulation of peroxisome degradation by macroautophagy, or pexophagy. The loss of AAA-complex function in cells results in the accumulation of ubiquitinated PEX5 on the peroxisomal membrane that signals pexophagy. Inhibiting autophagy by genetic or pharmacological approaches rescues peroxisome number, protein import and function. Our findings suggest that the peroxisomal AAA-complex is required for peroxisome quality control, whereas its absence results in the selective degradation of the peroxisome. Thus the loss of peroxisomes in PBD patients with mutations in their peroxisomal AAA-complex is a result of increased pexophagy. Our study also provides a framework for the development of novel therapeutic treatments for PBDs.

Modelling of the cancer cell cycle as a tool for rational drug development: A systems pharmacology approach to cyclotherapy.

The dynamic of cancer is intimately linked to a dysregulation of the cell cycle and signalling pathways. It has been argued that selectivity of treatments could exploit loss of checkpoint function in cancer cells, a concept termed “cyclotherapy”. Quantitative approaches that describe these dysregulations can provide guidance in the design of novel or existing cancer therapies. We describe and illustrate this strategy via a mathematical model of the cell cycle that includes descriptions of the G1-S checkpoint and the spindle assembly checkpoint (SAC), the EGF signalling pathway and apoptosis. We incorporated sites of action of four drugs (palbociclib, gemcitabine, paclitaxel and actinomycin D) to illustrate potential applications of this approach. We show how drug effects on multiple cell populations can be simulated, facilitating simultaneous prediction of effects on normal and transformed cells. The consequences of aberrant signalling pathways or of altered expression of pro- or anti-apoptotic proteins can thus be compared. We suggest that this approach, particularly if used in conjunction with pharmacokinetic modelling, could be used to predict effects of specific oncogene expression patterns on drug response. The strategy could be used to search for synthetic lethality and optimise combination protocol designs.

Abstract 3: Cellular Contributions of the Circadian Clock in Atherogenesis

Atherosclerosis is a leading cause of death despite the improvements in lipid and blood pressure control. The circadian clock, a molecular network of genes and proteins that controls 24-hour timing, has emerged to have a surprising role in the control of metabolic and vascular function. Herein we examined the impact of circadian rhythm dysfunction in atherogenesis by implementation of vascular transplant and PCSK9 based approaches to induce accelerated lesion development in mice. We find that atherogenesis is exacerbated in Bmal1-KO aortic grafts immersed in the hypercholesterolemic milieu of ApoE -/- mice. To assess if atherosclerosis was ‘circadian rhythm dependent’ we subjected wild-type mice to a shortened light cycle (4L/4D) and induced atherosclerosis by intravenous injection of a human PCSK-9 adeno associated virus. Atherosclerosis in the jet-lagged PCSK-9 mice was robustly increased relative to the atherosclerosis observed in WT mice on a normal light cycle (12L/12D), providing further evidence that circadian rhythm and the circadian clock contribute to atherosclerosis. However, atherosclerosis is a complex disease that is the net result of interplay between intrinsic (vascular cells) and extrinsic mechanisms (metabolism, blood pressure, and hormones) and the importance of clock function in individual cell types is poorly understood. We found that deletion or silencing of key circadian transcription factors resulted in an enhanced inflammatory and pro-oxidant phenotype with diminished NO production and greater lipid uptake in both macrophages and endothelial cells. Loss of circadian function in smooth muscle cells similarly resulted in enhanced production of reactive oxygen species and greater cell proliferation. Surprising, the silencing of Bmal2 in endothelial cells resulted in greater lipid uptake in oxLDL treated HAEC as well as increased expression of markers of autophagy, suggesting that Bmal2 may orchestrate numerous output functions in different cell types. In conclusion, we find that the circadian clock and circadian rhythm have a profound impact on atherosclerosis, to influence vascular cell inflammatory and lipid uptake responses, and identify an unexpectedly prominent role for the side-partner of Bmal1, Bmal2.

HnRNP I regulates neonatal immune adaptation and prevents colitis and colorectal cancer.

The intestinal epithelium plays a critical role in host-microbe homeostasis by sensing gut microbes and subsequently initiating proper immune responses. During the neonatal stage, the intestinal epithelium is under immune repression, allowing the transition for newborns from a relatively sterile intra-uterine environment to one that is rich in foreign antigens. The mechanism underlying such immune repression remains largely unclear, but involves downregulation of IRAK1 (interleukin-1 receptor-associated kinase), an essential component of toll-like receptor-mediated NF-κB signaling. We report here that heterogeneous nuclear ribonucleoprotein I (hnRNPI), an RNA binding protein, is essential for regulating neonatal immune adaptation. We generated a mouse model in which hnRNPI is ablated specifically in the intestinal epithelial cells, and characterized intestinal defects in the knockout mice. We found that loss of hnRNPI function in mouse intestinal epithelial cells results in early onset of spontaneous colitis followed by development of invasive colorectal cancer. Strikingly, the epithelium-specific hnRNPI knockout neonates contain aberrantly high IRAK1 protein levels in the colons and fail to develop immune tolerance to environmental microbes. Our results demonstrate that hnRNPI plays a critical role in establishing neonatal immune adaptation and preventing colitis and colorectal cancer.

Inflammatory Response and Barrier Dysfunction by Different e-Cigarette Flavoring Chemicals Identified by Gas Chromatography-Mass Spectrometry in e-Liquids and e-Vapors on Human Lung Epithelial Cells and Fibroblasts.

Recent studies suggest that electronic cigarette (e-cig) flavors can be harmful to lung tissue by imposing oxidative stress and inflammatory responses. The potential inflammatory response by lung epithelial cells and fibroblasts exposed to e-cig flavoring chemicals in addition to other risk-anticipated flavor enhancers inhaled by e-cig users is not known. The goal of this study was to evaluate the release of the proinflammatory cytokine (interleukin-8 [IL-8]) and epithelial barrier function in response to different e-cig flavoring chemicals identified in various e-cig e-liquid flavorings and vapors by chemical characterization using gas chromatography–mass spectrometry analysis. Flavorings, such as acetoin (butter), diacetyl, pentanedione, maltol (malt), ortho-vanillin (vanilla), coumarin, and cinnamaldehyde in comparison with tumor necrosis factor alpha (TNFα), were used in this study. Human bronchial epithelial cells (Beas2B), human mucoepidermoid carcinoma epithelial cells (H292), and human lung fibroblasts (HFL-1) were treated with each flavoring chemical for 24 hours. The cells and conditioned media were then collected and analyzed for toxicity (viability %), lung epithelial barrier function, and proinflammatory cytokine IL-8 release. Cell viability was not significantly affected by any of the flavoring chemicals tested at a concentration of 10 μM to 1 mM. Acetoin and diacetyl treatment induced IL-8 release in Beas2B cells. Acetoin- and pentanedione-treated HFL-1 cells produced a differential, but significant response for IL-8 release compared to controls and TNFα. Flavorings, such as ortho-vanillin and maltol, induced IL-8 release in Beas2B cells, but not in H292 cells. Of all the flavoring chemicals tested, acetoin and maltol were more potent inducers of IL-8 release than TNFα in Beas2B and HFL-1 cells. Flavoring chemicals rapidly impaired epithelial barrier function in human bronchial epithelial cells (16-HBE) as measured by electric cell surface impedance sensing. Our findings suggest that some of the e-cig liquids/aerosols containing flavoring chemicals can cause significant loss of epithelial barrier function and proinflammatory response in lung cells.

Isolevuglandins as a gauge of lipid peroxidation in human tumors

The cellular production of free radicals or reactive oxygen species (ROS) can lead to protein, lipid or DNA modifications and tumor formation. The cellular lipids undergo structural changes through the actions of enzymes (e.g. cyclooxygenases) or free radicals to form a class of compounds called Isolevuglandins (IsoLGs). The recruitment and continued exposure of tissue to ROS and IsoLGs causes increased cell proliferation, mutagenesis, loss of normal cell function and angiogenesis. The elevated concentration of ROS in cancerous tissues suggests that these mediators play an important role in cancer development. We hypothesized that tumors with elevated ROS levels would similarly possess an increased concentration of IsoLGs when compared with normal tissue. Using D11, an ScFv recombinant antibody specific for IsoLGs, we utilized immunohistochemistry to visualize the presence of IsoLG in human tumors compared to normal adjacent tissue (NAT) to the same tumor. We found that IsoLG concentrations were elevated in human breast, colon, kidney, liver, lung, pancreatic and tongue tumor cells when compared to NAT and believe that IsoLGs can be used as a gauge indicative of lipid peroxidation in tumors.

The Role of Sigma1R in Mammalian Retina.

This review article focuses on studies of Sigma 1 Receptor (Sigma1R) and retina . It provides a brief overview of the earliest pharmacological studies performed in the late 1990s that provided evidence of the presence of Sigma1R in various ocular tissues. It then describes work from a number of labs concerning the location of Sigma1R in several retinal cell types including ganglion, Müller glia , and photoreceptors . The role of Sigma1R ligands in retinal neuroprotection is emphasized. Early studies performed in vitro clearly showed that targeting Sigma1R could attenuate stress-induced retinal cell loss. These studies were followed by in vivo experiments. Data about the usefulness of targeting Sigma1R to prevent ganglion cell loss associated with diabetic retinopathy are reviewed. Mechanisms of Sigma1R-mediated retinal neuroprotection involving Müller cells , especially in modulating oxidative stress are described along with information about the retinal phenotype of mice lacking Sigma1R (Sigma1R -/- mice). The retina develops normally in Sigma1R -/- mice, but after many months there is evidence of apoptosis in the optic nerve head, decreased ganglion cell function and eventual loss of these cells. Additional studies using the Sigma1R -/- mice provide strong evidence that in the retina, Sigma1R plays a key role in modulating cellular stress. Recent work has shown that targeting Sigma1R may extend beyond protection of ganglion cells to include photoreceptor cell degeneration as well.

Inhibiting the SUMO Pathway Represses the Cancer Stem Cell Population in Breast and Colorectal Carcinomas.

SummaryMany solid cancers have an expanded CD44+/hi/CD24−/low cancer stem cell (CSC) population, which are relatively chemoresistant and drive recurrence and metastasis. Achieving a more durable response requires the development of therapies that specifically target CSCs. Recent evidence indicated that inhibiting the SUMO pathway repressed tumor growth and invasiveness, although the mechanism has yet to be clarified. Here, we demonstrate that inhibition of the SUMO pathway repressed MMP14 and CD44 with a concomitant reduction in cell invasiveness and functional loss of CSCs in basal breast cancer. Similar effects were demonstrated with a panel of E1 and E3 SUMO inhibitors. Identical results were obtained in a colorectal cancer cell line and primary colon cancer cells. In both breast and colon cancer, SUMO-unconjugated TFAP2A mediated the effects of SUMO inhibition. These data support the development of SUMO inhibitors as an approach to specifically target the CSC population in breast and colorectal cancer.

Establishment of the Muscle-Tendon Junction During Thorax Morphogenesis in Drosophila Requires the Rho-Kinase.

The assembly of the musculoskeletal system in Drosophila relies on the integration of chemical and mechanical signaling between the developing muscles with ectodermal cells specialized as "tendon cells." Mechanical tension generated at the junction of flight muscles and tendon cells of the notum epithelium is required for muscle morphogenesis, and is balanced by the epithelium in order to not deform. We report that Drosophila Rho kinase (DRok) is necessary in tendon cells to assemble stable myotendinous junctions (MTJ), which are required for muscle morphogenesis and survival. In addition, DRok is required in tendon cells to maintain epithelial shape and cell orientation in the notum, independently of chascon (chas). Loss of DRok function in tendon cells results in mis-orientation of tendon cell extensions and abnormal accumulation of Thrombospondin and βPS-integrin, which may cause abnormal myotendinous junction formation and muscle morphogenesis. This role does not depend exclusively on nonmuscular Myosin-II activation (Myo-II), indicating that other DRok targets are key in this process. We propose that DRok function in tendon cells is key to promote the establishment of MTJ attachment and to balance mechanical tension generated at the MTJ by muscle compaction.