The unfolded protein response (UPR), which is activated when unfolded or misfolded proteins accumulate in the endoplasmic reticulum, has been implicated in the normal physiology of immune defense and in several human diseases including diabetes, cancer, neurodegenerative disease, and inflammatory disease. In this study, we found that the nervous system controlled the activity of a non-canonical UPR pathway required for innate immunity in Caenorhabditis elegans. OCTR-1, a putative octopamine G protein-coupled catecholamine receptor (GPCR, G protein–coupled receptor), functioned in sensory neurons designated ASH and ASI to actively suppress innate immune responses by down-regulating the expression of non-canonical UPR genes pqn/abu in non-neuronal tissues. Our findings suggest a novel molecular mechanism by which the nervous system may sense inflammatory responses and respond by controlling stress-response pathways at the organismal level.

Photoacoustic (PA) imaging for biomedical applications has been under development for many years. Based on the many advances over the past decade, a new photoacoustic imaging system has been integrated into a micro-ultrasound platform for co-registered PA-ultrasound (US) imaging. The design and implementation of the new scanner is described and its performance quantified. Beamforming techniques and signal processing are described, in conjunction with in vivo PA images of normal subcutaneous mouse tissue and selected tumor models. In particular, the use of the system to estimate the spatial distribution of oxygen saturation (sO2) in blood and co-registered with B-mode images of the surrounding anatomy are investigated. The system was validated in vivo against a complementary technique for measuring partial pressure of oxygen in blood (pO2). The pO2 estimates were converted to sO2 values based on a standard dissociation curve found in the literature. Preliminary studies of oxygenation effects were performed in a mouse model of breast cancer (MDA-MB-231) in which control mice were compared with mice treated with a targeted antiangiogenic agent over a 3 d period. Treated mice exhibited a >90% decrease in blood volume, an 85% reduction in blood wash-in rate, and a 60% decrease in relative tissue oxygenation.

Endothelial cells (ECs) are aligned longitudinally under laminar flow, whereas they are polygonal and poorly aligned in regions of disturbed flow. The unaligned ECs in disturbed flow fields manifest altered function and reduced survival that promote lesion formation. We demonstrate that the alignment of the ECs may directly influence their biology, independent of fluid flow. We developed aligned nanofibrillar collagen scaffolds that mimic the structure of collagen bundles in blood vessels, and examined the effects of these materials on EC alignment, function, and in vivo survival. ECs cultured on 30-nm diameter aligned fibrils re-organized their F-actin along the nanofibril direction, and were 50% less adhesive for monocytes than the ECs grown on randomly oriented fibrils. After EC transplantation into both subcutaneous tissue and the ischemic hindlimb, EC viability was enhanced when ECs were cultured and implanted on aligned nanofibrillar scaffolds, in contrast to non-patterned scaffolds. ECs derived from human induced pluripotent stem cells and cultured on aligned scaffolds also persisted for over 28 days, as assessed by bioluminescence imaging, when implanted in ischemic tissue. By contrast, ECs implanted on scaffolds without nanopatterning generated no detectable bioluminescent signal by day 4 in either normal or ischemic tissues. We demonstrate that 30-nm aligned nanofibrillar collagen scaffolds guide cellular organization, modulate endothelial inflammatory response, and enhance cell survival after implantation in normal and ischemic tissues.

The nervous system regulates C. elegans defense against pathogen infection by controlling the dynamics of cuticle structure.

The increased demand on protein folding in the endoplasmic reticulum (ER) during bacterial infection activates the unfolded protein response (UPR). OCTR-1--a G protein-coupled catecholamine receptor expressed in neurons--suppresses innate immunity by downregulating a non-canonical UPR pathway and the p38 MAPK pathway. Here, we show that OCTR-1 also regulates the canonical UPR pathway, which is controlled by XBP-1, at the organismal level. Importantly, XBP-1 is not under OCTR-1 control during development, only at the adult stage. Our results indicate that the nervous system temporally controls the UPR pathway to maintain ER homeostasis during development and immune activation.

Diacetyl (DA), a component of artificial butter flavoring, has been linked to the development of bronchiolitis obliterans (BO), a disease of airway epithelial injury and airway fibrosis. The epidermal growth factor receptor ligand, amphiregulin (AREG), has been implicated in other types of epithelial injury and lung fibrosis. We investigated the effects of DA directly on the pulmonary epithelium, and we hypothesized that DA exposure would result in epithelial cell shedding of AREG. Consistent with this hypothesis, we demonstrate that DA increases AREG by the pulmonary epithelial cell line NCI-H292 and by multiple independent primary human airway epithelial donors grown under physiologically relevant conditions at the air-liquid interface. Furthermore, we demonstrate that AREG shedding occurs through a TNF-a-converting enzyme (TACE)-dependent mechanism via inhibition of TACE activity in epithelial cells using the small molecule inhibitor, TNF-a protease inhibitor-1, as well as TACE-specific small inhibitor RNA. Finally, we demonstrate supportive in vivo results showing increased AREG transcript and protein levels in the lungs of rodents with DA-induced BO. In summary, our novel in vitro and in vivo observations suggest that further study of AREG is warranted in the pathogenesis of DA-induced BO.

SummaryEpidemiological studies have demonstrated that co-infections of herpes simplex virus type 2 (HSV-2) and Chlamydia trachomatis occur in vivo. Data from a tissue culture model of C. trachomatis/HSV-2 co-infection indicate that viral co-infection stimulates the formation of persistent chlamydiae. Transmission electron microscopic (TEM) analyses demonstrated that in both HeLa and HEC-1B cells, co-infection caused developing chlamydiae to exhibit swollen, aberrantly shaped reticulate bodies (RBs), characteristically observed in persistence. Additionally, HSV-2 co-infection suppressed production of infectious chlamydial elementary bodies (EBs) in both host cell types. Co-infection with HSV type 1 (HSV-1) produced similar morphologic alterations and abrogated infectious EB production. These data indicate that virusinduced chlamydial persistence was neither host cellnor virus strain-specific. Purification of crude HSV-2 stocks demonstrated that viral particles were required for coinfection-induced chlamydial persistence to occur. Finally, co-infection with either UV-inactivated, replication-incompetent virus or replication-competent HSV-2 in the presence of cyclohexamide reduced chlamydial infectivity without altering chlamydial genomic DNA accumulation. These data demonstrate that productive viral replication is not required for the induction of chlamydial persistence and suggest that HSV attachment and entry can provide the necessary stimulus to alter C. trachomatis development.

Metronomic chemotherapy refers to the close, regular administration of conventional chemotherapy drugs at relatively low minimally toxic doses with no prolonged break periods; it is now showing encouraging results in various phase II clinical trials, and is currently undergoing phase III trial evaluation. It is thought to cause anti-tumor effects primarily by antiangiogenic mechanisms, both locally by targeting endothelial cells of the tumor neovasculature and systemically by effects on bone marrow derived cells, including circulating endothelial progenitor cells (CEPs). Previous studies have shown reduction of CEPs by metronomic administration of a number of different chemotherapeutic drugs, including vinblastine, cyclophosphamide, paclitaxel, topotecan, and tegafur plus uracil (UFT). However in addition to, or even instead of, anti angiogenic effects, metronomic chemotherapy may cause suppression of tumor growth by other mechanisms such as stimulating cytotoxic T cell responses, or by direct anti-tumor effects. Here we report results evaluating the properties of metronomic administration of an oral prodrug of gemcitabine LY2334737 (LY) in non tumor-bearing mice, and in preclinical models of human ovarian (SKOV3-13) and breast cancer (LM2-4) xenografts. Through daily gavage (at 6mg/kg/day) the schedules tested were devoid of toxicity and caused anti-tumor effects; however, a suppressive effect on CEPs was not detected. Unexpectedly metronomic LY administration caused increased blood flow in luciferase-tagged LM2-4 tumor xenografts; and this effect coincided with a relative increase in tumor bioluminescence. These results highlight the possibility of significant anti-tumor effects mediated by metronomic administration of some chemotherapy drugs without a concomitant inhibition of systemic angiogenesis.