Liver Sinusoid on a Chip: Long-Term Layered Co-Culture of Primary Rat Hepatocytes and Endothelial Cells in Microfluidic Platforms

We describe the generation of microfluidic platforms for the co-culture of primary hepatocytes and endothelial cells; these platforms mimic the architecture of a liver sinusoid. This paper describes a progressional study of creating such a liver sinusoid on a chip system. Primary rat hepatocytes (PRHs) were co-cultured with primary or established endothelial cells in layers in single and dual microchannel configurations with or without continuous perfusion. Cell viability and maintenance of hepatocyte functions were monitored and compared for diverse experimental conditions. When primary rat hepatocytes were co-cultured with immortalized bovine aortic endothelial cells (BAECs) in a dual microchannel with continuous perfusion, hepatocytes maintained their normal morphology and continued to produce urea for at least 30 days. In order to demonstrate the utility of our microfluidic liver sinusoid platform, we also performed an analysis of viral replication for the hepatotropic hepatitis B virus (HBV). HBV replication, as measured by the presence of cell-secreted HBV DNA, was successfully detected. We believe that our liver model closely mimics the in vivo liver sinusoid and supports long-term primary liver cell culture. This liver model could be extended to diverse liver biology studies and liver-related disease research such as drug induced liver toxicology, cancer research, and analysis of pathological effects and replication strategies of various hepatotropic infectious agents

Photoreception and vision in the ultraviolet

Ultraviolet (UV) light occupies the spectral range of wavelengths slightly shorter than those visible to humans. Because of its shorter wavelength, it is more energetic (and potentially more photodamaging) than 'visible light', and it is scattered more efficiently in air and water. Until 1990, only a few animals were recognized as being sensitive to UV light, but we now know that a great diversity, possibly even the majority, of animal species can visually detect and respond to it. Here, we discuss the history of research on biological UV photosensitivity and review current major research trends in this field. Some animals use their UV photoreceptors to control simple, innate behaviors, but most incorporate their UV receptors into their general sense of vision. They not only detect UV light but recognize it as a separate color in light fields, on natural objects or living organisms, or in signals displayed by conspecifics. UV visual pigments are based on opsins, the same family of proteins that are used to detect light in conventional photoreceptors. Despite some interesting exceptions, most animal species have a single photoreceptor class devoted to the UV. The roles of UV in vision are manifold, from guiding navigation and orientation behavior, to detecting food and potential predators, to supporting high-level tasks such as mate assessment and intraspecific communication. Our current understanding of UV vision is restricted almost entirely to two phyla: arthropods and chordates (specifically, vertebrates), so there is much comparative work to be done

Hyperpolarized 13C Spectroscopic Evaluation of Oxidative Stress in a Rodent Model of Steatohepatitis.

Nonalcoholic fatty liver disease (NAFLD) has become highly prevalent, now considered the most common liver disease in the western world. Approximately one-third of patients with NASH develop non-alchoholic steatohepatitis (NASH), histologically defined by lobular and portal inflammation, and accompanied by marked oxidative stress. Patients with NASH are at increased risk for cirrhosis and hepatocellular carcinoma, and diagnosis currently requires invasive biopsy. In animal models of NASH, particularly the methionine-choline deficient (MCD) model, profound changes are seen in redox enzymes and key intracellular antioxidants. To study antioxidant status in NASH non-invasively, we applied the redox probe hyperpolarized [1-13C] dehydroascorbic acid (HP DHA), which is reduced to Vitamin C (VitC) rapidly in the normal liver. In MCD mice, we observed a significant decrease in HP DHA to VitC conversion that accompanied hepatic fat deposition. When these animals were subsequently placed on a normal diet, resonance ratios reverted to those seen in control mice. These findings suggest that HP DHA, a potentially clinically translatable imaging agent, holds special promise in imaging NASH and other metabolic syndromes, to monitor disease progression and response to targeted therapies

Crustacean Larvae-Vision in the Plankton

We review the visual systems of crustacean larvae, concentrating on the compound eyes of decapod and stomatopod larvae as well as the functional and behavioral aspects of their vision. Larval compound eyes of these macrurans are all built on fundamentally the same optical plan, the transparent apposition eye, which is eminently suitable for modification into the abundantly diverse optical systems of the adults. Many of these eyes contain a layer of reflective structures overlying the retina that produces a counterilluminating eyeshine, so they are unique in being camouflaged both by their transparency and by their reflection of light spectrally similar to background light to conceal the opaque retina. Besides the pair of compound eyes, at least some crustacean larvae have a non-imaging photoreceptor system based on a naupliar eye and possibly other frontal eyes. Larval compound-eye photoreceptors send axons to a large and well-developed optic lobe consisting of a series of neuropils that are similar to those of adult crustaceans and insects, implying sophisticated analysis of visual stimuli. The visual system fosters a number of advanced and flexible behaviors that permit crustacean larvae to survive extended periods in the plankton and allows them to reach acceptable adult habitats, within which to metamorphose

Behavioural evidence for polychromatic ultraviolet sensitivity in mantis shrimp

Stomatopod crustaceans are renowned for their elaborate visual systems. Their eyes contain a plethora of photoreceptors specialized for chromatic and polarization detection, including several that are sensitive to varying wavelength ranges and angles of polarization within the ultraviolet (UV) range (less than 400 nm). Behavioural experiments have previously suggested that UV photoreception plays a role in stomatopod communication, but these experiments have only manipulated the entire UV range. Here, using a behavioural approach, we examine UV vision in the stomatopod Haptosquilla trispinosa Using binary trained choice assays as well as innate burrow-choice experiments, we assessed the ability of H. trispinosa to detect and respond to narrow-band LED stimuli peaking near 314 nm (UVB) versus 379 nm (UVA) in wavelength. We find that H. trispinosa can discriminate these stimuli and appears to display an aversive reaction to UVB light, suggesting segregated behavioural responses to stimuli within the UV range. Furthermore, we find that H. trispinosa can discriminate stimuli peaking near 379 nm versus 351 nm in wavelength, suggesting that their wavelength discrimination in the UV is comparable to their performance in the human-visible range

Differences in signal contrast and camouflage among different colour variations of a stomatopod crustacean, Neogonodactylus oerstedii

Animal colouration is often a trade-off between background matching for camouflage from predators, and conspicuousness for communication with con- or heterospecifics. Stomatopods are marine crustaceans known to use colour signals during courtship and contests, while their overall body colouration may provide camouflage. However, we have little understanding of how stomatopods perceive these signals in their environment or whether overall body coloration does provide camouflage from predators. Neogonodactylus oerstedii assess meral spot colour during contests, and meral spot colour varies depending on local habitat. By calculating quantum catch for N. oerstedii’s 12 photoreceptors associated with chromatic vision, we found that variation in meral spot total reflectance does not function to increase signal contrast in the local habitat. Neogonodactylus oerstedii also show between-habitat variation in dorsal body colouration. We used visual models to predict a trichromatic fish predator’s perception of these colour variations. Our results suggest that sandy and green stomatopods are camouflaged from a typical fish predator in rubble fields and seagrass beds, respectively. To our knowledge, this is the first study to investigate signal contrast and camouflage in a stomatopod. These results provide new insight into the function and evolution of colouration in a species with a complex visual system

Expression of ABCA4 in the retinal pigment epithelium and its implications for Stargardt macular degeneration.

Recessive Stargardt disease (STGD1) is an inherited blinding disorder caused by mutations in the Abca4 gene. ABCA4 is a flippase in photoreceptor outer segments (OS) that translocates retinaldehyde conjugated to phosphatidylethanolamine across OS disc membranes. Loss of ABCA4 in Abca4 -/- mice and STGD1 patients causes buildup of lipofuscin in the retinal pigment epithelium (RPE) and degeneration of photoreceptors, leading to blindness. No effective treatment currently exists for STGD1. Here we show by several approaches that ABCA4 is additionally expressed in RPE cells. (i) By in situ hybridization analysis and by RNA-sequencing analysis, we show the Abca4 mRNA is expressed in human and mouse RPE cells. (ii) By quantitative immunoblotting, we show that the level of ABCA4 protein in homogenates of wild-type mouse RPE is about 1% of the level in neural retina homogenates. (iii) ABCA4 immunofluorescence is present in RPE cells of wild-type and Mertk -/- but not Abca4 -/- mouse retina sections, where it colocalizes with endolysosomal proteins. To elucidate the role of ABCA4 in RPE cells, we generated a line of genetically modified mice that express ABCA4 in RPE cells but not in photoreceptors. Mice from this line on the Abca4 -/- background showed partial rescue of photoreceptor degeneration and decreased lipofuscin accumulation compared with nontransgenic Abca4 -/- mice. We propose that ABCA4 functions to recycle retinaldehyde released during proteolysis of rhodopsin in RPE endolysosomes following daily phagocytosis of distal photoreceptor OS. ABCA4 deficiency in the RPE may play a role in the pathogenesis of STGD1

Hyperpolarized 13C-pyruvate MRI detects real-time metabolic flux in prostate cancer metastases to bone and liver: a clinical feasibility study.

BackgroundHyperpolarized (HP) 13C-pyruvate MRI is a stable-isotope molecular imaging modality that provides real-time assessment of the rate of metabolism through glycolytic pathways in human prostate cancer. Heretofore this imaging modality has been successfully utilized in prostate cancer only in localized disease. This pilot clinical study investigated the feasibility and imaging performance of HP 13C-pyruvate MR metabolic imaging in prostate cancer patients with metastases to the bone and/or viscera.MethodsSix patients who had metastatic castration-resistant prostate cancer were recruited. Carbon-13 MR examination were conducted on a clinical 3T MRI following injection of 250 mM hyperpolarized 13C-pyruvate, where pyruvate-to-lactate conversion rate (kPL) was calculated. Paired metastatic tumor biopsy was performed with histopathological and RNA-seq analyses.ResultsWe observed a high rate of glycolytic metabolism in prostate cancer metastases, with a mean kPL value of 0.020 ± 0.006 (s-1) and 0.026 ± 0.000 (s-1) in bone (N = 4) and liver (N = 2) metastases, respectively. Overall, high kPL showed concordance with biopsy-confirmed high-grade prostate cancer including neuroendocrine differentiation in one case. Interval decrease of kPL from 0.026 at baseline to 0.015 (s-1) was observed in a liver metastasis 2 months after the initiation of taxane plus platinum chemotherapy. RNA-seq found higher levels of the lactate dehydrogenase isoform A (Ldha,15.7 ± 0.7) expression relative to the dominant isoform of pyruvate dehydrogenase (Pdha1, 12.8 ± 0.9).ConclusionsHP 13C-pyruvate MRI can detect real-time glycolytic metabolism within prostate cancer metastases, and can measure changes in quantitative kPL values following treatment response at early time points. This first feasibility study supports future clinical studies of HP 13C-pyruvate MRI in the setting of advanced prostate cancer

Developmental studies provide new insights into the evolution of sense organs in Sabellariidae (Annelida)

[Background] Sabellarids, also known as honeycomb or sandcastle worms, when building their tubes, produce chemical signals (free fatty acids) that are responsible for larval settlement and the formation of three-dimensional aggregations. The larval palps and the dorsal hump (becoming the median organ in adults) are presumed to participate in such a substrate selection during settlement. Notably, the sabellariid median organ is an apparently unique organ among annelids that has been attributed with a sensory function and perhaps with some affinities to the nuchal organs of other polychaetes. Nevertheless, detailed investigations of this prominent character complex including ultrastructural examinations are lacking so far.[Results] Our comprehensive investigations provide data about the anterior sensory organs in Sabellariidae and inform about their transformation during pelagic larval development. We used a comparative approach including immunostaining with subsequent confocal laser scanning microscopy (clsm), histological sections as well as electron microscopy in a range of larval and adult stages of two sabellariid species. We find that the neuronal innervation as well as the ultrastructure of the sabellariid ciliary structures along the median organ are highly comparable with that of nuchal organs known from other polychaetes. Furthermore, the myoinhibitory protein (MIP) – a protein known to be also involved into chemo-sensation - was detected in the region of the larval median organ. Moreover, we reveal the presence of an unusual type of photoreceptor as part of the median organ in Idanthyrsus australiensis with a corrugated sensory membrane ultrastructure unlike those observed in the segmental ocelli of other polychaetes.[Conclusions] We are describing for the first time the nuchal organ-like structures in different developmental stages of two species of Sabellariidae. The external morphology, neuronal innervation, developmental fate and ultrastructure of the newly-discovered median organ-based ciliary pits are comparable with the characteristics known for annelid nuchal organs and therefore indicate a homology of both sensory complexes. The presence of myoinhibitory peptide (MIP) in the respective region supports such a hypothesis and exhibits the possibility of an involvement of the entire sabellariid median organ complex, and in particular the prominent ciliated pits, in chemo-sensation.CH was financed by a personal research fellowship (“Rückkehrerstipendium”) from the DFG (HE 7224/2–1). MC was financed by ForBio – the Research School in Biosystematics, funded by the Norwegian Taxonomy Initiative (70184215), the Research Council of Norway (248799) and the Ramón y Cajal program (RYC-2016-20799) funded by Spanish Ministerio de Economía, Industria y Competitividad, Agencia Estatal de Investigación, Comunidad Autónoma de las Islas Baleares and the European Social Fund. The European Commission Taxonomic Initiative SYNTHESYS (ES-TAF-7033) financed a visit to Museo Nacional de Ciencias Naturales, Madrid, Spain. MJB is supported by the Biotechnology and Biological Sciences Research Council (BBSRC, BB/P011357/1) and the Knut and Alice Wallenberg Foundation and the Swedish Research Council (grants awarded to Dan-Eric Nilsson).Peer reviewe