Distinct Tissue Layers Research Articles

Novel technologies uncover novel 'anti'-microbial peptides in Hydra shaping the species-specific microbiome.

The freshwater polyp Hydra uses an elaborate innate immune machinery to maintain its specific microbiome. Major components of this toolkit are conserved Toll-like receptor (TLR)-mediated immune pathways and species-specific antimicrobial peptides (AMPs). Our study harnesses advanced technologies, such as high-throughput sequencing and machine learning, to uncover a high complexity of the Hydra's AMPs repertoire. Functional analysis reveals that these AMPs are specific against diverse members of the Hydra microbiome and expressed in a spatially controlled pattern. Notably, in the outer epithelial layer, AMPs are produced mainly in the neurons. The neuron-derived AMPs are secreted directly into the glycocalyx, the habitat for symbiotic bacteria, and display high selectivity and spatial restriction of expression. In the endodermal layer, in contrast, endodermal epithelial cells produce an abundance of different AMPs including members of the arminin and hydramacin families, while gland cells secrete kazal-type protease inhibitors. Since the endodermal layer lines the gastric cavity devoid of symbiotic bacteria, we assume that endodermally secreted AMPs protect the gastric cavity from intruding pathogens. In conclusion, Hydra employs a complex set of AMPs expressed in distinct tissue layers and cell types to combat pathogens and to maintain a stable spatially organized microbiome. This article is part of the theme issue 'Sculpting the microbiome: how host factors determine and respond to microbial colonization'.

Technical note: The design and validation of a multi-modality lung phantom.

Clinically relevant models that enable certain tasks such as calibration of medical imaging devices or techniques, device validation, training healthcare professionals, and more are vital to research throughout the medical field and are referred to as phantoms. Phantoms range in complexity from a vile of water to complex designs that emulate in vivo properties. Specific phantoms that model the lungs have focused on replication of tissue properties but lack replication of the anatomy. This limits the use across multiple imaging modalities and for device testing when anatomical considerations as well as tissue properties are needed. This work reports a lung phantom design utilizing materials that accurately mimic the ultrasound and magnetic resonance imaging (MRI) properties of in vivo lungs and includes relevant anatomical equivalence. The tissue mimicking materials were selected based on published studies of the materials, through qualitative comparisons of the materials with ultrasound imaging, and quantitative MRI relaxation values. A PVC ribcage was used as the structural support. The muscle/fat combined layer and the skin layer were constructed with various types of silicone with graphite powder added as a scattering agent where appropriate. Lung tissue was mimicked with silicone foam. The pleural layer was replicated by the interface between the muscle/fat layer and the lung tissue layer, requiring no additional material. The design was validated by accurately mimicking the distinct tissue layers expected with in vivo lung ultrasound while maintaining tissue-mimicking relaxation values in MRI as compared to reported values. Comparisons between the muscle/fat material and in vivo muscle/fat tissue demonstrated a 1.9% difference in T1 relaxation and a 19.8% difference in T2 relaxation. Qualitative US and quantitative MRI analysis verified the proposed lung phantom design for accurate modeling of the human lungs.

Tissue-specific mRNA profiling of the Brassica napus-Sclerotinia sclerotiorum interaction uncovers novel regulators of plant immunity.

White mold is caused by the fungal pathogen Sclerotinia sclerotiorum and leads to rapid and significant loss in plant yield. Among its many brassicaceous hosts, including Brassica napus (canola) and Arabidopsis, the response of individual tissue layers directly at the site of infection has yet to be explored. Using laser microdissection coupled with RNA sequencing, we profiled the epidermis, mesophyll, and vascular leaf tissue layers of B. napus in response to S. sclerotiorum. High-throughput tissue-specific mRNA sequencing increased the total number of detected transcripts compared with whole-leaf assessments and provided novel insight into the conserved and specific roles of ontogenetically distinct leaf tissue layers in response to infection. When subjected to pathogen infection, the epidermis, mesophyll, and vasculature activate both specific and shared gene sets. Putative defense genes identified through transcription factor network analysis were then screened for susceptibility against necrotrophic, hemi-biotrophic, and biotrophic pathogens. Arabidopsis deficient in PR5-like RECEPTOR KINASE (PR5K) mRNA levels were universally susceptible to all pathogens tested and were further characterized to identify putative interacting partners involved in the PR5K signaling pathway. Together, these data provide insight into the complexity of the plant defense response directly at the site of infection.

Spatially expressed WIP genes control Arabidopsis embryonic root development.

Development of plant organs is a highly organized process. In Arabidopsis, proper root development requires that distinct cell types and tissue layers are specified and formed in a restricted manner in space and over time. Despite its importance, genetic controls underlying such regularity remain elusive. Here we found that WIP genes expressed in the embryo and suspensor functionally oppose those expressed in the surrounding maternal tissues to orchestrate cell division orientation and cell fate specification in the embryonic root, thereby promoting regular root formation. The maternal WIPs act non-cell autonomously to repress root cell fate specification through SIMILAR TO RADICAL-INDUCED CELL DEATH ONE (SRO) family members. When losing all WIPs, root cells divide irregularly in the early embryo, but this barely alters their fate specification and the morphology of post-embryonic roots. Our results reveal cross-communication between the embryonic and maternal WIPs in controlling root development.

Gastrointestinal Tract and Accessory Organs in the Spotted Bent-toed Gecko, Cyrtodactylus peguensis (Boulenger, 1893): A Histological and Histochemical Study

AbstractThe spotted bent-toed gecko Cyrtodactylus peguensis is one of the exploited reptiles in Thailand. In order to provide basic information for the digestive system of this species, we have examined histologically the gastrointestinal and accessory organs of C. peguensis using routine methods. The gastrointestinal region of this reptile started from the stomach and the intestine. The stomach was separated into fundic and pyloric regions. In both regions, the stomach wall was formed by four distinct tissue layers, including mucosa, submucosa, muscularis, and serosa layers. Mucous neck cells and oxynticopeptic cells were identified as glycoprotein-producing cells in the stomach by Periodic acid-Schiff (PAS) staining. The small and large intestines shared many histological characteristics, but the former contained more intestinal folds, while the latter had more PAS-positive goblet cells. Histological characteristics of accessory organs, liver and pancreas, were also provided. Overall, the gastrointestinal and accessory organs of C. peguensis were largely similar to those from other reptiles, but fine structural information will open up considerable opportunities to further studies related to the endocrinology, the physiology, and the conservation of this species.

3D printed tissue engineered model for bone invasion of oral cancer.

Recent advances in three-dimensional printing technology have led to a rapid expansion of its applications in tissue engineering. The present study was designed to develop and characterize an in vitro multi-layered human alveolar bone, based on a 3D printed scaffold, combined with tissue engineered oral mucosal model. The objective was to incorporate oral squamous cell carcinoma (OSCC) cell line spheroids to the 3D model at different anatomical levels to represent different stages of oral cancer. Histological evaluation of the 3D tissue model revealed a tri-layered structure consisting of distinct epithelial, connective tissue, and bone layers; replicating normal oral tissue architecture. The mucosal part showed a well-differentiated stratified oral squamous epithelium similar to that of the native tissue counterpart, as demonstrated by immunohistochemistry for cytokeratin 13 and 14. Histological assessment of the cancerous models demonstrated OSCC spheroids at three depths including supra-epithelial level, sub-epithelial level, and deep in the connective tissue-bone interface. The 3D tissue engineered composite model closely simulated the native oral hard and soft tissues and has the potential to be used as a valuable in vitro model for the investigation of bone invasion of oral cancer and for the evaluation of novel diagnostic or therapeutic approaches to manage OSCC in the future.

Usefulness of a left ventricular assist device in patients with left ventricular noncompaction

ABSTRACTLeft ventricular noncompaction (LVNC) is a multifactorial structural abnormality of the myocardial wall characterized by prominent trabeculae and deep trabecular recesses. LVNC may present as a congenital or acquired defect characterized by 2 distinct tissue layers: a spongy, noncompacted inner myocardium and a thin, compacted outer myocardium. Patients with LVNC are prone to thromboembolic events, either due to deep trabeculations in the noncompacted myocardium or due to arrhythmias accompanying the defect. There are sparse data concerning treatment options for patients with LVNC who fail medical management. We present 2 such patients with LVNC who, following failed medical management, received a left ventricular assist device (LVAD): one for long-term management and one as a bridge to transplant. Both were managed successfully without thromboembolic events to date. The success of these cases suggests that LVAD placement is a viable therapy in patients with LVNC as a bridge to transplant or as long-term management.

More than Just a Simple Cardiac Envelope; Cellular Contributions of the Epicardium.

The adult pumping heart is formed by distinct tissue layers. From inside to outside, the heart is composed by an internal endothelial layer, dubbed the endocardium, a thick myocardial component which supports the pumping capacity of the heart and exteriorly covered by a thin mesothelial layer named the epicardium. Cardiac insults such as coronary artery obstruction lead to ischemia and thus to an irreversible damage of the myocardial layer, provoking in many cases heart failure and death. Thus, searching for new pathways to regenerate the myocardium is an urgent biomedical need. Interestingly, the capacity of heart regeneration is present in other species, ranging from fishes to neonatal mammals. In this context, several lines of evidences demonstrated a key regulatory role for the epicardial layer. In this manuscript, we provide a state-of-the-art review on the developmental process leading to the formation of the epicardium, the distinct pathways controlling epicardial precursor cell specification and determination and current evidences on the regenerative potential of the epicardium to heal the injured heart.

Characterizing and Simulating Needle Insertion Forces for Percutaneous Renal Access.

Percutaneous needle access is a critical step of performing percutaneous nephrolithotomy. Development of a synthetic model that accurately represents the forces encountered while gaining percutaneous renal access, allows for high ease of use and prevention of negative skill transfer. The objectives of this study were to define the needle insertion forces used during percutaneous renal access and to develop a multilayer synthetic physical simulator model based on human tissue data that is compatible with the SimPORTAL fluoro-less C-arm trainer (CAT) camera system. Needle insertion data were collected using in situ fresh human cadaveric tissue within 72 hours of death. Ultrasound guidance was used to place percutaneous reference needles into the kidney, and axial force vs displacement data was collected using a custom-built force measurement device. A novel multilayer model that includes several types of synthetic materials for simulation of distinct tissue layers was developed based on the human tissue reference data. The multilayer prototype model and an existing single material model were subsequently tested using the same needle insertion protocol and the results were compared with human tissue data. Average maximum forces for needle puncture into skin ranged from 2.75 to 2.80 N for human tissue and from 4.53 to 4.19 N for simulated human tissue. The overall slope for all force vs displacement data was 0.35 to 0.59 N/cm for human tissue. The overall slope was comparatively lower for the multilayer model (0.17 N/cm) and was highest in the existing single material model (5.08-9.79 N/cm). We have defined the forces for percutaneous renal access using fresh human cadaveric tissue and designed a multilayer synthetic simulator model that can be utilized for training percutaneous needle access to the renal collecting system using the CAT camera system.

Submucosal Elastic Laminae of the Middle and Lower Pharynx: A Histological Study Using Elderly Cadaveric Specimens.

Although the pharyngeal wall is well known to have high elasticity, the distribution of submucosal elastic fibers has not been described. Observations of histological sections of the mid and lower pharyngeal walls from 15 elderly donated cadavers were made. We found two distinct submucosal tissue layers with a high content of elastic fibers (tentatively termed the "submucosal elastic laminae"). The inferolateral elastic lamina was restricted to the level from the upper part of the arytenoid to the lower end of the inferior cornu of the thyroid cartilage. It originated from the pharyngeal submucosa, extended laterally along the inner aspect of the thyropharyngeal muscle, and inserted into the posterior margin of the thyroid cartilage including the cornu. The posteromedial lamina extended along the supero-inferior axis from a level above the greater horn of the hyoid bone to reach the muscularis mucosae of the cervical esophagus. The inferolateral and posteromedial laminae were connected at levels below the cricoarytenoid joint. Individual variations were evident in their thicknesses (ranging from almost absent to 0.3mm) as well as the extent of connection between them. In association with striated muscle function, the inferolateral lamina seemed to suspend the lower pharyngeal mucosa, while the posteromedial lamina seemed to provide mucosal fold forcing smoothly peristaltic conveyance of a bolus during swallowing.

Neuro diffusion MRI beyond myelinated white matter: cell structures that influence water diffusion in the fetal brain

Event Abstract Back to Event Neuro diffusion MRI beyond myelinated white matter: cell structures that influence water diffusion in the fetal brain Christopher Kroenke1 and Xiaojie Wang1* 1 Oregon Health & Science University, Advanced Imaging Research Center and Oregon National Primate Research Center, United States Distinct populations of neurons and glial cells are stratified in the fetal primate brain across several layers that span from the inner ventricular surface to the outer-most layer of the cerebral cortex. Changes in the sizes of these layers and the morphology of the cells within each layer accompany normal brain development, and the ability to detect deviations from the normal developmental trajectory is highly relevant to the study of many neurodevelopmental disorders. As an example, we have previously demonstrated that water diffusion is very anisotropic in the cerebral cortex at mid-gestation, and that diffusion anisotropy within this tissue layer decreases with maturation. In our modeling approach, we have linked anisotropy in water diffusion to dispersion in the orientation of axonal and dendritic cellular processes. In animal models of three neurodevelopmental disorders, fetal alcohol spectrum disorders, perinatal hypoxia/ischemia, and sensory deprivation, we have demonstrated that alterations in morphological development of cerebral cortical neurons can be revealed by diffusion MRI. Here, we focus on the remaining layers of the mid-gestation brain. A diffusion tensor analysis of post-mortem rhesus macaque brain reveals five distinct tissue layers, including the cerebral cortex. These layers are registered to previously-defined tissue zones identified by histological analyses. Interestingly, two of the MRI-defined zones, which exhibit strikingly different water apparent diffusion coefficients, reside within the subplate zone defined in histological studies. Subsequent to diffusion MRI performed histological analyses on the tissue. We find that multiple different cellular processes (axons, radial glial processes, and dendrites) contribute to diffusion anisotropy within each zone. For each of the five layers, these relationships are described, and unanswered questions related to the determinants of the water apparent diffusion coefficient are identified. Keywords: Diffusion, MRI, dMRI, multidimensional, diffusion encoding Conference: New dimensions in diffusion encoding, Fjälkinge, Sweden, 11 Jan - 14 Jan, 2016. Presentation Type: Oral presentation Topic: New Dimensions in Diffusion Encoding Citation: Kroenke C and Wang X (2016). Neuro diffusion MRI beyond myelinated white matter: cell structures that influence water diffusion in the fetal brain. Front. Phys. Conference Abstract: New dimensions in diffusion encoding. doi: 10.3389/conf.FPHY.2016.01.00027 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 07 Jul 2016; Published Online: 07 Jul 2016. * Correspondence: PhD. Xiaojie Wang, Oregon Health & Science University, Advanced Imaging Research Center and Oregon National Primate Research Center, Portland, Oregon, OR 97239, United States, waxia@ohsu.edu Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Christopher Kroenke Xiaojie Wang Google Christopher Kroenke Xiaojie Wang Google Scholar Christopher Kroenke Xiaojie Wang PubMed Christopher Kroenke Xiaojie Wang Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Blood flow mechanics and oxygen transport and delivery in the retinal microcirculation: multiscale mathematical modeling and numerical simulation.

The scientific community continues to accrue evidence that blood flow alterations and ischemic conditions in the retina play an important role in the pathogenesis of ocular diseases. Many factors influence retinal hemodynamics and tissue oxygenation, including blood pressure, blood rheology, oxygen arterial permeability and tissue metabolic demand. Since the influence of these factors on the retinal circulation is difficult to isolate in vivo, we propose here a novel mathematical and computational model describing the coupling between blood flow mechanics and oxygen ([Formula: see text]) transport in the retina. Albeit in a simplified manner, the model accounts for the three-dimensional anatomical structure of the retina, consisting in a layered tissue nourished by an arteriolar/venular network laying on the surface proximal to the vitreous. Capillary plexi, originating from terminal arterioles and converging into smaller venules, are embedded in two distinct tissue layers. Arteriolar and venular networks are represented by fractal trees, whereas capillary plexi are represented using a simplified lumped description. In the model, [Formula: see text] is transported along the vasculature and delivered to the tissue at a rate that depends on the metabolic demand of the various tissue layers. First, the model is validated against available experimental results to identify baseline conditions. Then, a sensitivity analysis is performed to quantify the influence of blood pressure, blood rheology, oxygen arterial permeability and tissue oxygen demand on the [Formula: see text] distribution within the blood vessels and in the tissue. This analysis shows that: (1) systemic arterial blood pressure has a strong influence on the [Formula: see text] profiles in both blood and tissue; (2) plasma viscosity and metabolic consumption rates have a strong influence on the [Formula: see text] tension at the level of the retinal ganglion cells; and (3) arterial [Formula: see text] permeability has a strong influence on the [Formula: see text] saturation in the retinal arterioles.

Homogenization of oxygen transport in biological tissues

In this paper, we extend previous work on the mathematical modeling of oxygen transport in biological tissues. Specifically, we include in the modeling process the arterial and venous microstructure within the tissue by means of homogenization techniques. We focus on the two-layer tissue architecture investigated in [Matzavinos et al., PNAS; 2009;106:12091] in the context of abdominal tissue flaps that are commonly used for reconstructive surgery. We apply two-scale convergence methods and unfolding operator techniques to homogenize the developed microscopic model, which involves different unit cell geometries in the two distinct tissue layers (skin layer and fat tissue) to account for different arterial branching patterns.

Developmental anatomy and immunocytochemistry reveal the neo-ontogenesis of the leaf tissues of Psidium myrtoides (Myrtaceae) towards the globoid galls of Nothotrioza myrtoidis (Triozidae).

The temporal balance between hyperplasia and hypertrophy, and the new functions of different cell lineages led to cell transformations in a centrifugal gradient that determines the gall globoid shape. Plant galls develop by the redifferentiation of new cell types originated from those of the host plants, with new functional and structural designs related to the composition of cell walls and cell contents. Variations in cell wall composition have just started to be explored with the perspective of gall development, and are herein related to the histochemical gradients previously detected on Psidium myrtoides galls. Young and mature leaves of P. myrtoides and galls of Nothotrioza myrtoidis at different developmental stages were analysed using anatomical, cytometrical and immunocytochemical approaches. The gall parenchyma presents transformations in the size and shape of the cells in distinct tissue layers, and variations of pectin and protein domains in cell walls. The temporal balance between tissue hyperplasia and cell hypertrophy, and the new functions of different cell lineages led to cell transformations in a centrifugal gradient, which determines the globoid shape of the gall. The distribution of cell wall epitopes affected cell wall flexibility and rigidity, towards gall maturation. By senescence, it provided functional stability for the outer cortical parenchyma. The detection of the demethylesterified homogalacturonans (HGAs) denoted the activity of the pectin methylesterases (PMEs) during the senescent phase, and was a novel time-based detection linked to the increased rigidity of the cell walls, and to the gall opening. Current investigation firstly reports the influence of immunocytochemistry of plant cell walls over the development of leaf tissues, determining their neo-ontogenesis towards a new phenotype, i.e., the globoid gall morphotype.

ATML1 promotes epidermal cell differentiation in Arabidopsis shoots

Molecular mechanisms that generate distinct tissue layers in plant shoots are not well understood. ATML1, an Arabidopsis homeobox gene, is expressed in the outermost cell layer, beginning at an early stage of development. The promoters of many epidermis-specific genes, including ATML1, contain an ATML1-binding site called an L1 box, suggesting that ATML1 regulates epidermal cell fate. Here, we show that overexpression of ATML1 was sufficient to activate the expression of epidermal genes and to induce epidermis-related traits such as the formation of stomatal guard cells and trichome-like cells in non-epidermal seedling tissues. Detailed observation of the division planes of these ectopic stomatal cells suggested that a near-surface position, as well as epidermal cell identity, were required for regular anticlinal cell division, as seen in wild-type epidermis. Moreover, analyses of a loss-of-function mutant and overexpressors implied that differentiation of epidermal cells was associated with repression of mesophyll cell fate. Collectively, our studies contribute new information about the molecular basis of cell fate determination in different layers of plant aerial organs.

K16 is a further new candidate for homotypic intermediate filament protein interactions

Keratin filaments form obligatory heterodimers consisting of one type I and one type II keratin that build the intermediate filaments (IF). These filaments mediate resilience and mechanical strength to epithelial cells and maintain tissue integrity. Specific type I/type II pairs are co-expressed in vivo and serve as markers for distinct tissue layers and cell differentiation states. Heterodimerization has been regarded the undisrupted hallmark of IF. We show now that recombinantly expressed cytokeratin 16 (K16) interacts with itself and forms homodimers even in denaturating SDS-PAGE analysis. Detailed FRET experiments in HaCaT keratinocytes were in accordance with our in vitro observations and showed clearly that K16 is able to form strong homodimers. Homotypic keratin interactions has been previously shown for keratin 17 (K17) and keratin 18 (K18) by Schnabel et al. (Biochim Biophys Acta, 1998: 1403: 158), and we now proved K16 to be the third type I keratin that is able to form homodimers.

Seasonal variation in photosynthesis and diel carbon balance under natural conditions in two Peperomia species that differ with respect to leaf anatomy1

Abstract The occurrence of crassulacean acid metabolism (CAM) and C3 photosynthetic activities in Peperomia species has been linked to the differential distribution of C3 and C4 photosynthetic enzymes within the distinct leaf tissue layers. Our objectives were to test if relative thickness of the spongy mesophyll layer is an indication of CAM and if the distribution of water in the leaf tissue changes in response to drought. Under field conditions we studied photosynthesis and water relations of Peperomia obtusifolia and P. macrostachya, species that differ in leaf anatomy. Gas exchange and titratable acidity were measured over a complete 24 hour photoperiod. Plant water status was determined at dawn while chlorophyll a fluorescence data were collected during the day (12 noon). Thickness of the distinct leaf layers was determined with the aid of a calibrated light microscope. Gas exchange and titratable acidity showed that when well-hydrated, P. obtusifolia has typical C3 activity and P. macrostachya has ...

Protein distribution of Kcnq1, Kcnh2, and Kcne3 potassium channel subunits during mouse embryonic development

Voltage-dependent potassium channels consist of a pore-forming alpha-subunit, which is modulated by additional beta-ancillary or regulatory subunits. Kcnq1 and Kcnh2 alpha-channel subunits play pivotal roles in the developing and adult heart. However, Kcnq1 and Kcnh2 have a much wider expression profile than strictly confined to the myocardium, similar to their putative regulatory Kcne1-5 beta-subunits. At present, the distribution of distinct potassium channel subunits has been partially mapped in adult tissues, whereas almost no information is available during embryonic development. In this study, we report a detailed analysis of Kcnq1, Kcnh2, and Kcne3 protein expression during mouse embryogenesis. Our results demonstrate that Kcnq1 and Kcnh2 are widely distributed. Coexpression of both alpha-subunits is observed in a wide variety of organs, such as heart and the skeletal muscle, whereas others display unique Kcnq1 or Knch2 expression. Interestingly, Kcne3 expression is also widely observed in distinct tissue layers during embryogenesis, supporting the notion that an exquisite balance of alpha- and beta-subunit expression is required for modulating potassium conductance in distinct organs and tissue layers.

Localization of Neogenin protein during morphogenesis in the mouse embryo

Neogenin, a close relative of the axon guidance receptor DCC, has been shown to be a receptor for members of the Netrin and Repulsive Guidance Molecule families. Recent studies have begun to uncover a role for Neogenin in organogenesis. Here we examine the localization of Neogenin protein in the developing mouse embryo (embryonic day 14.5) when organogenesis is progressing rapidly. We observe that Neogenin protein is restricted to distinct tissue layers within a given organ. In some embryonic epithelia such as the gut and pancreas, Neogenin protein is predominantly polarized to the basal surfaces of the epithelial cells. In contrast, Neogenin is restricted to mesenchymal cells within the lung and kidney. Neogenin is also seen in differentiating skeletal muscle and condensing cartilage throughout the embryo, and in the trigeminal and dorsal root ganglia of the peripheral nervous system. This study supports the emerging role for Neogenin as a key receptor in the establishment of the morphological architecture in many developing organ systems.

Spider silk protein refolding is controlled by changing pH.

Spidroins, the major silk proteins making up the spider's dragline silk, originate in two distinct tissue layers (A and B) in the spider's major ampullate gland. Formation of the complex thread from spidroins occurs in the lumen of the duct connected to the gland. Using pH-sensitive microelectrode probes, we showed that the spidroins traveling through the gland and duct experience a monotonic decrease in pH from 7.2 to 6.3. In addition, circular dichroism spectroscopy of material extracted from the gland showed a structural refolding concomitant with position in the gland and post-extraction changes in pH. We demonstrate that lowering the pH in vitro causes a dramatic conformational change in the protein from the A zone, converting it irreversibly from a coil to a predominantly beta-sheet structure. Furthermore, amino acid analyses have indicated that there are at least two distinct, though similar, proteins secreted in the A and B zones suggesting a potential factor in the progressive acidification as well as a pH sensitivity of the folding of spidroins in the gland. Thus, we provide, for the first time, a quantitative map of the pH value and position correlated with molecular structural folding in the silk gland characterizing the crucial role that pH plays in spider silk formation.