Basal Side Of Cells Research Articles

Elastic Contractile Stress in the Basement Membrane Generates Basal Tension in Epithelia

The shaping of epithelial tissues into functional organs often depends on dynamic changes of mechanical tension at the apical and basal sides of cells. While on the apical side, tension is known to be regulated by the apical actomyosin meshwork, basal tension regulation remains elusive due to the presence of a basal sheet of specialized extracellular matrix, the basement membrane. Here, we use atomic force microscopy to provide direct measurements of mechanical tension in the basal surface of the wing disk epithelium of in control and perturbed conditions. We find that next to actomyosin, the basement membrane is a key structure for the generation of basal tension. Furthermore, we show that the basement membrane is a solidlike sheet subject to an expansile stretch in epithelial homeostasis. Performing osmotic shocks and optogenetic actomyosin activation, we find evidence that basal stretch and tension can be generated by intracellular hydrostatic pressure and actomyosin tension in lateral cell faces of the epithelium. In conclusion, we propose that elastic contractile stress corresponding to the actomyosin-induced expansile stretch of the basement membrane generates a major contribution to basal tension. We thereby disclose fundamental differences in the mechanical regulation of apical and basal epithelial tension. Published by the American Physical Society 2024

Pulses of RhoA signaling stimulate actin polymerization and flow in protrusions to drive collective cell migration.

In animals, cells often move as collectives to shape organs, close wounds, or-in the case of disease-metastasize. To accomplish this, cells need to generate force to propel themselves forward. The motility of singly migrating cells is driven largely by an interplay between Rho GTPase signaling and the actin network. Whether cells migrating as collectives use the same machinery for motility is unclear. Using the zebrafish posterior lateral line primordium as a model for collective cell migration, we find that active RhoA and myosin II cluster on the basal sides of the primordium cells and are required for primordium motility. Positive and negative feedbacks cause RhoA and myosin II activities to pulse. These pulses of RhoA signaling stimulate actin polymerization at the tip of the protrusions and myosin-II-dependent actin flow and protrusion retraction at the base of the protrusions and deform the basement membrane underneath the migrating primordium. This suggests that RhoA-induced actin flow on the basal sides of the cells constitutes the motor that pulls the primordium forward, a scenario that likely underlies collective migration in other contexts.

The development and characterization of in vivo-like three-dimensional models of bronchial epithelial cell lines

In vitro models of differentiated respiratory epithelium that allow high-throughput screening are an important tool to explore new therapeutics for chronic respiratory diseases. In the present study, we developed in vivo-like three-dimensional (3-D) models of bronchial epithelial cell lines that are commonly used to study chronic lung disease (16HBE14o−, CFBE41o− and CFBE41o− 6.2 WT-CFTR). To this end, cells were cultured on porous microcarrier beads in the rotating wall vessel (RWV) bioreactor, an optimized suspension culture method that allows higher throughput experimentation than other physiologically relevant models. Cell differentiation was compared to conventional two-dimensional (2-D) monolayer cultures and to the current gold standard in the respiratory field, i.e. air-liquid interface (ALI) cultures. Cellular differentiation was assessed in the three model systems by evaluating the expression and localization of markers that reflect the formation of tight junctions (zonula occludens 1), cell polarity (intercellular adhesion molecule 1 at the apical side and collagen IV expression at the basal cell side), multicellular complexity (acetylated α-tubulin for ciliated cells, CC10 for club cells, keratin-5 for basal cells) and mucus production (MUC5AC) through immunostaining and confocal laser scanning microscopy. Results were validated using Western Blot analysis. We found that tight junctions were expressed in 2-D monolayers, ALI cultures and 3-D models for all three cell lines. All tested bronchial epithelial cell lines showed polarization in ALI and 3-D cultures, but not in 2-D monolayers. Mucus secreting goblet-like cells were present in ALI and 3-D cultures of CFBE41o− and CFBE41o− 6.2 WT-CFTR cells, but not in 16HBE14o− cells. For all cell lines, there were no ciliated cells, basal cells, or club cells found in any of the model systems. In conclusion, we developed RWV-derived 3-D models of commonly used bronchial epithelial cell lines and showed that these models are a valuable alternative to ALI cultures, as they recapitulate similar key aspects of the in vivo parental tissue.

Complement factor B is critical for sub-RPE deposit accumulation in a model of Doyne honeycomb retinal dystrophy with features of age-related macular degeneration.

EFEMP1 R345W is a dominant mutation causing Doyne honeycomb retinal dystrophy/malattia leventinese (DHRD/ML), a rare blinding disease with clinical pathology similar to age-related macular degeneration (AMD). Aged Efemp1 R345W/R345W knock-in mice (Efemp1ki/ki) develop microscopic deposits on the basal side of retinal pigment epithelial cells (RPE), an early feature in DHRD/ML and AMD. Here, we assessed the role of alternative complement pathway component factor B (FB) in the formation of these deposits. RNA-seq analysis of the posterior eyecups revealed increased unfolded protein response, decreased mitochondrial function in the neural retina (by 3months of age) and increased inflammatory pathways in both neural retina and posterior eyecups (at 17months of age) of Efemp1ki/ki mice compared with wild-type littermate controls. Proteomics analysis of eye lysates confirmed similar dysregulated pathways as detected by RNA-seq. Complement activation was increased in aged Efemp1ki/ki eyes with an approximately 2-fold elevation of complement breakdown products iC3b and Ba (P < 0.05). Deletion of the Cfb gene in female Efemp1ki/ki mice partially normalized the above dysregulated biological pathway changes and oral dosing of a small molecule FB inhibitor from 10 to 12 months of age reduced sub-RPE deposits by 65% (P = 0.029). In contrast, male Efemp1ki/ki mice had fewer sub-RPE deposits than age-matched females, no elevation of ocular complement activation and no effect of FB inhibition on sub-RPE deposits. The effects of FB deletion or inhibition on Efemp1ki/ki mice supports systemic inhibition of the alternative complement pathway as a potential treatment of dry AMD and DHRD/ML.

Confocal and Super-Resolution Imaging of Polarized Intracellular Trafficking and Secretion of Basement Membrane Proteins During Drosophila Oogenesis.

The basement membrane (BM) - a specialized sheet of extracellular matrix present at the basal side of epithelial cells - is critical for the establishment and maintenance of epithelial tissue morphology and organ morphogenesis. Moreover, the BM is essential for tissue modeling, serving as a signaling platform, and providing external forces to shape tissues and organs. Despite the many important roles that the BM plays during normal development and pathological conditions, the biological pathways controlling the intracellular trafficking of BM-containing vesicles and how basal secretion leads to the polarized deposition of BM proteins are poorly understood. The follicular epithelium of the Drosophila ovary is an excellent model system to study the basal deposition of BM membrane proteins, as it produces and secretes all major components of the BM. Confocal and super-resolution imaging combined with image processing in fixed tissues allows for the identification and characterization of cellular factors specifically involved in the intracellular trafficking and deposition of BM proteins. This article presents a detailed protocol for staining and imaging BM-containing vesicles and deposited BM using endogenously tagged proteins in the follicular epithelium of the Drosophila ovary. This protocol can be applied to address both qualitative and quantitative questions and it was developed to accommodate high-throughput screening, allowing for the rapid and efficient identification of factors involved in the polarized intracellular trafficking and secretion of vesicles during epithelial tissue development.

Subepithelial Serotonin Reduces Small Intestinal Epithelial Cell Tightness via Reduction of Occluding Expression.

The precise pathogenesis of irritable bowel syndrome (IBS) remains unresolved; however, recent studies have reported that patients with diarrhea-predominant IBS exhibit an increased small intestinal permeability and increased number of enterochromaffin cells containing high 5-hydroxytryptamine (5HT; serotonin) levels. In this study, we investigated whether 5HT has the potential to modulate small intestinal epithelial cell permeability, focusing on tight junction-associated proteins. The differentiated Caco-2 cell monolayer on porous filters (Millicell) was used. Then, 5HT was added to the lower Millicell compartment for 7 days. Intestinal epithelial cell permeability was assessed by measuring the flux of paracellular permeability markers. We further assessed the expression of occludin in the 5HT-stimulated Caco-2 monolayer. We found that 5HT did not affect the viability of Caco-2 cells at concentrations up to 100 µM during the experimental period. Administration of 5HT to the basal side of Caco-2 cells increased the flux of 3H-labeled mannitol (182 Da) but did not increase that of FITC-dextran (4000 Da). Among the tight junction proteins, the expression of occludin was specifically decreased by stimulation with 5HT at a concentration of 100 µM. In conclusion, excessive 5HT in the basal side increased the permeability of intestinal epithelial cells via reduction of occludin expression.

Wnt/β-Catenin Signaling Stabilizes Hemidesmosomes in Keratinocytes

Hemidesmosomes (HDs) are adhesion complexes that promote epithelial-stromal attachment in stratified and complex epithelia, including the epidermis. In various biological processes, such as differentiation and migration of epidermal keratinocytes during wound healing or carcinoma invasion, quick assembly and disassembly of HDs are prerequisites. In this study, we show that inhibition of Wnt/β-catenin signaling disturbs HD organization in keratinocytes. Screening with inhibitors identified the depletion of HD components and HD-like structures through Wnt inhibition, but keratinocyte differentiation was not affected. Wnt inhibition significantly diminished plectin and type XVII collagen expression in the basal side of Wnt-inhibited cells and the dermo-epidermal junction of the Wnt-inactive murine basal epidermis. Similar to Wnt inhibition, PLEC-knockout cells or cells with plectin-type XVII collagen binding defects showed type XVII collagen reduction in the basal side of the cells, implying the possible involvement of Wnt/β-catenin signaling in HD assembly. Atypical protein kinase C inhibition ameliorated the phenotypes of Wnt-inhibited cells. These findings show that Wnt/β-catenin signaling regulates the localization of HD components in keratinocytes and that the atypical protein kinase C pathway is involved in Wnt inhibition‒induced HD disarrangement. Our study suggests that the Wnt signaling pathway could be a potential therapeutic target for treating HD-defective diseases, such as epidermolysis bullosa.

A new hypothetical model for pancreatic development based on change in the cell division orientation.

Although lifelong renewal and additional compensatory growth in response to demand are undeniable facts, so far, no specific stem cells have been found for pancreatic cells. According to the consensus model, the development of pancreas results from the hierarchical differentiation of pluripotent stem cells towards the appearance of the first endocrine and exocrine cells at approximately 7.5 to 8th gestation week (GW) of human embryo. However, the primitive endocrine cells arising from the embryonic phase of development do not appear to be mature or fully functional. Asymmetric localization of cellular components, such as Numb, partition protein complexes (PAR), planar cell polarity components, and certain mRNAs on the apical and basal sides of epithelial cells, causes cellular polarization. According to our model, the equal distribution of cellular components during symmetric cell division yields similar daughter cells that are associated with duct expansion. In contrast, asymmetric cell division is associated with uneven distribution of cellular components among daughter cells, resulting in different fates. Asymmetric cell division leads to duct branching and the development of acinar and stellate cells by a daughter cell, as well as the development of islet progenitor cells through partial epithelial-to-mesenchymal transition (EMT) and delamination of another daughter cell. Recently, we have developed an efficient method to obtain insulin-secreting cells from the transdifferentiation of hESC-derived ductal cells inducing a partial EMT by treatment with Wnt3A and activin A in a hypoxic environment. Similar models can be offered for other tissues and organs such as mammary glands, lungs, prostate, liver, etc. This model may open a new horizon in the field of regenerative medicine and be useful in explaining the cause of certain abnormalities, such as the occurrence of certain cysts and tumors.

Increased lateral tension is sufficient for epithelial folding in Drosophila.

The folding of epithelial sheets is important for tissues, organs and embryos to attain their proper shapes. Epithelial folding requires subcellular modulations of mechanical forces in cells. Fold formation has mainly been attributed to mechanical force generation at apical cell sides, but several studies indicate a role of mechanical tension at lateral cell sides in this process. However, whether lateral tension increase is sufficient to drive epithelial folding remains unclear. Here, we have used optogenetics to locally increase mechanical force generation at apical, lateral or basal sides of epithelial Drosophila wing disc cells, an important model for studying morphogenesis. We show that optogenetic recruitment of RhoGEF2 to apical, lateral or basal cell sides leads to local accumulation of F-actin and increase in mechanical tension. Increased lateral tension, but not increased apical or basal tension, results in sizeable fold formation. Our results stress the diversification of folding mechanisms between different tissues and highlight the importance of lateral tension increase for epithelial folding.

Tissue distribution and subcellular localization of the family of Kidney Ankyrin Repeat Domain (KANK) proteins

Kidney Ankyrin Repeat-containing Proteins (KANKs) comprise a family of four evolutionary conserved proteins (KANK1 to 4) that localize to the belt of mature focal adhesions (FAs) where they regulate integrin-mediated adhesion, actomyosin contractility, and link FAs to the cortical microtubule stabilization complex (CMSC). The human KANK proteins were first identified in kidney and have been associated with kidney cancer and nephrotic syndrome. Here, we report the distributions and subcellular localizations of the four Kank mRNAs and proteins in mouse tissues. We found that the KANK family members display distinct and rarely overlapping expression patterns. Whereas KANK1 is expressed at the basal side of epithelial cells of all tissues tested, KANK2 expression is mainly observed at the plasma membrane and/or cytoplasm of mesenchymal cells and KANK3 exclusively in vascular and lymphatic endothelial cells. KANK4 shows the least widespread expression pattern and when present, overlaps with KANK2 in contractile cells, such as smooth muscle cells and pericytes. Our findings show that KANKs are widely expressed in a cell type-specific manner, which suggests that they have cell- and tissue-specific functions.

A novel DENND1B-localized structure found at the basal side of adherent cells.

Rab35 is a small G protein involved in various cellular events including clathrin-dependent endocytosis, phagocytosis, and autophagy. DENND1B, a DENN family member, acts as a guanine nucleotide exchange factor (GEF) for Rab35 to convert it to the GTP-bound active form from the GDP-bound inactive form. DENND1B contains the DENN domain which harbors GEF activity for Rab35 in the N-terminus, while the clathrin binding motif and adaptor protein-2-interaction motif are at the C-terminus. In this study, we investigated the intracellular localization of DENN1B in various cell types and found novel DENND1B-localized gathered line structures in BS-C-1 cells and in some other cell types. The localization of DENND1B to gathered line structures was dependent on a specific region located in the C-terminus of DENND1B protein. DENND1B-localized gathered lines were partially associated with microtubules but not with F-actin; instead, F-actin bundles surrounded the assembly of gathered lines. We also show that the gathered line structures appeared at the bottom of spreading lamellipodia and disappeared at the retracting site during cell motility in EGF-stimulated BS-C-1 cells. These results shed light on a new role for DENND1B in the regulation of cell migration.

Variations in Plasma Membrane Topography Can Explain Heterogenous Diffusion Coefficients Obtained by Fluorescence Correlation Spectroscopy.

Fluorescence correlation spectroscopy (FCS) is frequently used to study diffusion in cell membranes, primarily the plasma membrane. The diffusion coefficients reported in the plasma membrane of the same cell type and even within single cells typically display a large spread. We have investigated whether this spread can be explained by variations in membrane topography throughout the cell surface, that changes the amount of membrane in the FCS focal volume at different locations. Using FCS, we found that diffusion of the membrane dye DiI in the apical plasma membrane was consistently faster above the nucleus than above the cytoplasm. Using live cell scanning ion conductance microscopy (SICM) to obtain a topography map of the cell surface, we demonstrate that cell surface roughness is unevenly distributed with the plasma membrane above the nucleus being the smoothest, suggesting that the difference in diffusion observed in FCS is related to membrane topography. FCS modeled on simulated diffusion in cell surfaces obtained by SICM was consistent with the FCS data from live cells and demonstrated that topography variations can cause the appearance of anomalous diffusion in FCS measurements. Furthermore, we found that variations in the amount of the membrane marker DiD, a proxy for the membrane, but not the transmembrane protein TCRζ or the lipid-anchored protein Lck, in the FCS focal volume were related to variations in diffusion times at different positions in the plasma membrane. This relationship was seen at different positions both at the apical cell and basal cell sides. We conclude that it is crucial to consider variations in topography in the interpretation of FCS results from membranes.

The kinesin motor Klp98A mediates apical to basal Wg transport.

ABSTRACTDevelopment and tissue homeostasis rely on the tight regulation of morphogen secretion. In the Drosophila wing imaginal disc epithelium, Wg secretion for long-range signal transduction occurs after apical Wg entry into the endosomal system, followed by secretory endosomal transport. Although Wg release appears to occur from the apical and basal cell sides, its exact post-endocytic fate and the functional relevance of polarized endosomal Wg trafficking are poorly understood. Here, we identify the kinesin-3 family member Klp98A as the master regulator of intracellular Wg transport after apical endocytosis. In the absence of Klp98A, functional mature endosomes accumulate in the apical cytosol, and endosome transport to the basal cytosol is perturbed. Despite the resulting Wg mislocalization, Wg signal transduction occurs normally. We conclude that transcytosis-independent routes for Wg trafficking exist and demonstrate that Wg can be recycled apically via Rab4-recycling endosomes in the absence of Klp98A.

A Cdc42-mediated supracellular network drives polarized forces and Drosophila egg chamber extension

Actomyosin supracellular networks emerge during development and tissue repair. These cytoskeletal structures are able to generate large scale forces that can extensively remodel epithelia driving tissue buckling, closure and extension. How supracellular networks emerge, are controlled and mechanically work still remain elusive. During Drosophila oogenesis, the egg chamber elongates along the anterior-posterior axis. Here we show that a dorsal-ventral polarized supracellular F-actin network, running around the egg chamber on the basal side of follicle cells, emerges from polarized intercellular filopodia that radiate from basal stress fibers and extend penetrating neighboring cell cortexes. Filopodia can be mechanosensitive and function as cell-cell anchoring sites. The small GTPase Cdc42 governs the formation and distribution of intercellular filopodia and stress fibers in follicle cells. Finally, our study shows that a Cdc42-dependent supracellular cytoskeletal network provides a scaffold integrating local oscillatory actomyosin contractions at the tissue scale to drive global polarized forces and tissue elongation.

Cell-Substrate Patterns Driven by Curvature-Sensitive Actin Polymerization: Waves and Podosomes.

Cells adhered to an external solid substrate are observed to exhibit rich dynamics of actin structures on the basal membrane, which are distinct from those observed on the dorsal (free) membrane. Here we explore the dynamics of curved membrane proteins, or protein complexes, that recruit actin polymerization when the membrane is confined by the solid substrate. Such curved proteins can induce the spontaneous formation of membrane protrusions on the dorsal side of cells. However, on the basal side of the cells, such protrusions can only extend as far as the solid substrate and this constraint can convert such protrusions into propagating wave-like structures. We also demonstrate that adhesion molecules can stabilize localized protrusions that resemble some features of podosomes. This coupling of curvature and actin forces may underlie the differences in the observed actin-membrane dynamics between the basal and dorsal sides of adhered cells.

Integrin Binding Dynamics Modulate Ligand-Specific Mechanosensing in Mammary Gland Fibroblasts.

SummaryThe link between integrin activity regulation and cellular mechanosensing of tissue rigidity, especially on different extracellular matrix ligands, remains poorly understood. Here, we find that primary mouse mammary gland stromal fibroblasts (MSFs) are able to spread efficiently, generate high forces, and display nuclear YAP on soft collagen-coated substrates, resembling the soft mammary gland tissue. We describe that loss of the integrin inhibitor, SHARPIN, impedes MSF spreading specifically on soft type I collagen but not on fibronectin. Through quantitative experiments and computational modeling, we find that SHARPIN-deficient MSFs display faster force-induced unbinding of adhesions from collagen-coated beads. Faster unbinding, in turn, impairs force transmission in these cells, particularly, at the stiffness optimum observed for wild-type cells. Mechanistically, we link the impaired mechanotransduction of SHARPIN-deficient cells on collagen to reduced levels of collagen-binding integrin α11β1. Thus integrin activity regulation and α11β1 play a role in collagen-specific mechanosensing in MSFs.

Distribution of tight junctions in the primate cochlear lateral wall

The blood-labyrinth barrier (BLB) is a major structure that separates the inner ear from the systemic blood circulation. Many drugs cannot access the inner ear because of their inability to cross the BLB. In the cochlea, the BLB is mainly distributed in the lateral wall. However, the ultrastructure of the cochlear lateral wall, including the distribution of tight junctions (TJs), which are its main component, has not been thoroughly examined in primates. This study investigated the distribution of TJs in the cochlear lateral wall of the common marmoset by performing immunohistochemistry for TJ markers and transmission electron microscopy. As previously reported in rodents, TJs were distributed at the lumenal side of stria marginal cells and basal cells. In outer sulcus cells, which are more developed in primates than in rodents, TJs were distributed at the side with the endolymph but not at the side with the spiral ligament, where many capillaries were located. These findings indicate that drugs and small compounds circulating systemically in the blood can easily access outer sulcus cells, but have a limited ability to enter endolymph. No structural differences were detected between species, indicating that the in vivo distribution of drugs in cochlear lateral wall cells, including outer sulcus cells, in primates can be predicted by performing rodent experiments.

“Chip-on-a-Transwell” Devices for User-Friendly Control of the Microenvironment of Cultured Cells

We present microfabricated poly(dimethylsiloxane) templates assembled atop Transwell membranes ("Chip-on-a-Transwell") for focal delivery of soluble reagents to cells. This modified Transwell operates based on area-selective diffusion of soluble species to the basal side of cells. The device was capable of addressing small groups of cells, with a maximum spatial resolution of 20 μm, while sustaining focal signal for up to 48 hours. Furthermore, we developed a fluorosilane-based cell micropatterning method that complements the Transwell system, creating well-defined arrays of muscle cell cultures and enabling automated microscopy and image analysis protocols. The micropatterning method is based on plasma-etching hydrophilic cell-adhesive tracks against a hydrophobic fluorosilane cell-repellent background. We applied the platform as a model of the neuromuscular junction (NMJ), in which we effectively substituted motor neurons with apertures delivering agrin stimuli to subcellular regions of micropatterned myotubes. With the Transwell system, we demonstrated that focal agrin application to subcellular sections of myotubes induced localized acetylcholine receptor microclustering, mimicking the in vivo NMJ. We also demonstrated spatially selective transfection of a monolayer of cells using Lipofectamine carrying the plasmid DNA for pMax GFP, which is ubiquitously expressed in transfected cells. In summary, we present a user-friendly cell culture tool that provides spatiotemporal control of the fluidic and substrate microenvironments with potential applications to developmental biology and tissue engineering.

Molecular profiling of the basement membrane of pluripotent epiblast cells in post-implantation stage mouse embryos

IntroductionThe basement membrane (BM) is a sheet-like extracellular matrix (ECM) lining the basal side of epithelial and endothelial cells. The molecular composition of the BM diversifies as embryonic development proceeds, providing optimized microenvironments for individual cell types. In post-implantation stage embryos, the embryonic BMs are essential for differentiation of the epiblast, a layer of multipotent embryonic stem cells, and subsequent embryogenesis. To better understand the role of BMs and cell–BM interactions in early embryogenesis, it is imperative to accumulate information on the molecular entities of the embryonic BMs.MethodsWe analyzed the expressions and localizations of 20 major BM proteins (11 laminin subunits, 6 type IV collagen subunits, nidogen-1 and -2, and perlecan) and other ECM-related proteins such as fibronectin and integrins in post-implantation stage embryos by immunohistochemistry.ResultsWe found that a set of BM proteins, laminin α5, β1, and γ1 (comprising laminin-511), type IV collagen α1 and α2 (yielding type IV collagen α12α2 [IV]), nidogen-1 and -2, and perlecan, were consistently present in the epiblast/ectoderm BMs throughout the early post-implantation stages. In contrast, laminin α1 was detected in the epiblast BM at E5.5 but decreased in later stages, suggesting that laminin-511 is a major laminin isoform in the early embryonic BM. In addition, fibronectin, a mesenchymal ECM protein, was enriched in the endoderm BM, indicating that the BM compositions differ between the ectoderm and the endoderm. Consistent with these observations, integrin α5, a high-affinity receptor for fibronectin, was localized in the endoderm, while integrin α6, a receptor for laminin-511, was localized in the ectoderm.ConclusionsThe embryonic BMs underlying the epiblast/ectoderm contain a common toolkit comprising laminin-511, type IV collagen (α12α2 [IV]), nidogen-1 and -2, and perlecan, providing a physiological basis for the utility of laminin-511 as a culture substrate for pluripotent stem cells. The distinctive association of laminin-511 and fibronectin with endodermal and ectodermal cells, together with the differential expression of integrin α5 and α6 in these cells, suggests that the ectodermal and endodermal cells rely on their integrin-dependent interactions with laminin-511 and fibronectin, respectively, to ensure their fate specification in embryonic development.

Digestibility in the gastrointestinal tract and migration to blood of β′-component (Onk k 5), a major salmon roe IgE-binding protein

The major allergen of chum salmon (Oncorhynchus keta) roe is the β′-component (Onc k 5, β′-c), which is a yolk protein and a fragment of vitellogenin. When yolk content containing β′-c was orally administered to mice, β′-c passed through the gastrointestinal tract and was excreted in feces without marked degradation. The direct administration of β′-c to ligated jejunal and ileal loops showed that β′-c was absorbed through the small intestine and transferred into the blood. Immunohistochemical staining showed that orally administered β′-c was distributed from the apical side to the basal side of intestinal epithelial cells, suggesting that endocytosis may be involved in the intestinal absorption of β′-c. In conclusion, β′-c is absorbed along a large portion of the small intestine and circulates in the blood stream without significant digestion. The resistance of β′-c to gastrointestinal digestion seems to contribute to its strong allergenicity.