Restricted distribution of laminin α1 chain in normal adult mouse tissues

The distribution of laminin α1 chain in adult mouse tissue was determined by immunofluorescence using monoclonal antibody 200, reacting with the globular carboxyterminus E3 fragment of α1 chain. Strong reactivity was noted only in a few tissues. Reactivity was restricted to epithelial basement membranes. Expression was noted in several epithelial basement membranes of the urinary tract, and male and female reproductive organs. In addition, expression was seen in some parts of the nervous system. Expression was seen in pia mater which surrounds the brain, and in the extracellular matrices covering the vitreous chamber and the lens of the eye. Staining was seen in the adrenal gland cortex, with strongest staining in the zona glomerulosa. Staining was negative in all other studied epithelial basement membranes, such as the lung (trachea or lung epithelium), epidermis, and all parts of the gastrointestinal tract (liver, gut) except for weak staining in the ventricle and Brunner’s glands. No expression was seen in basement membranes of fat, Schwann, or endothelial cells in any studied parts of the body. Both small- and large-size vessel walls were negative both in endothelial basement membranes and blood vessel walls, with the exception of some larger brain blood vessels in locations where epithelial cells have invaginated. Neither smooth muscle, myocardium or striated muscle expressed α1 chain. We conclude that α1-containing heterotrimers including laminin-1 (α1β1γ1) have a very restricted tissue distribution.     

Localization of laminin alpha4-chain in developing and adult human tissues.

Recent studies suggest important functions for laminin-8 (Ln-8; alpha4beta1gamma1) in vascular and blood cell biology, but its distribution in human tissues has remained elusive. We have raised a monoclonal antibody (MAb) FC10, and by enzyme-linked immunoassay (EIA) and Western blotting techniques we show that it recognizes the human Ln alpha4-chain. Immunoreactivity for the Ln alpha4-chain was localized in tissues of mesodermal origin, such as basement membranes (BMs) of endothelia, adipocytes, and skeletal, smooth, and cardiac muscle cells. In addition, the Ln alpha4-chain was found in regions of some epithelial BMs, including epidermis, salivary glands, pancreas, esophageal and gastric glands, intestinal crypts, and some renal medullary tubules. Developmental differences in the distribution of Ln alpha4-chain were detected in skeletal muscle, walls of vessels, and intestinal crypts. Ln alpha4- and Ln alpha2-chains co-localized in BMs of fetal skeletal muscle cells and in some epithelial BMs, e.g., in gastric glands and acini of pancreas. Cultured human pulmonary artery endothelial (HPAE) cells produced Ln alpha4-chain as M(r) 180,000 and 200,000 doublet and rapidly deposited it to the growth substratum. In cell-free extracellular matrices of human kidney and lung, Ln alpha4-chain was found as M(r) 180,000 protein.     

Structural and functional analysis of the recombinant G domain of the laminin alpha4 chain and its proteolytic processing in tissues

The C-terminal G domains of laminin alpha chains have been implicated in various cellular and other interactions. The G domain of the alpha4 chain was now produced in transfected mammalian cells as two tandem arrays of LG modules, alpha4LG1-3 and alpha4LG4-5. The recombinant fragments were shown to fold into globular structures and could be distinguished by specific antibodies. Both fragments were able to bind to heparin, sulfatides, and the microfibrillar fibulin-1 and fibulin-2. They were, however, poor substrates for cell adhesion and had only a low affinity for the alpha-dystroglycan receptor when compared with the G domains of the laminin alpha1 and alpha2 chains. Yet antibodies to alpha4LG1-3 but not to alpha4LG4-5 clearly inhibited alpha(6)beta(1) integrin-mediated cell adhesion to laminin-8, indicating the participation of alpha4LG1-3 in a cell-adhesive structure of higher complexity. Proteolytic processing within a link region between the alpha4LG3 and alpha4LG4 modules was shown to occur during recombinant production and in endothelial and Schwann cell culture. Cleavage could be attributed to three different peptide bonds and is accompanied by the release of the alpha4LG4-5 segment. Immunohistology demonstrated abundant staining of alpha4LG1-3 in vessel walls, adipose, and perineural tissue. No significant staining was found for alpha4LG4-5, indicating their loss from tissues. Immunogold staining demonstrated an association of the alpha4 chain primarily with microfibrillar regions rather than with basement membranes, while laminin alpha2 chains appear primarily associated with various basement membranes.     

Cyclic modulation of integrin expression in bovine endometrium.

Integrins are heterodimeric glycoproteins involved in cell-cell and cell-extracellular matrix adhesion. In this study, the spatial and temporal distribution of selected integrins and extracellular matrix proteins was determined in bovine endometrium from cycling and ovariectomized animals using indirect immunohistochemistry. The expression of integrins alpha(6) and alpha(v)beta(3) was estrous cycle-dependent. Strong immunostaining for alpha(v)beta(3) occurred in the basement membrane region of intercaruncular luminal epithelium except on Day 16 (P<0.05). Staining of subepithelial stromal cells declined in diestrous samples (P<0.05). In all samples, there was reduced alpha(v)beta(3) reactivity in the caruncles. Staining for alpha(6) decreased in the epithelial basement membrane at proestrus through estrus (Days 18-0). Expression of integrin subunits alpha(3) and alpha(4) was cycle-independent. Moderate staining for alpha(3) was detected on epithelium and alpha(4) was present on stromal cells. The distribution of beta(1) suggested dimerization with alpha(3), alpha(4), and alpha(6). Laminin was detected in the epithelial and vasculature basement membranes. Collagen IV was present in the glandular epithelium basement membrane and subepithelial stromal cells, whereas fibronectin was found only in the stroma. Estrous cycle-dependent distribution and expression of alpha(v)beta(3) and alpha(6) suggest their regulation by ovarian steroids, growth factors, and prostaglandins.     

Structural analysis and mutation detection strategy for the human LAMC3 gene

Laminins are heterotrimeric extracellular matrix molecules, present in a wide range of basement membranes within human tissues. They consist of a combination of different alpha, beta, and gamma subunits. Three different gamma subunits have been described to date. Two of them, the gamma1 and gamma2 chains are constituents of basement membrane related laminins, while the gamma3 chain was detected in skin, heart, lung, reproductive tract, brain, and in the retina. Unlike other laminins, the expression of the gamma3 chain was localized to peripheral nerves and to the apical surface of ciliated epithelial cells and in the retina. To further investigate the function and the possible pathogenic role of laminin gamma3 in human disease, we elucidated the structure of the corresponding LAMC3 gene which encodes this polypeptide. Here we report the genomic organization of the LAMC3 gene and a mutation detection strategy for use in genetic studies.     

The relation between connective tissue cells and intercellular substances, including basement membranes.

Basement membranes are distributed widely in the body forming an extracellular matrix for epithelial and endothelial cells. The collagenous and glycoprotein constituents of basement membranes are synthesized by these two cell types. Disturbance of the interactions between basement membranes and their associated epithelial and endothelial cells can lead to the pathological changes seen in diseases involving basement membranes. These changes are illustrated here by reference to glomerulonephritis induced by the deposition of immune complexes in the glomerulus of the kidney, and chronic inflammatory changes occurring in the lung after inhalation of asbestos. In these diseases basement membrane changes can occur in several ways. Hydrolytic enzymes released from inflammatory cells degrade basement membranes while other constituents by epithelial and endothelial cells. Alternatively the physical separation of epithelial and endothelial cells from their basement membrances by space-occupying substances such as immune complexes can interfere with feedback mechanisms leading to synthesis of basement membrane constituents and cell proliferation. Studies of these pathological changes at a cellular level should shed new light on the ways in which cells interact with their pericellular environment.     

Ontogenesis of glomerular basement membrane: structural and functional properties

Protein A-gold immunocytochemistry was applied in combination with morphometrical approaches to reveal the alpha 1(IV), alpha 2(IV), and alpha 3(IV) chains of type IV collagen as well as entactin on renal basement membranes, particularly on the glomerular one, during maturation. The results have indicated that a heterogeneity between renal basement membranes appears during the maturation process. In the glomerulus at the capillary loop stage, both the epithelial and endothelial cell basement membranes were labeled for the alpha 1(IV) and alpha 2(IV) chains of type IV collagen and entactin. After fusion, both proteins were present on the entire thickness of the typical glomerular basement membrane. At later stages, the labeling for alpha 1(IV) and alpha 2(IV) chains of type IV collagen decreased and drifted towards the endothelial side, whereas the labeling for the alpha 3(IV) chain increased and remained centrally located. Entactin remained on the entire thickness of the basement membrane during maturation and in adult stage. The distribution of endogenous serum albumin in the glomerular wall was studied during maturation, as a reference for the functional properties of the glomerular basement membrane. This distribution, dispersed through the entire thickness of the basement membrane at early stages, shifted towards the endothelial side of the lamina densa with maturation, demonstrating a progressive acquisition of the permselectivity. These results demonstrate that modifications in the content and organization of the different constituents of basement membranes occur with maturation and are required for the establishment of the filtration properties of the glomerular basement membrane.     

Heterogeneity of basement membranes of the human genitourinary tract revealed by sequential immunofluorescence staining with monoclonal antibodies to laminin

We used monoclonal antibodies specific for human laminin to analyze immunohistochemically the heterogeneity of the basement membranes in various parts of the genitourinary tract. By indirect immunofluorescence microscopy we show that antibody 3H11 reacts with all epithelial basement membranes in the kidneys, testes, epididymis, prostate, uterus, oviduct, and ovary, as well as the smooth muscle cells, blood vessels, and nerves. Antibody 4E10 reacted with most epithelial basement membranes in these organs but was unreactive with the basement membranes of peripheral glomerular capillary loops and the basement membranes of the oviductal mucosa, seminiferous tubules, straight tubules, and rete testis. Hilar seminiferous tubules were reactive with 4E10. In contrast to 3H11, which reacted with all vascular, subendothelial, and muscular basement membranes, 4E10 reacted only with the subendothelial basement membrane of capillaries and veins. The difference in the distribution of epitopes could be demonstrated in tissue sections sequentially reacted with two monoclonal antibodies, but only if the antibody of restricted reactivity (4E10) was used first. These data show that the heterogeneous expression of distinct epitopes of laminin in basement membranes can be demonstrated in the same tissue section by sequential staining. This heterogeneity of basement membranes most likely reflects conformational differences in the expression of epitopes on the laminin molecule in various anatomic structures.     

Laminin alpha1-chain shows a restricted distribution in epithelial basement membranes of fetal and adult human tissues

Two novel monoclonal antibodies were raised and used to study the expression of laminin (Ln) alpha1-chain in developing and adult human tissues. In both fetal and adult kidney, a distinct immunoreactivity was seen in basement membranes (BM) of most proximal tubules but not in the distal tubular or glomerular BM or in the basal laminae of blood vessels. Immunoprecipitation of metabolically labeled cultured human renal proximal tubular cells showed an abundant production and deposition of Ln alpha1-chain to the extracellular matrix, suggestive of an epithelial origin of kidney Ln-1. Quantitative cell adhesion experiments with JAR choriocarcinoma cells showed that purified human Ln-1 is a good substrate for cell adhesion that it is differently recognized by integrin receptors when compared to mouse Ln-1. In fetal and adult testes immunoreactivity was solely confined to BM of the seminiferous epithelium. In the airways BM-confined reaction was only seen in fetal budding bronchial tubules (16-19 weeks) at the pseudoglandular stage of development. In the skin a distinct immunoreactivity was confined to BM of developing hair buds but not in epithelial BMs of adult epidermis or of epidermal appendages. In other adult tissues, immunoreactivity was found in BMs of thyroid, salivary, and mammary glands as well as in BMs of endometrium and endocervix, but not of ectocervix or vagina. No immunoreactivity was found in BMs of most of the digestive tract, including the liver and pancreas, except for BMs of esophageal submucosal glands and duodenal Brunner's glands. In fetal specimens, BMs of the bottoms of the intestinal and gastric glands were positive. Basal laminae of blood vessels were generally negative for Ln alpha1 chain with the exception of specimens of both fetal and adult central nervous system in which immunoreactivity for Ln alpha1 chain was prominently confined to capillary walls. The results suggest that outside the central nervous system, Ln alpha1 chain shows a restricted and developmentally regulated expression in BMs of distinct epithelial tissues.     

Defective glomerulogenesis in the absence of laminin alpha5 demonstrates a developmental role for the kidney glomerular basement membrane

Laminins are major components of all basement membranes. They are a diverse group of alpha/beta/gamma heterotrimers formed from five alpha, three beta, and three gamma chains. Laminin alpha5 is a widely expressed chain found in many embryonic and adult basement membranes. During embryogenesis, alpha5 has a role in disparate developmental processes, including neural tube closure, digit septation, and placentation. Here, we analyzed kidney development in Lama5 mutant embryos and found a striking defect in glomerulogenesis associated with an abnormal glomerular basement membrane (GBM). This correlates with failure of the developmental switch in laminin alpha chain deposition in which alpha5 replaces alpha1 in the GBM at the capillary loop stage of glomerulogenesis. In the absence of a normal GBM, glomerular epithelial cells were in disarray, and endothelial and mesangial cells were extruded from within the constricting glomerulus, leading to a complete absence of vascularized glomeruli. In addition, a minority of Lama5 mutant mice lacked one or both kidneys, indicating that laminin alpha5 is also important in earlier kidney development. Our results demonstrate a dual role for laminin alpha5 in kidney development, illustrate a novel defect in glomerulogenesis, and indicate a heretofore unappreciated developmental role for the GBM in influencing the behavior of epithelial and endothelial cells.     

Laminin alpha5 chain is required for intestinal smooth muscle development

Laminins (comprised of alpha, beta, and gamma chains) are heterotrimeric glycoproteins integral to all basement membranes. The function of the laminin alpha5 chain in the developing intestine was defined by analysing laminin alpha5(-/-) mutants and by grafting experiments. We show that laminin alpha5 plays a major role in smooth muscle organisation and differentiation, as excessive folding of intestinal loops and delay in the expression of specific markers are observed in laminin alpha5(-/-) mice. In the subepithelial basement membrane, loss of alpha5 expression was paralleled by ectopic or accelerated deposition of laminin alpha2 and alpha4 chains; this may explain why no obvious defects were observed in the villous form and enterocytic differentiation. This compensation process is attributable to mesenchyme-derived molecules as assessed by chick/mouse alpha5(-/-) grafted associations. Lack of the laminin alpha5 chain was accompanied by a decrease in epithelial alpha3beta1 integrin receptor expression adjacent to the epithelial basement membrane and of Lutheran blood group glycoprotein in the smooth muscle cells, indicating that these receptors are likely mediating interactions with laminin alpha5-containing molecules. Taken together, the data indicate that the laminin alpha5 chain is essential for normal development of the intestinal smooth muscle and point to possible mesenchyme-derived compensation to promote normal intestinal morphogenesis when laminin alpha5 is absent.     

Laminin isoforms and lung development: all isoforms are not equal

Laminins are a major component of basement membranes. Each laminin molecule is a heterotrimeric glycoprotein composed of one alpha, one beta, and one gamma chain. Fifteen laminin isoforms exist, assembled from various combinations of 5alpha, 3beta, and 3gamma chains. The embryonic lung has abundant laminin isoforms. Increasing evidence suggests that different laminin isoforms have unique functions in lung development. Studies of embryonic lung explants and organotypic co-cultures show that laminin alpha1 and laminin 111 are important for epithelial branching morphogenesis and that laminin alpha2 and laminin 211 have a role in smooth muscle cell differentiation. In vivo studies of laminin alpha5-deficient mice indicate that this laminin chain, found in laminins 511 and 521, is essential for normal lobar septation in early lung development and normal alveolization and distal epithelial cell differentiation and maturation in late lung development. However, not all of the laminin chains present in the developing lung appear to be necessary for normal lung development since laminin alpha4 null mice do not have obvious lung abnormalities and laminin gamma2 null mice have only minimal changes in lung development. The mechanisms responsible for the lung phenotypes in mice with laminin mutations are unknown, but it is clear that multiple laminin isoforms are crucial for lung development and that different laminin isoforms exhibit specific, non-overlapping functions.     

Differential expression of two basement membrane collagen genes, COL4A6 and COL4A5, demonstrated by immunofluorescence staining using peptide- specific monoclonal antibodies

Genes for the human alpha 5(IV) and alpha 6(IV) collagen chains have a unique arrangement in that they are colocalized on chromosome Xq22 in a head-to-head fashion and appear to share a common bidirectional promoter. In addition we reported a novel observation that the COL4A6 gene is transcribed from two alternative promoters in a tissue-specific manner (Sugimoto, M., T. Oohashi, and Y. Ninomiya. 1994. Proc. Natl. Acad. Sci. USA. 91:11679-11683). To know whether the translation products of both genes are colocalized in various tissues, we raised alpha 5(IV) and alpha 6(IV) chain-specific rat monoclonal antibodies against synthetic peptides reflecting sequences near the carboxy terminus of each noncollagenous (NC)1 domain. By Western blotting alpha 6(IV) chain-specific antibody recognized 27-kD monomers and associated dimers of the human type IV collagen NC1 domain, which is the first demonstration of the presence in tissues of the alpha 6(IV) polypeptide as predicted from its cDNA sequence. Immunofluorescence studies using anti-alpha 6(IV) antibody demonstrated that in human adult kidney the alpha 6(IV) chain was never detected in the glomerular basement membrane, whereas the basement membranes of the Bowman's capsules and distal tubules were positive. The staining pattern of the glomerular basement membrane was quite different from that obtained with the anti- alpha 5(IV) peptide antibody. The alpha 5(IV) and alpha 6(IV) chains were colocalized in the basement membrane in the skin, smooth muscle cells, and adipocytes; however, little if any reaction was seen in basement membranes of cardiac muscles and hepatic sinusoidal endothelial cells. Thus, both genes are expressed in a tissue-specific manner, perhaps due to the unique function of the bidirectional promoter for both genes, which is presumably different from that for COL4A1 and COL4A2.     

Deletion of the Laminin α4 Chain Leads to Impaired Microvessel Maturation

The laminin alpha4 chain, a component of laminin-8 and -9, is expressed in basement membranes, such as those beneath endothelia, the perineurium of peripheral nerves, and around developing muscle fibers. Laminin alpha4-null mice presented with hemorrhages during the embryonic and neonatal period and had extensive bleeding and deterioration of microvessel growth in experimental angiogenesis, as well as mild locomotion defects. Histological examination of newborn mice revealed delayed deposition of type IV collagen and nidogen into capillary basement membranes, and electron microscopy showed discontinuities in the lamina densa. The results demonstrate a central role for the laminin alpha4 chain in microvessel growth and, in the absence of other laminin alpha chains, in the composition of endothelial basement membranes.     

Cellular origin of fibronectin in interspecies hybrid kidneys

The cellular origin of fibronectin in the kidney was studied in three experimental models. Immunohistochemical techniques that use cross-reacting or species-specific antibodies against mouse or chicken fibronectin were employed. In the first model studied, initially avascular mouse kidneys cultured on avian chorioallantoic membranes differentiate into epithelial kidney tubules and become vascularized by chorioallantoic vessels. Subsequently, hybrid glomeruli composed of mouse podocytes and avian endothelial-mesangial cells form. In immunohistochemical studies, cross-reacting antibodies to fibronectin stained vascular walls, tubular basement membranes, interstitium, and glomeruli of mouse kidney grafts. The species-specific antibodies reacting only with mouse fibronectin stained interstitial areas and tubular basement membranes, but showed no reaction with hybrid glomeruli and avian vascular walls. In contrast, species-specific antibodies against chicken fibronectin stained both the interstitial areas and the vascular walls as well as the endothelial-mesangial areas of the hybrid glomeruli, but did not stain the mouse-derived epithelial structures of the kidneys. In the second model, embryonic kidneys cultured under avascular conditions in vitro develop glomerular tufts, which are devoid of endothelial cells. These explants showed fluorescence staining for fibronectin only in tubular basement membranes and in interstitium. The avascular, purely epithelial glomerular bodies remained unstained. Finally, in outgrowths of separated embryonic glomeruli, the cross-reacting fibronectin antibodies revealed two populations of cells: one devoid of fibronectin and another expressing fibronectin in strong fibrillar and granular patterns. These results favor the idea that the main endogenous cellular sources for fibronectin in the embryonic kidney are the interstitial and vascular cells. All experiments presented here suggest that fibronectin is not synthesized by glomerular epithelial cells in vivo.     

Expression of lynx1 in developing lung and its modulation by prenatal nicotine exposure

The expression of nicotinic acetylcholine receptors (nAChR) in fetal lung suggests maternal smoking during pregnancy effects newborn lung structure and function by the direct interaction of nicotine with nAChR in the developing lung. The recent identification of the lynx1 nAChR modulator protein in nicotinic neurons in the brain suggests that lynx1 may be similarly expressed in the lung. To study this, cDNAs encoding lynx1 were cloned from rhesus monkey lung. The temporal expression of lynx1 was studied in pre- and postnatal monkey lungs by in situ hybridization, immunohistochemistry, and realtime polymerase chain reaction (PCR). Lynx1 mRNA signal and lynx1 immunohistochemical staining were localized predominantly in airway epithelial cells, submucous glands, and smooth muscle cells, in endothelial and smooth muscle cells in vessel walls, and in alveolar type II cells. The distribution of lynx1 was similar to that of 4, 2, and 4 nAChR expression as determined by immunohistochemistry. Immunohistochemical staining also co-localized choline acetyltransferase, the enzyme that synthesizes acetylcholine, with lynx1 expression. Lynx1 expression was first observed in 71-day fetal lungs and increased with age. Immunohistochemistry, Western analysis, and realtime PCR analysis showed increased lynx1 expression in lungs following prenatal nicotine exposure. Thus, lynx1 is co-expressed with nAChR in the lung. Alteration of lynx1 levels is a potential new mechanism by which nicotine affects lung development. This research was supported by National Institute of Health grants RR-00163 and HD/HL-37131.     

Extraocular muscle is spared upon complete laminin alpha2 chain deficiency: comparative expression of laminin and integrin isoforms.

Mutations in the gene encoding laminin (LM) alpha2 chain cause congenital muscular dystrophy. Here, we show that extraocular muscle (EOM) is spared upon complete LMalpha2 chain absence. The major LM chains in limb muscle basement membranes are alpha2, beta1, beta2 and gamma1 whereas alpha2, alpha4, beta1, beta2 and gamma1 chains are expressed in EOM. Expression of LMalpha4 chain mRNA is further increased in LMalpha2 chain deficient EOM. Mainly integrin alpha7X1 subunit, which binds to laminin-411, is expressed in EOM and in contrast to dystrophic limb muscle, sustained integrin alpha7B expression is seen in LMalpha2 chain deficient EOM. We propose that LMalpha4 chain, possibly by binding to integrin alpha7BX1beta1D, protects EOM in LMalpha2 chain deficient muscular dystrophy.     

Antibodies against domain E3 of laminin-1 and integrin alpha 6 subunit perturb branching epithelial morphogenesis of submandibular gland, but by different modes

Branching epithelial morphogenesis requires interactions between the surrounding mesenchyme and the epithelium, as well as interactions between basement membrane components and the epithelium. Embryonic submandibular gland was used to study the roles of two mesenchymal proteins, epimorphin and tenascin-C, as well as the epithelial protein laminin-1 and one of its integrin receptors on branching morphogenesis. Laminin-1 is a heterotrimer composed of an alpha 1 chain and two smaller chains (beta 1 and gamma 1). Immunofluorescence revealed a transient expression of laminin alpha 1 chain in the epithelial basement membrane during early stages of branching morphogenesis. Other laminin-1 chains and alpha 6, beta 1, and beta 4 integrin subunits seemed to be expressed constitutively. Expression of epimorphin, but not tenascin-C, was seen in the mesenchyme during early developmental stages, but a mAb against epimorphin did not perturb branching morphogenesis of this early epithelium. In contrast, inhibition of branching morphogenesis was seen with a mAb against the carboxy terminus of laminin alpha 1 chain, the E3 domain. An inhibition of branching was also seen with a mAb against the integrin alpha 6 subunit. The antibodies against laminin alpha 1 chain and integrin alpha 6 subunit perturbed development in distinct fashions. Whereas treatment with the anti-E3 resulted in discontinuities of the basement membrane at the tips of the branching epithelium, treatment with the mAb against alpha 6 integrin subunit seemed to leave the basement membrane intact. We suggest that the laminin E3 domain is involved in basement membrane formation, whereas alpha 6 beta 1 integrin binding to laminin-1 may elicit differentiation signals to the epithelial cells.     

Localization of human interleukin 13 receptor in non-haematopoietic cells

Although the functional roles of interleukin (IL-)13 in haematopoietic cells are well investigated, those in non-haematopoietic cells remain to be addressed. IL-13 exerts its actions by binding to the IL-13 receptor (IL-13R) on target cells, which is composed of IL-13Ralpha1 and the IL-4 receptor alpha chain (IL-4Ralpha). However, there has been no study of localization of IL-13R in each tissue. To address this question, we generated monoclonal anti-IL-13Ralpha1 antibody, and performed immunohistochemistry using this antibody and anti-IL-4Ralpha antibody. Distribution of these two components was the same in all examined tissues. Staining was positive in keratinocytes, hair follicles, and sebaceous and sweat glands in skin; in ciliated respiratory epithelial cells in nasal tissue; in heart muscle cells; in foveola cells, gastric glands, and the smooth muscle layer in stomach; and in hepatocytes in liver. However, staining was undetectable in brain and bone marrow. Fibroblasts and endothelial cells were stained in some tissues. These results provide clues to elucidate the known pathological roles of IL-13 in atopic dermatitis and allergic rhinitis, as well as its unknown physiological roles.