Lumican mRNA Research Articles

M1746 Identification of the CCK-C Spliced Receptor in Human Pancreatic Cancer Cells

negative cancer cells. Lumican mRNA and the protein were detected in all cancer cell lines. The phosphorylation of AKT, known as an important factor in cellular behaviors such as growth, survival, and drug resistance, was up-regulated in lumican knocked down cancer cells. Conclusion: Lumican in cancer cells might inhibit AKT signal pathway and play important roles in effectiveness of anti-cancer drugs in pancreatic cancer patients. It may be a novel candidate for anti-cancer drug via AKT signal pathway.

Rho-Mediated Regulation of TGF-β1– and FGF-2–Induced Activation of Corneal Stromal Keratocytes

To investigate the role of Rho GTPase signaling in FGF-2- and TGF-beta1-induced activation of corneal keratocytes. Keratocytes isolated from rabbit corneal stroma and plated in a serum-free medium were treated with FGF-2/heparin or TGF-beta1 in the presence or absence of Rho inhibitor (C3 exoenzyme) or ROCK (Rho kinase) inhibitor (Y27632). Specific phenotypic changes were analyzed by immunocytochemistry and Western blot analysis, and the relative abundance of specific mRNAs was estimated by quantitative RT-PCR. TGF-beta1-induced expression of alpha-SMA and transcription of alpha-SMA mRNA in activated keratocytes were reduced by Rho or ROCK inhibition during the activation. In nonactivated keratocytes, the expression of alpha3(IV) collagen was downregulated by Rho-inhibition. TGF-beta1- or FGF-2-induced downregulation of the expression of alpha3(IV) collagen and its mRNA was not significantly altered by Rho or ROCK inhibition. TGF-beta1- and FGF-2-induced decreases in cell-associated and secreted KS, and lumican mRNA levels were prevented by Rho or ROCK inhibition. However, FGF-2-induced decreases in keratocan mRNA levels were prevented by Rho inhibition but not by ROCK inhibition. Whereas Rho inhibition downregulated both TGF-beta1- and FGF-2-induced tenascin-C expression, ROCK inhibition was found to downregulate only TGF-beta1-induced expression. Rho signaling has a significant role in the activation of keratocytes. Rho, via ROCK-independent and/or -dependent pathways differentially regulates the TGF-beta1-induced expression of alpha-SMA and TGF-beta1- and FGF-2-induced de novo expression of tenascin-C and the downregulation of alpha3(IV) collagen and KSPGs, lumican and keratocan.

Gene expressions of small leucine-rich repeat proteoglycans and fibulin-5 are decreased in pelvic organ prolapse

Few studies are performed on the sustainability of the pelvic floor extracellular matrix important for preventing development of pelvic organ prolapse (POP). Collagens I and III, the elastin-associated proteins fibrillin-1 and fibulin-5 and the small leucine-rich repeat proteoglycans (SLRPs) decorin, lumican and fibromodulin are involved in giving the tissue its mechanical properties. Para-urethral biopsies were obtained from 15 women, 6 pre- and 9 post-menopausal, with POP. Real-time reverse transcription-polymerase chain reaction and immunohistochemistry for collagen I, collagen III, fibrillin-1, fibulin-5, decorin, lumican and fibromodulin were performed and compared with 14 controls, 8 pre- and 6 post-menopausal. Statistical comparisons controlled for age changes in gene expressions. A 16-fold decrease in decorin mRNA expression, P = 0.0001, and 8-fold in lumican mRNA expression, P = 0.001, were discovered in premenopausal POP compared with matched controls. In all women with POP, there were lower gene expressions of fibromodulin, P = 0.004, and fibulin-5, P = 0.001, compared with all controls. All proteins were detectable by immunohistochemistry, showing a weaker staining for decorin in premenopausal POP. For the first time, we show substantially decreased gene signal for production of SLRPs, regulators of collagen fiber assembly and impairment in elastic fiber assembly by down-regulation of fibulin-5 in POP.

Role of lumican in cancer cells and adjacent stromal tissues in human pancreatic cancer

Lumican is a member of a small leucine-rich proteoglycan family and its overexpression has been reported in carcinoid tumor, breast, colorectal, neuroendocrine cell, uterine cervical and pancreatic cancers. The expression of lumican in stromal tissues in breast cancer is associated with a high tumor grade, a low estrogen receptor expression level and young age. Lumican expression in the cytoplasm in advanced colorectal cancer is correlated with a poor prognosis. Lumican expression was previously reported in pancreatic cancer, but the role of lumican in pancreatic cancer is still not well understood. In this study, we aimed to clarify the role of lumican in pancreatic cancer. Reverse-transcription polymerase chain reaction and Western blot analyses revealed lumican mRNA and protein expression in six pancreatic ductal adenocarcinoma cell lines (i.e. PANC-1, MIA PaCa-2, KLM-1, Capan-1, PK-1 and PK-8). On the basis of its immunoreactivity, lumican was found to be localized in islet cells of normal pancreatic tissues, but not in exocrine cells. In pancreatic cancer tissues, lumican was predominantly localized in the cytoplasm of cancer cells in 30 out of 53 (56.6%) cancer patients, whereas lumican was detected in stromal tissues in 36 out of 53 (67.9%) cancer patients. Lumican expression in pancreatic cancer cells did not correlate with clinicopathological factors, whereas lumican expression in stromal tissues correlated with the female gender, advanced stage, retroperitoneal and duodenal invasion and residual tumor (p=0.030, 0.038, 0.049, 0.049 and 0.048, respectively). Patients with lumican-positive cancer cells tended to survive longer than those with lumican-negative cancer cells (p=0.286), but patients with lumican-positive stromal tissues had shorter survival than those with lumican-negative stromal tissues (p=0.062). These results suggest that lumican in stromal tissues plays an important role in the growth and invasion of pancreatic cancer.

Loss of Alpha3(IV) Collagen Expression Associated with Corneal Keratocyte Activation

To determine whether changes in the expression of type IV alpha1, alpha2, or alpha3 collagen isoforms are stringently associated with corneal stromal cell activation. Keratocytes isolated from rabbit corneal stroma by collagenase digestion were plated in serum-free or insulin-, bFGF/heparin sulfate (HS)-, TGF-beta1-, or fetal bovine serum (FBS)-supplemented DMEM/F12 medium. Expression of type IV collagen isoforms and keratan sulfate proteoglycans (KSPGs) was evaluated by immunocytochemical analysis, Western blot analysis, or both. Concentrations of mRNAs were estimated by quantitative RT-PCR using SYBR Green RT-PCR reagents. Immunohistochemical analysis indicated that type IV alpha1, alpha2, and alpha3 collagens were expressed in normal rabbit corneal stroma and in keratocytes cultured in serum-free and insulin-supplemented media. However, alpha3(IV) collagen was not detectable in the regenerating stroma after photorefractive keratectomy (PRK) in rabbit or in corneal stromal cells cultured in media supplemented with FBS, bFGF/HS, or TGF-beta1. alpha3(IV) collagen mRNA levels were also diminished in the stromal cells cultured in these growth factor-supplemented media. KSPGs (lumican and keratocan) were expressed and secreted in serum-free medium. Although the expression of KSPGs was promoted by insulin, the expression and intracellular levels of lumican and keratocan mRNAs were downregulated by TGF-beta1 and FBS. bFGF/HS promoted the downregulation of intracellular keratocan but not lumican mRNA levels. The loss in the expression of alpha3(IV) collagen is a stringent phenotypic change associated with activation of keratocytes in vivo and in vitro. This phenotypic change in activated corneal stromal cells is induced by bFGF/HS and by TGF-beta1, and it accompanies the downregulation of keratocan expression.

Lumican, a small leucine‐rich proteoglycan substituted with keratan sulfate chains is expressed and secreted by human melanoma cells and not normal melanocytes

Melanoma is a frequent and therapy-resistant human disease. Malignant melanocytes modulate their microenvironment in order to penetrate the dermal/epidermal junction and eventually invade the dermis. The small leucine-rich proteoglycans (SLRPs) constitute important constituents of the dermis extracellular matrix (ECM), participating in both the structural and the functional organization of the skin. The role of a keratan sulphate SLRP lumican, has recently been investigated in the growth and metastasis of several cancers. In this study, the expression of lumican was studied in two human melanoma cell lines (WM9, M5) as well as in normal neonatal human melanocytes (HEMN) using real time PCR, western blotting with antibodies against the protein core and keratan sulfate, and treatments with specific enzymes. Both human metastatic melanoma cell lines were found to express lumican mRNA and effectively secrete lumican in a proteoglycan form, characterized to be substituted mostly with keratan sulfate chains. Lumican mRNA was not detected in normal melanocytes. This is the first time that the synthesis and secretion of lumican in human melanoma cell lines is reported. The role of this proteoglycan in the development and progression of malignant melanoma has to be further investigated.

TNF-α and IFN-γ inversely modulate expression of the IL-17E receptor in airway smooth muscle cells

The interleukin-17B receptor (IL-17BR) is expressed in a variety of tissues and is upregulated under inflammatory conditions. This receptor binds both its cognate ligand IL-17B and IL-17E/IL-25, a novel cytokine known to promote Th2 responses. The present study shows that airway smooth muscle cells express IL-17BR in vitro and that its expression is upregulated by TNF-alpha and downregulated by IFN-gamma. Our data indicate that TNF-alpha upregulates IL-17BR mainly through nuclear factor-kappaB as assessed with the IkappaB kinase 2 inhibitor AS-602868. In addition, both IFN-gamma and dexamethasone are able to antagonize a TNF-alpha-induced IL-17BR increase in mRNA expression. The mitogen-activated protein kinase kinase inhibitor U0126 totally reversed the inhibition observed with IFN-gamma, suggesting the involvement of the extracellular signal-regulated kinase pathway in this effect. In addition, on stimulation with IL-17E, airway smooth muscle cells increase their expression of ECM components, namely procollagen-alphaI and lumican mRNA. Furthermore, immunohistochemical analysis of biopsies from asthmatic subjects reveals that this receptor is abundant in smooth muscle layers. This is the first report showing IL-17BR receptor in structural cells of the airways. Our results suggest a potential proremodeling effect of IL-17E on airway smooth muscle cells through the induction of ECM and that its receptor is upregulated by proinflammatory conditions.

EXPRESSION AND ROLE OF LUMICAN IN PANCREATIC CANCER CELLS AND STROMAL TISSUES

Background: Lumican is a member of the small leucine-rich proteoglycan family and is overexpressed during the wound healing process in the cornea and in human breast, colon and pancreatic cancer. In breast cancer tissues, lumican mRNA is overexpressed in stromal fibroblasts but not in cancer cells. The high expression level of lumican in stromal tissues is associated with a high pathological tumor grade, low estrogen receptor levels and young patient age. In the present study, we attempted to clarify whether lumican overexpression affects the clinicopathological features of pancreatic cancer. Methods: Lumican protein expression was examined in MIA-PaCa-2, Panc-1, PK-45H and PK-8 pancreatic cancer cell lines by Western blot analysis. Immunohistochemical analysis with anti-lumican antibody was performed in 54 pancreatic cancer cases. Lumican immunoreactivity in the pancreatic cancer cells and stromal tissues were estimated and the correlation of lumican expression and clinicopathological factors, and prognosis were examined. Results: Lumican protein was detected in pancreatic cancer cell lines. Immunohistochemically, lumican was localized in the alpha cells of islets in control pancreas. In chronic pancreatitis-like lesions adjacent to cancer cells, lumican was localized in acinar and islet cells. In 18 of 56 cases (33%), lumican was strongly expressed in the cytoplasm of cancer cells. In stromal tissue close to cancer cells, lumican was localized in 36 of 56 cases (71%). The overall survival rate was worse in patients showing lumican-negative cytoplasm in cancer cells than for patients with lumican-positive cancer cells. In contrast, lumican in stromal tissues close to cancer cells correlated with retroperitoneal and duodenal invasion and the overall survival rate was worse for patients with lumican-positive stroma than for patients with lumican-negative stroma. Conclusion: Lumican in pancreatic cancer cells and stromal tissues may play different roles in cancer cell growth and invasion.

Influence of aging on glycosaminoglycans and small leucine-rich proteoglycans production by skin fibroblasts

Skin aging is characterised by a progressive deterioration of its functional properties, linked to alterations of dermal connective tissue. Whereas many studies have been devoted to collagen alterations during aging, the situation is less clear concerning glycosaminoglycans and proteoglycans. Particularly, the alterations of the expression of small leucine-rich proteoglycans (SLRPs), a family of proteoglycans strongly implicated in cell regulation, have never been studied. In the present study we measured glycosaminoglycans and small leucine-rich proteoglycans synthesis by skin fibroblasts from donors of 1 month to 83 years old. [3H]-glucosamine and [35S]-sulfate incorporation did not show significant differences of sulfated GAG synthesis during aging. On the other hand, a significant positive correlation was found between hyaluronan secretion and donor's age. Northern blot analysis of SLRPs mRNAs showed a significant negative correlation of lumican mRNA with donor's age, whereas decorin and biglycan mRNAs were not significantly altered. Immunohistochemical study and quantitative image analysis confirmed a decreased lumican accumulation in aged human skin. Taken together, our results suggest that impairment of glycosaminoglycans and SLRPs synthesis might be involved in the functional alterations of aged skin.

Different expression levels of lumican in human carcinoid tumor and neuroendocrine cell carcinoma

Lumican is a member of the small leucine-rich proteoglycan (SLRP) family and participates in the maintenance of tissue structures and tumor growth. Neuroendocrine cell tumors (NETs) including carcinoid tumors and NE cell carcinomas (NECs) possess numerous neuroendocrine (NE) granules, and differences between these tumors are in terms of their biological and metastatic aggressiveness during tumor progression. The purpose of this study was to examine the expression of lumican in NETs, and to determine whether the presence of lumican may be associated with the growth of NETs by comparing its expression between carcinoid tumors and NECs. Immunohistochemically, the positivity rates of lumican expression in the cytoplasm of the tumor cells were 87.5% in carcinoid tumors and 37.5% in NECs. Those of lumican expression in the stroma adjacent to the tumors were 90.1% in carcinoid tumors and 79.2% in NECs. In situ hybridization analysis revealed the lumican mRNA expression in the cytoplasm of carcinoid tumor and NEC cells. Ultrastructurally, the lumican protein was observed in the rough endoplasmic reticulum and NE granules of NETs and interspaces of collagen fibers. Furthermore, RT-PCR analysis revealed the presence of lumican mRNA in NEC cell lines. These results indicate that the higher expression level of cytoplasmic lumican in carcinoid tumors than in NECs may play a role in the slow growth of these tumors.

The Role of Small Leucine-rich Proteoglycan (SLRP) Family in Pathological Lesions and Cancer Cell Growth

The roles of lumican, a member of the small-leucine-rich-proteoglycan (SLRP) family, in pathological fibrosis, cancer tissues and tumor cell growth were reviewed. Lumican is predominantly localized in the areas of pathological fibrosis including the thickened intima of human coronary arteries, ischemic and reperfused hearts, and acute pancreatitis and chronic pancreatitis (CP)-like lesions adjacent to pancreatic cancer nests. In these lesions, lumican mRNA and protein were transiently and ectopically overexpressed in most of the vascular smooth muscle cells (VSMCs) that migrated into the thickened intima, myocardial cells adjacent to an ischemic lesion, acinar cells, islet cells and fibroblasts of pathological pancreatic tissues. The low expression level of lumican in breast cancer is associated with rapid progression and poor survival. Lumican mRNA in breast cancer is overexpressed in fibroblasts adjacent to cancer cells but not in cancer cells. Furthermore, the high expression level of lumican is associated with a high pathological tumor grade, a low estrogen receptor level in the cancer tissues, and young age of patients. The suppression of lumican expression in culture cells induces their cell growth. Lumican-transfected tumor cells are characterized by a strong suppression of their anchorage-independent growth and capacity of invasion. Lumican significantly suppressed subcutaneous tumor formation in syngenic mice, with a concomitant decrease in cyclin D1 expression level, and induced and/or enhanced the apoptosis of these cells. The autocrine mechanism in cancer cells and the paracrine mechanism in cancer cells and fibroblasts via transforming growth factor (TGF)-beta and Smad signals may play important roles in the regulation of tumor growth by SLRPs.

Lumican expression is associated with the formation of mesenchyme-like elements in salivary pleomorphic adenomas.

Pleomorphic adenomas are the most common salivary gland tumour. Although this tumour is considered to be of epithelial origin, it contains 'mesenchyme'-like elements histologically. Lumican is a keratan sulphate proteoglycan that belongs to the small leucine-rich repeat (LRR) proteoglycans and has been reported to be associated with cartilage formation. These findings suggest that lumican expression may be related to the chondroid component in pleomorphic adenomas. To investigate this hypothesis, the present study investigated the expression and localization of lumican in 20 normal human salivary glands and 35 pleomorphic adenomas. Firstly, immunohistochemistry for lumican was performed with pepsin pretreatment. In normal salivary glands, lumican was deposited in the periductal regions. In pleomorphic adenomas, it was predominantly deposited in the hyaline (100%) and fibrous areas (89.4%). In 16 tumours (66.7%), lumican was also deposited in the chondroid areas. Without pepsin pretreatment, lumican was identified in myoepithelial cells in myxoid areas, lacuna cells in chondroid areas, and in the cytoplasm of inner ductal cells. In situ hybridization revealed lumican mRNA expression mainly in the inner cells, the neoplastic myoepithelial cells, and the lacuna cells. These results suggest that lumican is associated with the formation of 'mesenchyme'-like structures in pleomorphic adenomas. In conclusion, normal salivary glands express lumican, which appears to be related to stromal maintenance, and pleomorphic adenomas express lumican mRNA and protein, which may play important roles in the formation of 'mesenchyme'-like areas in this type of tumour.

Effect of morpholino antisense oligonucleotide against lumican mRNA in human embryonic kidney (HEK) 293 cells.

Lumican is a member of the small-leucine-rich proteoglycan (SLRP) family and is overexpressed during wound healing of the cornea, in ischemic and reperfused heart, and in several cancer tissues. Lumican is considered to regulate the collagen fibril diameter and interfibrillar spacing. However, the effect of lumican on cell growth has not been adequately examined. In the present study, we attempted to clarify whether lumican contributes to human embryonic kidney (HEK) 293 cell growth, using the morpholino antisense oligonucleotide (m-anti oligo) against lumican mRNA. M-anti oligo is a novel oligonucleotide and exhibits a higher antisense activity, higher water solubility, and greater resistance to nucleases in target cells than phosphorothioate types of oligonucleotide. After delivery of m-anti oligo against lumican mRNA, the fluorescein 5-isothiocyanate (FITC) conjugated oligonucleotides were observed in the cytoplasm and nucleus of HEK 293 cells at 24 h by confocal laser microscopy. M-anti oligo for lumican mRNA strongly inhibited the synthesis of lumican protein in the HEK 293 cells, and the HEK cell growth rate was higher than those in the control groups. These findings may indicate that lumican protein has an inhibitory effect on HEK 293 cell growth in vitro.

Transient and ectopic expression of lumican by acinar cells in l-arginine-induced acute pancreatitis

Lumican is a member of a small leucine-rich proteoglycan family. We previously found that lumican mRNA and its protein were ectopically and highly expressed in acinar cells in chronic pancreatitis (CP)-like lesions close to pancreatic cancer cells. CP-like lesions are characterized by acinar and ductal-ductular cell proliferation with expanding fibrosis. This finding suggests that lumican is ectopically synthesized by acinar cells under chronic inflammatory conditions and plays a role in fibrosis of the pancreas. However, the expression and role of lumican in acute inflammatory changes of the pancreas are not completely elucidated. In the present study, we aim to clarify whether lumican mRNA and its protein are expressed in exocrine or endocrine components in acute pancreatitis (AP). For experimental AP, Wistar rats received an intraperitoneal injection of L-arginine. Western blot analysis showed an intense 50-kDa band corresponding to the lumican protein in normal and L-arginine-treated rat pancreas. After L-arginine injection, three intense bands at 42, 57, and 92 kDa were detected on day 1. Immunohistochemically, the lumican protein was localized in ductal and a few centroacinar cells in the normal pancreas. After L-arginine injection, an immature fibrosis with fragmented and loose collagen fibers was observed in AP on day 4 and lumican immunoreactivity was detected in the collagen fibers. Lumican mRNA was faintly detected in islet cells in the normal pancreas, but it was strongly expressed in acinar and islet cells on day 1. Furthermore, lumican mRNA was expressed in many proliferating fibroblasts on day 4 by in situ hybridization. These findings indicate that lumican is transiently synthesized by acinar cells and fibroblasts in AP. Lumican proteins synthesized by acinar cells, islet cells, and fibroblasts may contribute to immature and transient fibrosis of AP.

Expression of lumican in human colorectal cancer cells.

Lumican is a member of a small leucine-rich proteoglycan family and its overexpression in human breast cancer tissues is reported to influence the growth of cancer cells. In the present study, we aimed to clarify the expression of lumican mRNA and its protein in human colorectal cancer cell lines and their localization in normal and cancerous colorectal tissues. Reverse transcription-polymerase chain reaction and western blot analysis revealed lumican mRNA and its protein expression in COLO 205, DLD-1, HCT-15, SW 480 and WiDr colorectal cancer cell lines. The lumican in colorectal cancer cells had non-sulfated or poorly sulfated polylactosamine side chains. Based on its immunoreactivity, the lumican protein was found to be localized in fibroblasts and stromal tissues of normal colorectal tissues, but not in colorectal epithelial cells. In colorectal cancer tissues, the lumican was strongly localized in cancer cells in eight of 12 cancer cases. The lumican protein was also localized in epithelial cells with mild reactive dysplasia and fibroblasts adjacent to cancer cells. Lumican mRNA was expressed in cancer cells and adjacent fibroblasts, and epithelial cells. These findings may indicate that the lumican protein synthesized by cancer cells, fibroblasts and epithelial cells with mild reactive dysplasia found adjacent to cancer cells may affect the growth of human colorectal cancer cells.

Expression and accumulation of lumican protein in uterine cervical cancer cells at the periphery of cancer nests

Lumican is a member of a small leucine-rich proteoglycan (SLRP) family and is reported to be overexpressed during the wound healing process of the cornea, and ischemic and reperfused heart. In the carcinomatous tissues, lumican is overexpressed in human breast and pancreatic cancer tissues. In the present study, we aimed to clarify the expression of lumican mRNA and its protein in human cervical cancer cell lines (CaSki, ME-180 and HeLa cells) and their localization in normal and cancerous human cervical tissues. Reverse transcription-polymerase chain reaction and Western blot analysis revealed the expression of lumican mRNA and its protein in CaSki, ME-180 and HeLa cells. No or weak immunoreactivity of the lumican protein was observed in stroma but not in squamous and ductal cells of non-cancerous uterine cervical tissues. In 21 of 28 (75%) cervical cancer cases, the lumican protein was strongly expressed in cancer cells, and accumulated particularly in cancer cells at the periphery of the cancer nests. It was also expressed in the fibroblasts adjacent to the cancer cells. In situ hybridization analysis revealed that lumican mRNA was not expressed in squamous or ductal epithelial cells in non-cancerous tissues, but was expressed in most cancer cells and stromal fibroblasts in uterine cervical cancer tissues. The lumican protein was not localized in normal squamous or ductal cells close to cancer cells, but its mRNA was strongly expressed in the same cells. To our knowledge, this is the first report on lumican synthesized by squamous cell carcinomas. These findings may indicate that the accumulated lumican protein in cancer cells at the periphery of cancer nests may play roles in the growth or invasion of human cervical cancer cells.

Expression of Lumican in Thickened Intima and Smooth Muscle Cells in Human Coronary Atherosclerosis

Lumican is a member of a small leucine-rich proteoglycan family. Members of this family play an important role in cell migration and proliferation during embryonic development, tissue repair, and tumor growth. Lumican is reported to be overexpressed during the wound-healing process in the cornea and ischemic and reperfused heart. Recently, we found that lumican mRNA and its protein are expressed in cultured vascular smooth muscle cells (VSMCs) from the rat aorta. However, the expression and role of lumican in human atherosclerotic tissues are not clearly elucidated. In the present study, we aimed to clarify whether lumican is expressed in VSMCs and its localization in human coronary atherosclerotic tissues. The lumican protein and its mRNA were expressed in a small number of VSMCs in the media of normal coronary artery, but the lumican protein was not localized in the medial stroma. In contrast, the lumican protein and its mRNA were expressed in most of VSMCs that migrated into the thickened intima, but not in infiltrating foamy macrophages. The lumican protein was prominently localized in the thickened intimal stroma. The lumican protein and its mRNA were also expressed in VSMCs in the inner layer of the media and its protein was localized in medial stromal tissues. These findings indicate that the lumican protein is mainly synthesized by intimal and medial VSMCs in coronary atherosclerosis and that lumican contributes to collagen fibrillogenesis of coronary atherosclerosis.

Lumican expression in alpha cells of islets in pancreas and pancreatic cancer cells.

Lumican is a member of a small leucine-rich proteoglycan family, members of which play an important role in cell migration and proliferation during embryonic development, tissue repair, and tumour growth. Lumican is reported to be overexpressed during the wound healing process in the cornea and in human breast cancer tissues, but its expression and localization in normal pancreas and pancreatic cancer tissues are not known. The present study aimed to clarify the expression of lumican protein and its mRNA in human pancreatic cancer cell lines and their localization in normal and cancerous human pancreatic tissues. Reverse transcription-polymerase chain reaction (RT-PCR) and western blot analysis revealed lumican mRNA and its protein expression in PK-8 and MIA-PaCa-2 human pancreatic cancer cells. The tumour lumican had non- or poorly sulphated polylactosamine side-chains rather than highly sulphated keratan sulphate chains. Immunoreactivity of the lumican protein was localized in alpha cells of islets and stromal tissues of a normal pancreas. In pancreatic cancer tissues, the lumican protein was strongly localized in cancer cells, and in acinar and islet cells in chronic pancreatitis-like lesions adjacent to the cancer cells. It was also localized in fibroblasts and collagen fibres close to cancer cells. Lumican mRNA was expressed in cancer cells, in acinar and islet cells in chronic pancreatitis-like lesions, and in stromal fibroblasts in the pancreatic cancer tissues. This is the first report that lumican is synthesized in endocrine and cancer cells. Lumican protein may play a role in the maintenance of islet cell function in normal pancreas and the lumican protein synthesized by cancer cells, acinar and islet cells, and stromal fibroblasts may play a role in the growth of human pancreatic cancer cells.

Effects of the extracellular matrix on lumican expression in rat aortic smooth muscle cells in vitro.

Lumican is a small leucine-rich proteoglycan (SLRP), which contributes to cell migration, proliferation, tissue hydration, and collagen fibrillogenesis. Whether lumican is localized in rat aortic smooth muscle cells (SMCs) and what its relationships might be to other extracellular matrix components have not yet been elucidated. In this study, using reverse transcription-polymerase chain reaction (RT-PCR), competitive RT-PCR, and western blot, lumican messenger ribonucleic acid (mRNA) was expressed in cultured rat aortic SMCs. SMCs cultured in serum-free medium showed four bands at 68, 62, 50, and 37 kD. The 68 and 62 kD bands corresponded to proteoglycan, the 50 kD band to glycoprotein, and the 37 kD band to the core protein form of lumican. The relationships of lumican to fibronectin and laminin were also investigated. The lumican mRNA level in SMCs cultured on fibronectin was highest at day 1, but it increased at day 3 in SMCs cultured on laminin. On the fibronectin or laminin-coated plates, SMCs expressed only the 68 and 62 kD bands, corresponding to proteoglycan. Pretreatment with anti-beta1 integrin receptor antibody revealed a decrease in the proteoglycan forms of lumican protein and an additional two bands at 50 and 37 kD, indicating glycoprotein and the core protein of lumican. These results show that lumican was synthesized in cultured rat aortic SMCs as proteoglycan, glycoprotein, and core protein. The extracellular matrix (ECM) affected lumican protein production and restricted the lumican protein form to proteoglycan via the beta1 integrin receptor in SMCs.

Expression and localization of lumican in the ischemic and reperfused rat heart.

Lumican belongs to the small leucine-rich proteoglycan family and has an important role in the regulation of corneal and dermal collagen fiber assembly. Recently, lumican mRNA was found in the heart and its high expression was reported during wound healing of the cornea. In the present study, the expression and role of lumican in fibrosis of an ischemic and reperfused rat heart were examined. The expression level of lumican mRNA increased in the ischemic and reperfused rat heart and was highest on the fourth week. Lumican protein existed in the forms of core protein and proteoglycan in the control rat heart. The amount of lumican in the form of proteoglycan increased, and that as the form glycoprotein was newly detected in the ischemic and reperfused rat heart on the fourth week. In the control heart, lumican was weakly expressed in the collagen fibers of the perivascular area, but it was expressed strongly in many capillary endothelial cells in the ischemic lesion on day 1. After 3 days, lumican was localized in collagen fibers and in the fibroblasts of fibrotic lesions. A few myocardial cells close to the ischemic lesion expressed lumican mRNA. Lumican is considered to play an important role in the fibrillogenesis of the ischemic and reperfused rat heart.