| The tumor microenvironment plays a vital role in tumor development and metastasis. Extracellular matrix proteins is an important part of the tumor microenvironment, the abnormal expression of matrix proteins are often found in tumors, and which contribute to tumor progression and metastasis. Recent studies have shown that the expression of Lumican closely related to the growth and metastasis of many malignant tumors, but its exact role in tumor development is unclear. We herein evaluated the effect of Lumican, a novel member of SLRP family, on human gastric cancer (GC). Reverse transcription polymerase chain reaction (RT-PCR), real-time quantitative PCR and immunohistochemistry staining on tissue array were employed to assess the expression of Lumican in GC samples. Lumican overexpression and specific interference cell line was established by lentiviral vector system. CCK-8method, colony formation and in vivo tumor formation test were used to analyze the growth of gastric cancer cells after Lumican overexpression and interference; Transwell and Matrigel method were to analyze the migration and invasion abilitie of gastric cancer cells after Lumican overexpression and interference. Cell cycle and Cell anoikis rate were analyzed by flow cytometry. Immunoblotting and immunofluorescence to detect smad2, samd3and samd4of expression and activation. The pQE30-HLumican Lumcian recombinant protein prokaryotic expression vector system and V152-HLumican of eukaryotic expression vector system were constructed. Two recombinant proteins affect cell viability of gastric cancer cells were used by CCK-8method; Transwell method was to analyze the migration abilitie of gastric cancer cells after the treatment of the two recombinant proteins; Flow cytometry analysis the V152-HLumican recombinant protein on gastric cancer cell anoikis impact; Soft agar colony formation assay to observe the the V152-HLumican recombinant protein in gastric cancer cell growth of non-anchor; The tumorigenicity experiments analyzed the impact of the V152-HLumican recombinant proteins on the growth of gastric cancer cells in vivo. The experiments results showed that: the expression of mRNA was3.15-fold higher in gastric cancer than in adjacent normal tissue (P<0.01). Lumican primarily a molecular weight of about55kDa size glycoprotein present in the gastric cancer tissues,40kDa size of the core protein also have a small amount of expression. Immunohistochemistry analys showed that the expression level of Lumican in gastric cancer was higher than that in corresponding paracancerous tissue (91.67%vs.7.5%, P<0.01). There was no significant difference in the Lumican expression among gender, age, Tumor location, Laren sub-type, signet ring cell carcinoma, vascular invasion, tumor thrombus (P>0.05). But significant difference in the Lumican expression among invasion depth (P<0.01), lymph node metastasis (P<0.01), distant metastasis (P<0.05), TNM stage (P<0.01), tumor size (P<0.05) and early gastric cancer(P<0.01). Overall survival rates of gastric cancer patients after five years in Lumican expression negative group, weak positive group, positive group and strong positive group were89.48%,68.48%,53.56%and48.17%respectively, as Lumican enhanced expression, patients with gastric cancer after5-year survival rate gradually decreased ((P<0.01). Lumican overexpression can be significantly enhanced gastric cancer cell within the40kDa size of the core protein, while no significant enhancement of55kDa size of glycoprotein; lumican expression of40kDa size of core protein expression was significantly decreased after the disturbance, while the55kDa size of glycoprotein did not decline significantly. Lumican overexpression can inhibit cell growth in vitro and in vivo, inhibit cell migration and invasion, increase the rate of anoikis. Lumican overexpression able to inhibit cell cycle progression and caused G1phase arrest. Lumican interference can promote cell proliferation, migration, invasion and resistance to anoikis ability. Lumcian regulation of gastric cancer cell proliferation, migration, invasion and resistance to anoikis may be due Lumican can suppress the expression of smad4, and Smad2and Smad3activation. The pQE30-HLumican recombinant protein can inhibit the gastric cancer cell proliferation and migration. V152-HLumican recombinant protein can promote the gastric cancer cell proliferation and migration, and enhance the ability of gastric cancer cells resistant to anoikis. Our results show that the expression level of Lumican core protein and glycoprotein in gastric cancer tissues were significantly increased, Lumcian core protein can inhibit gastric cancer cell growth, migration and invasion. Lumican glycoprotein can promote gastric cancer proliferation, migration and invasion. Lumican and its glycosylating plays an important role in the development of gastric cancer. |
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