| Lung cancer is one of the most common serious malignant tumors and has the highest incidence and mortality in malignant tumors.And there is a rising trend year by year. Non-small cell lung cancer (NSCLC) comprises about80-85%of all lung cancers, which include four major types:including adenocarcinoma, squamous cell carcinoma, adenosquamous carcinoma and large cell carcinoma. As known, early diagnosis is thought to be an important means to reduce the mortality of NSCLC. Metastasis is an important factor to affect the poor prognosis of lung cancer. Most patients with non small cell lung cancer are diagnosed at advanced stages with metastasis. The5years survival rate is very low. In decades, there have been advances in many aspects of the clinical diagnosis and treatment especially molecular targeted therapy in NSCLC. However, our understanding of the pathogenesis of lung cancer is still insufficient, and the effective indicators for early diagnosis and prognosis of NSCLC were lacked. Therefore, looking for effective indicators for early diagnosis and prevention of lung cancer which can prolong the survival rate and improve the quality of patients become a hot topic in the current research of lung cancer.High mobility group protein B1(HMGB1) is a non-histone with a series of highly conserved. It was involved in many important biological processes, such as transcription, DNA preparation, cell growth and differentiation, and extracellular signal transduction of tumor cells. The HMGB1protein is involved in certain types of solid tumor, such as colon, melanoma, prostate, breast cancer, hepatocellular carcinoma and pancreatic cancer. Over expression of HMGB1is closely related to invasion and lymph node metastasis of tumor. Research has shown that level of HMGB1in patients with chronic obstructive pulmonary disease is5times of the normal controls. Invasion and metastasis are the basic characteristic of malignant tumor. The extracellular matrix is a biochemical barrier of cell migration and its degradation is an important process of the metastasis of tumor cells. In this process, matrix metalloproteinases play an important role. But the relationship between HMGB1and matrix metalloproteinase-9(MMP-9) in non small cell lung cancer is not clear. Our study aimed to investigate the expression of HMGB1in lung cancer from nucleic acid, protein and serum levels, and the correlation between HMGB1and MMP-9in lung cancer tissues.Part1:Expression of HMGB1and MMP-9in non-small cell lung cancer and analysis of their correlationObjective:The role of HMGB1in metastasis of non small cell lung cancer is not clear. This study aimed to evaluate the expression level of High mobility group box-B1(HMGB-1) and Matrix metalloproteinase-9(MMP-9) in Non-small cell lung cancer (NSCLC), and reveal the relationship between HMGB-1and MMP-9. And analyze the relationship between HMGB1expression and tumor staging and clinical histology in lung cancer.Methods:NSCLC and the corresponding adjacent normal tissues were selected from30patients, which were used for RT-PCR and Western blot analyses of HMGB1to determine the expression of NSCLC tissue in the mRNA and protein. After extraction of RNA and detection by using qPCR instrument, the relative expression of HMGB1was calculated. Expression of HMGB1protein in lung cancer tissues was tested by Western blot.100paraffin embedding NSCLC tissues were isolated from patients through surgical resection, and used for detection of HMGB-1by immunohistochemistry. In which50samples were also used for MMP-9detection, and30normal tissues were considered to be control. Correlation analysis of HMGB-1and MMP-9was carried out.Statistical analyses:All data were expressed as mean±SD. Statistical analysis was treated by SPSS version17.0(SPSS Inc., Chicago, IL). Comparison between different groups was performed using t test (LSD-t). A p-value less than0.05was considered to be significantly different. Correlation analysis of HMGB-1and MMP-9 was carried out by Pearson.Results:In order to observe the expression level of HMGB-1in NSCLC,30NSCLC tissues were analyzed by both RT-PCR and Western blot. As a result, the average expression level of HMGB-1in NSCLC patients was significantly higher than the normal lung tissues, respectively (relative to the beta-actin)8.62±3.32,3.27±1.74, t=2.219,p<0.05. The expression of HMGB1protein in lung cancer tissue was determined by Western blot method. As a result, the expression of HMGB1was significantly higher than that in corresponding normal lung tissues, respectively0.58±0.12,0.21±0.72, t=11.7, P<0.001.Expression of HMGB-1was analyzed by immunohistochemistry in100NSCLC tissues and30normal tissues. As a result, HMGB1was mainly expressed in the cytoplasm of tumor cells. There were71positive expression cases (positive rate71%) in NSCLC tissues. In normal tissues,10cases exhibit positive (positive rate33.3%) expression. No difference was found among the various gender, age and pathological type of patients. There was no significant difference between the expression of HMGB1in squamous cell carcinoma and adenocarcinoma tissues, P>0.05. However, patients in different stages showed different expression intensity. In42I-II period cases, there were24cases exhibt positive expression (positive rate51.7%). But in58III-IV period cases, the positive rate increased to81.0%(47positive expression cases), we also analyzed the expression of MMP-9in NSCLC tissues and normal tissues. As a result, there were38positive expression cases (positive rate76%) in NSCLC tissues.and7positive expression cases (positive rate23.3%) normal tissues, P<0.05.We speculate that there maybe some relationship between HMGB-1and MMP-9because of the similar expression analysis on HMGB-1and MMP-9. As a result, a positive significant correlation with HMGB-1was found in the expression of MMP-9, R-0.399,P<0.01.Conclusion:The expression of HMGB-1in NSCLC was high and it may be one of prognostic and predictive indicators of NSCLC. In addition, there was positive correlation between the expression of MMP-9and HMGB-1. HMGB1and MMP-9may play an important role in invasion and metastasis of non-small cell lung cancer.Part2:The value of combined detection of serum HMGB1and CEA in diagnosis of non small cell lung cancerObjective:High mobility group box-B1(HMGB-1) has a wide range of biological functions, Previous studies have suggested that it plays an important role in tumor development and angiogenesis through a variety of mechanisms. However, the level of serum HMGB-1in non-small-cell lung carcinomas (NSCLC) is unclear. Few research report serum the level of HMGB1in patients with non small cell lung cancer. Carcinoembryonic antigen (CEA) is an important tumor maker and is commonly used in lung cancer. The aim of this study was to test serum HMGB-1and CEA levels in non-small-cell lung cancer (NSCLC) and to investigate correlation between serum level of HMGB1and CEA.Methods:The study entered178patients and32healthy volunteers (control group) from the Affiliated Hospital of Qingdao University between March2012and March2013. The serum HMGB-1and CEA levels were measured by enzyme-linked immunosorbent assay.71patients of pathological-stage I-II underwent operation. Peripheral fasting blood from all patients was collected in the early morning and serum HMGB-1and CEA were assessed by electro-chemiluminescence immunoassay (ECLIA) before and2weeks after operation. Correlation analysis of HMGB-1and CEA was carried out by Pearsons correlation. In accordance with the declaration of Helsinki, this study was approved by the local ethics committee.Results:The baseline serum CEA level of178patients was higher than those in controls, respectively13.36±2.26ng/mL,1.76±0.52ng/mL,P<0.05. The baseline serum HMGB-1level of178patients was higher than those in controls, respectively3.21±0.81ng/mL,2.07±0.65ng/mL,P<0.05.In order to determine whether the serum level of CEA and HMGB1in post-operration patients differ from those in pre-operration patients, we detected the serum levels of CEA and HMGB1before and2weeks after surgery of the same patients. The serum CEA level of71patients after operation was lower than the same patiens before operation, respectively6.08±1.48ng/mL,6.8±1.57ng/mL, P<0.05. The serum HMGB1level of71patients after operation was lower than those before operation, respectively2.12±0.54ng/mL,2.69±0.71ng/mL, P<0.05.The serum CEA level of Patients with lung adenocarcinoma was higher than that of lung squamouscarcinoma, respectively19.2±3.45ng/mL,5.6±1.62ng/mL,P<0.05. The serum HMGB1level of patients with lung adenocarcinoma was no different from that of lung squamouscarcinoma, respectively3.18±0.72ng/mL,3.23±0.78ng/mL, P>0.05.The serum CEA level of Patients of stage Ⅲ-Ⅳ was higher than that of stage Ⅰ-Ⅱ, respectively17.71±3.12ng/mL,6.8±1.43ng/mL, P<0.05. The serum HMGB1level of patients of stage Ⅲ-Ⅳ was higher than that of stageⅠ-Ⅱ,respectively3.55±0.65ng/mL,2.69±0.57ng/mL,P<0.05.Both the serum CEA and HMGB1levels in patients with lung cancer were elevated.Therefore, we performed Pearson analysis on HMGB-1and CEA. But no significant correlation was found in the expression of HMGB-1and CEA as shown in Figurel (t=-0.21,P=0.856)Conclusion:Our data indicated that the serum levels of HMGB1and CEA could discriminate lung cancer patients from health donors, and that HMGB1and CEA might be a useful marker in early diagnosis of NSCLC. There is no correlation between the serum HMGB1and CEA level.The conclusions:1. Expression of HMGB1in non small cell lung cancer was higher than that in paracancerous matched control tissue in nucleic acid, protein level. Serum HMGB1level was higher than that of healthy controls. Our data showed that the level of HMGB1and clinical staging of lung cancer tissue was correlated.But there was no difference between adenocarcinoma and adenocarcinoma.2. Expression of MMP-9in non-small cell lung cancer tissues was elevated.There was significant correlation between the expression of HMGB1and MMP-9in lung cancer tissues.3.The serum level of CEA in patients with non small cell lung cancer was significantly higher than that in healthy control subjects,.But at present no correlation was found between the serum HMGB and CEA levels. |
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