Serum-hepatocyte Growth Factor (S-HGF) in Diagnosis of SPNs

Serum-hepatocyte Growth Factor (S-HGF) in Diagnosis of SPNsValue of hepatocyte emersion factor in the runner derivative diagnosis of lonesome pulmonic nodulesF1Haixin Yu, Yan Wang*, Wenduan Ma, Haixiang Yu, Shengtao ShangAbstractPurpose To evaluate blood serum-hepatocyte growth factor (S-HGF) in the differentiation of solitary pulmonary nodules(SPNs)F2.F3Methods The study comprised 42 serum samples from SPN patients and 10 brawny samples as see to it. The HGF was measured by the commercially available immunoassayF4.F5 Serum directs of HGF of 42 patients with SPN was measured by ELISA kit, and compargond with the control meeting of 10 normal subjects. The nodules were diagnosed by operation and pathology.Results The median level of S-HGF was 180( range from one hundred to 300) pg/ ml in the sun-loving control chemical sort out, 165( range from 100 to 400) pg/ ml in kind SPNs crowd and while 395( range from 100 to 1550) pg/ ml in cancerous SPNs group, The S-HGF mean l evel of cancerous group was momentously uplifteder than the with solid diversion observed between the malignant group and control group(P. Moreover, the malignant group was also authoritatively mettlesomeer than the , and between the malignant group and the benign group(Pwhile no significant difference between the benign , but not between the benign group and the control group(P0.05). upgrademore, the S-HGF was also shown no statistically significant difference was observed(P0.05) in different pathologic types of the limited number of lung crab louse patients.In addition, when S-HGF in different pathologic types of the limited number of lung crabmeat patients were comp ared, no statistically significant difference was observed(P0.05).Conclusion S-HGF is valuable in the differential diagnosis of solitary pulmonary nodules. It was suggest that the patients with SPNs should consider an operation when the S-HGF level 250pg/ml, and malignant SPNs are highly suspected while S-H GF level 400pg/ml, surgical intercession should be taken immediately.S-HGF is valuable in the differential diagnosis of solitary pulmonary nodules. An el S-HGF level250pg/ml in patients with SPNs may strongly speak for malignant nodules and operation is suggested. If S-HGF level 400pg/ml, malignant SPNs are highly suspected, active surgical interpolation should be taken.Key words diagnosis, hepatocyte growth factor, solitary pulmonary nodule, NSCLClung cancer1. IntroductionThe solitary pulmonary nodules (SPNs) is a single mass in the lung less(prenominal) than or equal to 3 cm in diameter, without concomitant pneumonia and atelectasis of involved lung segments and lobes 1. In the general population, its reported that approximately 5% of SPN patients show lung cancer by radiology 2, which is considered one of the most common forms of cancer with a high death incidence ratio in the world 3. Diagnoses of benign and malignant SPN has been concerned and become a challenge in these de cades 4, 5. Therefore, it is utmost important to improve the method in the characterization of SPNs6.With the development of modern medical scholarship and technology, several detecting and monitoring method were used in screening the SPNs and lung cancer 2, 7, 8, Momen9 et al. have compared three detection methods for identifying malignant SPNs for the sensitivity and specificity. The positron waiver tomography (favourite) imaging was consistently higher (80 to 100%) for its sensitivity, while was with lower specificity and larger variation (40 to 100%). Also, they comprise the similar results in dynamic CT with enhancement (sensitivity, 98 to 100% specificity, 54 to 93%). In studies of CT-guided needle biopsy, sensitivity and specificity performed excellent, but nondiagnostic results were seen approximately 20%. Dalli8 et al. also showed the similar result in 2013. While Carsten10 et al. suggested that routine flexible bronchoscopy should be included in the pre-operative work-u p of patients with SPNs in his study. Even so, it seems to find a better detection method of long cancer and characterization of SPNs is still necessary. Serum-hepatocyte growth factor (S-HGF, Serum-HGF) is an important fibroblast-secreted protein that mediates development and progression of cancers11. Nagio et al. 12 gave the evidence that the S-HGF levels of patients with small cell lung cancer (SCLC) were significantly higher than those of patients with benign SPNs and thinking(a) subjects. Ujiie et al13 had proved that the levels of HGF in serum could be used as prognostic indicators of the patients with stage III non-small cell lung cancer (NSCLC) undergoing military operation and chemotherapy. Kasahara et al. 14 found that higher HGF levels were significantly associated with a shorter progression-free survival (PFS) and overall survival (OS) in patients with non-small cell lung adenocarcinoma. The expression level of S-HGF could be a sensitive indicator and an independent bi omarker for evaluating the therapeutic effects and the prognosis in patients with lung cancer. Therefore, we give the hypothesis that S-HGF may be a potential target in diagnoses of benign and malignant SPNs associated with lung cancer. In our study, we used Enzyme relate immunospot assay (ELISA) method to detect the S-HGF levels between different serum samples from SPNs patients and healthy subjects. The solitary pulmonary nodule(SPN) is defined as a round opacity 3 cm in diameter surrounded by lung parenchyma1.There should be no associated with hilar lymphadenopathy, atelectasis, pneumonia or chest wall pathologies. With more importance attached to bodily examination and the development of medical imaging examination technology, the detection rate of SPN is on the increase.In the general population, approximately 5% of all SPNs shown by radiology are reported to be carcinomas2. In eight large trials of lung cancer screening, Momen et al3 have compared the sensitivity and speci ficity in three detection methods for identifying malignant SPNs. The sensitivity of PET imaging was consistently high (80 to 100%), whereas specificity was lower and more variable (40 to 100%). They found similar results in dynamic CT with enhancement(sensitivity, 98 to 100% specificity, 54 to 93%).In studies of CT-guided needle biopsy, sensitivity and specificity were excellent, but nondiagnostic results were seen approximately 20% of the time. Carsten et al4, in a study of 225 patients with SPN of unknown origin, observed that the bronchoscopic biopsy results were positive in 84(46.5%) patients with lung cancer. The differential diagnosis between malignant and benign solitary pulmonary nodules (SPNs) is always a difficult point in clinical practice. In this study, we check the clinical significance of the serum level of hepatocyte growth factor(HGF) in patients with SPNs.2. Methods2.1. PatientsAccording to the definition, inclusion criteria was set1On computed tomography (CT), S PN is a round opacity 3 cm in diameter surrounded by lung parenchyma.2There should be no associated with hilar lymphadenopathy, atelectasis, pneumonia or chest wall pathologies.3Regardless of age and gender. In consideration of some influences, exclusion criteria was set(1)Inflammation or infection within a month. (2)Surgery or trauma within 6 months. (3)Various liver diseases. (4)Chronic renal failure. (5)Arteriosclerosis. (6)Rheumatoid arthritis and osteoarthritis. (7)Diabetes mellitus.The case group included 42 patients with SPNs, mean age 60.7 years (range, 42 to 72). Besides, 10 healthy adult subjects were chosen as control.2.2. Specimen collectionThe morning fasting venous blood of all subjects was collected in sterile polypropylene tubes, containing ethylenediamine tetraacetic acid (EDTA), and immediately centrifuged at 400rpm for 10min. Then, the blood plasma was stored at -70C until the assays were performed.2.3. Assay for S-HGFWe used Sandwich enzyme-linked immunosorbent assay(ELISA) to measure S-HGF. The HGF monoclonal antibody and standard substance for the assays were purchased from American RD systems. Goat-anti-human HGF polyclonal antibody as the primary antibody and donkey-anti-goat IgG polyclonal antibody labeled with horse radish peroxidase as the secondary antibody were both purchased from British biotech company Abcam.2.4. Pathological diagnosesAll the 42 patients with SPNs were pathological diagnosed postoperatively. 12 cases were benign nodules(4/12 were tuberculoma, 6/12 were rabble-rousing pseudotumor, 2/12 were hamartoma) and 30 cases were malignant nodules(17/30 were adenocarcinoma, 13/30 were squamous carcinoma).2.5. Statistical methodsAll selective information were analyzed by SPSS 19.0. Because the measured data manifested as skewed distribution, geometrical mean GlogGswas calculated in for each one group after logarithmic transmutation had been carried out on each datum. Then, Students t test was performed on both sides. Dif ferences were considered statistically significant at P0.05.3. ResultThe S-HGF data measured of healthy control group, benign SPNs group and malignant SPNs group is shown in Table 1Table 1 are the measured S-HGF data of healthy control group, benign SPNs group and malignant SPNs group. All the data manifest as skewed distribution(All P0.05). Geometrical mean GlogGswas calculated in each group after logarithmic transformation had been carried out on each datum(Table 2).TABLE 1 The S-HGF levelpg/ml of healthy control group, benign SPNs group and malignant SPNs group.TABLE 2 The analogy of S-HGF level of each group after logarithmic transformation had been carried out on each datum.aBenign SPNs group vs healthy control group, P0.05bMalignant SPNs group vs healthy control group, P0.05cMalignant SPNs group vs benign SPNs group, P0.05The S-HGF level of benign SPNs group compared with the healthy control group, there were no significant differences (P0.05). The S-HGF levels of malignant SPNs group were significantly higher than those of healthy control group(Psignificant differences (P0.05, Table 3).TABLE 3 The comparison of S-HGF level of adenocarcinoma and squamous carcinomaaSquamous carcinoma vs adenocarcinoma, P0.054. DiscussionHepatocyte growth factor/ spreadhead factor (HGF/SF) from the serum of hepatectomized rats was first partially purified and described by Nakamura in 198415. HGF receptor encoded by the c-met proto-oncogene is a piece of the tyrosine kinase class of cell surface receptors. As a kind of cytokine, the hepatocyte growth factor(HGF) has widely biological activities, including regeneration, antifibrosis, cytoprotection, and differentiation16. Moreover, HGF is a predominant fibroblast-derived factor that stimulates mitogenesis, motogenesis, and the invasion and metastasis of human carcinoma cells 17. The growth and metastasis of tumors depend on angiogenesis which is the result of the derangement of promoters and inhibitors.The S-HGF levels in patients with acute hepatitis, chronic hepatitis and cirrhosis of the liver were found to be slightly higher than those in normal subjects18. So the patients with various liver and gall diseases were first excluded. So far, some studies showed the S-HGF levels were significantly increased in patients with Inflammation, infection, underwent surgery or trauma. Therefore, the patients with inflammation or infection within a month and the patients underwent surgery or trauma within 6 months were both excluded. Johanna et al. 19 had concluded that patients with chronic renal failure (CRF) have a systemic HGF profile reflecting a chronic incendiary condition with high concentration, but low biological activity, of HGF. Therefore, the patient samples with CRF were also excluded. The S-HGF levels in patients with arteriosclerosis, rheumatoid arthritis, osteoarthritis, and diabetes mellitus were reported to be significantly higher than that in healthy population. So, the patients with these diseases were excluded as well.Tsao et al.20 showed the HGF messenger RNA(mRNA) and protein were preponderantly expressed by the tumor cells in a high percentage of primary NSCLC. Our study showed serum of the healthy control group contained trace amounts of S-HGF, the S-HGF levels of the patients with benign SPNs were nearly close to the healthy control group(PHGF levels of the patients with malignant SPNs were significantly higher than the healthy control group(P0.05) and the benign SPNs group(P0.05). It illustrated that the high level of S-HGF was associated with lung cancer. And it was further confirmed that S-HGF could be expressed by the carcinoma cells in NSCLC.The S-HGF levels of part of patients with squamous carcinoma in the malignant SPNs group were observed to be higher(700pg/ml) and the S-HGF statistical analytic thinking by the statistical difference between the squamous carcinoma group and adenocarcinoma group, for the S-HGF, the median level of the squamous ca rcinoma group was 370(100-1500)pg/ml while the adenocarcinoma group was 420(100-1550)pg/ml, no statistically significant difference between the two groups(P0.05). No further conclusions could be made, in case of the number limitation of the samples. The result confirmation should be amortized awaits further research. get ahead analysis of the 20 patients with high levels of S-HGF(250pg/ml), there are 3 patients(15%) with benign SPNs and 17 patients(85%) with malignant SPNs. Furthermore, for the 20 patients, the result shows that 1 patients (6.25%) with benign SPNs and 15 patients (93.75%) with malignant SPNs in the 16 patients with high levels of S-HGF(400pg/ml), It reveals that an distinguished S-HGF level 250pg/ml in patients with SPNs are more likely to be malignant and when the S-HGF level 400pg/ml, malignant SPNs are highly suspected.ConclusionIn conclusion, our study shows significant in the differential diagnosis between malignant and benign solitary pulmonary nodules (SPNs) for the S-FGF assay. The S-HGF levels of malignant SPNs group are significantly higher than the healthy control group(P SPNs group(P0.05). The differences between benign SPNs group and healthy control group have no statistically significant(P0.05). An elevated S-HGF level 250pg/ml in patients with SPNs are more likely to be malignant, surgical therapy should be considered. If S-HGF level 400pg/ml, malignant SPNs are highly suspected, surgical intervention is recommended without delay. Hepatocyte growth factor/scatter factor (HGF/SF) from the serum of hepatectomized rats was partially purified and described by Nakamura for the first time in 1984. HGF receptor encoded by the c-met proto-oncogene is a member of the tyrosine kinase class of cell surface receptors. As a kind of cytokine, the hepatocyte growth factor(HGF) has widely biological activities, including regeneration, antifibrosis, cytoprotection, and differentiation5. Moreover, HGF is a predominant fibroblast-derived factor t hat stimulates mitogenesis, motogenesis, and the invasion and metastasis of human carcinoma cells6. The growth and metastasis of tumors depend on angiogenesis which is the result of the imbalance of promoters and inhibitors. Sengupta et al7 had demonstrated that HGF/SF could induce angiogenesis independently of VEGF, possibly through the direct activation of the Akt and ERKs.The S-HGF levels in patients with acute hepatitis, chronic hepatitis and cirrhosis were found to be slightly higher than those in normal subjects8. So the patients with various liver and gall diseases were first excluded. So far, some studies have found the S-HGF levels were significantly increased in patients with Inflammation or infection, or underwent surgery or trauma. Therefore, the patients with inflammation or infection within a month and the patients underwent surgery or trauma within 6 months were both excluded. Johanna et al9 had concluded that patients with CRF have a systemic HGF profile reflecting a chronic inflammatory condition with high concentration, but low biological activity, of HGF. Therefore, the patients with CRF were also excluded. The S-HGF levels in patients with arteriosclerosis, rheumatoid arthritis, osteoarthritis, and diabetes mellitus were reported to be significantly higher than that in healthy population. So, the patients with these diseases were all excluded. Tsao et al10 had showed that HGF messenger RNA(mRNA) and protein were predominantly expressed by the tumor cells in a high percentage of primary NSCLC. It indicated in our research that the serum of the healthy control group further contained trace amounts of S-HGF, the levels of S-HGF of the patients with benign SPNs were close to those of the healthy control group(P0.05) and the benign SPNs group(P0.05). It illustrated the fact that high level of S-HGF was associated with lung cancer. And, it was further confirmed that S-HGF could be expressed by the carcinoma cells in NSCLC. In addition, Nagio et al11 had proved that the levels of S-HGF of patients with SCLC were significantly higher than those of patients with benign SPNs and healthy subjects.The levels of S-HGF of a portion of patients with squamous carcinoma in the malignant SPNs group were observed to be higher(700pg/ml) and statistical analysis was conducted to fond the statistical difference of S-HGF between the squamous carcinoma group and the adenocarcinoma group. The S-HGF median of the squamous carcinoma group was 370(100-1500)pg/ml and the adenocarcinoma group was 420(100-1550)pg/ml, no statistically significant difference was found between the two groups(P0.05). No firm conclusions could be made, possibly due to the limited number of cases. It is of concern and remains to be further studied.Further analysis was taken in 20 patients with high levels of S-HGF(250pg/ml), 3 patients(15%) had benign SPNs and 17 patients(85%) had malignant SPNs. Further observation was made, among the 20 patients, there were 16 patien ts with high levels of S-HGF(400pg/ml), 1 patients(6.25%) had benign SPNs and 15 patients(93.75%) had malignant SPNs. It reveals that an elevated S-HGF level 250pg/ml in patients with SPNs are more likely to be malignant and if S-HGF level 400pg/ml, malignant SPNs are highly suspected.Ujiie et al11 had proved that the levels of HGF in serum could be useful prognostic indicators of the survival of patients with stage III NSCLC undergoing surgery and chemotherapy. Kasahara et al12 had shown that higher HGF levels were significantly associated with a shorter progression-free survival (PFS) and overall survival (OS) in patients with non-small cell lung adenocarcinoma. The expression level of S-HGF could be a sensitive indicator and an independent judgement standard for evaluating the therapeutic effects and the prognosis in patients with lung cancer. Furthermore, understanding the role of HGF in the tumor progression may help in designing new therapeutic strategies for lung cancer.In co nclusion, the assay for S-HGF may be of some significance in the differential diagnosis between malignant and benign solitary pulmonary nodules(SPNs). The S-HGF levels of malignant SPNs group were significantly higher than those of healthy control group(P0.05). The differences between benign SPNs group and healthy control group had no statistically significant(P0.05). An elevated S-HGF level 250pg/ml in patients with SPNs are more likely to be malignant, surgical therapy should be suggested. If S-HGF level 400pg/ml, malignant SPNs are highly suspected, active surgical intervention should be taken.References1.Hansell, D.M., et al., Fleischner Society glossary of terms for thoracic imaging. Radiology, 2008. 246(3) p. 697-72213.2.Klein, J.S. and M.A. Zarka, Transthoracic needle biopsy an overview. 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