Role of relaxin-2 in human primary osteosarcoma

Cancer Cell International

Yongming Xi¹  Lina Zang³  Wenjiu Yang¹  Min Niu²  Jinfeng Ma¹ 

[1] Department of Spine, the Affiliated Hospital of Qingdao Medical College, Qingdao University, Qingdao 266003, R.P China;Department of pharmacy, the Affiliated Hospital of Qingdao Medical College, Qingdao University, Qingdao 266003, R.P China;Department of Internal medicine-cardiovascular, the Affiliated Hospital of Qingdao Medical College, Qingdao University, Qingdao 266003, R.P China

关键词: Gene treatment;    Relaxin-2;    Angiogenesis;    Metastases;    Prognosis;    Osteosarcoma;   

Others  :  793595
DOI  :  10.1186/1475-2867-13-59

 received in 2013-04-05, accepted in 2013-06-05,  发布年份 2013

【 摘 要 】

Background

The aim of this study was to clarify the clinicopathological outcome of serum relaxin-2 and tissues relaxin-2 expression levels in human primary osteosarcoma (OS), and to explore the roles of relaxin-2 inhibition and determine its possibility as a therapeutic target in human osteosarcoma.

Methods

Real-time quantitative RT-PCR assay was performed to detect the expression of relaxin-2 mRNA in 36 cases of human osteosarcoma tissue samples. Serum relaxin-2 levels was measured in ELISA-based method in the 36 cases of osteosarcoma and 50 cases of controls. MTT and TUNEL assay was used to detect cell proliferation and apoptosis after relaxin-2 knockdown with siRNA transfection for 48 hs in vitro. Matrigel invasion and angiogenesis formation assay was used to detect cell metastasis and angiogenesis with HMEC-1 endothelial cells after relaxin-2 knockdown with siRNA transfection for 48 hs in vitro. The effects of relaxin-2 knockdown with anti- relaxin-2 mAb treatment on growth, apoptosis angiogenesis formation and lung metastasis in vivo was analyzed.

Results

The results showed the levels of relaxin-2 mRNA expression in osteosarcoma tissue samples were significantly higher than those in the corresponding non-tumor tissue samples (P < 0.01), and the serum relaxin-2 levels were significantly higher in OS patients than in healthy controls (P < 0.01). The incidence of advanced stage cancer and hematogenous metastasis cancer in the high relaxin-2 mRNA expression group and high serum relaxin-2 levels groups was significantly higher than that in the low relaxin-2 expression group and low serum relaxin-2 levels groups, respectively. Knockdown of relaxin-2 by siRNA transfection in vitro inhibited proliferation, invasion and angiogenesis in vitro in MG-63 OS cells. In vivo, knockdown of relaxin-2 with anti- relaxin-2 mAb treatment inhibited tumor growth by 62% (P < 0.01) and the formation of lung metastases was inhibited by 72.4% (P < 0.01). Microvascular density was reduced more than 60% due to anti- relaxin-2 mAb treatment (P < 0.01).

Conclusions

Our study suggests that overexpression of relaxin-2 is critical for the metastasis of human osteosarcoma. Detection of relaxin-2 mRNA expression or serum relaxin-2 levels may provide the first biological prognostic marker for OS. Furthermore, relaxin-2 is the potential molecular target for osteosarcoma therapy.

【 授权许可】

   
2013 Ma et al.; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Broadhead ML, Clark JC, Myers DE, Dass CR, Choong PF: The molecular pathogenesis of osteosarcoma: a review. Sarcoma 2011, 2011:959248-959252.
• [2]Jaffe N: Osteosarcoma: review of the past, impact on the future. The American experience. Cancer Treat Res 2009, 152:239-262.
• [3]Chou AJ, Geller DS, Gorlick R: Therapy for osteosarcoma: where do we go from here? Paediatr Drugs 2008, 10:315-327.
• [4]Trent JC: Rapid evolution of the biology and treatment of sarcoma. Curr Opin Oncol 2008, 20:393-394.
• [5]Siegel HJ, Pressey JG: Current concepts on the surgical and medical management of osteosarcoma. Expert Rev Anticancer Ther 2008, 8:1257-1269.
• [6]Chou AJ, Gorlick R: Chemotherapy resistance in osteosarcoma: current challenges and future directions. Expert Rev Anticancer Ther 2006, 6:1075-1085.
• [7]Sherwood OD: Relaxin’s physiological roles and other diverse actions. Endocr Rev 2004, 25:205-234.
• [8]Bathgate RA, Ivell R: SanbornBM, et al. International Union of Pharmacology LVII: recommendations for the nomenclature of receptors for relaxin family peptides. Pharmacol Rev 2006, 58:7-31.
• [9]Feng S, Agoulnik IU, Bogatcheva NV, Kamat AA, Kwabi-Addo B, Li R, Ayala G, Ittmann MM, Agoulnik AI: Relaxin promotes prostate cancer progression. Clinical Cancer Res 2007, 13:1695-1702.
• [10]Shabanpoor F, Separovic F, Wade JD: The human insulin superfamily of polypeptide hormones. Vitam Horm 2009, 80:1-31.
• [11]Johnson MR, Abdalla H, Allman AC, Wren ME, Kirkland A, Lighman SL: Relaxin levels in ovum donation pregnancies. Fertil Steril 1991, 56:59-61.
• [12]Brener SHL, Schoenfeld C, Amelar RD, Dubin L, Weiss G: Stimulation of human sperm cervical mucus penetration in vitro by relaxin. Fertil Steril 1984, 42:92-96.
• [13]Conrad KP, Novak J: Emerging role of relaxin in renal and cardiovascular function. Am J Physiol Regul Integr Comp Physiol 2004, 287:250-261.
• [14]Silvertown JD, Symes JC, Neschadim A, Nonaka T, Kao JCH, Summerlee AJ, Medin JA: Analog of H2 relaxin exhibits antagonistic properties and impairs prostate tumor growth. FASEB J 2007, 21:754-765.
• [15]Silvertown JD, Ng J, Sato T, Summerlee AJ, Medin JA: H2 relaxin overexpression increases in vivo prostate xenograft tumor growth and angiogenesis. Int J Cancer 2006, 118:62-73.
• [16]Silvertown JD, Geddes BJ, Summerlee AJ: Adenovirusmediated expression of human prorelaxin promotes the invasive potential of canine mammary cancer cells. Endocrinology 2003, 144:3683-3691.
• [17]Binder C, Simon A, Binder L: Elevated concentrations of serum relaxinare associated with metastatic disease in breast cancer patients. Breast Cancer Res Treat 2004, 87:157-166.
• [18]Einspanier A: Relaxin enhances in-vitro invasiveness of breast cancer cell lines by up-regulation of matrix metalloproteases. Mol HumReprod 2002, 8:789-796.
• [19]Kamat AA, Feng S, Agoulnik IU: The role of relaxin in endometrial cancer. Cancer Biol Ther 2006, 5:71-77.
• [20]Hombach-Klonisch S, Bialek J, Trojanowicz B: Relaxin enhances the oncogenic potential of human thyroid carcinoma cells. Am J Pathol 2006, 169:617-632.
• [21]Facciolli A, Ferlin A, Gianesello L, Pepe A, Foresta C: Role of relaxin in human osteoclastogenesis. Ann N Y Acad Sci 2009, 1160:221-225.
• [22]Ferlin A, Pepe A, Facciolli A, Gianesello L, Foresta C: Relaxin stimulates osteoclast differentiation and activation. Bone 2010, 46:504-513.
• [23]Vinall RL, Tepper CG, Shi XB: The R273H p53 mutation can facilitate the androgen-independent growth of LNCaP by a mechanism that involves H2 relaxin and its cognate receptor LGR7. Oncogene 2006, 25:2082-2093.
• [24]Thompson VC, Morris TG, Cochrane DR: Relaxin becomes upregulated during prostate cancer progression to androgen independence and is negatively regulated by androgens. Prostate 2006, 66:1698-1709.
• [25]Goldsmith LT, Weiss G, Palejwala S: Relaxin regulation of endometrial structure and function in the rhesus monkey. Proc Natl Acad Sci USA 2004, 101:4685-4689.
• [26]Palejwala S, Tseng L, Wojtczuk A, Weiss G, Goldsmith LT: Relaxin gene and protein expression and its regulation of procollagenase and vascular endothelial growth factor in human endometrial cells. Biol Reprod 2002, 66:1743-1748.