TFE3 is a Novel Biomarker of Ovarian Sclerosing Stromal Tumours

TFE3 is a Novel Biomarker of Ovarian Sclerosing Stromal Tumours | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article TFE3 is a Novel Biomarker of Ovarian Sclerosing Stromal Tumours Li Zhao, Yan Zhou, Yuping Liu, Qiuping Luo, Qingping Jiang, Hui Wang, and 1 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-2563971/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 01 Aug, 2023 Read the published version in Journal of Ovarian Research → Version 1 posted 4 You are reading this latest preprint version Abstract Sclerosing stromal tumours of the ovary are benign and tend to occur in young women with lobular structures at low frequencies. Three types of cells, luteinized cells, short spindle myoid cells, and intermediate cells, are found in lobules, which are rich in blood vessels. Currently, immunohistochemistry and fluorescence in situ hybridization are used to detect normal follicles, sclerosing stromal tumours, granulosa cell tumours, and theca fibromas. Our research found the expression of transcription factor enhancer 3 (TFE3) was moderately and strongly positive in the inner thecal cell layer of normal follicles. It was expressed in seven out of eight sclerosing stromal tumours, mainly in luteinized cells, but not in 20 granulosa cell tumours and 1 microcystic stromal tumour. In nine cases of theca cell tumours and theca fibromas, TFE3 was not expressed, except in two cases of weak TFE3 expression. Eight cases of sclerosing stromal tumours were analysed by FISH using a TFE3 separation probe, and the results were negative. In a word, as a nuclear transcription protein, TFE3 was specifically expressed in sclerosing stromal tumours and could serve as a new marker for the diagnosis and differential diagnosis of sclerosing stromal tumours. Ovarian Sclerosing Stromal Tumours TFE3 Novel Biomarker Figures Figure 1 Figure 2 Figure 3 Figure 4 Introduction Sclerosing stromal tumour (SST) is a rare sex cord-stromal ovarian tumour that was first reported by Chalvaridjian and Scully in 1973 [ 1 ] . SST occurs predominantly in young women of 20–30 years of age [ 2 , 3 ] , and its clinical symptoms include pelvic pain, menstrual irregularity [ 4 ] , and nonspecific symptoms associated with ovarian cysts. In a few cases, patients had elevated serum levels of CA125 [ 5 ] . However, the levels of hormones in these patients were not affected. Focal adenoid hyperplasia was diagnosed in only 1 of the 10 cases, but endometrial biopsy could not be performed in this case [ 5 ] . In addition, a case of ovarian SST complicated by endometrial adenocarcinoma has been reported [ 6 ] . Characteristic feature of SST is the abundance of blood vessels in the nodules. Computed tomography (CT) and magnetic resonance imaging (MRI) have been employed to diagnose SSTs, particularly to assess their vascularization [ 7 – 9 ] . New ultrasound technologies have been developed to facilitate the exploration of adnexal masses, such as the diagnosis of tissue vascularization via colour Doppler [ 10 ] . This blood flow feature of SST is relatively unique, which is helpful to differentiate from other sex cord stromal tumors and ovarian malignant tumors. This method not only obtains examination results quickly but also reduces the economic burden on patients. In general, SST is unilateral, mostly 5–10 cm, with well-defined boundaries and sometimes a thin fibrous envelope. It is a grey‒white to grey‒yellow nodular oedema, and unilocular cystic cases are rare [ 11 ] . According to previous studies, SST consists of three types of cells, lipid-rich cells, fibroblast-like cells, and undifferentiated stromal mesenchymal cells, with intermediate morphology showing different degrees of differentiation [ 12 ] . In addition, some studies indicate that tumour cells in SST have the characteristics of muscle-like cells and express SMA or desmin [ 13 , 14 ] . Growing evidence shows that cytoplasm-rich cells express calretinin and inhibin [ 15 ] but do not express SMA, desmin, CK, and CK7 [ 16 – 18 ] . In some cases, lipid-rich cells express CD10 [ 19 ] and Melan A [ 20 , 21 ] . These cells are similar to normal cells in the ovary but have not been clearly defined. A study found that TFE3 was highly expressed in sclerosing stromal tumors and pointed out TFE3 is highly expressed in the nucleus of lutein cells and polygonal-to-round tumour cells in 7 out of 9 patients with SST [ 22 ] . However, neither luteinized fibromas nor thecomas express appreciable levels of TFE3 [ 22 ] . Fluorescence in situ hybridization (FISH) analysis revealed the presence of trisomy 12 in > 20% of SST cells [ 12 ] . In 2020, using whole-exome, targeted capture, and RNA sequencing, Sarah et al. reported that 65% (17/26) of SST patients had recurrent FHL2-GLI2 fusion genes and 15% (4/26) had other GLI2 rearrangements [ 23 ] . Specifically, these genetic abnormalities were not detected in other types of sex cord-stromal tumours (n = 48) and common cancers (n = 9,950). In this study, the origin of SSTs was discussed. TFE3 immunohistochemical and molecular analyses were performed on SSTs and other types of ovarian sex cord-stromal tumours. TFE3 staining was used to distinguish SST from other sex cord-stromal tumours. Materials And Methods Patients The records of 38 patients who had undergone surgical resection of sex cord-stromal ovarian tumours at The Third Affiliated Hospital of Guangzhou Medical University (Guangzhou, China) between December 2013 and December 2021. The focus of this study is sclerosing stromal tumor (8 cases) , and its main differential diagnosis is also involved, which are theca cell tumor and theca fibroma tumour (9 cases) , granulosa cell tumour (20 cases), and microcystic stromal tumour (1 case)in turn. The age of patients ranged from 17 to 70 years and they had not received any pre-operative therapy. Ethical approval was obtained from the institutional review board of the ethics committee. Lipid stains (Oil Red O) The biopsy samples were placed in a tissue tek container (Sakura Finetek, CA, USA) and then filled with tissue tek OCT compound gel. After being cut into 7-µm slices, the samples were snap-frozen in liquid nitrogen and stained with Oil Red O according to standard procedures. The oil red O 【 24 】 fat staining method is usually used to detect the fat in tissues or cells. Oil red O is a fat-soluble dye, which is a strong fat solvent and fat dye, and can be highly dissolved in fat. Its dyeing principle is that oil red O can specifically adsorb with the neutral triglycerides, lipids and lipoproteins in tissues and cells to make fat dye. The solubility of dye in intracellular lipids is greater than that of solution. Immunohistochemistry (IHC) Collected tumours or normal tissue after removal of paraffin as the experimental subject. Consecutive 4-μm thick unstained sections for immunohistochemical staining, which was performed using the Leica automatic immunostaining device (Leica Microsystems, Inc.). Primary antibodies against CD10 (1:100; no. 563871; DAKO; DK), α-inhibin (1:100; no. GT230202; CHN), SMA (1:50; no. MAB-0980; MXB; CHN), desmin (1:300; no. GT225202; Gene tech; CHN), TFE3 (1:100; no. ZA-0657; ZSGB-BIO; CHN), calretinin (1:100; no. ZM-0063; ZSGB-BIO; CHN), WT-1 (1:100; no. ZM-0269; ZSGB-BIO; CHN), and EMA (1:300; no. GM061302; Gene tech; CHN). Appropriate positive and negative controls were simultaneously stained to validate the staining method. Immunohistochemistry were conducted according to previously described methods. All slides were reviewed and scored independently by three pathologists. The pathologists were blinded to the experiment. The scoring method based on both the intensity (0, no staining; 1+, weak staining; 2+, medium staining; 3+, strong staining). Fluorescence in Situ Hybridization ( FISH ) analysis The TFE3 isolation probe was provided by Guangzhou LBP Medicine Science and Technology Co., Ltd. (China). Specific operations were performed according to the manufacturer’s protocol. The results showed that there was no fracture of TFE3 gene in females (2 yellow) and males (1 yellow). In females, 1 red 1 green 1 yellow, and in males, 1 red 1 green indicated that TFE3 had a balanced translocation and the gene was fused. In females, 1 red and 2 yellow, and in males, 1 red and 1 yellow, showed unbalanced translocation and fracture of TFE3 gene. Results Clinical findings As shown in Table 1 , the age range of patients with SSTs was 17–39 years old. The levels of hormones were normal in all patients. However, menstruation was irregular in cases 5 and 7. In case 5, an adnexal mass was found during physical examination, and no abnormality was found in the endometrium. Endometrial biopsy showed atypical hyperplasia, and an adnexal mass was found in case 7. In case 4, in which the patient presented with abdominal distension for 1-year, an adnexal tumor was found on B-ultrasound with peritoneal effusion. Adnexal tumours were found in other patients during physical examinations. Table 1 Clinical findings of SSTs Patient with SSTs age gross Size (mm) menstruation endometrium Case 1 23 cystic 60 normal normal Case 2 17 solid 14 normal normal Case 3 26 solid 160 normal normal Case 4 26 solid 45 normal normal Case 5 39 solid 80 irregular normal Case 6 29 solid 60 normal normal Case 7 33 solid 50 irregular atypical hyperlasia Case 8 23 solid 25 normal normal Gross Findings The size of the eight tumours ranged from 14 to 160 mm. All tumours were well-circumscribed nodules, and except for one cystic case (Fig. 1 A), all tumours were solid (Fig. 2 A), which ranged from soft to tough. The cut surfaces of the solid tumours were typically white, slightly leafy, with scattered yellow nodules and were most abundant at the periphery. Focal haemorrhage was observed in one patient. In case 1, the tumour was cystic, with cysts containing thick gelatinous material (Fig. 1 A). Histologic Findings And Immunohistochemical Results Of Sclerosing Stromal Tumours All tumours, including cystic tumours, showed overt, often discrete, cellular and hypocellular regions, resulting in a pseudolobular appearance (Fig. 1 B). The cellular areas and lower cellular intervals were always collagenous, loose collagenous, and markedly oedematous. Cellular foci consisted of a mixture of round and spindle cells (Fig. 1 C). The cytoplasm of the former was pale, eosinophilic, vacuolated, or foamy to varying degrees (Fig. 1 C). Thin-walled, dilated, focally branching (“staghorn”) blood vessels were prominently found in all cases, and markedly conspicuous labyrinthine and haemangiopericytoma-like tumours were evident in some cases (Fig. 2 B). Oil Red O staining revealed that the cytoplasm was rich in lipids but not in mucus (Fig. 1 D). Spindle cells expressed SMA (Fig. 1 E) but not desmin (Fig. 1 F). CD34 staining showed abundant blood vessels in the cell lobules (Fig. 1 G). Only cells with abundant cytoplasm in the tumours expressed TFE3, and nuclear expression was moderately and strongly positive (Fig. 1 H). Expression Of Tfe3 In Ovarian Sex Cord Stromal Tumours As shown in Table 2 , TFE3 was highly expressed in sclerosing stromal tumours (7/8), weakly expressed in two cases of thecoma (2/9), and not expressed in other tumours (0/21, including 20 cases of granulosa cell tumours and one case of microcystic stromal tumours). As shown in Fig. 2 , TFE3 was mainly expressed in sclerosing stromal tumours (Fig. 2 A, B, C) but not in ovarian granulosa cell tumours (Fig. 2 D, E, F), thecoma (Fig. 2 G, H, I), or microcystic stromal tumours (Fig. 2 J, K, L). Moreover, TFE3 mainly expressed in the nucleus of cells with abundant cytoplasm, and the staining intensity was medium to strong (Fig. 2 C). Table 2 Immunohistochemical Results of TFE3 Tumor Type No. of casess 3+ 2+ 1+ 0 Total(%) Theca cell tumor and theca fibroma tumor 9 2 7 22.2% Granulosa cell tumor 20 20 0 microcystic stromal tumor 1 1 0 Sclerosing stromal tumor 8 7 1 87.5% Immunophenotypes Of Theca Cells In Normal Follicles No literature has reported the expression mode of TFE3 in normal ovary. In order to explore the source of TFE3 immunohistochemistry positive cells in sclerosing stromal tumors, we collected 10 normal ovarian tissues. Normal ovarian follicles were obtained from specimens removed due to ovarian endometriosis or teratoma. We tested a total of 30 follicles. Normal follicular structure was observed at low and high magnifications. The boundary between the inner and outer theca cell layers was not very clear, but the cytoplasm of inner theca cells is rich and clear (Fig. 3 A and B). A mostly consistent immunophenotype was observed in the rich cytoplasm cells: each demonstrated diffuse TFE3, TFE3 expressed in the inner theca cells, but not in the granular layer, outer theca layer, or fibrous tissue in normal follicles (Fig. 3 H). FISH detection Using the separation probe of TFE3, TFE3 was detected by FISH in 7 cases of SSTs expressing TFE3. The results showed no separation of TFE3 in these seven cases (Fig. 4 ). Discussion TFE3 is located at Xp11.23, and its protein belongs to the microphthalmia-associated transcription factor (MiTF) family, which plays an important role in the regulation of lysosomal biogenesis and autophagy [ 25 ] . The TFE/MiTF family consists of four important members: (i) TFEB, (ii) TFEC, (iii) TFE3, and (iv) MITF [ 26 ] . By searching the human protein atlas webtool ( https://www.proteinatlas.org/ ), TFE3 was found to be expressed in adipose tissue, urinary bladder, ovary, testis, breast, etc. TFE3 protein can be transported to the nucleus when cells are starved and/or stressed. The expression and activity of TFE3 are upregulated in many types of human cancers, and are associated with enhanced proliferation and motility of cancer cells. The main tumours related to TFE3 gene fusion include epithelioid haemangioendothelioma, alveolar soft-part sarcoma (ASPS), renal cell carcinoma, malignant chondroid syringoma, rare ossifying fibromyxoid tumours, and perivascular epithelioid cell tumor [ 27 – 34 ] . Park CK and Kim HS had reported that TFE3 was expressed in sclerosing stromal tumors, but there was no abnormality in TFE3 gene [ 22 ] . Our research got the same result that TFE3 was specifically expressed in sclerosing stromal tumours, but not in other sex cord stromal tumours. Moreover, TFE3 was specifically expressed in luteinized cells, but not in other two cells. Sclerosing stromal tumours often occur in young women, and a few cases have the secretion of estrogen and/or androgen. The clinical results of all cases were benign. This study analyzed 8 cases of sclerosing stromal tumours, ranging in age from 17 to 39 years old. No hormone abnormality was found clinically, but 2 patients had irregular menstruation. Follow-up results showed no recurrence.The lobulated structure and three types of cell composition are the unique morphological characteristics of ovarian sclerosing stromal tumours. Due to the young age of SST and good prognosis, accurate pathological diagnosis is very important in order to avoid over-treatment. In most cases, we can obtain a positive pathological diagnosis based on the above findings. However, due to the diversity and variety of ovarian sex cord stromal tumours, we need more specific indicators to assist the pathologist in diagnosis in a few cases.For example, the cystic case in Fig. 1 . Our experimental results show that the luteinized tumor cells in sclerosing stromal tumors have abundant intracellular lipids, and Oil red O staining is obvious. Although microencapsulated stromal tumors and granulosa cell tumors have no intracellular lipids, and Oil red O is negative, there can also be intracellular lipids in thecal cell tumors. Meanwhile, there was no significant difference in immunohistochemical results between SSTs and other ovarian sexual cord stromal tumours, because SF-1, calretinin and inhibin were also expressed in these tumours. Therefore, the positive expression of TFE3 immunohistochemistry is of great significance in the diagnosis and differential diagnosis of sclerosing stromal tumors [5,7,11,14−16] . Because there is no abnormality of TFE3 gene in sclerosing stromal tumors, in order to explored the source of TFE3 positive cells, we analyzed the expression of TFE3 in normal ovaries. To our knowledge, this is the first study to demonstrate that TFE3 nuclei are expressed in the inner membrane cells of follicles, but not in the granulosa cell layer, outer membrane cell layer, or fibroblasts. However, it remains unclear whether the nuclear localization of TFE3 is involved in egg maturation and plays a role in intrathecal cells, which will be the focus of our next study. In our study, seven out of eight cases of SSTs expressed luteinized cells with moderate-to-severe staining of TFE3, but it was not expressed in the ovarian granulosa, theca, or microcystic stromal tumor. Moreover, the FISH analysis revealed that the TFE3 gene was not broken, indicating that there was no possibility of TFE3 fusion with other genes. These results are consistent with those previously reported [ 22 ] . Combined with the results of Chamberlain et al. [ 17 ] and Schoolmeester et al. [ 18 ] , these findings indicate that unlike ASPS, TFE3 overexpression in SST was not caused by genetic translocation, suggesting that other types of genetic alterations may be involved. Chamberlain et al. [ 21 ] proposed that the abnormal accumulation of TFE3 in the nucleus could be caused by the dysfunction of organelles or intracellular metabolic signalling pathways, thus leading to the accumulation of phagosomes typical of granulosa cell tumor of soft tissue (GrCTS) in the cytoplasm. Other studies have reported that TFE3 is involved in the regulation of lysosome/phagosome synthesis and the Golgi stress response [ 22 – 27 ] . Thus, aberrant TFE3 expression can result in degenerative changes due to its lysosomal or cytoplasmic accumulation in these tumours and may not be a unique event for SSTs. Tumours with abnormal TFE3 expression and gene fusion have some common morphological characteristics, such as abundant cytoplasm and obvious nucleoli. However, SSTs, GrCTSs and other tumours that only express TFE3 without TFE3 gene abnormalities have abundant cytoplasm, but no obvious cell nucleoli. These results indicate that the phenotype of cells is not caused by TFE3 genetic abnormalities. In conclusion, our results show that TFE3 is expressed in the inner theca cells of normal follicles. Meanwhile, this study also suggests that the immunohistochemical detection of TFE3 is helpful for the diagnosis of difficult cases of sclerosing stromal tumor (such as cystic SST). The Lipid-rich SST cells mimic the inner theca cells of normal ovary and express TFE3 without disrupting the gene structure. Declarations Acknowledgements We thank Guangzhou LBP Medicine Science & Technology Co. Ltd. (China) for providing the TFE3 isolation probe. Conflicts of interest The authors have no relevant conflicts of interest to disclose. Patient Anonymity and Informed Consent We ensured that the patients’ anonymity was carefully protected and verified that any experimental investigation with human subjects reported in the manuscript was performed with informed consent and followed all the guidelines for experimental investigation with human subjects required by the institution(s) with which all the authors are affiliated. Funding: This work was supported by the Youth Fund of the Third Affiliated Hospital of Guangzhou Medical University in China (NO.80022018Q14) and Guangdong Natural Science Foundation (2020A1515011389) References Chalvardjian A, Scully RE. Sclerosing stromal tumours of the ovary. Cancer . 1973; 31:664–670. Kaygusuz EI, Cesur S, Cetiner H, et al. Sclerosing stromal tumor in young women: Clinicopathologic and immunohistochemical spectrum. J Clin Diagn Res . 2013; 7:1932–1935. Özdemir Ö, Sarı ME, Şen E, et al. Sclerosing stromal tumor of the ovary: a case report and the review of literature. Niger Med J .2014; 55:432–437. Zhao Z, Yan L, Lv H, Liu H, Rong F. Sclerosing stromal tumor of the ovary in a postmenopausal woman with estrogen excess: A case report. Medicine (Baltimore). 2019;98:47(e18171). Liou JH, Su TC, Hsu JC. Meigs' syndrome with elevated serum cancer antigen 125 levels in a case of ovarian sclerosing stromal tumor. Taiwan J Obstet Gynecol . 2011;50(2):196-200. Katsube Y, Iwaoki Y, Silverberg SG, Fujiwara A. Sclerosing stromal tumor of the ovary associated with endometrial adenocarcinoma: a case report. Gynecol Oncol. 1988;29(3): 392-398. Joja I, Okuno K, Tsunoda M, Takeda Y, Sugita K, Mizutani Y, Miyagi Y, Kudo T, Notohara K, Hiraki Y. Sclerosing stromal tumor of the ovary: US, MR, and dynamic MR findings. J Comput Assist Tomogr . 2001; 25: 201–206. Ihara N, Togashi K, Todo G, Nakai A, Kojima N, Ishigaki T, Suginami N, Kinoshita M, Shintaku M. Sclerosing stromal tumor of the ovary: MRI. J Comput Assist Tomogr . 1999; 23: 555–557. Matsubayashi R, Matsuo Y, Doi J, Kudo S, Matsuguchi K, Sugimori H. Sclerosing stromal tumor of the ovary: radiologic findings. Eur Radiol. 1999; 9: 1335–1338. Deval B, Rafii A, Darai E, Hugol D, Buy JN. Sclerosing stromal tumor of the ovary: color Doppler findings. Ultrasound Obstet Gynecol . 2003; 22: 531–534. Kyle M. Devins, Robert H. Young, Jaclyn C. Watkins. Sclerosing Stromal Tumor: A Clinicopathologic Study of 100 Cases of a Distinctive Benign Ovarian Stromal Tumor Typically Occurring in the Young. Histopathology . 2021; DOI: 10.1111/his.14554. Lam RMY, Geittmann P. Sclerosing stromal tumor of the ovary: The light, electron microscopic and enzyme histochemical study. Int J Gynecol Pathol . 1988; 7:280–290. Santini D, Ceccarelli C, Leone D, et al. Smooth muscle differentiation in normal human ovaries, ovarian stromal hyperplasia and ovarian granulosa-stroma cell tumours. Mod Pathol. 1995; 8:25-30. Saitoh A, Tsutsumi Y, Osamura Y, Watanabe K. Sclerosing stromal tumor of the ovary: immunohistochemical and electron microskopic demonstration of smooth muscle differantiation. Arch Pathol Lab Med. 1989; 113:372-376 Kommoss F, Oliva E, Bhan K, Young RH, Scully RE. Inhibin expression in ovarian tumours and tumor-like lesion: an immunohistochemical study. Mod Pathol . 1998; 11:656-661. Kawauchi S, Tsujı T, Kaku T, et al. Sclerosing stromal tumor of the ovary: a clinicopathologic, immunohistochemical, ultrastructual and cytogenetic analysis with special reference to its vasculature. Am J Surg Pathol . 1998; 22:83-92. Kaygusuz EI, Cesur S, Cetiner H, Yavuz H, Koc N. Sclerosing stromal tumour in young women: clinicopathologic and immunohistochemical spectrum. J Clin Diagn Res. 2013;7(9):1932-1935. McCluggage WG, Maxwell P. Immunohistochemical staining for calretinin is useful in the diagnosis of ovarian sex-cord stromal tumours. Histopathology . 2001; 38:403-408. Oliva E, Garcia-Miralles N, Vu Q Young RH. CD10 expression in pure stromal and sex cord-stromal tumours of the ovary: an immunohistochemical analysis of 101 cases. Int J Gynecol Pathol . 2007; 26:359-367. Stewart CJR, Nandini CL, Richmond JA. Value of A 103 (melan-A) immunostaining in the differantial diagnosis of ovarian sex cord-stromal tumours. J Clin Pathol . 2000; 53:206-211. Busam KJ, Iversen K, Coplan KA, et al. Immunoreactivity for A103, an antibody to melan-A (mart-1), in adrenocortical and other steroid tumours. Am J Surg Pathol . 1998; 22:57-63. Park CK, Kim HS. Clinicopathological Characteristics of Ovarian Sclerosing Stromal Tumor with an Emphasis on TFE3 Overexpression. Anticancer Res . 2017; 37: 5441-5447. Kim SH, Da Cruz Paula A, Basili T, Dopeso H, Bi R, Pareja F, da Silva EM, Gularte-Mérida R. Identification of recurrent FHL2-GLI2 oncogenic fusion in sclerosing stromal tumours of the ovary. Nat Commun . 2020; 11:44. Jia Q, Cao H, Shen D, Li S, Yan L, Chen C, Xing S, Dou F. Quercetin protects against atherosclerosis by regulating the expression of PCSK9, CD36, PPARγ, LXRα and ABCA1. Int J Mol Med. 2019 Sep;44(3):893-902. artina JA, Diab HI, Lishu L, et al. The nutrient-responsive transcription factor TFE3 promotes autophagy, lysosomal biogenesis, and clearance of cellular debris. Sci Signal. 2014;7:ra9. erera RM, Di Malta C, Ballabio A. MiTF/TFE family of transcription factors, lysosomes, and cancer. Annu Rev Cancer Biol . 2019; 3:203–222. Sun G, Chen J, Liang J, Yin X, Zhang M, Yao J, et al. Integrated exome and RNA sequencing of TFE3-translocation renal cell carcinoma. Nat Commun. 2021; 12:5262. Fang R, Wang X, Xia Q, et al. Nuclear translocation of ASPL-TFE3 fusion protein creates favorable metabolism by mediating autophagy in translocation renal cell carcinoma. Oncogene . 2021;40(18):3303-3317. O'Sullivan Coyne G, Naqash AR, Sankaran H, Chen AP. Advances in the management of alveolar soft part sarcoma. Curr Probl Cancer. 2021; 45(4):100775. Monga V, Skubitz KM, Maliske S, Mott SL, Dietz H, Hirbe AC, Van Tine BA, Oppelt P, Okuno S. A retrospective analysis of the efficacy of immunotherapy in metastatic soft-tissue sarcomas. Cancers (Basel) 2020 Jul 11;12(7): 1873. Antonescu CR, Le Loarer F, Mosquera J-M, et al. Novel YAP1-TFE3 fusion defines a distinct subset of epithelioid hemangioendothelioma. Genes Chromosomes Cancer. 2013; 52:775–84. Schoolmeester JK, Dao LN, Sukov WR, et al. TFE3 translocation-associated perivascular epithelioid cell neoplasm (PEComa) of the gynecologic tract: morphology, immunophenotype, differential diagnosis. Am J Surg Pathol . 2015; 39:394–404. Suurmeijer AJH, Song W, Sung Y-S, et al. Novel recurrent PHF1-TFE3 fusions in ossifying fibromyxoid tumours. Genes Chromosomes Cancer. 2019; 58:643–9. Panagopoulos I, Gorunova L, Lund-Iversen M, et al. Fusion of the Genes PHF1 and TFE3 in Malignant Chondroid Syringoma. Cancer Genomics Proteomics . 2019; 16:345–51. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Published Journal Publication published 01 Aug, 2023 Read the published version in Journal of Ovarian Research → Version 1 posted Editorial decision: Major revision 21 Feb, 2023 Editor assigned by journal 13 Feb, 2023 Submission checks completed at journal 10 Feb, 2023 First submitted to journal 08 Feb, 2023 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-2563971","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":175030086,"identity":"ed31f94b-6d87-4320-9f8c-c751600969a4","order_by":0,"name":"Li Zhao","email":"","orcid":"","institution":"The Third Affiliated Hospital of Guangzhou Medical University","correspondingAuthor":false,"prefix":"","firstName":"Li","middleName":"","lastName":"Zhao","suffix":""},{"id":175030087,"identity":"9b2f4502-7a03-4a3d-ad37-29c9cb005bd1","order_by":1,"name":"Yan Zhou","email":"","orcid":"","institution":"Affiliated Cancer Hospital \u0026 Institute of Guangzhou Medical University","correspondingAuthor":false,"prefix":"","firstName":"Yan","middleName":"","lastName":"Zhou","suffix":""},{"id":175030088,"identity":"cafbaf6b-1347-4eee-9ad3-07a526c25326","order_by":2,"name":"Yuping Liu","email":"","orcid":"","institution":"The Fourth Affiliated Hospital of Guangzhou Medical University","correspondingAuthor":false,"prefix":"","firstName":"Yuping","middleName":"","lastName":"Liu","suffix":""},{"id":175030091,"identity":"5db20bf2-882b-4585-936b-c086ec0d31c5","order_by":3,"name":"Qiuping Luo","email":"","orcid":"","institution":"The Third Affiliated Hospital of Guangzhou Medical University","correspondingAuthor":false,"prefix":"","firstName":"Qiuping","middleName":"","lastName":"Luo","suffix":""},{"id":175030093,"identity":"850b58c2-b1ca-4eb0-910f-b8c06017aba9","order_by":4,"name":"Qingping Jiang","email":"","orcid":"","institution":"The Third Affiliated Hospital of Guangzhou Medical University","correspondingAuthor":false,"prefix":"","firstName":"Qingping","middleName":"","lastName":"Jiang","suffix":""},{"id":175030094,"identity":"cbfb98d7-03ab-49b8-aa1c-07af09626af6","order_by":5,"name":"Hui Wang","email":"","orcid":"","institution":"Affiliated Cancer Hospital \u0026 Institute of Guangzhou Medical University","correspondingAuthor":false,"prefix":"","firstName":"Hui","middleName":"","lastName":"Wang","suffix":""},{"id":175030097,"identity":"7dd270f3-8354-480b-9404-78edc24cb272","order_by":6,"name":"Na Wang","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA40lEQVRIiWNgGAWjYPACCR429v6PDxIqbIjWYiHHx3PA2ODBmTSitVQYy0kkmEk+bDtEWK3B8bOHXxfUSCS2SSSkVSSwHWDgb+9OwK/lTF6a9YxjQC08D47dSOC5wyBx5uwGvFrMDuSYGfOwAbWwJ7bdSJB4xmAgkUtAy/k3QC3/gFoYktkKEgwOE6HlRo7xY942CWM2jjQ2hoQEIrTY33hjxszbJyHHxnOGWSLhQBoPQb9I9ucYf+b5Vscj397D+PHnPxs5/vZe/FqAgE0CmcdDSDkIMH8gRtUoGAWjYBSMYAAAfFRIRtgsKLsAAAAASUVORK5CYII=","orcid":"","institution":"The Third Affiliated Hospital of Guangzhou Medical University","correspondingAuthor":true,"prefix":"","firstName":"Na","middleName":"","lastName":"Wang","suffix":""}],"badges":[],"createdAt":"2023-02-08 10:14:28","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-2563971/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-2563971/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1186/s13048-023-01241-y","type":"published","date":"2023-08-01T21:52:29+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":32877483,"identity":"ccc80e1c-8e06-4748-bebf-0472dbe8e71f","added_by":"auto","created_at":"2023-02-13 23:34:10","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":2044737,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eOne cystic case of SST. \u003c/strong\u003eThe cut surface of SST was cystic (A). The cellular foci were consistedof roundcells admixed with spindle cells (B and C). Oil Red Ostaining was positive in lipid-rich cells (D). The tumor cells weakly expressed inhibin (E). The round cells and spindle cells did notexpress desmin (F). The short spindle cells in the tumor expressingSMA (G). TFE3 was diffuse positive in the lipid-rich cells (H).\u003c/p\u003e","description":"","filename":"Figure1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-2563971/v1/6dde5ae7c47369142587dbb7.jpg"},{"id":32876588,"identity":"6c56b22a-712e-42cd-a0fb-cc268afe5670","added_by":"auto","created_at":"2023-02-13 23:26:10","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":2237365,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eExpression of TFE3 in different types of sex cord stromal tumours. \u003c/strong\u003eThe cut surfaces of SST were solid with yellow (A). The cytoplasm of most cells in the lobules is rich, eosinophilic and granular (×200) (B). Cytoplasm-rich cells express TFE3 (C). The cut surface of the adult granulosa cell tumor is nodular and light yellow (D). Granulosa cells usually have scanty cytoplasm and pale, uniform, angular to oval, often grooved nuclei that are typically arranged haphazardly to each other, and typical Call-Exner bodies can be seen (E). TFE3 is not expressed in AGCT (F). Thecoma. The typical sectioned surface of a thecoma showing a yellow appearance (G). Thecoma. A high-power view shows the characteristic appreciable pale grey cytoplasm, ill-defined cytoplasmic membranes, and scattered collagen bundles (H). TFE3 is not expressed in the tumor cells of thecoma (I). Microcystic stromal tumor. The sectioned surface showing a solid, yellow appearance (J). Microcystic stromal tumor. Characteristic small cysts and hyaline plaques are seen (K). The tumor cells do not express TFE3 (L).\u003c/p\u003e","description":"","filename":"Figure2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-2563971/v1/81740b62d666e2a345bc3522.jpg"},{"id":32876589,"identity":"8fb963d6-e456-4373-9136-79f952018eac","added_by":"auto","created_at":"2023-02-13 23:26:10","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":2358827,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eNormal follicular tissue\u003c/strong\u003e. The follicle was observed at low magnification (×40) (A) and high magnification (×200) (B). Reticular fibers surrounded the inner theca cells and were absent around granulosa cells (C). CD10 was negative (D). The inner theca cells and granulosa cells expressed calretinin and inhibin (E and F). SMA was mainly expressed in the outer theca cell layer (G). TFE3 was expressed in the inner theca cells (H).\u003c/p\u003e","description":"","filename":"Figure3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-2563971/v1/1fe45a612cddfcf2370f09dc.jpg"},{"id":32876590,"identity":"58d7d69d-5dbc-474e-82ee-536396bf144a","added_by":"auto","created_at":"2023-02-13 23:26:10","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":147629,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eThe results of FISH. There was no separation of TFE3 in SST.\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"Figure4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-2563971/v1/6d15c50c96c9628ee9063fac.jpg"},{"id":44735515,"identity":"f61cf307-535c-43f5-9754-3c6b5cb73c87","added_by":"auto","created_at":"2023-10-16 22:25:41","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1146076,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-2563971/v1/818f68db-f6da-4f48-b27d-bfdd73ef3b20.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"TFE3 is a Novel Biomarker of Ovarian Sclerosing Stromal Tumours","fulltext":[{"header":"Introduction","content":"\u003cp\u003eSclerosing stromal tumour (SST) is a rare sex cord-stromal ovarian tumour that was first reported by Chalvaridjian and Scully in 1973 \u003csup\u003e[\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]\u003c/sup\u003e. SST occurs predominantly in young women of 20\u0026ndash;30 years of age \u003csup\u003e[\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]\u003c/sup\u003e, and its clinical symptoms include pelvic pain, menstrual irregularity \u003csup\u003e[\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]\u003c/sup\u003e, and nonspecific symptoms associated with ovarian cysts. In a few cases, patients had elevated serum levels of CA125 \u003csup\u003e[\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]\u003c/sup\u003e. However, the levels of hormones in these patients were not affected. Focal adenoid hyperplasia was diagnosed in only 1 of the 10 cases, but endometrial biopsy could not be performed in this case \u003csup\u003e[\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]\u003c/sup\u003e. In addition, a case of ovarian SST complicated by endometrial adenocarcinoma has been reported \u003csup\u003e[\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eCharacteristic feature of SST is the abundance of blood vessels in the nodules. Computed tomography (CT) and magnetic resonance imaging (MRI) have been employed to diagnose SSTs, particularly to assess their vascularization \u003csup\u003e[\u003cspan additionalcitationids=\"CR8\" citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]\u003c/sup\u003e. New ultrasound technologies have been developed to facilitate the exploration of adnexal masses, such as the diagnosis of tissue vascularization via colour Doppler \u003csup\u003e[\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]\u003c/sup\u003e. This blood flow feature of SST is relatively unique, which is helpful to differentiate from other sex cord stromal tumors and ovarian malignant tumors. This method not only obtains examination results quickly but also reduces the economic burden on patients.\u003c/p\u003e \u003cp\u003eIn general, SST is unilateral, mostly 5\u0026ndash;10 cm, with well-defined boundaries and sometimes a thin fibrous envelope. It is a grey‒white to grey‒yellow nodular oedema, and unilocular cystic cases are rare \u003csup\u003e[\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]\u003c/sup\u003e. According to previous studies, SST consists of three types of cells, lipid-rich cells, fibroblast-like cells, and undifferentiated stromal mesenchymal cells, with intermediate morphology showing different degrees of differentiation \u003csup\u003e[\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]\u003c/sup\u003e. In addition, some studies indicate that tumour cells in SST have the characteristics of muscle-like cells and express SMA or desmin \u003csup\u003e[\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]\u003c/sup\u003e. Growing evidence shows that cytoplasm-rich cells express calretinin and inhibin \u003csup\u003e[\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]\u003c/sup\u003e but do not express SMA, desmin, CK, and CK7 \u003csup\u003e[\u003cspan additionalcitationids=\"CR17\" citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e]\u003c/sup\u003e. In some cases, lipid-rich cells express CD10 \u003csup\u003e[\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]\u003c/sup\u003e and Melan A \u003csup\u003e[\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]\u003c/sup\u003e. These cells are similar to normal cells in the ovary but have not been clearly defined.\u003c/p\u003e \u003cp\u003eA study found that TFE3 was highly expressed in sclerosing stromal tumors and pointed out TFE3 is highly expressed in the nucleus of lutein cells and polygonal-to-round tumour cells in 7 out of 9 patients with SST \u003csup\u003e[\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]\u003c/sup\u003e. However, neither luteinized fibromas nor thecomas express appreciable levels of TFE3 \u003csup\u003e[\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]\u003c/sup\u003e. Fluorescence in situ hybridization (FISH) analysis revealed the presence of trisomy 12 in \u0026gt;\u0026thinsp;20% of SST cells \u003csup\u003e[\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]\u003c/sup\u003e. In 2020, using whole-exome, targeted capture, and RNA sequencing, Sarah et al. reported that 65% (17/26) of SST patients had recurrent FHL2-GLI2 fusion genes and 15% (4/26) had other GLI2 rearrangements \u003csup\u003e[\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e]\u003c/sup\u003e. Specifically, these genetic abnormalities were not detected in other types of sex cord-stromal tumours (n\u0026thinsp;=\u0026thinsp;48) and common cancers (n\u0026thinsp;=\u0026thinsp;9,950).\u003c/p\u003e \u003cp\u003eIn this study, the origin of SSTs was discussed. TFE3 immunohistochemical and molecular analyses were performed on SSTs and other types of ovarian sex cord-stromal tumours. TFE3 staining was used to distinguish SST from other sex cord-stromal tumours.\u003c/p\u003e"},{"header":"Materials And Methods","content":"\u003cp\u003e\u003cstrong\u003ePatients\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe records of\u0026nbsp;38\u0026nbsp;patients who had undergone surgical resection of\u0026nbsp;sex cord-stromal\u0026nbsp;ovarian tumours\u0026nbsp;at The Third Affiliated Hospital of Guangzhou Medical University (Guangzhou, China) between December 2013 and December 2021. The focus of\u0026nbsp;this study is sclerosing stromal tumor\u0026nbsp;(8 cases)\u0026nbsp;, and its main differential diagnosis is also involved, which\u0026nbsp;are\u0026nbsp;theca cell tumor and theca fibroma tumour\u0026nbsp;(9 cases)\u0026nbsp;, granulosa cell tumour\u0026nbsp;(20 cases), and microcystic stromal tumour \u0026nbsp;(1 case)in turn. The age of patients ranged from 17 to 70 years and they\u0026nbsp;had not received any pre-operative\u0026nbsp;therapy.\u003c/p\u003e\n\u003cp\u003eEthical approval was obtained\u0026nbsp;from\u0026nbsp;the institutional review board of the ethics committee.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eLipid stains (Oil Red O)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe biopsy samples were\u0026nbsp;placed\u0026nbsp;in\u0026nbsp;a tissue tek container (Sakura Finetek, CA, USA)\u0026nbsp;and\u0026nbsp;then filled with tissue tek OCT compound gel.\u0026nbsp;After\u0026nbsp;being cut into 7-\u0026micro;m slices, the samples were snap-frozen in liquid nitrogen and\u0026nbsp;stained with\u0026nbsp;Oil Red O\u0026nbsp;according to standard procedures.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe oil red O\u003csup\u003e【\u003c/sup\u003e\u003csup\u003e24\u003c/sup\u003e\u003csup\u003e】\u003c/sup\u003e fat staining method is usually used to detect the fat in tissues or cells. Oil red O is a fat-soluble dye, which is a strong fat solvent and fat dye, and can be highly dissolved in fat. Its dyeing principle is that oil red O can specifically adsorb with the neutral triglycerides, lipids and lipoproteins in tissues and cells to make fat dye. The solubility of dye in intracellular lipids is greater than that of solution.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eImmunohistochemistry\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;(IHC)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eCollected tumours or normal tissue after removal of paraffin as the experimental subject.\u0026nbsp;Consecutive 4-\u0026mu;m thick unstained sections for immunohistochemical staining, which was performed using the Leica automatic immunostaining device (Leica Microsystems, Inc.). Primary antibodies against\u0026nbsp;CD10\u0026nbsp;(1:100; no. 563871; DAKO; DK),\u0026nbsp;\u0026alpha;-inhibin\u0026nbsp;(1:100; no.\u0026nbsp;GT230202; CHN),\u0026nbsp;SMA\u0026nbsp;(1:50; no. MAB-0980; MXB; CHN),\u0026nbsp;desmin (1:300; no.\u0026nbsp;GT225202; Gene tech; CHN), TFE3 (1:100; no. ZA-0657; ZSGB-BIO; CHN), calretinin (1:100; no. ZM-0063; ZSGB-BIO; CHN), WT-1 (1:100; no. ZM-0269; ZSGB-BIO; CHN),\u0026nbsp;and EMA (1:300; no.\u0026nbsp;GM061302; Gene tech; CHN). Appropriate\u0026nbsp;positive and negative controls were simultaneously\u0026nbsp;stained to validate the staining method.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eImmunohistochemistry were conducted according to previously described methods.\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll slides were reviewed and scored independently by three pathologists. The pathologists were blinded to the experiment. The scoring method based on both the intensity (0, no staining; 1+, weak staining; 2+, medium staining; 3+, strong staining).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFluorescence in Situ Hybridization\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e(\u003c/strong\u003e\u003cstrong\u003eFISH\u003c/strong\u003e\u003cstrong\u003e)\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe TFE3 isolation probe was provided by Guangzhou LBP Medicine Science and Technology Co., Ltd. (China). Specific operations were performed according to the manufacturer\u0026rsquo;s protocol. The results showed that there was no fracture of TFE3 gene in females (2 yellow) and males (1 yellow). In females, 1 red 1 green 1 yellow, and in males, 1 red 1 green indicated that TFE3 had a balanced translocation and the gene was fused. In females, 1 red and 2 yellow, and in males, 1 red and 1 yellow, showed unbalanced translocation and fracture of TFE3 gene.\u003c/p\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eClinical findings\u003c/h2\u003e \u003cp\u003eAs shown in Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e, the age range of patients with SSTs was 17\u0026ndash;39 years old. The levels of hormones were normal in all patients. However, menstruation was irregular in cases 5 and 7. In case 5, an adnexal mass was found during physical examination, and no abnormality was found in the endometrium. Endometrial biopsy showed atypical hyperplasia, and an adnexal mass was found in case 7. In case 4, in which the patient presented with abdominal distension for 1-year, an adnexal tumor was found on B-ultrasound with peritoneal effusion. Adnexal tumours were found in other patients during physical examinations.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eClinical findings of SSTs\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"6\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePatient with SSTs\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eage\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003egross\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eSize (mm)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003emenstruation\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eendometrium\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCase 1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e23\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ecystic\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003enormal\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003enormal\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCase 2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e17\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003esolid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e14\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003enormal\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003enormal\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCase 3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e26\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003esolid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e160\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003enormal\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003enormal\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCase 4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e26\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003esolid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e45\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003enormal\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003enormal\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCase 5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e39\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003esolid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e80\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eirregular\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003enormal\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCase 6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e29\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003esolid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003enormal\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003enormal\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCase 7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e33\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003esolid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e50\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eirregular\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eatypical hyperlasia\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCase 8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e23\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003esolid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e25\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003enormal\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003enormal\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eGross Findings\u003c/h3\u003e\n\u003cp\u003eThe size of the eight tumours ranged from 14 to 160 mm. All tumours were well-circumscribed nodules, and except for one cystic case (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eA), all tumours were solid (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eA), which ranged from soft to tough. The cut surfaces of the solid tumours were typically white, slightly leafy, with scattered yellow nodules and were most abundant at the periphery. Focal haemorrhage was observed in one patient. In case 1, the tumour was cystic, with cysts containing thick gelatinous material (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eA).\u003c/p\u003e \n\u003ch3\u003eHistologic Findings And Immunohistochemical Results Of Sclerosing Stromal Tumours\u003c/h3\u003e\n\u003cp\u003eAll tumours, including cystic tumours, showed overt, often discrete, cellular and hypocellular regions, resulting in a pseudolobular appearance (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eB). The cellular areas and lower cellular intervals were always collagenous, loose collagenous, and markedly oedematous. Cellular foci consisted of a mixture of round and spindle cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eC). The cytoplasm of the former was pale, eosinophilic, vacuolated, or foamy to varying degrees (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eC). Thin-walled, dilated, focally branching (\u0026ldquo;staghorn\u0026rdquo;) blood vessels were prominently found in all cases, and markedly conspicuous labyrinthine and haemangiopericytoma-like tumours were evident in some cases (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eB). Oil Red O staining revealed that the cytoplasm was rich in lipids but not in mucus (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eD). Spindle cells expressed SMA (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eE) but not desmin (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eF). CD34 staining showed abundant blood vessels in the cell lobules (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eG). Only cells with abundant cytoplasm in the tumours expressed TFE3, and nuclear expression was moderately and strongly positive (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eH).\u003c/p\u003e\n\u003ch3\u003eExpression Of Tfe3 In Ovarian Sex Cord Stromal Tumours\u003c/h3\u003e\n\u003cp\u003eAs shown in Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e, TFE3 was highly expressed in sclerosing stromal tumours (7/8), weakly expressed in two cases of thecoma (2/9), and not expressed in other tumours (0/21, including 20 cases of granulosa cell tumours and one case of microcystic stromal tumours). As shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e, TFE3 was mainly expressed in sclerosing stromal tumours (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eA, B, C) but not in ovarian granulosa cell tumours (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eD, E, F), thecoma (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eG, H, I), or microcystic stromal tumours (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eJ, K, L). Moreover, TFE3 mainly expressed in the nucleus of cells with abundant cytoplasm, and the staining intensity was medium to strong (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eC).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eImmunohistochemical Results of TFE3\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"7\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTumor Type\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eNo. of casess\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3+\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003e2+\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1+\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eTotal(%)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTheca cell tumor and theca fibroma tumor\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e22.2%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGranulosa cell tumor\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003emicrocystic stromal tumor\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSclerosing stromal tumor\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e87.5%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e\n\u003ch3\u003eImmunophenotypes Of Theca Cells In Normal Follicles\u003c/h3\u003e\n\u003cp\u003eNo literature has reported the expression mode of TFE3 in normal ovary. In order to explore the source of TFE3 immunohistochemistry positive cells in sclerosing stromal tumors, we collected 10 normal ovarian tissues. Normal ovarian follicles were obtained from specimens removed due to ovarian endometriosis or teratoma. We tested a total of 30 follicles. Normal follicular structure was observed at low and high magnifications. The boundary between the inner and outer theca cell layers was not very clear, but the cytoplasm of inner theca cells is rich and clear (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eA and B). A mostly consistent immunophenotype was observed in the rich cytoplasm cells: each demonstrated diffuse TFE3, TFE3 expressed in the inner theca cells, but not in the granular layer, outer theca layer, or fibrous tissue in normal follicles (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eH).\u003c/p\u003e \u003cp\u003eFISH detection\u003c/p\u003e \u003cp\u003eUsing the separation probe of TFE3, TFE3 was detected by FISH in 7 cases of SSTs expressing TFE3. The results showed no separation of TFE3 in these seven cases (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e).\u003c/p\u003e "},{"header":"Discussion","content":"\u003cp\u003eTFE3 is located at Xp11.23, and its protein belongs to the microphthalmia-associated transcription factor (MiTF) family, which plays an important role in the regulation of lysosomal biogenesis and autophagy \u003csup\u003e[\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]\u003c/sup\u003e. The TFE/MiTF family consists of four important members: (i) TFEB, (ii) TFEC, (iii) TFE3, and (iv) MITF \u003csup\u003e[\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e]\u003c/sup\u003e. By searching the human protein atlas webtool (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.proteinatlas.org/\u003c/span\u003e\u003cspan address=\"https://www.proteinatlas.org/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e), TFE3 was found to be expressed in adipose tissue, urinary bladder, ovary, testis, breast, etc. TFE3 protein can be transported to the nucleus when cells are starved and/or stressed. The expression and activity of TFE3 are upregulated in many types of human cancers, and are associated with enhanced proliferation and motility of cancer cells. The main tumours related to TFE3 gene fusion include epithelioid haemangioendothelioma, alveolar soft-part sarcoma (ASPS), renal cell carcinoma, malignant chondroid syringoma, rare ossifying fibromyxoid tumours, and perivascular epithelioid cell tumor \u003csup\u003e[\u003cspan additionalcitationids=\"CR28 CR29 CR30 CR31 CR32 CR33\" citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e]\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003ePark CK and Kim HS had reported that TFE3 was expressed in sclerosing stromal tumors, but there was no abnormality in TFE3 gene \u003csup\u003e[\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]\u003c/sup\u003e. Our research got the same result that TFE3 was specifically expressed in sclerosing stromal tumours, but not in other sex cord stromal tumours. Moreover, TFE3 was specifically expressed in luteinized cells, but not in other two cells.\u003c/p\u003e \u003cp\u003eSclerosing stromal tumours often occur in young women, and a few cases have the secretion of estrogen and/or androgen. The clinical results of all cases were benign. This study analyzed 8 cases of sclerosing stromal tumours, ranging in age from 17 to 39 years old. No hormone abnormality was found clinically, but 2 patients had irregular menstruation. Follow-up results showed no recurrence.The lobulated structure and three types of cell composition are the unique morphological characteristics of ovarian sclerosing stromal tumours. Due to the young age of SST and good prognosis, accurate pathological diagnosis is very important in order to avoid over-treatment. In most cases, we can obtain a positive pathological diagnosis based on the above findings. However, due to the diversity and variety of ovarian sex cord stromal tumours, we need more specific indicators to assist the pathologist in diagnosis in a few cases.For example, the cystic case in Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e.\u003c/p\u003e \u003cp\u003eOur experimental results show that the luteinized tumor cells in sclerosing stromal tumors have abundant intracellular lipids, and Oil red O staining is obvious. Although microencapsulated stromal tumors and granulosa cell tumors have no intracellular lipids, and Oil red O is negative, there can also be intracellular lipids in thecal cell tumors. Meanwhile, there was no significant difference in immunohistochemical results between SSTs and other ovarian sexual cord stromal tumours, because SF-1, calretinin and inhibin were also expressed in these tumours. Therefore, the positive expression of TFE3 immunohistochemistry is of great significance in the diagnosis and differential diagnosis of sclerosing stromal tumors \u003csup\u003e[5,7,11,14\u0026minus;16]\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eBecause there is no abnormality of TFE3 gene in sclerosing stromal tumors, in order to explored the source of TFE3 positive cells, we analyzed the expression of TFE3 in normal ovaries. To our knowledge, this is the first study to demonstrate that TFE3 nuclei are expressed in the inner membrane cells of follicles, but not in the granulosa cell layer, outer membrane cell layer, or fibroblasts. However, it remains unclear whether the nuclear localization of TFE3 is involved in egg maturation and plays a role in intrathecal cells, which will be the focus of our next study.\u003c/p\u003e \u003cp\u003eIn our study, seven out of eight cases of SSTs expressed luteinized cells with moderate-to-severe staining of TFE3, but it was not expressed in the ovarian granulosa, theca, or microcystic stromal tumor. Moreover, the FISH analysis revealed that the TFE3 gene was not broken, indicating that there was no possibility of TFE3 fusion with other genes. These results are consistent with those previously reported \u003csup\u003e[\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]\u003c/sup\u003e. Combined with the results of Chamberlain et al. \u003csup\u003e[\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]\u003c/sup\u003e and Schoolmeester et al. \u003csup\u003e[\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e]\u003c/sup\u003e, these findings indicate that unlike ASPS, TFE3 overexpression in SST was not caused by genetic translocation, suggesting that other types of genetic alterations may be involved. Chamberlain et al. \u003csup\u003e[\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]\u003c/sup\u003e proposed that the abnormal accumulation of TFE3 in the nucleus could be caused by the dysfunction of organelles or intracellular metabolic signalling pathways, thus leading to the accumulation of phagosomes typical of granulosa cell tumor of soft tissue (GrCTS) in the cytoplasm. Other studies have reported that TFE3 is involved in the regulation of lysosome/phagosome synthesis and the Golgi stress response \u003csup\u003e[\u003cspan additionalcitationids=\"CR23 CR24 CR25 CR26\" citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]\u003c/sup\u003e. Thus, aberrant TFE3 expression can result in degenerative changes due to its lysosomal or cytoplasmic accumulation in these tumours and may not be a unique event for SSTs.\u003c/p\u003e \u003cp\u003eTumours with abnormal TFE3 expression and gene fusion have some common morphological characteristics, such as abundant cytoplasm and obvious nucleoli. However, SSTs, GrCTSs and other tumours that only express TFE3 without TFE3 gene abnormalities have abundant cytoplasm, but no obvious cell nucleoli. These results indicate that the phenotype of cells is not caused by TFE3 genetic abnormalities.\u003c/p\u003e \u003cp\u003eIn conclusion, our results show that TFE3 is expressed in the inner theca cells of normal follicles. Meanwhile, this study also suggests that the immunohistochemical detection of TFE3 is helpful for the diagnosis of difficult cases of sclerosing stromal tumor (such as cystic SST). The Lipid-rich SST cells mimic the inner theca cells of normal ovary and express TFE3 without disrupting the gene structure.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe thank Guangzhou LBP Medicine Science \u0026amp; Technology Co. Ltd. (China) for providing the TFE3 isolation probe.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflicts of interest\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors have no relevant conflicts of interest to disclose.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ePatient Anonymity and Informed Consent\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe ensured that the patients\u0026rsquo; anonymity was carefully protected and verified that any experimental investigation with human subjects reported in the manuscript was performed with informed consent and followed all the guidelines for experimental investigation with human subjects required by the institution(s) with which all the authors are affiliated.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding: \u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis work was supported by the Youth Fund of the Third Affiliated Hospital of Guangzhou Medical University in China (NO.80022018Q14) and Guangdong Natural Science Foundation (2020A1515011389)\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n \u003cli\u003eChalvardjian A, Scully RE. Sclerosing stromal tumours of the ovary. \u003cem\u003eCancer\u003c/em\u003e. 1973; 31:664\u0026ndash;670.\u003c/li\u003e\n \u003cli\u003eKaygusuz EI, Cesur S, Cetiner H, et al. Sclerosing stromal tumor in young women: Clinicopathologic and immunohistochemical spectrum. \u003cem\u003eJ Clin Diagn Res\u003c/em\u003e. 2013; 7:1932\u0026ndash;1935.\u003c/li\u003e\n \u003cli\u003e\u0026Ouml;zdemir \u0026Ouml;, Sarı ME, Şen E, et al. Sclerosing stromal tumor of the ovary: a case report and the review of literature. \u003cem\u003eNiger Med J\u003c/em\u003e.2014; 55:432\u0026ndash;437.\u003c/li\u003e\n \u003cli\u003eZhao Z, Yan L, Lv H, Liu H, Rong F. Sclerosing stromal tumor of the ovary in a postmenopausal woman with estrogen excess: A case report.\u003cem\u003e\u0026nbsp;Medicine (Baltimore).\u003c/em\u003e2019;98:47(e18171).\u003c/li\u003e\n \u003cli\u003eLiou JH, Su TC, Hsu JC. Meigs\u0026apos; syndrome with elevated serum cancer antigen 125 levels in a case of ovarian sclerosing stromal tumor.\u003cem\u003e\u0026nbsp;Taiwan J Obstet Gynecol\u003c/em\u003e. 2011;50(2):196-200.\u003c/li\u003e\n \u003cli\u003eKatsube Y, Iwaoki Y, Silverberg SG, Fujiwara A. Sclerosing stromal tumor of the ovary associated with endometrial adenocarcinoma: a case report. \u003cem\u003eGynecol Oncol.\u003c/em\u003e 1988;29(3): 392-398.\u003c/li\u003e\n \u003cli\u003eJoja I, Okuno K, Tsunoda M, Takeda Y, Sugita K, Mizutani Y, Miyagi Y, Kudo T, Notohara K, Hiraki Y. Sclerosing stromal tumor of the ovary: US, MR, and dynamic MR findings. \u003cem\u003eJ Comput Assist Tomogr\u003c/em\u003e. 2001; 25: 201\u0026ndash;206.\u003c/li\u003e\n \u003cli\u003eIhara N, Togashi K, Todo G, Nakai A, Kojima N, Ishigaki T, Suginami N, Kinoshita M, Shintaku M. Sclerosing stromal tumor of the ovary: MRI. \u003cem\u003eJ Comput Assist Tomogr\u003c/em\u003e. 1999; 23: 555\u0026ndash;557.\u003c/li\u003e\n \u003cli\u003eMatsubayashi R, Matsuo Y, Doi J, Kudo S, Matsuguchi K, Sugimori H. Sclerosing stromal tumor of the ovary: radiologic findings.\u003cem\u003e\u0026nbsp;Eur Radiol.\u003c/em\u003e 1999; 9: 1335\u0026ndash;1338.\u003c/li\u003e\n \u003cli\u003eDeval B, Rafii A, Darai E, Hugol D, Buy JN. Sclerosing stromal tumor of the ovary: color Doppler findings. \u003cem\u003eUltrasound Obstet Gynecol\u003c/em\u003e. 2003; 22: 531\u0026ndash;534.\u003c/li\u003e\n \u003cli\u003eKyle M. Devins, Robert H. Young, Jaclyn C. Watkins. Sclerosing Stromal Tumor: A Clinicopathologic Study of 100 Cases of a Distinctive Benign Ovarian Stromal Tumor Typically Occurring in the Young. \u003cem\u003eHistopathology\u003c/em\u003e. 2021; DOI: 10.1111/his.14554.\u003c/li\u003e\n \u003cli\u003eLam RMY, Geittmann P. Sclerosing stromal tumor of the ovary: The light, electron microscopic and enzyme histochemical study. \u003cem\u003eInt J Gynecol Pathol\u003c/em\u003e. 1988; 7:280\u0026ndash;290.\u003c/li\u003e\n \u003cli\u003eSantini D, Ceccarelli C, Leone D, et al. Smooth muscle differentiation in normal human ovaries, ovarian stromal hyperplasia and ovarian granulosa-stroma cell tumours. \u003cem\u003eMod Pathol.\u003c/em\u003e 1995; 8:25-30.\u003c/li\u003e\n \u003cli\u003eSaitoh A, Tsutsumi Y, Osamura Y, Watanabe K. Sclerosing stromal tumor of the ovary: immunohistochemical and electron microskopic demonstration of smooth muscle differantiation. \u003cem\u003eArch Pathol Lab Med.\u003c/em\u003e 1989; 113:372-376\u003c/li\u003e\n \u003cli\u003eKommoss F, Oliva E, Bhan K, Young RH, Scully RE. Inhibin expression in ovarian tumours and tumor-like lesion: an immunohistochemical study. \u003cem\u003eMod Pathol\u003c/em\u003e. 1998; 11:656-661.\u003c/li\u003e\n \u003cli\u003eKawauchi S, Tsujı T, Kaku T, et al. Sclerosing stromal tumor of the ovary: a clinicopathologic, immunohistochemical, ultrastructual and cytogenetic analysis with special reference to its vasculature.\u003cem\u003e\u0026nbsp;Am J Surg Pathol\u003c/em\u003e. 1998; 22:83-92.\u003c/li\u003e\n \u003cli\u003eKaygusuz EI, Cesur S, Cetiner H, Yavuz H, Koc N. Sclerosing stromal tumour in young women: clinicopathologic and immunohistochemical spectrum.\u003cem\u003e\u0026nbsp;J Clin Diagn Res.\u003c/em\u003e 2013;7(9):1932-1935.\u003c/li\u003e\n \u003cli\u003eMcCluggage WG, Maxwell P. Immunohistochemical staining for calretinin is useful in the diagnosis of ovarian sex-cord stromal tumours. \u003cem\u003eHistopathology\u003c/em\u003e. 2001; 38:403-408.\u003c/li\u003e\n \u003cli\u003eOliva E, Garcia-Miralles N, Vu Q Young RH. CD10 expression in pure stromal and sex cord-stromal tumours of the ovary: an immunohistochemical analysis of 101 cases.\u003cem\u003e\u0026nbsp;Int J Gynecol Pathol\u003c/em\u003e. 2007; 26:359-367.\u003c/li\u003e\n \u003cli\u003eStewart CJR, Nandini CL, Richmond JA. Value of A 103 (melan-A) immunostaining in the differantial diagnosis of ovarian sex cord-stromal tumours. \u003cem\u003eJ Clin Pathol\u003c/em\u003e. 2000; 53:206-211.\u003c/li\u003e\n \u003cli\u003eBusam KJ, Iversen K, Coplan KA, et al. Immunoreactivity for A103, an antibody to melan-A (mart-1), in adrenocortical and other steroid tumours. \u003cem\u003eAm J Surg Pathol\u003c/em\u003e. 1998; 22:57-63.\u003c/li\u003e\n \u003cli\u003ePark CK, Kim HS. Clinicopathological Characteristics of Ovarian Sclerosing Stromal Tumor with an Emphasis on TFE3 Overexpression. \u003cem\u003eAnticancer Res\u003c/em\u003e. 2017; 37: 5441-5447.\u003c/li\u003e\n \u003cli\u003eKim SH, Da Cruz Paula A, Basili T, Dopeso H, Bi R, Pareja F, da Silva EM, Gularte-M\u0026eacute;rida R. Identification of recurrent FHL2-GLI2 oncogenic fusion in sclerosing stromal tumours of the ovary. \u003cem\u003eNat Commun\u003c/em\u003e. 2020; 11:44.\u003c/li\u003e\n \u003cli\u003eJia Q, Cao H, Shen D, Li S, Yan L, Chen C, Xing S, Dou F. Quercetin protects against atherosclerosis by regulating the expression of PCSK9, CD36, PPAR\u0026gamma;, LXR\u0026alpha; and ABCA1. Int J Mol Med. 2019 Sep;44(3):893-902.\u003c/li\u003e\n \u003cli\u003eartina JA, Diab HI, Lishu L, et al. The nutrient-responsive transcription factor TFE3 promotes autophagy, lysosomal biogenesis, and clearance of cellular debris. \u003cem\u003eSci Signal.\u003c/em\u003e2014;7:ra9.\u003c/li\u003e\n \u003cli\u003eerera RM, Di Malta C, Ballabio A. MiTF/TFE family of transcription factors, lysosomes, and cancer. \u003cem\u003eAnnu Rev Cancer Biol\u003c/em\u003e. 2019; 3:203\u0026ndash;222.\u003c/li\u003e\n \u003cli\u003eSun G, Chen J, Liang J, Yin X, Zhang M, Yao J, et al. Integrated exome and RNA sequencing of TFE3-translocation renal cell carcinoma. \u003cem\u003eNat Commun.\u003c/em\u003e 2021; 12:5262.\u003c/li\u003e\n \u003cli\u003eFang R, Wang X, Xia Q, et al. Nuclear translocation of ASPL-TFE3 fusion protein creates favorable metabolism by mediating autophagy in translocation renal cell carcinoma. \u003cem\u003eOncogene\u003c/em\u003e. 2021;40(18):3303-3317.\u003c/li\u003e\n \u003cli\u003eO\u0026apos;Sullivan Coyne G, Naqash AR, Sankaran H, Chen AP. Advances in the management of alveolar soft part sarcoma. \u003cem\u003eCurr Probl Cancer.\u003c/em\u003e 2021; 45(4):100775.\u003c/li\u003e\n \u003cli\u003eMonga V, Skubitz KM, Maliske S, Mott SL, Dietz H, Hirbe AC, Van Tine BA, Oppelt P, Okuno S. A retrospective analysis of the efficacy of immunotherapy in metastatic soft-tissue sarcomas. \u003cem\u003eCancers (Basel)\u003c/em\u003e 2020 Jul 11;12(7): 1873.\u003c/li\u003e\n \u003cli\u003eAntonescu CR, Le Loarer F, Mosquera J-M, et al. Novel YAP1-TFE3 fusion defines a distinct subset of epithelioid hemangioendothelioma. \u003cem\u003eGenes Chromosomes Cancer.\u003c/em\u003e 2013; 52:775\u0026ndash;84.\u003c/li\u003e\n \u003cli\u003eSchoolmeester JK, Dao LN, Sukov WR, et al. TFE3 translocation-associated perivascular epithelioid cell neoplasm (PEComa) of the gynecologic tract: morphology, immunophenotype, differential diagnosis.\u003cem\u003e\u0026nbsp;Am J Surg Pathol\u003c/em\u003e. 2015; 39:394\u0026ndash;404.\u003c/li\u003e\n \u003cli\u003eSuurmeijer AJH, Song W, Sung Y-S, et al. Novel recurrent PHF1-TFE3 fusions in ossifying fibromyxoid tumours. \u003cem\u003eGenes Chromosomes Cancer.\u003c/em\u003e 2019; 58:643\u0026ndash;9.\u003c/li\u003e\n \u003cli\u003ePanagopoulos I, Gorunova L, Lund-Iversen M, et al. Fusion of the Genes PHF1 and TFE3 in Malignant Chondroid Syringoma.\u003cem\u003e\u0026nbsp;Cancer Genomics Proteomics\u003c/em\u003e. 2019; 16:345\u0026ndash;51.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"journal-of-ovarian-research","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"jovr","sideBox":"Learn more about [Journal of Ovarian Research](http://ovarianresearch.biomedcentral.com)","snPcode":"13048","submissionUrl":"https://submission.nature.com/new-submission/13048/3","title":"Journal of Ovarian Research","twitterHandle":"@BioMedCentral","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Ovarian Sclerosing Stromal Tumours, TFE3, Novel Biomarker","lastPublishedDoi":"10.21203/rs.3.rs-2563971/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-2563971/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eSclerosing stromal tumours of the ovary are benign and tend to occur in young women with lobular structures at low frequencies. Three types of cells, luteinized cells, short spindle myoid cells, and intermediate cells, are found in lobules, which are rich in blood vessels. Currently, immunohistochemistry and fluorescence in situ hybridization are used to detect normal follicles, sclerosing stromal tumours, granulosa cell tumours, and theca fibromas. Our research found the expression of transcription factor enhancer 3 (TFE3) was moderately and strongly positive in the inner thecal cell layer of normal follicles. It was expressed in seven out of eight sclerosing stromal tumours, mainly in luteinized cells, but not in 20 granulosa cell tumours and 1 microcystic stromal tumour. In nine cases of theca cell tumours and theca fibromas, TFE3 was not expressed, except in two cases of weak TFE3 expression. Eight cases of sclerosing stromal tumours were analysed by FISH using a TFE3 separation probe, and the results were negative. In a word, as a nuclear transcription protein, TFE3 was specifically expressed in sclerosing stromal tumours and could serve as a new marker for the diagnosis and differential diagnosis of sclerosing stromal tumours.\u003c/p\u003e","manuscriptTitle":"TFE3 is a Novel Biomarker of Ovarian Sclerosing Stromal Tumours","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2023-02-13 23:26:05","doi":"10.21203/rs.3.rs-2563971/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Major revision","date":"2023-02-21T14:55:57+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2023-02-14T03:02:18+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2023-02-10T10:10:36+00:00","index":"","fulltext":""},{"type":"submitted","content":"Journal of Ovarian Research","date":"2023-02-08T10:12:32+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"journal-of-ovarian-research","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"jovr","sideBox":"Learn more about [Journal of Ovarian Research](http://ovarianresearch.biomedcentral.com)","snPcode":"13048","submissionUrl":"https://submission.nature.com/new-submission/13048/3","title":"Journal of Ovarian Research","twitterHandle":"@BioMedCentral","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"e4fd61b6-e92b-485a-8e06-25849cde751c","owner":[],"postedDate":"February 13th, 2023","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2023-10-16T22:11:10+00:00","versionOfRecord":{"articleIdentity":"rs-2563971","link":"https://doi.org/10.1186/s13048-023-01241-y","journal":{"identity":"journal-of-ovarian-research","isVorOnly":false,"title":"Journal of Ovarian Research"},"publishedOn":"2023-08-01 21:52:29","publishedOnDateReadable":"August 1st, 2023"},"versionCreatedAt":"2023-02-13 23:26:05","video":"","vorDoi":"10.1186/s13048-023-01241-y","vorDoiUrl":"https://doi.org/10.1186/s13048-023-01241-y","workflowStages":[]},"version":"v1","identity":"rs-2563971","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-2563971","identity":"rs-2563971","version":["v1"]},"buildId":"_2-kVJe1T_tPrBINL-cwx","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}