The Role of Fibroblast Activation in Uterine Fibroid

Study Overview
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Brief Summary
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Uterine fibroids (UFs), also called uterine leiomyomas or myomas, are steroid hormone-responsive, benign tumors of the smooth muscle compartment (myometrium) of the uterus. They are the most common neoplasm affecting women in their reproductive age. It is estimated that up to 77% of women develop UF in their life. UFs are one of the leading causes of hospitalisations for gynaecological disorders and are the most frequent reason for hysterectomy. According to relevant literature, 40%-60% of all the hysterectomies performed are due to the presence of UFs.

Detailed Description
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Uterine fibroids (UFs) are steroid hormone-responsive, benign tumors of the smooth muscle compartment (myometrium) of the uterus .They are the most common neoplasm affecting women in their reproductive age. UFs are mainly composed of fibroid cells and a significant ECM component which principally consists of fibroblasts. Previous studies on the pathogenesis of uterine UF have mainly focused on the differentiation and proliferation of fibroid cells. However, the histologic features of fibroid tissue suggest that fibroblasts may play an important role in the generation of UF. Fibroblast activation protein (FAP), fibroblast-specific marker, is a 95 kDa cell surface glycoprotein. It is a type II transmembrane serine protease and a member of proline-specific proteases family. Recent studies showed that the high expression of FAP is closely related to the occurrence of UF .Luo et al 2015 were the first who suggested that estrogen could stimulate fibroblast activation. In addition, they revealed that proliferative activity of fibroblast and the expression of FAP were significantly increased after estrogen stimulation. They also found that estrogen could promote the release of cytokines (TGFβ and IGF-1) and ECM components (collagen I, fibronectin, and laminin) from fibroblasts. Furthermore they found that silencing of FAP expression significantly decreased promotion effects of estrogen on TAF suggesting that FAP plays an important role in estrogen-mediated fibroblast activation. Autophag (eating of self) is a collection of processes that enables the cells to digest and recycle their cytoplasmic contents, such as toxic protein aggregates, disused organelles and invading microorganisms. Dysregulation in autophagy process have been recently described in many neoplasms. However the role of autophagy in the pathogenesis of UFs is still unclear and further understanding of its regulation and significance will be needed. The PI3K/AKT/mTOR signalling pathway is considered the main pathway involved in the initiation and regulation of autophagy. Previous studies found that reduced FAP significantly decreased the expression of phosphorylated AKT suggesting that FAP is an upstream factor modulating the PI3K/AkT. This study will be the first to study the link between fibroblast activation and autophagy in pathogenesis of UF through PI3K/AKT signaling pathway. Although several types of drugs (mostly antiproliferative agents) are available for UF treatment, none of them were introduced specifically as antifibrotic agents. Targeting such novel signaling pathway may be considered useful for future non surgical treatment of UF affecting both proliferative and fibrotic changes.

Official Title
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A Novel Insight Into the Role of Fibroblast Activation and Autophagy in Uterine Fibroid

Conditions
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Uterine Fibroid

Intervention / Treatment
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* Genetic: Measurement of protein expression in tissues.


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: i. Premenopausal women (age ˂ 50) with uterine fibroids who are diagnosed through clinical gynecological, ultrasound and other auxiliary examinations. ii. Patients who exhibit a normal coagulation function. Exclusion Criteria i. Patients who have a previous history of myomectomy or with ovarian malignancy and borderline tumors ii. Patients who are subsequently diagnosed with uterine adenomyosis. iii. Pregnant women. iv. Patients who receive hormonal treatment within three months before surgery. v. Patients with history of coronary artery disease, hypertension, liver cirrhosis or hematologic disorders.

Ages Eligible for Study
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Maximum Age: 50 Years

Sexes Eligible for Study
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Female

Accepts Healthy Volunteers
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Accepts Healthy Volunteers

Study Plan
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How is the study designed?
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Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| patients group<br>include 35 pre-menopausal women (age ˂ 50 years) enrolled to undergo hysterectomy for symptomatic UF at the women health hospitals, Assiut University. The protein expression of the followings markers will be estimated in tumor tissue samples: Fibroblast activation protein (FAP) will be measured by quantitative real time polymerase chain reaction qRTPCR (mRNA level) and ELISA (protein level). Autophagy markers level (LC3 and p62) will be measured by qRTPCR (mRNA level) and by immunohistochemical analysis. Phosphorylated protein kinase B (pAKT ) by ELISA (protein level). analysis. Markers of oxidative stress (Malondialdehyde as lipid peroxide) by a colorimetric method. Reduced glutathione, an antioxidant marker by a colorimetric method . | Genetic: Measurement of protein expression in tissues.<br>* The followings markers will be estimated in tissue FAP, a marker of fibroblast activation, using quantitative real time polymerase chain reaction qRTPCR (mRNA level) and ELISA (protein level). Markers of autophagy including LC3 and p62 using qRTPCR (mRNA level) and by immunohistochemical analysis. Phosphorylated protein kinase B (pAKT) level by ELISA (protein level). Markers of oxidative stress (Malondialdehyde as lipid peroxide) by a colorimetric method. Reduced glutathione, an antioxidant marker by a colorimetric method<br>|
| Control group<br>include 35 normal myometrial tissue samples obtained 1 cm away from the fibroid capsule from the same patients. The protein expression followings markers will be estimated in normal myometrial tissue samples Fibroblast activation protein (FAP) will be measured by quantitative real time polymerase chain reaction qRTPCR (mRNA level) and ELISA (protein level). Autophagy markers level (LC3 and p62) will be measured by qRTPCR (mRNA level) and by immunohistochemical analysis. Phosphorylated protein kinase B (pAKT) by ELISA (protein level). Markers of oxidative stress (Malondialdehyde as lipid peroxide) by a colorimetric method. Reduced glutathione, an antioxidant marker by a colorimetric method . | Genetic: Measurement of protein expression in tissues.<br>* The followings markers will be estimated in tissue FAP, a marker of fibroblast activation, using quantitative real time polymerase chain reaction qRTPCR (mRNA level) and ELISA (protein level). Markers of autophagy including LC3 and p62 using qRTPCR (mRNA level) and by immunohistochemical analysis. Phosphorylated protein kinase B (pAKT) level by ELISA (protein level). Markers of oxidative stress (Malondialdehyde as lipid peroxide) by a colorimetric method. Reduced glutathione, an antioxidant marker by a colorimetric method<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Comparing FAP and autophagy markers in patient and control groups. | Explore difference in level of fibroblast activation marker (FAP) and autophagy markers (LC3 and P62) in UF tissue samples compared to normal myometrial samples (1 cm from the UF lesion). | 1 year | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Study signaling pathway linking FAP and autophagy which ma considered as a therapeutic target | Investigate the correlation between fibroblast activation and autophagy through PI3/AKT signaling pathway and their role in the pathogenesis of UF through estimation of AKT protein expression in UF tissue samples compared to normal myometrial samples 1 cm from the UF lesion. | 1 year | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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Uterine fibroid, FAP, Autophagy