A Study Protocol to Assess the Correlation of Endometrial Morphology with Concerned Hormone Levels in Patients of Abnormal Uterine Bleeding

Background: Abnormal-uterine-bleeding (AUB) is caused due to numerous systemic or functional etiologies. Patients may come with metrorrhagia, menorrhagia, intermenstrual bleeding and polymenorrhea. The underlying pathology can be ascertained by histological variations of endometrium considering the age, the phase of menstrual cycle and the use of any exogenous hormones. This study aims to assess the correlation between endometrial changes and concerned hormone levels. Methodology: This will be a Prospective study conducted for two years in Histopathology Department of Pathology, JNMC, Wardha. Total 65 female patients with complaint of irregular uterine bleeding will be included in this study. The histomorphology patterns of endometrial lesions will be studied and classified according to WHO classification. The collected data will be analysed using appropriate statistical methods. Results: A Significant correlation is expected between endometrial changes and concerned hormone levels. Conclusion: Conclusion will be drawn from the results obtained from the study. Study Protocol Bhardwaj et al.; JPRI, 33(64B): 246-252, 2021; Article no.JPRI.81047 247


INTRODUCTION
The definition of Abnormal-uterine-bleeding (AUB) is any form of bleeding which is not within the normal range of quantity, frequency, duration or cyclicity. AUB, which involves both acute and chronic AUB, is thought to be a symptom of potential uterine or systemic illness. In all age categories, it is the most prevailing complaint [1]. In adolescents, vaginal bleeding which occurs before menarche is referred to abnormal, while in females of child bearing and premenopausal age, any variation in menstrual period, frequency, duration or amount of flow, as well as bleeding between cycles, are considered as an abnormal sign.
There are systemic or functional etiologies for the AUB. Patients may come with metrorrhagia, menorrhagia, intermenstrual bleeding and polymenorrhea. The underlying pathology can be ascertain by histological variations of endometrium considering the age , the phase of menstrual cycle and the use of any exogenous hormones [1]. Other factors may be psychological stress, weight (obesity, anorexia, or rapid change), exercise, endocrinopathy, neoplasm, medication, or unexplained etiology, according to the studies carried out [2]. The frequency of abnormal uterine bleeding in patients is more prevalent in the age group of 40 to 49 years, where the prevalence in the population pool was around 64 percent [3].
The word abnormal-uterine-bleeding (AUB) encompasses both organic and non-organic causes. Dysfunctional-uterine-bleeding (DUB) is actually a subtype of AUB that, due to nonorganic etiology, includes irregular bleeding. In 50 percent of women with AUB, it is present . For abnormal-uterine bleeding, endometrial biopsy is generally indicated to rule out organic pathology. Age and menstrual history are of particular significance since, according to age and menstrual status, the causes of abnormaluterine-bleeding vary. In women of the reproductive age group, pregnancy complications, including abortion, are normal, while atrophy and organic pathologies are seen more commonly in postmenopausal women [1][2][3].
Only by ruling out the organic causes after the histopathological exam can a diagnosis of dysfunctional-uterine-bleeding be made. In DUB, there are three trends usually seen. The first is referred to as "estrogen breakthrough bleeding," which happens from a "persistent follicle" in the continuous production of estrogen. The proliferative endometrium grows in size, thus increasing its own blood supply, and there is breakthrough bleeding. The second is known as "estrogen withdrawal bleeding," which is due to "failed follicle," i.e. 2, subnormal estrogen is released by the follicles. Both of these causes are anovulation-accredited. The third one is "ovulatory endometrium" due to defects in the follicular/luteal process [2][3][4].
Hypothyroidism is a common cause of multiple disorders in women of reproductive age, ranging from sexual dysfunction, menstrual defects and infertility. The first menstrual symptoms for women with hypothyroidism are disordered cycles and irregular blood flow [5].
As part of a hysterectomy specimen, the endometrium may be examined . DUB cases, depending significantly on the age and clinical history of the patient, may have structural or functional causes. Some studies have shown infertility as the most common sign of endometrial biopsy and endometrium in the secretory process as the most common morphological pattern [6].
A lot of women with fibroids are completely asymptomatic. However, in many women, gynaecological services with AUB and associated iron-deficiency anaemia are most frequently present. Everyday life is frequently interrupted for women with uterine fibroids, and fibroids are a leading sign of hysterectomy [7].
The length and heaviness of bleeding episodes in the form of variance, perimenopausal women facing menstrual issues experience cycle irregularity. Follicular growth is irregular during this process, leading to variability in the amount of oestrogen. Therefore in order to understand its potential consequences for reproductive cancers and other health outcomes, detailed characterization of sex hormone dynamics during perimenopause is important [8].
Some adolescents do not know that their patterns of bleeding are abnormal, although it is recognized that menstrual cycles during puberty are sometimes irregular. AUB and/or AUB alone may have long-term health effects, affect the quality of life and have an effect on school attendance [9].
The irreversible cessation of menstruation arising from the absence of follicular function of the ovary is menopause. Bleeding is labeled as postmenopausal bleeding 12 months after the last period of menstruation (PMB). Around 3 percent of menopausal women are affected by this condition and require prompt and thorough evaluation [10].
Abnormal uterine bleeding can occur in different forms at any age and has various modes of presentation. Excessive uterine bleeding can result from a wide range of conditions during reproductive age, from physiological processes to malignant lesions that include organic, systemic, and hormonal responses. Adenomyoma, endometrial polyps, ovarian tumors, pelvic inflammatory disease (PID), endometrial hyperplasia, endometrial carcinoma, hormonal dysfunction (such as hypothyroidism), and hypothalamic-pituitary disorders can be induced by fibromyoma. Abnormal uterine bleeding happens without any systemic causes or any organic lesions of the genital tract in a significant number of patients and the term dysfunctional uterine bleeding is used for this [11].
The normal cyclic physiological changes that occur during the reproductive cycle in the endometrium of females are regulated by the cyclical release of ovarian estrogen and progesterone. Concentration of receptors for these hormones also varies respectively. In hyperplasia's and endometrial cancers, especially type I Estrogen and progesterone hormones are also expressed. The level of these hormones gives a prognostic information. They also offer the option of hormone therapy. The key factor involved in the pathogenesis of Dysfunctional-Uterine-Bleeding is hormonal imbalance. This imbalance is best studied by endometrium histopathological assessment. Abnormal bleeding can be caused by welldefined pathological conditions such as chronic endometritis, endometrial polyp, submucosal leiomyomas [12].

Research Gap
The present study aimsto study the correlation of morphological findings of endometrium with concerned hormone levels in patients of abnormal-uterine-bleeding at AVBRH and to correlate the histopathological findings in endometrium with concerned hormonal level in patients of AUB.

Research Question
With the understanding of changes in endometrium in patients of AUB in correlation with concerned hormone level, the following research question is framed-"Does correlation of morphological findings of endometrium with concerned hormone levels in patients of abnormal-uterine-bleeding."

Study Design
Observational, analytical and prospective.

Methods
 A detailed clinical history regarding age, clinical features, radiological findings and probable diagnosis of the patients will be obtained who have abnormal uterine bleeding.  The grossed specimens will be then processed in tissue processor and paraffin blocks will be prepared. Sections will be cut and stained by H and E stains. The slides will be assessed under microscopy.  The histomorphology patterns of endometrial lesions will be studied and classified according to WHO classification.  The collected data will be tabulated and results will be calculated and analyzed by using appropriate statistical methods.

Sample Size -65 Patients
The sample size was calculated by using Krejci and Morgan formula with desired error of margin: • All patients clinically and USG diagnosed cases of AUB, endometrial sampling, hormonal assessment, will be selected in this study group.

Exclusion criteria
• Patients with non-endometrial conditions of abnormal-uterine-bleeding.
• Patients with uterine-bleeding due to Intrauterine Devices.
• Patients presenting with abnormal-uterinebleeding due to Pregnancy and its related complications.

Statistical Analysis
Statistical analysis will be carried out by using 'chi-square-test' where a value of P<0.05 will be considered to indicate statistical-significance.

EXPECTED RESULTS
The study will be conducted for a period of 2 years and all the observations will be depicted in a well-tabulated master chart.

DISCUSSION
Tara et al. conducted a cross-sectional prospective study in the outpatient clinic of Maternity Teaching Hospital in Erbil city from 1st of September, 2017 to 30th of June, 2018 on 100 women. The aim of the study was to highlight the significant association between high thyroid stimulating hormone level and women with abnormal uterine bleeding (p=0.002). The study concludes that the thyroid dysfunction is a common cause of abnormal-uterine-bleeding among females of reproductive age group [5].
Ifeyinwa Mary Asuzu et al. A cross-sectional research was performed on specimens collected over a one-year period at the Department of Pathology of Premier Hospital, Abuja. Four hundred and eighty-six endometrial biopsy and curettage samples were examined from women with irregular uterine bleeding submitted to the histopathology laboratory. The research concludes that retained products of pregnancy are the most common source of abnormaluterine-bleeding in this population [6].
Lucy Whitaker et al. conducted a prospective study with the aim that AUB also co-exists with fibroids, but the relationship between the two remains incompletely understood and a menstrual bleeding complaint may be incidental to the identification of fibroids in many women. The study concludes. The care must remain individualized and involve the effect of pressure symptoms, the need for fertility retention and the need for contraceptives, as well as the control of their AUB to achieve enhanced quality of life [7].
MaltiKumariMaurya et al. A prospective research was performed to assess the clinical characteristics and histopathological results of irregular uterine bleeding and to compare the hormone profile in perimenopausal women. The study concludes that intermenstrual bleeding can represent a troubling symptom of endometrial malignancy in perimenopausal females. Further management and result can be predicted by histopathological endometrium trend and clinicopathological variables [8].
Selin Elmaoğulları et al. A prospective research was performed to establish that because of the immature hypothalamic-pituitary-ovarian axis and a related bleeding disorder in about 20 percent of HMB cases, anovulatory cycles are the leading etiology of HMB. In general, AUB is a likely cause of endocrine disorders such as hypothyroidism, polycystic ovarian syndrome and hyperprolactinemia. The study concludes that the AUB has been one of the key problems of adolescent gynecology, and the AUB is the most frequent anovulatory HMB presentation.. Anovulatory processes in most teenagers are normally hormonal and resolve spontaneously as the hypothalamic-pituitary-ovarian axis matures [9]. Pushpa Singh et al. conducted a prospective study withThe purpose of this research was to study endometrial thickness by transvaginal sonography and to compare it with the cytological pattern assessed by endometrial aspiration and the histopathological pattern of hysteroscopic biopsy. The study concludes that the role of endometrial thickness in the diagnosis of patients at high risk, especially with comorbid conditions, cannot be undermined. In selected cases of PMB by hysteroscopy, histopathological examination is mandatory for ruling out malignancy [10].
Mitali Mahapatra et al. conducted a prospective study with the aim study the various aetiopathological factors responsible for AUB in women ages 20-55 years, and to correlate the clinical presentation with ultrasonography, hysteroscopy, laparoscopy and histopathological finding.The study concludes that it is seen that incidence of AUB is more common in 5 th decade of life and in multiparous women. Menorrhagia is the most common bleeding pattern followed by metrorrhagia. Histopathological examination of the endometrium revealed that whatever may be the pathology, proliferative endometrium is the most common pattern [11].

CONCLUSION
Conclusion will be drawn from the results obtained from the study.

CONSENT
As per international standard or university standard, patients' written consent will be collected and preserved by the author(s).

ETHICAL APPROVAL
As per international standard or university standard written ethical approval will be collected and preserved by the author(s).