37 Su r g ic a l D iS e a Se S iSSN 2413-6077. iJMMr 2022 Vol. 8 issue 1 *Corresponding author: Thokchom Bishwajit Singh, Assistant Professor (General Surgery), Department of Surgery, Military Hospital, Haryana, 133001, India. E-mail: bishwajitsingh6068@gmail.com. International Journal of Medicine and Medical Research 2022, Volume 8, Issue 1, p. 37-43 copyright © 2022, TNMU, All Rights Reserved T. B. Singh et al. DOI 10.11603/ijmmr.2413-6077.2022.1.13083 AN OBSERVATIONAL STUDY TO FIND RELATIONSHIP OF ACUTE APPENDICITIS TO MENSTRUATION CYCLE IN NORTHERN AND NORTHEASTERN PART OF INDIA *T. B. Singh1, R. Kumar2, A. Nigam3, T. R. Devi4 1 – MILITARY HOSPITAL, HARYANA, INDIA 2 – MILITARY HOSPITAL, TEZPUR, ASSAM, INDIA 3 – PUBLIC HEALTH DEPARTMENT GANGTOK, SIKKIM, INDIA 4 – JAWAHARLAL NEHRU INSTITUTE OF MEDICAL SCIENCES, IMPHAL EAST, INDIA Background. Diagnosing acute appendicitis correctly in a female patient is a challenge for a practicing surgeon. Rate of misdiagnosis of acute appendicitis is very high among female patients. There are many studies carried out to find incidence of acute appendicitis as per various phases of menstruation cycle but the results were conflicting. Objective. The study was conducted to find the relationship of acute appendicitis with the different phases of the menstruation cycle. Methods. This research was an observational study carried out in two regional hospitals in northern and north-eastern part of India. Duration of study was 24 months between 01 June 2019 and 31 May 2021. Inclusion Criteria were for all female patients, non-pregnant and menstruating, who were histopathologically confirmed as acute appendicitis. All pathologically proven negative appendectomy patients were excluded from this study. All female patients taking oral contraceptive pills (OCP) were excluded from the study. Results. A total of 96 females were hospitalized in both hospitals during the study period; 12 of them did not attain menarche (12.5%), 6 had menopause (6.3%) and 78 were menstruating (81.25%). Of those 78 patients, who had physiological menstrual cycles, 6 were in menstrual phase (7.6%), 18 were in proliferative phase (23%), and 54 in luteal phase (69.2%). In our study, there were only 6 cases of acute appendicitis during menstruation. therefore, the expected ratio of cases was 6/14=0.42. The corresponding expected rate for the proliferative phase was 2.78×9=25 cases, whereas for the luteal phase it was 2.78×14=38.92. The expected ratio was 18/25=0.72 for the proliferative phase and 54/39=1.38 for the luteal phase. There was a significant increase in number of cases of acute appendicitis among the menstruating women in luteal phase with p value <0.05. Mean age of the study participants was 28.31±9.56. Conclusions. According to the result of the study, the incidence of acute appendicitis significantly differs in different phases of menstruation cycle with highest incidence in luteal phase. Hence, female hormones (estrogen and progesterone) are significant in causing acute appendicitis. KEYWORDS: acute appendicitis; menstruation phase; proliferative phase; luteal phase. Introduction Diagnosing an acute appendicitis in female patients is a challenge for a practicing surgeon. The rate of misdiagnosis is comparatively high in female patients especially in reproductive age group. The lifetime risk of appendicitis is 6.7% for females [1]. Diseases of the female i n t e r n a l re p ro d u c t i v e o r g a n s t h a t m a y erroneously be diagnosed as appendicitis in approximately descending order of frequency include pelvic inflammatory disease, ruptured graffian follicle, twisted ovarian cyst or tumor, endometriosis and ruptured ectopic pregnancy [1]. Timely diagnosis and early treatment can prevent complications associated with the disease. The sonographic diagnosis of acute appendicitis has reported sensitivity of 55% to 96% and a specificity of 85% to 98% [1]. Ap­ pendicitis is also associated with gastrointestinal symptoms like nausea (sensitivity, 58%; spe- cificity, 36%), vomiting (sensitivity, 51%; speci­ ficity, 45%), and anorexia (sensitivity, 68%; specificity, 36%) [1]. Gastrointestinal symptoms that develop before the onset of pain suggest a different etiology such as gastroenteritis [1]. 38 Su r g ic a l D iS e a Se S iSSN 2413-6077. iJMMr 2022 Vol. 8 issue 1 The ratio of cases of appendicitis to cases of pelvic inflammatory disease is low in females in the early phase of the menstrual cycle and high during the luteal phase [1]. Many studies showed different results. Arnbjo rnsson et al. showed that appendicitis is more common during the luteal phase of the cycle [2] whereas Robinson et. al. showed that acute appendicitis is equally common in all phases of the cycle [3]. The aim of the current study was to find out the relationship of acute appendicitis to mens- truation cycle. Methods This study was an observational study carried out in two regional hospitals in northern and north­eastern parts of India. The study was aimed to assess the relationship of phases of menstruation cycle to acute appendicitis. Duration of study was 24 months between 01 June 2019 and 31 May 2021. Inclusion Criteria: all female patients, non- pregnant and menstruating, who were his­ topathologically confirmed as acute appen­ dicitis. Exclusion Criteria: all pathologically proven negative appendectomy patients were excluded from this study. All female patients, who were con suming in OCP, were excluded from the study. Menstrual cycle was divided into 3 phases: 1. Menstruation phase; 2. Proliferative phase; 3. Luteal phase. Duration of two phases was considered as constant (a) menstruation phase: 05 days of menstruation in physiological cycle from the beginning of bleed; (b) luteal phase: last 14 days of cycle (calculated retrospectively 14 days from the first day of menstruation bleed of next cycle excluding the day 1 of bleed). Hence, the period of proliferative phase was variable. It was also considered that ovulation occurred at the 14th day of the cycle. Results A total of 96 females were hospitalized in both hospitals during the study period (Table 1); 12 of them did not attained menarche (12.5%), 06 had menopause (6.3%), and 78 were mens­ truating (81.25%). Of those seventy­eight pa­ tients, who had physiological menstrual cycles, 6 were in menstrual phase (7.6%), 18 were in proliferative phase (23%), and 54 – in luteal phase (69.2%). Fig. 1. Phases of menstruation cycle. Menstruation phase: 5 days Proliferative phase: [n-(5+14) days] Luteal phase: 14 days Total days in a Cycle: n days Table 1. Acute appendicitis cases (total number and percentage) as per phases of menstruation Number of females Percent Not attained menarche 12 12.5 Menstrual phase 6 6.3 Proliferative phase 18 18.8 Luteal phase 54 56.3 Post menopause 6 6.3 Total 96 100.0 T. B. Singh et al. 39 Su r g ic a l D iS e a Se S iSSN 2413-6077. iJMMr 2022 Vol. 8 issue 1 T. B. Singh et al. Four out of seventy-eight acute appendicitis cases (Table 2) in menstruating females were complicated type, whereas none in non­menst­ ruating female group had any complications. The expected daily occurrence of acute appendicitis for patients with physiological menstrual cycles (for a 28­day cycle) would have been 78/28 = 2.78 cases/day. The expected rate for cases of acute appendicitis during menst- ruation (accepting that menstruation lasts for 5 days), would have been 2.78×5=14 cases. In current study there were only 06 cases of acute appendicitis during menstruation. The obser- ved: expected ratio of cases therefore was 06/14=0.42. The corresponding expected rate for the proliferative phase was 2.78×9=25 ca­ ses, whereas for the luteal phase it was 2.78×14=38.92. The observed: expected ratio therefore was 18/25=0.72 for the proliferative phase and 54/39=1.38 for the luteal phase. There was a significant increase in number of cases of acute appendicitis among the menstruating women in luteal phase with p value <0.05. The mean age of the study participants was 28.31±9.56. Table 2. Total complicated and non-complicated cases Menstruating Non Menstruating Total Complicated 4 0 4 Non Complicated 74 18 92 Total 78 18 96 Table 3. Observed frequency, expected frequency and P value with respect to different phases of menstruation cycle Phase of menstrual cycle Observed frequency Expected frequency P value Menstrual 6 14 P value 0.034066Proliferative 18 25 Luteal 54 39 Chi-square statistic 6.7589 Table 4. Mean, Median, Std deviation and Ranges Statistics N Valid 78 Mean 28.31 Median 28.00 Std. Deviation 9.558 Range 39 Minimum 13 Maximum 52 Discussion The previous studies by Arnbjornsson [2] and Eldar et. al. [4] reported that incidence of histologically confirmed acute appendicitis was less common during menstruation phase [1] but the difference were not statistically sig­ nificant. On literature search before these cur­ rent studies, there were conflicting observations. Some studies [1, 4] showed higher incidence of acute appendicitis in luteal phase whereas some other [2] – in proliferative and menstrua­ tion phase. Another study by Robinson et. al. failed to show any difference [3]. The study had established that there was a significant increase in number of cases of acute appendicitis among the menstruating women in luteal phase with p value <0.05. Similar result was shown in the studies by Eldar et. al. The cause of this significant raise in incidence of acute appendicitis may be because of low estrogen level and high progesterone level modulating immune response and increasing susceptibility for gut infections. Souza et. al. [5] reported increased NK activity in follicular phase as compared with luteal phase. Progesterone receptors and pro- gesterone induced apoptosis of NK cells and suppresses IL-12-induced IFN-g production of Killer Ig­like receptor (KIR)+ NK cells [6]. Whereas estrogen has been reported in many stu dies to prevent B cell apoptosis, enhance sur vival and activation of autoreactive cells [7], and increase expression of CD4+ T cell che­ mokine receptors [8]. Estrogen exerts immune regu lation via estrogen receptors (ERs) on the lymphocytes, and receptors for estrogen, pro- gesterone, androgen, and glucocorticoid are found in lymphoid organs and/or lymphocytes [9, 10]. 40 Su r g ic a l D iS e a Se S iSSN 2413-6077. iJMMr 2022 Vol. 8 issue 1 Hall et. al described that concentrations of progesterone fluctuated over the life course in females, with increased concentrations at puberty, cyclical changes during the menstrual cycle, and a steady rise during pregnancy followed by a sharp decline post­partum [11]. At menopause, concentrations steadily declined to levels that are similar to those prior to puberty [11]. Progesterone hormone level started rising after ovulation on the 14th day of cycle and reached its highest level during mid-luteal phase (Fig. 2). Speroff et al. in his study mentioned that serum levels of progesterone fluctuate during the menstrual cycle with a peak of 20 ng ml-1 during the luteal phase and the nadir (<1ng ml-1) during the follicular phase [12]. Progesterone generally inhibits inflam- matory innate immune responses [11]. In vitro studies by Hardy et. al. and Jones et. al. showed that progesterone can suppress activation of macrophages and dendritic cells [13, 14]. When progesterone is bound to its receptor, it directly interferes with the transcription factor nuclear factor­kappa B (NF­kB) through transrepression and inhibits gene transcription downstream of the NF­kB pathway, including cyclooxygenase­2 to decrease inflammation [13, 15]. Progesterone can also decrease inflammation by inhibiting the production of proinflammatory cytokines (e.g., TNF­a, IFN­g, and IL­12) and increasing production of anti-inflammatory cytokines, including IL-10 [14, 16]. Raised level of progesterone also suppresses Th1 response and enhances IL-10 producing Th2 cells. Miyaura et. al. in his study mentioned that progesterone was established to suppress Th1 response and enhance IL-10 producing Th2 cells [17]. Ehring et. at. in his study had showed that progesterone suppressed immune func- tions of T cells by a non-genomic mechanism, which was the blocking of K+ channel, Ca2+ signaling and NF of activated T cells driven gene expression [18]. Siiteri et. al. had demonstrated in vivo the immunosuppressive effects of progesterone in his study by prolonged survival of xenografts near silastic implants containing progesterone at concentrations typically found in the placenta [19]. Hence, such high pro­ gesterone level in luteal phase may have caused unfavorable immunity changes leading to more susceptibility to gut infection and causing highest incidence of acute appendicitis in luteal phase. Estrogen hormone increases its level from menstruation phase and reaches its peak at late proliferative phase. Estrogen then decline its level and reaches minimum level at late luteal phase. Hence, maximum proliferation of both lymphocytes and macrophages occurs during proliferative phase whereas minimum proliferation of both lymphocytes and macro- phages occurs during luteal phase. Fig. 2. Relationship between different level of estrogen and progesterone with percentage of acute appendicitis in different phases of menstruation cycle. T. B. Singh et al. 41 Su r g ic a l D iS e a Se S iSSN 2413-6077. iJMMr 2022 Vol. 8 issue 1 T. B. Singh et al. Whitacre et. al. stated in his study that the effects of estrogen were pleotropic on immune cell function and estrogen had been shown to significantly stimulate the proliferation of both lymphocytes and macrophages [20]. In many studies, estrogen has been shown to regulate immune response by impairing negative selection of high affinity auto­reactive B cells, modulating B cell function and leading to Th2 response [21, 22]. Estrogen also influences phy­ siological functions via ERs which are expressed in brain, gut epithelial cells, lymphoid tissue cells as well as immune cells [23, 24]. R. Mo et. al has stated that estrogen also induces T cell homing by enhancing the expression of C-C chemokine receptor type 5 (CCR5), a homing marker [8]. Many studies has also shown that estrogen regulates immune res ponse via modulation of endosomal TLRs and TLR8 expression thus hormonal balance determines the overall response to infection in females [25, 26, 27]. Hence, such low estrogen level with high progesterone level in luteal phase may cause unfavorable immunity changes leading to more susceptibility to gut infection and causing highest incidence of acute appendicitis in luteal phase and vice versa in proliferative phase. Conclusions According to the results of the study, it can be concluded that the incidence of acute appendicitis significantly differs in different phases of menstruation cycle with highest incidence in luteal phase. Hence, female hormones (estrogen and progesterone) are significant in causing acute appendicitis. This relationship can be used as additional guiding tool or information when there is diagnostic dilemma for acute appendicitis in females. Limitations Small sample size. Conflict of Interests Authors declare no conflict of interest. Acknowledgements I express thanks to all my interns Mr. Rameez, Mr. Praveen, Mr. Suryakant, Mr. Ashok, Mr. Sryakant and Mr. Agyastha for helping me in collection of data, tabulation and statistical analysis. Author’s Contributions Thokchom Bishwajit Singh, Ranjan Kumar – conceptualization, methodology, formal ana­ lysis, writing – original draft, writing – reviewing and editing; Ankur Nigam – investigation, formal analysis; Tekcham Roshini Devi – data curation, writing – reviewing and editing. ВЗАЄМОЗВ'ЯЗОК ГОСТРОГО АПЕНДИЦИТУ ТА ТА фАЗИ МЕНСТРУАльНОГО ЦИКлУ У ЖИТЕлІВ ПІВНІЧНОЇ ТА ПІВНІЧНО-СХІДНОЇ ЧАСТИНІ ІНДІЇ *T. B. Singh1, R. Kumar2, A. Nigam3, T. R. Devi4 1 – MILITARY HOSPITAL, HARYANA, INDIA 2 – MILITARY HOSPITAL, TEZPUR, ASSAM, INDIA 3 – PUBLIC HEALTH DEPARTMENT GANGTOK, SIKKIM, INDIA. 4 – JAWAHARLAL NEHRU INSTITUTE OF MEDICAL SCIENCES, IMPHAL EAST, INDIA Вступ. Діагностування гострого апендициту у пацієнта може бути проблемною для практикуючого хірурга. Частота неправильної діагностики гострого апендициту дуже висока серед пацієнтів. Існує багато досліджень, проведених для виявлення частоти гострого апендициту відповідно до різних фаз циклу менструації, але їх результати є суперечливими. Мета. Дане дослідження проводилося для встановлення та оцінки взаємозв'язку гострого апендициту з різними фазами менструального циклу. Методи. Дослідження було спостережним, проведеним у двох регіональних лікарнях у північній та північно-східній частині Індії. Тривалість дослідження становила 24 місяці з 01 червня 2019 року по 31 травня 2021 року. Критерії включення: Усі пацієнти, які не порушень менструального циклу та які мали гістопатологічне підтвердження діагнозу гострий апендицит. Критерії виключення: Усі пацієнти яким не проводилася апендектомія чи які приймали оральні контрацептиви, були виключені з дослідження. Результати. Всього 96 жінок були госпіталізовані в обох лікарнях протягом періоду дослідження. З них 12 не досягли менархе (12,5%), 6 мали менопаузу (6,3%), а 78 – мали менструальні цикли (81,25%). З тих 78 пацієнтів, які мали фізіологічні менструальні цикли; 6 були в менструальній фазі (7,6%), 18 - у 42 Su r g ic a l D iS e a Se S iSSN 2413-6077. iJMMr 2022 Vol. 8 issue 1 проліферативній фазі (23%) та 54 у лютеїновій фазі (69,2%). У нашому дослідженні під час менструації було лише 6 випадків гострого апендициту. Таким чином, спостережуване: очікуване співвідношення випадків становило 6/14 = 0,42. Відповідна очікувана цифра для проліферативної фази становила 2,78×9 = 25 випадків, тоді як для лютеїнової фази було 2,78×14 = 38,92. Таким чином, спостережуване/очікуване співвідношення становило 18/25 = 0,72 для проліферативної фази та 54/39 = 1,38 для лютеїнової фази. Спостерігалось значне збільшення кількості випадків гострого апендициту серед менструальних жінок у лютеїновій фазі (р<0,05). Середній вік учасників дослідження становив 28,31 ± 9,56. Висновки. За результатами нашого дослідження можна зробити висновок, що захворюваність на гострий апендицит значно відрізняється різними фазами циклу менструації з найвищою частотою в лютеїновій фазі. Отже, жіночі гормони (естроген та прогестерон) відіграють важливу роль у виникненні гострого апендициту. КЛЮЧОВІ СЛОВА: гострий апендицит; фаза менструації; проліферативна фаза; лютеїнова фаза. Information about the authors Thokchom Bishwajit Singh, Assistant Professor (General Surgery), Department of Surgery, Military Hospital, Haryana, India. https://orcid.org/0000­0001­5849­5999, e­mail: bishwajitsingh6068@gmail.com Ranjan Kumar, Assistant Professor (General Surgery), Department of Surgery, Military Hospital Tezpur, Assam, India. https://orcid.org/0000­0002­4573­2446, e­mail: ranjan_afmc@yahoo.co.in Ankur Nigam, Deputy Assistant Director of Health, Public health department Gangtok, Sikkim, India. https://orcid.org/0000­0003­3689­0327, e­mail: ankurnigam8511@yahoo.com Tekcham Roshini Devi, Junior Resident, Department of Anesthesia, Jawaharlal Nehru Institute of Medical Sciences, Imphal East, India. https://orcid.org/0000­0003­1048­9682, e­mail: tekchamroshini12@gmail.com References 1. Liang M, Andersson R, Jaffe B and Berger D. The Appendix | Schwartz's Principles of Surgery, 10th ed, New York; McGraw Hill Education Medical; 2015. 1241-62. 2. Arnbjörnsson E. 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