Elucidation of the biological roles of Wnt5a signaling in follicle development

La santé folliculaire est déterminée par un nombre de facteurs endocriniens, paracrines et autocrines. Les gonadotrophines hypophysaires sont les principaux moteurs du développement du follicule, mais leurs actions sont modulées localement par les hormones et des facteurs de croissance. Les glycoprotéines de la famille des WNTs représentent une grande famille de molécules impliquées dans différentes voies de signalisation. Ils sont sécrétés dans le but de moduler et coordonner la réponse des follicules aux gonadotrophines, et leurs activités sont indispensables à la fonction ovarienne et à la fertilité féminine. Les WNTs sont généralement classés en fonction de la (des) voie(s) qu’ils activent. Le rôle des membres de la voie canonique WNT et de ses composants tels que CTNNB1, WNT4, WNT2, FZD1 et FZD4 est bien établi au cours du développement du follicule chez les rongeurs. Un rôle similaire des WNTs dans les espèces mono-ovulatoires demeure essentiellement inconnu. De plus, le rôle des WNT non canoniques dans l'ovaire de rongeurs est méconnu. Les objectifs de cette thèse sont (1) d'élucider la régulation hormonale de l'expression de WNT5A et le rôle physiologique de WNT5A dans les cellules de la granulosa bovine in vitro et (2) d'identifier les rôles physiologiques de WNT5A dans l'ovaire de souris par inactivation génique conditionnelle. Chacun de ces objectifs a mené à la publication d’un article à partir des résultats obtenus au cours de cette thèse. Dans le premier article, le rôle de WNT5A dans les cellules de la granulosa bovine a été étudié in vitro. Nous avons constaté que WNT5A est un régulateur négatif de la stéroïdogenèse stimulée par la FSH issue des cellules de la granulosa, et qu'il agit en supprimant l'activité de signalisation des WNTs canoniques tout en induisant la voie de signalisation MAPK8/JUN. le deuxième article, afin d’examiner le rôle de deux WNTs non-canoniques, WNT5A et WNT11, à différents stades de développement folliculaire, nous avons généré des modèles de souris knock-out conditionnels ciblant les cellules de la granulosa pour chacun de ces WNTs. Les résultats obtenus ont permis de mettre en évidence que WNT5A est nécessaire pour assurer la fertilité normale chez la femelle, le développement folliculaire et la stéroïdogenèse ovarienne. Il est aussi un antagoniste de la réponse aux gonadotrophines, agissant par l’intermédiaire de la suppression de la signalisation canonique des WNTs. Chez les souris knock-out pour WNT11, nous ne constatons aucun défaut important dans la fertilité des femelles. L’ensemble de notre travail met en évidence que WNT5A est essentiel pour le développement normal du follicule et qu’il agit pour inhiber la différenciation des cellules de la granulosa. En résumé, nous avons fourni une étude novatrice et approfondie, utilisant plusieurs modèles et techniques pour déterminer les mécanismes par lesquels WNT5A régule le développement des follicules.Follicle health is determined by an array of endocrine, paracrine and autocrine factors. Pituitary gonadotropins (LH and FSH) are major drivers of follicle development, but their actions are modulated by local hormones and growth factors. The WNT family of secreted glycoproteins are signaling molecules that act to modulate and coordinate follicular responses to the gonadotropins, and whose activities are indispensable for ovarian function and female fertility. WNTs are normally categorized according to pathway(s) via which they signal. The role of canonical WNT members and components such as CTNNB1, WNT4, WNT2, FZD4 and FZD1 is well established during follicle development in rodents. Whether WNTs play similar roles in monovular species remains essentially unknown. Moreover, the role of non-canonical WNTs in the rodent ovary is unclear. The objectives of the present thesis were (1) to elucidate the hormonal regulation of WNT5a expression and the physiological role of WNT5a in bovine granulosa cells in vitro and (2) to identify the physiological roles of WNT5a in the mouse ovary by conditional gene inactivation. The results of this thesis are presented in two articles. In the first article, the role of WNT5a in bovine granulosa cells was investigated in vitro. We found that WNT5A is a negative regulator of FSH-stimulated granulosa cell steroidogenesis, and that it acts by suppressing canonical WNT signaling activity and inducing the noncanonical MAPK8/JUN pathway. In the second article, we generated granulosa-specific knockout mouse models to examine the roles of two non-canonical WNTs, WNT5a and WNT11, in different stages of the follicular development. Our results showed that WNT5a is required for normal female fertility, follicle development and ovarian steroidogenesis, and is an antagonist of gonadotropin responsiveness that acts via the suppression of canonical WNT signaling. However we did not observe any significant defects in fertility of WNT11 knockout mice. Together, our work demonstrates that WNT5a is essential for normal follicle development and acts to inhibit differentiation in granulosa cells. In overview, we provided a novel and comprehensive investigation, using multiple models and techniques, to determine the mechanisms by which WNT5a regulates follicle development

Differential diagnosis of tuberculosis, sarcoidosis, and anthracosis by CD8, CD3, and CD4 levels by flow-cytometry

Background: Determination of a desirable diagnostic tests is an issue of importance especially to differentiate between tuberculosis) TB(, sarcoidosis, and anthracosis. The purpose of the present study was to determine the differential diagnosis of tuberculosis, sarcoidosis and anthracosis with CD8, CD3, and CD4 by flow-cytometry. Materials and Methods: In this descriptive cross-sectional comparative survey, 40 consecutive patients attending to Masih Daneshvari Hospital in Tehran were enrolled and CD4/CD8 ratio, CD8, CD3, and CD4 were determined by flow-cytometry and compared across patients with three diseases including tuberculosis, sarcoidosis, and anthracosis.Results: The results demonstrated that CD4 was significantly higher in anthracosis cases (P<0.007) and the CD8 was significantly higher in patients with TB (P<0.008).Conclusion: It was attained ultimately that CD4 and CD8 levels could be a desirable diagnostic markers for anthracosis and TB, respectively

Evaluating the relationship between high-resolution computed tomography findings and their extent in eosinophilic lung diseases with peripheral blood eosinophil level

Purpose: Eosinophilic lung diseases are a group of pulmonary disorders associated with peripheral or tissue eosinophilia. They can be classified into primary (idiopathic) and secondary groups based on their aetiology. On the other hand, blood eosinophilia is a common clinical problem with various causes. In many cases the lung infiltrations in patients with peripheral eosinophilia are non-eosinophilic. Herein, we aim to assess the relationship between blood eosinophilia and abnormal high-resolution computed tomography (HRCT) features and their extent in idiopathic chronic eosinophilic pneumonia (ICEP) (the most common form of idiopathic eosinophilic lung disease). This can help in differentiating eosinophilic from non-eosinophilic lung infiltrations in patients with blood eosinophilia. Material and methods: In this descriptive-correlational study, all patients with proven ICEP, who were referred to Masih Daneshvari Hospital, Tehran, Iran from 2012 to 2019, were included. The ICEP diagnosis was based on lung infiltrations on imaging, in addition to increased numbers of eosinophils in bronchoalveolar lavage fluid, blood or lung biopsy samples, and rapid response to corticosteroids. Patients with known aetiologies for eosinophilic lung diseases were excluded. The HRCT findings and their extent in each patient were compared with the blood eosinophil level. Results: Positive correlation was found only between blood eosinophil level and frequency and extent of consolidation, and with frequency of lymphadenopathy. Conclusions: HRCT can help to differentiate eosinophilic from non-eosinophilic lung infiltrations in patients with blood eosinophilia by comparing the extent of consolidation with the blood eosinophil level

Diagnostic values of bronchodilator response versus 9-question questionnaire for asthma

Introduction: Several studies have investigated different tools for asthma diagnosis in order to reduce the cost and improve its early recognition. The goal of this study is to establish a short questionnaire to be used in practice for asthma screening and compare diagnostic values between this method and spirometric response to bronchodilators.Material and method: 208 patients presenting with chronic stable dyspnea (> 6 months) and definite clinical diagnosis of chronic obstructive pulmonary disease, bronchiectasis, pulmonary fibrosis or asthma, were enrolled. 9 questions out of 43 based on the literature search were selected by regression analysis. Patients were asked to complete the questionnaire and then their spirometric responses to bronchodilators were evaluated. Results: Of all, 53.8% of cases were diagnosed clinically to have asthma. For establishing diagnosis of asthma, the bronchodilator test had 48.2% sensitivity, 78.1% specificity, 72% positive, 56.4% negative predictive values, 2.2 positive, 0.66 negative likeli-hood ratios, and false positive, false negative and accuracy of 21.9%, 51.8% and 62.01%, respectively. The revised 9 questions from the questionnaire had 97.3% sensitivity, 77.1% specificity, 83.2% positive, 96.1% negative predictive values, 4.24 positive,  0.03 negative likelihood ratios, 22.9% false positive, 2.7% false negative and 87.98% accuracy.Conclusions: The 9-question questionnaire had better diagnostic values in defining asthma in patients with chronic dyspnea than reversibility of airway obstruction to salbutamol and can be used as a useful screening test for diagnosis of asthma in clinical practice and for investigational purposes

The incidence of mTOR marker in tracheal adenoid cystic carcinoma by immunohistochemical staining

Introduction: There is an association between the activation of mammalian target of rapamycin (mTOR) signaling and aggressive tumor growth in multiple forms of cancer, including adenoid cystic carcinoma (ACC). ACCs are uncommon yet a malignant form of neoplasms that arises within the secretory glands. Therefore, the aim of this study was to investigate the increase of mTOR in the ACC tumors in order to survey the possibility of treating these tumors with mTOR inhibitors.Material and methods: Samples from known cases of the lung and tracheal ACC were retrieved from the archives of the pa-thology department of Masih Daneshvari hospital, and immunohistochemical (IHC) staining for mTOR was performed on them. After preparation of the blocks with specific antibodies, tumor cells with cytoplasmic and/or nuclear expression of mTOR were considered as positive cells by applying a specific scoring method introduced in this study.Results: The paraffin blocks of 26 patients were surveyed and the IHC marker of mTOR was positive in the tumors of 10 patients (38.5%). Out of 10 mTOR positive cases, 5 were females and 5 were males. The primary site of the surveyed tumors was the trachea and bronchus in 12 cases (46%), salivary glands in 7 individuals (27%), and lung tissue in 7 cases (27%), and there was no significant correlation between the primary site of the ACC tumors and the existence of the mTOR markers in them (P = 0.67). From all cases, 13 patients (50%) had cribriform and tubular cells without solid components, 9 cases (34.6%) had cribriform and tubular with less than 30% of solid components, and 4 cases (15.4%) had cribriform and tubular cells with more than 30% of solid com-ponents. There was no significant difference between the morphologies and the existence of mTOR markers in them (P = 0.741). Conclusions: As the incidence of mTOR markers is seen in patients with tracheal ACC, evaluation and scoring of mTOR in these persons can be helpful as further studies can distinguish the use of it in the treatment of the disease.

The diagnostic accuracy of endobronchial ultrasound and spiral chest computed tomography scan in the prediction of infiltrating and non-infiltrating lymph nodes in patients undergoing endobronchial ultrasound

Purpose: Endobronchial ultrasound (EBUS) and spiral chest computed tomography (CT) scan are important methods in the prediction of infiltrating and non-infiltrating lymph nodes, and the determination of their diagnostic accuracy would result in a reduction of the burden of problems and an improvement in prognosis. The purpose in this study was to determine the diagnostic accuracy of endobronchial ultrasound and spiral chest CT scan in the prediction of infiltrating and non-infiltrating lymph nodes in patients undergoing endobronchial ultrasound. Material and methods: In this observational prospective study, 40 consecutive patients with infiltrating and non-infiltrating lymph nodes in Masih-Daneshvari Hospital in 2017 and 2018 were enrolled, and the sensitivity, specificity, and accuracy of EBUS and CT-scan versus fine needle aspiration pathology results were determined in them. Results: The results in this study demonstrated that the congruence between EBUS and CT scan was 80.5% (p = 0.0001). The sensitivity, specificity, and accuracy for CT scan were 100%, 22.6%, and 40%, respectively, and the sensitivity, specificity, and accuracy for EBUS were 100%, 16.1%, and 35%, respectively. Conclusion: According to the obtained results, it may be concluded that CT scan and EBUS results have good congruence and high sensitivity to differentiate infiltrating and non-infiltrating lymph nodes. Hence, these methods are useful for screening methods, but due to their low specificity and accuracy the use of them for a confirmative approach is not beneficial. However, regarding the accessibility and less invasive nature, use of chest CT scan is more rational and is recommended in these patients

An eleven-year retrospective cross-sectional study on pulmonary alveolar proteinosis

Introduction: Pulmonary alveolar proteinosis (PAP) is a rare disease in the field of pulmonary medicine. The efficacy of whole-lunglavage (WLL) as the treatment of PAP had never been evaluated in the Iranian population. Therefore, there is a real need to investigatethe characteristics of PAP and also to evaluate the efficacy of WLL in this rare disease. The study aimed to investigatedemographic features, clinical presentation and treatment outcomes of the disease in Iranian PAP patients. Material and methods: Data of 45 patients with definite diagnosis of PAP, who had regular follow-ups from March 2004 to March2015 at an Iranian referral respiratory hospital, were collected. Whole-lung lavages (WLL) efficacy was assessed by comparingspirometric, arterial blood gas parameters and six-minute walk test (6MWT) results before and after all lavages. Results: Mean age at diagnosis of disease was 30.33 ± 14.56 years. Four patients (8.8%) reported non-massive hemoptysis and threesubjects (6.6%) had concomitant pulmonary tuberculosis. In 71.1% of cases, transbronchial lung biopsy and bronchoalveolar lavage weresufficient for diagnosis. Spirometric results and arterial blood gas parameters and 6MWD improved significantly after all the lavages. Fourpatients (8.8%) died because of respiratory failure. The only variable capable of predicting treatment failure was the history of hemoptysis. Conclusion: The study revealed sufficiency of WLL as the PAP patients’ treatment. Also hemoptysis was found to be the independentfactor that can predict treatment failure

Procalcitonin and Proinflammatory Cytokines in Early Diagnosis of Bacterial Infections after Bronchoscopy

BACKGROUND: Fiberoptic bronchoscopy (FOB) guided bronchoalveolar lavage (BAL) remains as the chief diagnostic tool in respiratory disorders. 1.2-16% of patients frequently experience fever after bronchoscopy. To exclude the need for multiple antibiotic prescribing in patients with post-bronchoscopy fever, the presence of the self-limiting inflammatory responses should be excluded. AIM: The current study was conducted to test the serum of patients undergoing bronchoscopy for some proinflammatory cytokines including Tumor Necrosis Factor-alpha (TNF-ɑ), Interleukin-1beta (IL-1β), Interleukin-8 (IL-8) and Interleukin-6 (IL-6) and the value of Procalcitonin (PCT). MATERIAL AND METHODS: Current case-control study was conducted at the National Research Institute of Tuberculosis and Lung Disease in Iran. Nineteen patients (48.72%) that attended with a reasonable sign for a diagnostic bronchoscopy from January 2016 to December 2017 were included in the case group. The control group consisted of 20 patients who underwent a simple bronchoscopy and without FOB-BAL. The laboratory findings for PCT concentrations and cytokine levels in the three serum samples (before FOB-BAL (t0), after 6 hr. (t1), and at 24 hr. past (t2) FOB-BAL) were compared between two groups. RESULTS: The frequency of post-bronchoscopy fever was 5.12, and the prevalence of post-bronchoscopy infectious fever was 2.56%. PCT level was considerably higher in the patient with a confirmed bacterial infection when compared to other participants (p-value < 0. 05). Interestingly, IL-8 level in the bacterial infection proven fever patient was higher than in other patients (p < 0.001). IL-8 levels displayed a specificity of 72.7% and a sensitivity of 100%, at the threshold point of 5.820 pg/ml. PCT levels had a specificity of 84% and a sensitivity of 81%, at the threshold point of 0.5 ng/ml. CONCLUSION: The present findings show that in patients with fever after bronchoscopy, PCT levels and IL-8 levels are valuable indicators for antibiotic therapy, proving adequate proof for bacterial infection. The current findings also illustrate that to monitor the serum levels of PCT and proinflammatory cytokines in the patients undergoing FOB-BAL, the best time is the 24-hour postoperative bronchoscopy

Abdominal Compartment Syndrome in Critically Ill Patients

BACKGROUND: Abdominal compartment syndrome patients suffer severe obstacles such as kidney failure and shock. To evade further complications, identifying the abdominal compartment syndrome (ACS) and Intra-abdominal hypertension (IAH), in critically ill individuals and hospitalised in the intensive care unit (ICU) is obligated. AIM: The current study intended to study the abdominal compartment syndrome and the concomitant risk factors among hospitalised patients in ICU, by using the Intra-abdominal pressure test. MATERIAL AND METHODS: One hundred and twenty-five hospitalised patients at ICU entered the current survey. Abdominal pressure was measured by standard intravesical technique. The SPSS 21 analysed the preoperative and intraoperative factors such as demographic records and comorbidities. RESULTS: Seventy-three (58.4%) participants were males, and 52 (41.6%) were women in the mean age of 55.1 ± 18.3 years. Eighty-nine patients (71.2%) showed normal intra-abdominal pressure since 31 patients (24.8%), and 5 patients (4%) developed IAH and ACS. The intra-abdominal pressure (IAP) applied to Glasgow Coma Scale (GCS), Acute Physiology, shock, Systemic Inflammatory Response Syndrome (SIRS), central venous oxygen saturation and Chronic Health Evaluation (APACHE II) score (P < 0.05). Patients with high IAP have shown a higher mortality frequency, compared to others (P < 0.05). CONCLUSION: Current findings showed a correlation between IAP hospitalised patients in ICU and shock, SIRS, APACHE II, central venous oxygen saturation and GCS. Intra-abdominal pressure test, as a valuable prognosis test for the abdominal compartment syndrome (ACS) and Intra-abdominal hypertension (IAH), may offer better results when added to the routine medical checkup of ICU patients