Investigation of the molecular genetic causes of non-syndromic primary ovarian insufficiency by next generation sequencing analysis.

Objective: The aim of this study is to investigate the molecular genetic causes of non-syndromic primary ovarian insufficiency (POI) cases with the gene panel basedon next generation sequencing analysis and to establish the relationship between genotype and phenotype. Materials and methods: Twenty three cases aged 14-40 years followed up with POI were included. Patients with a karyotype of 46, XX, primary or secondary amenorrhea before the age of 40, with elevated FSH (>40 IU/mL) and low AMH levels (<0.03 ng/mL) were included in the study. Molecular genetic analyzes were performed by the next generation sequencing analysis method targeted with the TruSight TM Exome panel. Results: Median age of the cases was 17.8 (14.0-24.3) years, and 12 (52%) cases admitted before the age of 18. Fifteen (65%) patients had consanguineous parents. In2 (8.6%) cases, variants detected were in genes that have been previously proven to cause POI. One was homozygous variant in FIGLA gene and the other was homozygous variant in PSMC3IP gene. Heterozygous variants were detected in PROK2, WDR11 and CHD7 associated with hypogonadotropic hypogonadism, but these variants are insufficient to contribute to the POI phenotype. Conclusion: Genetic panels based on next generation sequencing analysis technologies can be used to determine the molecular genetic diagnosis of POI, which has a highly heterogeneous genetic basis.

The expression of HDAC9 and P300 in papillary thyroid carcinoma cell line.

PURPOSE: Papillary thyroid carcinoma (PTC) is the most common type of thyroid cancer and accounts for 85-90% of all thyroid cancers. Metastatic differentiated thyroid cancer, radioiodine-refractory thyroid cancer, and anaplastic thyroid cancer still lack effective therapeutic options. Here, we aimed to assess HDAC9 and P300 expression in the papillary thyroid carcinoma cell line and compare them with normal thyroid cells. METHODS: Nthy-ori-3-1, a normal thyroid cell line, and BCPAP, a PTC cell line, were cultured for 24 and 48 h and immunofluorescence staining was used to determine the levels of HDAC9 and P300 protein expression. HDAC9 paracrine release was assessed using an ELISA assay. RESULTS: HDAC9 protein expression was higher in both cell groups at the 48th hour than at the 24th hour; however, P300 protein expression was lower in BCPAP cells at the 48th hour than at the 24th hour. In comparison to Nthy-ori-3-1, BCPAP expressed more HDAC9 and P300 proteins. HDAC9 secretion slightly increased in Nthy-ori-3-1 cells from 24 to 48 h. Furthermore, HDAC9 secretion in BCPAP cells dramatically decreased from 24 to 48 h. CONCLUSION: Our findings revealed that the expression of HDAC9 and P300 was higher in the PTC cell line than in normal thyroid cells. This indicates that the acetylation mechanism in thyroid cancer cells is not the same as it is in healthy cells. Epigenetic studies may reveal the mechanisms underlying PTC with further analysis.

Stemness potency and structural characteristics of thyroid cancer cell lines.

BACKGROUND: Thyroid cancer is the most frequent type of endocrine malignancy. Thyroid carcinomas are derived from the follicular epithelium and classified as papillary (PTC) (85%), follicular (FTC) (12%), and anaplastic (ATC) (<3%). Thyroid cancer could arise from thyroid cancer stem-like cells (CSCs). CSCs are cancer cells that feature stem-like properties. Kruppel-like factor (KLF4) and Stage-spesific embryonic antigen 1 (SSEA-1) are types of stem cell markers. Filamentous actin (F-actin) is an essential part of the cellular cytoskeleton. The purpose of this study was to evaluate the stem cell potency and the spatial distribution of the cytoskeletal element F-actin in PTC, FTC, and ATC cell lines. MATERIALS AND METHODS: Normal thyroid cell line (NTC) Nthy-ori-3-1, PTC cell line BCPAP, FTC cell line FTC-133 and ATC cell line 8505c were stained with SSEA-1 and KLF4 for stem cell potency and F-actin for cytoskeleton. The morphological properties of cells were assessed by a scanning electron microscope (SEM) and elemental ratios were compared with EDS. RESULTS: PTCs had greater percentages of SSEA-1 and KLF4 protein intensity (0.32% and 0.49%, respectively) than NTCs. ATCs had a greater proportion of KLF4 expression (0.8%) than NTCs. NTCs and FTCs had increased F-actin intensity across the cell, but PTCs had the lowest among these four cell lines. NTCs and PTCs, as well as NTCs and FTCs, have statistically identical aspect ratios and round values. These values, however, were statistically different in ATCs. CONCLUSION: The study of stem cell markers and the cytoskeletal element F-actin in cancer and normal thyroid cell lines may assist in the identification of new therapeutic targets and contribute in the understanding of treatment resistance mechanisms.

High chitotriosidase and AGE levels in acromegaly: a case-control study.

PURPOSE: Acromegaly is associated with oxidative stress and inflammation parameters. Chitotriosidase (CHITO) is a marker of macrophage activation and plays a pivotal role in the activation of inflammatory and immunological responses. Our study aimed to determine CHITO,YKL-40, advanced glycation end product (AGE), and high-sensitivity C-reactive protein (hsCRP) levels to investigate malondialdehyde (MDA), catalase, superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px) activities and to evaluate any association of these parameters with carotid intima media thickness (cIMT) in patients with controlled acromegaly. METHODS: Thirty controlled acromegaly patients and 41 age- and sex-matched control cases were studied. We obtained demographic data, hormonal and metabolic parameters, and cIMT. CHITO activity was measured with the fluorometric method of Chamoles et al. YKL-40 and hsCRP levels were measured using ELISA. AGEs were measured based on spectrofluorimetric detection. GSH-Px activity was determined by a colorimetric assay. MDA, SOD, and catalase activities were determined in hemolysis. RESULTS: Higher CHITO, AGE, and hsCRP concentrations were observed in patients with acromegaly compared to controls. SOD levels were non-significantly higher in the acromegaly group, while catalase activities were lower in patients with acromegaly. Correlation analyses of CHITO, AGEs, YKL-40, hsCRP, MDA, catalase, GSH-Px, and SOD with metabolic, anthropometric, and laboratory parameters did not demonstrate any significant correlation (p > 0.05). There was no significant difference between groups with regard to cIMT levels. CONCLUSION: This is the first study investigating CHITO and AGE levels in patients with acromegaly. Serum CHITO, AGE, and hsCRP levels in acromegalic patients were significantly increased. It may be important to evaluate CHITO, AGE, and hsCRP levels in acromegalic patients who are already under cardiometabolic surveillance due to risk of developing cardiovascular disease.

Two challenging cases of pituicytoma.

INTRODUCTION: Pituicytoma is a rare tumor of the pituitary gland derived from neurohypophyseal pituicytes. CASE 1: A 58-year-old female presented with decreased vision; she was admitted to the neurosurgery department of Ege University after the detection of a pituitary macroadenoma. Magnetic resonance imaging (MRI) showed a 28 * 18 * 17-mm suprasellar mass, and laboratory tests revealed hypopituitarism. Hydrocortisone and L-thyroxine treatment were initiated, and the patient underwent resection through the endoscopic endonasal approach (EEA). The histopathological examination revealed a pituicytoma. The recurrence of tumor was detected during the 1-year follow-up, and the patient is awaiting surgery. CASE 2: A 70-year-old woman presented with visual changes; she had a past medical history of hypophyseal macroadenoma and pituicytoma resected through an EEA in 2012 and 2017, respectively. During follow-up, 2 years after the second surgery, MRI showed progression of the pituicytoma then measuring 38 × 23 × 22 mm; it had invaded the cavernous sinus and was causing hydrocephaly and panhypopituitarism. The patient underwent the third resection through the transcranial approach in order to minimize bleeding. After this surgery, the patient developed diabetes insipidus and underwent treatment with desmopressin. Histopathological examination revealed a pituicytoma. At 6-month follow-up, imaging showed a sellar suprasellar mass 37 × 22 × 24 mm invading the cavernous sinus, indicative of recurrence. In the postoperative period, the patient applied to the department of radiation oncology to have fractionated radiotherapy. DISCUSSION: Pituicytomas are known to be low-grade tumors; because of their rarity, they are a real challenge. These patients should be followed up closely.

The effect of serum lipid levels on peripheral blood hematopoietic stem cell levels.

INTRODUCTION: There are limited numbers of available retrospective studies on various hematological diseases treated with stem cell mobilization therapy. In the present study, we aimed to demonstrate the effects of serum lipid levels on peripheral blood CD34+ (PBCD34+) cell counts as well as the changes in serum lipid levels during stem cell mobilization process. METHOD: PBCD34+ cell counts were compared between hypercholesterolemic patients and healthy individuals. Additionally, total cholesterol (TChol), LDL-cholesterol (LDL-C), HDL-cholesterol (HDL-C), and triglyceride (TG) levels were measured from healthy donors who underwent stem cell mobilization, at different time points (prior to filgrastim [phase 1], prior to apheresis [phase II], and the first week following apheresis [phase III]. RESULTS: In the hypercholesterolemia group, the PBCD34+ cell count was found to be higher among patients with elevated LDL-C (2.6 ± 0.35/µL vs. 1.7 ± 0.17/µL, p = 0.003) and TChol (2.6 ± 0.34/µL vs. 1.7 ± 0.14/µL, p = 0.006) in comparison to the healthy controls. In the mobilization group, phase II HDL-C levels (35.3 ± 2.8 mg/dL) were found to be lower than both phase I (45.6 ± 2.1 mg/dL) and phase III (44.5 ± 2.6 mg/dL) (p = 0.007). Phase II TChol levels (183.5 ± 10.0 mg/dL) were lower than both phase I (216.8 ± 8.5 mg/dL) and phase III (212.2 ± 8.4 mg/dL) (p = 0.02). At phase II, there was an inverse correlation between PBCD34+ cell count and HDL-C (r = - 0.57, p = 0.003). DISCUSSION: Our results indicate that, while increased LDL-C level is the determinant of baseline PBCD34+ cell count, reduced HDL-C is the determinant of PBCD34+ cell count during mobilization process.

Metreleptin replacement treatment improves quality of life and psychological well-being in congenital generalized lipodystrophy.

The near total lack of subcutaneous fat in congenital generalized lipodystrophy (CGL) leads to the accumulation of fat in ectopic organs and severe insulin resistance, which are associated with serious metabolic abnormalities. Cosmetic aspects of the disease are likely to affect the quality of life (QoL) and physiological well-being in these individuals. Metreleptin, recombinant human leptin, replacement treatment has been shown to have benefits in treating the metabolic abnormalities of CGL. In a patient with CGL caused by a homozygous AGPAT2 pathogenic variant, we examined QoL and mood alterations (depression and anxiety) caused by this chronic disease. Metreleptin replacement treatment led to dramatic metabolic improvement in our patient. It was also was associated with improvements in QoL, depression and anxiety scores. We suggest that there is need for studies to document the benefit of metreleptin replacement treatment on QoL and physiological well-being in patients with CGL.

Therapeutic plasma exchange in hypertriglyceridemic patients

Background/aim: High triglyceride (TG) levels are associated with increases in atherosclerotic cardiovascular disease (CVD), hepatic steatosis, and pancreatitis. Acute pancreatitis is a condition with high mortality. Therapeutic plasma exchange (TPE) in the treatment of hypertriglyceridemic pancreatitis (HTGP) is a rapid and effective treatment modality. In this study, the results of TPE were evaluated and the frequency of lipoprotein lipase (LPL) mutation in these patients was determined. Materials and methods: TPE was performed in 31 patients with HTGP at the Adult Therapeutic Apheresis Center. Results: A TG level under 500 mg/dL was achieved by applying apheresis at a median of 2 times (IQR 2­2, min 1, max 6) in the 31 cases. LPL mutation was detected in 8 (25.8%) of the 31 hypertriglyceridemia cases. When TG levels before and after TPE were evaluated, the mean TG level before TPE was significantly higher (3132 ± 1472 mg/dL) than the mean TG level afterwards (948 ± 465 mg/dL, P < 0.001). This result represented a decrease of 69.7% TG after TPE. Conclusion: TPE is a safe, fast, and effective treatment modality in experienced centers.

Plasma Levels of Antioxidant Vitamins in Patients with Acromegaly: A Case-Control Study.

OBJECTIVE: The aim of this study was to compare antioxidant vitamin C and vitamin E levels in the non-acromegaly control group and in patients with acromegaly with and without remission. MATERIAL AND METHODS: In this study, 100 cases, acromegaly patients of 57% (n=57, 29F, 28M, mean ages of 49.5±12.1) and control subjects of 43% (n=43, 29F, 14M, mean ages of 49.6±9.2). Acromegaly patients were classified into two groups; active acromegaly (AA; n=33) and controlled acromegaly (CA; n=24). RESULTS: Vitamin C levels were significantly lower in the acromegaly group [7.6 (4.7) mg/L, as median (IQR)] when compared to the control group [12.2 (5.5) mg/L, as median (IQR)] (p <0.001). Vitamin E levels didn't show a significant difference between the acromegaly and the control groups (14.2±3.6 vs. 14.8±3.7, as mean±SD, respectively, p = 0.439). Correlation analysis showed that vitamin C levels were not significantly associated with clinical, anthropometric and laboratory parameters in the acromegaly group. Vitamin E levels were significantly associated with the total cholesterol, triglyceride, LDL-C, HDL-C, APO A1, APO B both in the acromegaly and the control groups. CONCLUSION: This study is the first one to investigate the relationship between the levels of vitamin C & E and anthropometric & metabolic parameters in acromegaly patients and control group. In our study, vitamin C level was significantly lower in the acromegaly group compared to the level in the control group. There was no significant difference in vitamin E levels between the acromegaly and control group.

Obstructive sleep apnea characteristics in patients with well-controlled acromegaly and their compliance with positive airway pressure therapy

Background/aim: Acromegaly is often associated with obstructive sleep apnea syndrome (OSAS) with a frequency between 40% and 80%. The aim of the present study was to evaluate the clinical and polysomnographic characteristics of acromegaly patients with sleep apnea symptoms and to identify positive airway pressure (PAP) adherence in acromegaly patients with OSAS diagnosis.Materials and methods: Twenty-eight well-controlled acromegaly patients (17 males, mean age 48.7 ± 10.1 years) with sleep apnea symptoms were included in this prospective study. Demographic data, anthropometric measurements, and medical history were evaluated. Full-night in-laboratory polysomnography was performed.Results: Polysomnography results showed that 25 patients (89.3%) had OSAS with a mean apnea-hypopnea index (AHI) of 37.7 ± 28.8/h. All 17 male patients were diagnosed with OSAS, whereas 8 female patients (72.7%) had OSAS (P = 0.05). Male patients also had more severe OSAS than females (AHI 48.3 ± 29.0 vs. 21.3 ± 20.1 events/h, respectively; P = 0.012). Twenty-two patients out of 28 were considered to be eligible candidates for PAP therapy. The PAP adherence rate was found to be 50% during follow-up. Conclusion: Our results confirm OSAS as a common disorder in acromegaly patients as well as PAP therapy being required for a majority of patients. Therefore, all acromegaly patients should be assessed in terms of OSAS and be followed closely for the evaluation of PAP adherence.

Testicular Adrenal Rest Tumor (TART) in congenital adrenal hyperplasia.

Congenital adrenal hyperplasia is one of the most common autosomal recessive genetic disorders. Testicular adrenal tumors are significant complications of congenital adrenal hyperplasia. We would like to present two patients of testicular adrenal rest tumors. Patient 1 24 year-old male, he was diagnosed with congenital adrenal hyperplasia at the age of 8 due to precocious puberty. He received hydro-cortisone treatment until the age of 18. Testicular mass had been detected and right radical orchiectomy had been applied 6 months ago and reported as testicular adrenal rest tumor. In scrotal ultrasound, a mixed type mass lesion (6 × 4x3 cm) covering a large part of left testis was observed. The imaging findings were consistent with adrenal rest tumor. The patient took adrenocorticotropic hormone supressive therapy with dexamethasone 0.75 mg once a day. Patient 2, 38 year-old male, he had been followed-up as adrenal insufficiency for 35 years. He underwent right orchiectomy operation due to the testicular mass in 2010 and the pathological examination revealed Leydig cell tumor. In scrotal ultrasound, small multifocal lesions were detected on the left testis and resection was done. It was reported as testicular adrenal rest tumor. He is being followed-up with glucocorticoid treatment according to androgen and adrenocorticotropic hormone levels. Early diagnosis of testicular adrenal rest tumor is significant in preventing irreversible testicular damage and infertility. In the differential diagnosis, we should keep in mind that testicular adrenal rest tumor can mimic other testicular tumors such as primary germ cell tumors.

Natural History of Congenital Generalized Lipodystrophy: A Nationwide Study From Turkey.

CONTEXT: Congenital generalized lipodystrophy (CGL) is a rare autosomal recessive disorder characterized by near-total lack of body fat. OBJECTIVE: We aimed to study natural history and disease burden of various subtypes of CGL. DESIGN: We attempted to ascertain nearly all patients with CGL in Turkey. SETTING: This was a nationwide study. PATIENTS OR OTHER PARTICIPANTS: Participants included 33 patients (22 families) with CGL and 30 healthy controls. MAIN OUTCOME MEASURE(S): We wanted to ascertain genotypes by sequencing of the known genes. Whole-body magnetic resonance imaging was used to investigate the extent of fat loss. Metabolic abnormalities and end-organ complications were measured on prospective follow-up. RESULTS: Analysis of the AGPAT2 gene revealed four previously reported and four novel mutations (CGL1; c.144C>A, c.667_705delinsCTGCG, c.268delC, and c.316+1G>T). Analysis of the BSCL2 gene revealed four different homozygous and one compound heterozygous possible disease-causing mutations (CGL2), including four novel mutations (c.280C>T, c.631delG, c.62A>T, and c.465-468delGACT). Two homozygous PTRF mutations (c.481-482insGTGA and c.259C>T) were identified (CGL4). Patients with CGL1 had preservation of adipose tissue in the palms, soles, scalp, and orbital region, and had relatively lower serum adiponectin levels as compared to CGL2 patients. CGL4 patients had myopathy and other distinct clinical features. All patients developed various metabolic abnormalities associated with insulin resistance. Hepatic involvement was more severe in CGL2. End-organ complications were observed at young ages. Two patients died at age 62 years from cardiovascular events. CONCLUSIONS: CGL patients from Turkey had both previously reported and novel mutations of the AGPAT2, BSCL2, and PTRF genes. Our study highlights the early onset of severe metabolic abnormalities and increased risk of end-organ complications in patients with CGL.

A case of familial partial lipodystrophy caused by a novel lamin A/C (LMNA) mutation in exon 1 (D47N).

BACKGROUND: Familial partial lipodystrophy (FPL) is a rare genetic disorder characterized by selective lack of subcutaneous fat which is associated with insulin resistant diabetes. The Dunnigan variety (FPL2) is caused by several missense mutations in the lamin A/C (LMNA) gene, most of which are typically located in exon 8 at the codon position 482. CASE REPORT: Here, we report on a Turkish family with FPL2 which is caused by a novel heterozygous missense LMNA mutation in exon 1 (D47N, c.139G>A), in the rod domain of lamins A/C. Fat distribution and metabolic features of LMNA D47N mutation were similar to typical codon 482 mutation. Metabolic abnormalities were observed as a form of insulin resistant diabetes, hypertriglyceridemia, low HDL cholesterol and hepatic steatosis. There was no evidence for neuromuscular and cardiac involvement. CONCLUSION: Although it is previously known that alterations in the rod domain of type A lamins are involved in cardiac and neuromuscular diseases, our current observation shows that exon 1 LMNA mutations may be associated with partial lipodystrophy without any cardiac and neurological abnormalities, at least at the time of the presentation.

An Association Study Between Gene Polymorphisms of Folic Acid Metabolism Enzymes and Biochemical and Hormonal Parameters in Acromegaly.

AIM: Folate metabolism is fundamental to several biological functions and required for cell replication, division, and survival. The mammalian folic acid cycle is highly complex and the enzymes, methylenetetrahydrofolate reductase (MTHFR), methionine synthase (MTR), and methionine synthase reductase (MTRR), have crucial roles in this metabolic pathway. The common polymorphisms of the MTHFR (C677T and A1298C), MTRR (A66G), and MTR (A2756G) enzymes are well documented as folate deficiency-related disorders, but their roles have not been examined in acromegalic patients. The aim of this study was to compare the genotypic distribution of these gene polymorphisms between patients with acromegaly and controls and explore whether these polymorphisms were associated with biochemical and hormonal parameters in acromegaly. We examined 91 acromegaly patients and 112 healthy subjects who were compared in terms of age and gender. Blood specimens of the subjects were collected in tubes containing ethylenediaminetetraacetic acid. Genomic DNA was isolated from peripheral blood leukocytes and genotyping of the MTHFR (C677T and A1298C) gene polymorphisms was assessed by melting temperature analyses after real-time polymerase chain reaction (PCR), whereas MTRR A66G and MTR A2756G gene polymorphism analyses were performed by PCR/restriction fragment length polymorphism from the isolated DNA of the subjects. RESULTS: MTHFR-677TT genotype frequency was significantly higher in the acromegaly group than the control group (p=0.017), and a significant increase was found in fibrinogen (p=0.032) levels in 677TT-carrying acromegaly patients. MTRR-66AA genotype was significantly higher in the control group than the acromegaly group (p=0.004). Total cholesterol (p=0.048) and C-reactive protein (p=0.046) levels decreased significantly in 66AA genotypes. Although MTR-2756AG genotype frequency was not different between the control and acromegaly groups, 2756AG genotype-carrying individuals have higher left carotid intima-media thickness levels within the patient group. CONCLUSION: Our results suggest that polymorphisms of the genes encoding the folic acid metabolism enzymes affect biochemical parameters in acromegaly and this may result in predispositions to some complications associated with folate metabolism and acromegaly.

Characteristics and Treatment Results of 5 Patients with Fibrous Dysplasia and Review of the Literature.

Aim. Fibrous dysplasia is a rare bone disease caused by missense mutation leading to abnormal fibroblast and osteoblast proliferation and increased bone resorption. FD can present in monostotic or polyostotic forms. About 3% of FD could be in association with McCune-Albright syndrome (MAS). Because FD is a rare disease, there is limited data in the literature about characteristics of disease and response to treatment. Methods. We present our five cases of FD with general properties and their responses to medical treatment. Results. Two of our patients had polyostotic and three had monostotic FD. One of the polyostotic patients had MAS. One of our patients had surgery for femur fractures, facial asymmetry, and findings of compression. Four patients were given pamidronate; one was given zoledronic acid as bisphosphonate treatment. Bone pain was relieved in all patients with medical treatment. Conclusion. There was a decrease in bone turnover markers to some degree with medical treatment but no radiological improvement was observed.

Medullary carcinoma of the thyroid with axillary metastasis: a case report.

We report a case of axillary lymph node metastasis as a consequence of medullary thyroid carcinoma (MTC) in a 42-year-old man. On January 2009, the patient was referred to us for the management of right cervical lymph node enlargement. Total thyroidectomy was performed with right-sided functional neck dissection. Postoperative histopathology revealed MTC in the right lobe of the thyroid, with extrathyroidal extension and right-sided neck metastases. Multiple left cervical, mediastinal, and right axillary lymphadenopathies were detected at the third year follow-up exam. Left-sided functional neck dissection, axillary lymph node dissection, and mediastinal lymph node dissection were performed, and the pathologic outcomes revealed as the metastatic dissemination of MTC. After a disease-free term for 1 year, multiple metastatic lesions were detected in the patient.