VARIATIONS IN MONOGENIC DIABETES AND DIABETES SUSCEPTIBILITY GENES IN PEDIATRIC CASES: SINGLE CENTER EXPERIENCE.

Context: Diabetes is a chronic disorder with a complex pathogenetic background including monogenic, polygenic, and environmental causes. Objective: The aim of the present paper is to share the information related to genetic and clinical data of large pediatric diabetes cohort. Design: The present study retrospectively analyzes genetic and clinical findings of subjects diagnosed with diabetes under the age of 18 year and are in follow-up in a pediatric diabetes referral center. Subjects and Methods: Out of 1205 children with diabetes (902 treated with insulin) 246 underwent genetic tests on the basis of clinical selection criteria since 2007. Results: One hundred and ten variants related to diabetes were found in 89 of them. Age at presentation was 9.5±4.02 years (F/M 44/45). In total 49 pathogenic and likely pathogenic, 11 "hot and warm" of unknown significance variants were found in fourteen MODY and fifteen non-MODY genes according to criteria developed by American College of Medical Genetics. Thirty novel mutations were found. GCK (26.6%) and ABCC8 (10%) were two most frequently affected genes. Antibody testing revealed negative results in 80% of cases. Conclusions: Genetic interpretation in selected cases is important to understand the nature of the disease better. Improvement in testing opportunity and awareness might increase the prevalence of genetically explained diabetes cases. The distribution of subtypes differs between countries and even regions of the same country.

The detection of curcumins' antitumoral effects via argyrophilic nucleolar organizing region-associated protein synthesis in mice with ehrlich's ascitic carcinoma.

BACKGROUND: Curcumin is a polyphenol compound that has antioxidant, anticancer, anti-inflammatory, anti-hyperlipidemic and antimicrobial effects. Nucleolar-organizing regions are the sites of the gene on chromosomes. The present study was aimed to show the antitumoral effect of curcumin via AgNOR protein synthesis in Ehrlich's ascitic carcinoma (EAC) bearing mice. METHODS: Twenty three mice with EAC were randomly divided into 3 groups as positive control (n = 7), group 2 (n = 8) and 3 (n = 8) treated intraperitoneally with curcumin (25 mg/kg) and (50 mg/kg), respectively. The animals were sacrificed on Day 16, the solid tumors were removed out. Then, total AgNOR area/nuclear area (TAA/NA) and the mean AgNOR number were estimated for each mice. RESULT: Statistically significant differences were determined among the whole groups for TAA/NA ratio (p = 0.000), conversely mean AgNOR number (p = 0.361). When comparingthe two groups; while no difference was determined between the control and curcumin (25 mg/kg) groups (p = 0.061), the significant differences were detected between the control and curcumin (50 mg/kg) groups (p = 0.000) and between curcumin (25 mg/kg) and curcumin (50 mg/kg) groups (p = 0.000) for TAA/NA ratio. However, there was no significant difference for the mean AgNOR number in double comparison of the groups. CONCLUSIONS: The current study showed that curcumin had a crucial function against cancer development. Also, both AgNOR values might be used as biomarkers for detection of the most reliable therapeutic dose selection of cancer treatment (Tab. 3, Fig. 2, Ref. 27).

Investigation of argyrophilic nucleolar organizing region.

BACKGROUND: Ischemia/reperfusion (I/R) injury is a complex event frequently observed in vascular surgery and can cause functional and structural cell damage. Nucleolar-organizing regions (NORs) are sites of the ribosomal genes located on chromosomes and can be stained with silver when they are active. Thus these proteins are named as argyrophilic-NOR (AgNOR)-associated proteins. We aimed to investigate any possible effects of renal I/R injury on the NOR protein synthesis and association between the AgNOR proteins amount and histopathological injuring score. METHODS: Nine female wistar-albino rats with weight of 200-250g were included into the study. The animals were randomly divided in two groups, a Control Group and an I/R Group. In I/R group, rats were subjected to 45 minutes of renal pedicle occlusion followed by 24 hours of reperfusion. In the control group no drug injections or ischemia reperfusion were performed in animals. Then histopathological injury score, mean AgNOR number and total AgNOR area/nuclear area (TAA/NA) were detected for each rat. RESULTS: The differences between control and I/R groups were significant for histopathological injury scores (p = 0.016). Also the differences between control group and I/R group were significant for mean AgNOR number (p = 0.000) and TAA/NA ratio (p = 0.000). Additionally, there was a positive correlation between TAA/NA ratio and histopathological injury score (r = 0.728; p = 0.026) and between mean AgNOR number and histopathological injury score (r = 0.670; p = 0.048). CONCLUSION: The detection of AgNOR proteins amount may be used as an indicator to obtain information about the cellular behaviour (self-protective mechanism of tubular epithelial cells) against I/R injury and cellular damage levels (Tab. 2, Fig. 4, Ref. 24).

THE RELATIONSHIP BETWEEN CONGENITAL HEART DEFECTS AND e-NOS GENE IN DOWN SYNDROME.

The aim of the study was to compare the effects of three eNOS gene polymorphisms associated with congenital heart defects, between Down syndrome patients with and without cardiac anomalies. Transthoracic echocardiographic examinations and eNOS single-nucleotide polymorphisms were investigated on seventy-five patients, prospectively. The frequencies of mutant alleles in the eNOS promoter (the -786T/C polymorphism) and exon 7 mutant alleles (the 894G--->T polymorphism) were significantly higher in Down syndrome patients with and without cardiac anomalies. The frequency of the intron GIOT polymorphism did not significantly differ between patients with and without cardiac anomalies. We found a significant relationship between eNOS gene polymorphisms and the congenital heart defects in patients with Down syndrome. Screening for the presence or absence of eNOS polymorphisms may be useful to obtain preliminary data on the risk of congenital heart defects in patients with Down syndrome.

Are there any effects of chronic carbon monoxide exposure on argyrophilic nucleolar-organizing region-associated protein synthesis in rat myocardium?

The aims of the study are to detect whether there are any possible effects of chronic carbon monoxide (CO) exposure on the argyrophilic nucleolar-organizing region (AgNOR)-associated protein synthesis and evaluate any possible relationship between the amount of AgNOR protein and the level of myocardial injury also and between AgNOR and histopathological evaluation methods. Adult male albino Wistar rats (n = 18) were randomly divided into three groups (groups A, B, and C). Group A served as control, while groups B and C were rats exposed to CO gas chronically (1000 and 3000 ppm CO concentration with a flow rate of 4 L/min for 30 min/day for 7 days, respectively). Total AgNOR area/nuclear area (TAA/NA) and the mean AgNOR numbers for each myocyte nucleus were determined. There were significant differences among all groups for TAA/NA ratio. These differences were not significant for mean AgNOR numbers. According to the histopathological evaluation scores, there were significant differences between the groups. The differences were significant among the groups for loss of sarcomere pattern. A strong positive correlation between histopathological injury scores and TAA/NA ratio was found (Rsq = 0.48; p = 0.002), however, the correlation was not significant for mean AgNOR numbers (Rsq = 0.08; p = 0.25). In conclusion, TAA/NA ratio can be used as an indicator for obtaining information about the level of myocardial damage instead of histopathological evaluation scores.

Can argyrophilic nucleolar organizing region-associated protein amount be used for the detection of cardiac damage?

INTRODUCTION: Carbon monoxide (CO) is a colorless, tasteless, odorless, nonirritant gas and CO poisoning affects all organ systems. AIM: We aimed to detect any possible effects of CO exposure on the argyrophilic nucleolar organizing region (AgNOR)-associated protein synthesis of heart cells and whether there is any relationship between AgNOR protein amount and both carboxyhemoglobin (COHb) level and histopathological evaluation methods used for the detection of damage in heart tissue after CO exposure. MATERIALS AND METHODS: The rats were divided into four groups (control, 1000, 3000, and 5000 ppm), each containing six rats. After CO intoxication, COHb levels were measured and the animals were killed on the 7th day. AgNOR staining was performed in the heart tissue. One hundred nuclei per rat were evaluated, and total AgNOR area/nuclear area and mean AgNOR number were analyzed for each nucleus. The CO exposure groups had significantly higher AgNOR values than the control group (p < 0.0001). According to cardiomyopathy (CMY) scoring methods, the differences between groups 3 and 4 and groups 1 and 2 were significant (p < 0.05). A significant positive correlation between AgNOR values and both CMY and COHb levels were detected. CONCLUSION: The detection of AgNOR protein amount may give information about the CMY levels and be used to detect the CO intoxication levels instead of COHb in later periods.

May argyrophilic nucleolar organizing region-associated protein synthesis be used for selecting the most reliable dose of drugs such as rhamnetin in cancer treatments?

BACKGROUND: Rhamnetin is a flavonoid that has antioxidant, anti-inflammatory and anti-cancer effects. Nucleolar-organizing regions are the ribosomal genes region. We aimed to identify whether rhamnetin has an effect on cell proliferation and whether AgNOR proteins may be used for the detection of therapeutic benefits of the drugs and new metabolites, which have the potential of being used for cancer treatments. METHODS: Twenty-four mice with Ehrlich's ascites carcinoma (EAC) were randomly assigned to three main groups as positive control, and groups 2 and 3 treated intraperitoneally with rhamnetin (100 µg/kg and 200 µg/kg, respectively). All the animals were sacrificed on day16, 24 h after the last dose; the tumors, which developed at the site of injection were removed. Then, mean AgNOR number and total AgNOR area/nuclear area (TAA/NA) were detected for each mouse. RESULTS: Significant differences were detected among all groups for mean AgNOR number (p = 0.000) and TAA/NA ratio (p = 0.000). While the difference between positive control and Rhamnetin (100 µg/kg) group was not significant (p = 0.387), there are significant differences between positive control and Rhamnetin (200 µg/kg) group (p = 0.000) and between Rhamnetin (100 µg/kg) and Rhamnetin (200 µg/kg) groups (p = 0.000) for TAA/NA ratio. CONCLUSION: Rhamnetin has an important role in preventing cancer formation. Our study showed that mean AgNOR numbers and TAA/NA values may be used also as biomarkers for evaluating the success rate of the performed therapeutic strategy and accurate dose selection for the management of the disease (Tab. 3, Fig. 3, Ref. 45).

MEGALENCEPHALIC LEUKOENCEPHALOPATHY WITH SUBCORTICAL CYSTS WITH HOMOZYGOUS MUTATION (C.448DELC, P.LEU150 SER FSX11) ON EXON 6 OF MLC1 GENE.

MLC or Van der Knaap disease is a rare entity, a rare and genetically heterogeneous cerebral white matter disease. It is characterized by the presence of macrocephaly, epilepsy and a slowly progressive spastic cerebellar syndrome. It is an autosomal recessive disease caused from mutations of MLC1 gene. In the current case report, a case with MLC who had a homozygous mutation (c.448delC, p.Leul50 ser fsX11) on exon 6 of MLC1 gene is presented.

Comparison of fine needle aspiration biopsy and paraffin embedded tissue sections for measuring AgNOR proteins.

Paraffin embedded tissue sections and fine needle aspiration biopsy (FNAB) are important methods for diagnosis. We compared thyroid tissue obtained by FNAB to paraffin embedded sections to determine whether there were differences in detection of the amounts of argyrophilic nucleolar organizing region (AgNOR) proteins. Twenty-two patients with papillary thyroid carcinoma were included in the study. Slides were prepared with both FNAB tissue and 3 µm sections of paraffin embedded tissue, and stained for AgNOR. One hundred nuclei per individual were evaluated; total AgNOR number/nucleus (TAn/TNn) and total AgNOR area/nuclear area (TAa/TNa) of individual cells were determined. Mean TAn/TNn and TAa/TNa values were 4.800 ± 1.118 and 13.382 ± 2.612, respectively, for FNAB samples; corresponding values were 2.406 ± 0.649 and 8.49 ± 0.893, respectively, for paraffin embedded sections. The differences between FNAB materials and paraffin embedded tissue sections were significant for the mean TAn/TNn and TAa/TNa values. Significant differences in the amounts of AgNOR protein detected were found between FNAB and paraffin embedded tissue sections.

Argyrophilic nucleolar organizing region associated protein synthesis for cytologic discrimination of follicular thyroid lesions.

Fine needle aspiration biopsy (FNAB) of the thyroid gland is an important tool for preoperative diagnosis; however, its benefit is limited for follicular lesions. Nucleolar organizer regions (NORs) are ribosomal gene regions that stain with silver (Ag) when they are active. These regions can be used to differentiate neoplastic and non-neoplastic lesions. We used a new AgNOR technique to investigate FNAB of cases diagnosed as follicular adenoma and carcinoma. Fourteen cases of follicular thyroid carcinoma (FTC) and 28 cases of thyroid follicular adenomas (FA) were stained using the silver NOR-associated protein (AgNOR) technique. One hundred nuclei per sample were examined, AgNORs were counted, and the total AgNOR area/nuclear area (TNORa/Na) ratio of each cell was calculated. We found that cases with FTC had significantly higher TNORa/Na than cases of FA. Also, cases with FTC had significantly higher AgNOR counts than cases with FA. AgNOR counting may help discriminate FTC and FA by routine cytopathology before surgery.

Is one-time carbon monoxide intoxication harmless? Evaluation by argyrophilic nucleolar-organizing regions staining method.

In carbon monoxide (CO) poisoning, CO affects the oxygen-carrying capacity of the hemoglobin molecule. Nucleolar-organizing regions (NORs) are genetic loci on chromosomes that are composed of ribosomal DNA and proteins. NORs can be stained with silver. A total of 18 rats were exposed to CO in three different concentrations (1000, 3000, and 5000 ppm) with 6 rats as controls. The animals were euthanized 7 days after CO intoxication. Lung tissues were taken, embedded in paraffin blocks, and sectioned at 5 µm thickness. Argyrophilic nucleolar-organizing region (AgNOR) staining was carried out. One hundred nuclei per individual were evaluated, and total AgNOR number per total nuclear number and total AgNOR area per nuclear area (TAA/NA) for each nucleus were analyzed. The CO exposure groups had significantly higher TAA/NA values and AgNOR numbers than the control group (p < 0.05). Although the differences between 1000 ppm and the other two CO-exposed groups were meaningful (p < 0.05) in the TAA/NA values, there were no differences among the CO exposure groups for the AgNOR number (p > 0.05). The increase in TAA/NA value depends on the increase in the CO exposure. Significant correlations between both the AgNOR values and histopathological scoring methods were found. Therefore, AgNOR staining method may be used as an indirect indicator for evaluating the degree of cell damage rate.

Anophthalmia-plus syndrome with unusual findings. A clinical report and review of the literature.

We present a male child at 3 years old with Anophthalmia-Plus Syndrome (APS). He has asymmetry of the face and head, left choanal atresia, a sunken facial appearance, microphthalmia in the right eye, severe microphthalmia in the left eye, bilateral low-set ears, scarring from cleft palate surgery. Magnetic resonance imaging (MRI) sections revealed decreased right globe volume, an undeveloped left globe, decreased left optical nerve thickness, Chiari type 2 malformation, left choanal atresia and cleft palate. Echocardiography and abdominal ultrasonography were normal. The patient has a 45 dB conductive hearing loss in the left ear. Repeated thyroid function tests were evaluated as compatible with central hypothyroidism. We report a Fryns Anophthalmia-Plus Syndrome in a child with unusual findings including central hypothyroidism, chiari type 2 malformation, conductive hearing loss and developmental regression. Summary of the features reported in the present case and all 14 previous cases that might be defined as APS.

Argyrophilic nucleolar organizing region associated protein synthesis in hair root cells of humans at different developmental stages and sex.

Argyrophilic nucleolar organizing region (AgNORs) associated proteins are important for cell proliferation and various diseases. We investigated AgNOR protein synthesis in hair root cells of males and females at different ages using two-dimensional image analysis. Experiments were performed on 58 healthy male and 24 healthy female volunteers in three groups according to age and sex. Hair root cells obtained from hair follicles were stained with silver. Total AgNOR number/total nuclear number (TAN/TNN) and total AgNOR area/nuclear area (TAA/NA) for each nucleus were analyzed. The only significant difference was observed in TAA/NA values for males and females from 6 to 12 years old. We suggest that the difference is due to high NOR activity caused by increased growth hormone production in hair root cells.

Does higher NORs expression affect the developmental stages of Down syndrome infants?

BACKGROUND: The nucleolar organizer regions (NORs) are localized at the secondary constriction of the five pairs of acrocentric chromosomes (13, 14, 15, 21 and 22) in human. MATERIALS AND METHODS: To evaluate whether increasing AgNOR protein synthesis effects or not the development of babies/children, 25 Down syndrome patients were included in this study. Firstly, the Ankara Development Screening Inventory (AGTE) test was performed. Then the buccal epithelial cells of patients were taken via a sterile toothpick on clean glass slides and spreaded and AgNOR staining technique was applied to the slides of each individual. Mean NOR area/Total nucleus area (NORa/TNa) were evaluated for each nucleus using a special computer program. RESULTS: The mean NORa/TNa was found to be 3.8+/-1.16. According to these data, a significant correlation was not evident between the NORa/TNa and developmental stages (p>0.05). CONCLUSIONS: There is no correlation between extra energy spending for NOR protein synthesis and developmental deficiency.

Is there any relationship between decreased AgNOR protein synthesis and human hair loss?

Argyrophilic nucleolar organizing region associated proteins (AgNORs) play roles in cell proliferation and a variety of diseases. We attempted to determine whether decreased NOR protein synthesis causes human hair loss. We studied 21 healthy males who suffered hair loss on the frontal/vertex portion of the head. Hair root cells from normal and hair loss sites were stained for AgNOR. One hundred nuclei per site were evaluated and the AgNOR number and NORa/TNa proportions of individual cells were determined using a computer program. The cells from normal sites had significantly higher AgNOR counts than those from hair loss sites. Also, the cells from the normal sites had significantly higher NORa/TNa than cells from the hair loss sites. In the normal sites, the cells demonstrated more NOR protein synthesis than cells in hair loss sites. Therefore, decreased NOR protein synthesis appears to be related to hair loss in humans.

EEC syndrome with a de novo mutation (c.953g > a) on exon 7 of P63 gene: a case report.

EEC syndrome is characterized by ectodermal dysplasia, ectrodactyly and cleft lip and/or palate and associated anomalies such as lacrimal duct obstruction, urinary tract anomaly, and hearing loss. This syndrome is a rare autosomal dominant disorder caused by heterozygous mutations in the p63 gene. Herein, a newborn infant with EEC syndrome with secundum atrial septal defect who had a de novo mutation (c.953G > A) on exon 7 of p63 gene is presented.