Molecular Genetic Diagnosis with Targeted Next Generation Sequencing in a Cohort of Turkish Osteogenesis Imperfecta Patients and Their Genotype-phenotype Correlation.

Introduction: Osteogenesis imperfecta (OI) is a group of phenotypically and genetically heterogeneous connective tissue disorders that share similar skeletal anomalies causing bone fragility and deformation. This study aimed to investigate the molecular genetic etiology and determine the relationship between genotype and phenotype in OI patients with targeted next-generation sequencing (NGS). Method: In patients with OI, a targeted NGS analysis panel (Illumina TruSight One) containing genes involved in collagen/bone synthesis was performed on the Illumina Nextseq550 platform. Results: Fifty-six patients (female/male: 25/31) from 46 different families were enrolled in the study. Consanguinity between parents was noted in 15 (32.6%) families. Clinically according to Sillence classification; 18(33.1%) patients were considered to type I, 1(1.7%) type II, 26(46.4%) type III and 11(19.6%) type IV. Median body weight was -1.1 (-6.8, - 2.5) SDS, and height was -2.3 (-7.6, - 1.2) SDS. Bone deformity was detected in 30 (53.5%) of the patients, while 31 (55.4%) were evaluated as mobile. Thirty-six (60.7%) patients had blue sclera, 13 (23.2%) had scoliosis, 12 (21.4%) had dentinogenesis imperfecta (DI), and 2 (3.6%) had hearing loss. Disease-causing variants in COL1A1 and COL1A2 genes were found in 24 (52.1%) and 6 (13%) families, respectively. In 8 (17.3%) of the remaining 16 (34.7%) families, the NGS panel revealed disease-causing variants in three different genes (FKBP10, SERPINF1, and P3H1). Nine (23.6%) of the variants detected in all investigated genes were not previously reported in the literature and were classified to be pathogenic according to ACMG guidelines pathogenity scores. In ten (21.7%) families, a disease-related variant was not found in a total of 13 OI genes included in the panel. Conclusion: Genetic etiology was found in 38 (82.6%) of 46 families by targeted NGS analysis. In addition, 9 new variants were assessed in known OI genes which is a significant contribution to the literature.

A case-control study of early-stage radiological markers of endothelial dysfunction and cardiovascular findings in patients with osteogenesis imperfecta: genotype-phenotype correlations.

OBJECTIVES: Osteogenesis imperfecta (OI) is a disease caused by defective collagen synthesis. Collagen type 1 is found in many structures in the cardiovascular system. Endothelial dysfunction, which develops prior to the emergence of structural and clinical signs of atherosclerosis, is believed to play a key role in atherogenesis. Endothelial dysfunction may be detected presymptomatically by non-invasive radiologic methods, such as flow-mediated dilatation (FMD) and carotid intima-media thickness (CIMT). These modalities may provide early indicators of endothelial dysfunction. This cross-sectional comparative study aimed to investigate early-stage radiological markers of endothelial dysfunction and cardiovascular diseases in OI patients and healthy controls and to investigate the correlation of findings with OI genotype. METHODS: Thirty patients diagnosed with OI were paired with thirty healthy age- and gender-matched controls and echocardiogram findings were compared. RESULTS: None of the patients had known underlying cardiovascular disease. The mean age was 13.18 ± 2.91 years. According to Sillence classification, 15 patients had type 1 OI, 10 had type III, and 5 had type IV. Mean CIMT in the OI group was higher in the control group (OI group: 0.42 ± 0.06 vs. healthy controls: 0.34 ± 0.04 mm, p<0.01), and mean FMD percent was lower in the patient group (p<0.01). Left ventricular ejection fraction was 78.97 ± 10.32 vs. 77.56 ± 8.50 %, (OI group: 7.00 ± 3.06 vs. healthy controls: 12.14 ± 1.99, p=0.56), and fractional shortening was 42.68 ± 11.94 vs. 40.23 ± 7.99 %, (p=0.35), in OI patients and controls, respectively. CONCLUSIONS: Pediatric patients with OI without clinical signs of cardiovascular abnormality had significantly worse CIMT and FMD findings than healthy controls. However, no difference was determined when comparing left ventricular ejection fraction or fractional shortening. OI patients may need to be screened for cardiovascular system complications starting from an early age.

Molecular diagnosis in patients with monogenic diabetes mellitus, and detection of a novel candidate gene.

AIM: We aimed to investigate molecular genetic basis of monogenic diabetes (DM) and novel responsible candidate genes with targeted Next Generation Sequencing (NGS) and Whole Exome Sequencing (WES). METHODS: A hundred cases presenting with clinical findings and a family history of monogenic DM were included in the study. Molecular analysis was performed using an NGS panel including 14 genes. Following targeted NGS, WES was planned in cases in whom no variant was detected. RESULTS: Thirty different disease-causing variants in seven different genes were detected in thirty-five (35 %) cases with targeted NGS approach. Most common pathogenic variant was found in GCK gene in 25 (25 %) cases. Four different variants were detected in 4 (4 %) patients in ABCC8 gene. In 45 of 65 cases; WES analyses were done. A heterozygous c.2635C > T(p.Gln879Ter) variant was detected in IFIH1 gene in a patient with incidental hyperglycemia. In the segregation analysis affected mother was shown to be heterozygous for the same variant. CONCLUSION: Molecular etiology was determined in 35 % cases with the NGS targeted panel. Seventeen novel variants in monogenic DM genes have been identified. A candidate gene determined by WES analysis in a case that could not be diagnosed with NGS panel in this study.

ABCC8-related maturity-onset diabetes of the young: switching from insulin to sulphonylurea therapy: how long do we need for a good metabolic control?

OBJECTIVES: Activating variants of the ABCC8 gene cause neonatal diabetes or maturity-onset diabetes of the young (MODY). We report three cases of MODY type 12 caused by variants in the ABCC8 encoding sulphonylurea receptor 1, and the experience of switching from insulin therapy to sulphonylurea therapy. CASE PRESENTATIONS: We describe a 12.5-year-old girl with permanent neonatal diabetes mellitus, and two diabetes mellitus cases with variants in the ABCC8 gene. Two of these cases were successfully switched from subcutaneous insulin to oral glibenclamide, with a marked improvement in glycemic control. In permanent neonatal diabetes case, glibenclamide dose was progressively increased to achieve a full dose (2 mg/kg/day) in 9 days. Nine months after starting oral sulphonylurea therapy, her blood glucose control dramatically improved and insulin therapy was discontinued. CONCLUSIONS: We conclude that patients with ABCC8 gene variants can successfully switch from insulin to sulphonylureas.

Molecular genetic etiology by whole exome sequence analysis in cases with familial type 1 diabetes mellitus without HLA haplotype predisposition or incomplete predisposition.

OBJECTIVES: Familial transmission is observed in approximately 10% of cases with type 1 diabetes mellitus (T1DM). The most important gene determining susceptibility is the human leukocyte antigen complex (HLA) located on chromosome 6. More than 50 susceptible loci are associated with T1DM susceptibility have been identified in genes other than HLA. In this study, it was aimed to investigate the molecular genetic etiology by whole-exome sequence (WES) analysis in cases with familial T1DM with no or weakly detected HLA tissue type susceptibility. We aimed to identify new genes responsible for the development of type 1 diabetes and to reveal new genes that have not been shown in the literature before. METHODS: Cases with at least one T1DM diagnosis in first-degree relatives were included in the study. In the first step, HLA DQ2 and DQ8 loci, which are known to be associated with T1DM susceptibility, were investigated by. In the second step, the presence of variants that could explain the situation was investigated by WES analysis in patients who were negative for both HLA DQ2 and HLA DQ8 haplotypes, HLA DQ2 negative, HLA DQ8 positive, and HLA DQ2 positive and HLA DQ8 negative patients. RESULTS: The mean age and duration of diabetes of the 30 cases (Girl/Male: 17/13) were 14.9 ± 6 and 7.56 ± 3.84 years, respectively. There was consanguineous marriage in 5 (16%) of the families. As a result of filtering all exome sequence analysis data of two cases with DQ2 (DQB1*02) (-) and DQ8 (DQB1*03:02) (-), seven cases with DQ2 (DQB1*02) (+) and DQ8 (DQB1*03:02) (-), and one case with DQ2 (DQB1*02) (-) and DQ8 (DQB1*03:02) (+), seven different variants in seven different genes were detected in five cases. The pathogenicity of the detected variants were determined according to the "American College of Medical Genetics and Genomics (ACMG)" criteria. These seven variants detected were evaluated as high-score VUS (Variants of unknown/uncertain significance). In the segregation study conducted for the mutation in the POLG gene detected in case 5, this variant was detected in the mother of the case and his brother with T1DM. Segregation studies are ongoing for variants detected in other affected individuals in the family. CONCLUSIONS: In conclusion, in this study, seven different variants in seven different genes were detected in five patients by WES analysis in familial T1DM patients with no or weak HLA tissue type susceptibility. These seven variants detected were evaluated as high-score VUS. POLG might be a novel candidate gene responsible for susceptibility to T1DM. Non-HLA genes directly responsible for the development of T1DM were not detected in any of the cases.

Treatment and long-term follow-up of patients diagnosed with type 1 diabetes mellitus before age 5.

OBJECTIVES: This study aimed to determine the effects of continuous subcutaneous insulin infusion (CSII) treatment on anthropometric measurements, mean HbA1c, and insulin dosage in patients diagnosed under 5 years of age and compare with multiple-dose injection therapy (MDI). METHODS: Children with type 1 diabetes mellitus, diagnosed <5 years since 2000 and their 19-year follow-up were evaluated retrospectively. Weight, height, body mass index (BMI), blood pressure, and HbA1c values were recorded for each visit. RESULTS: Hundred and five patients (58.1% female, 41.9% male) were included in the study. Sixty-three (60 %) patients were treated by CSII and 42 (40%) by MDI. Mean age at diagnosis was 2.68 ± 1.42 and 3.29 ± 1.30 years respectively. Mean follow-up was 7.42 ± 4.76 and 6.01 ± 4.41 years respectively. For each group, weight standard deviation score (SDS) increased significantly in the first year after the diagnosis (p<0.001), and with the onset of puberty weight SDS decreased significantly (p<0.001). The trend of weight and BMI SDS changes over the years showed similar characteristics in both groups. During follow-up height SDS was similar in both groups except in Tanner stage 5. When puberty was completed, mean height SDS was 0.51 ± 1.03 in CSII and -0.31 ± 0.75 in the MDI group (p: 0.029). Mean HbA1c was significantly lower in the CSII group (7.62 ± 0.82 and 8.17 ± 1.22 respectively). Systolic and diastolic blood pressure change trends during the follow-up were also similar in both groups. CONCLUSIONS: CSII treatment had positive effects on metabolic control and height SDS in patients with early-onset diabetes without increasing BMI.

The Value of Telemedicine for the Follow-up of Patients with New Onset Type 1 Diabetes Mellitus During COVID-19 Pandemic in Turkey: A Report of Eight Cases

The current Coronavirus disease-2019 (COVID-19) pandemic has forced health care teams to look for alternative approaches to manage a great number of children with diabetes, not only in rural but also in urban locations. The aim was to assess the provision of information about follow-up of new-onset pediatric type 1 diabetes (T1D) patients, and to investigate the integration of telemedicine into routine clinical care in the long term. The changes in coefficient of variation (CV), standard deviation and percentages of time in range (TIR), time below range (TBR) and time above range were evaluated in eight children with new-onset T1D, diagnosed during the COVID-19 pandemic. The study period was two-months of follow-up using a telemedicine system. Median follow-up time was 51 (24-66) days. Two of the patients were using low glucose suspend system and six were on multiple daily injection therapy. Target TIR values were achieved in seven patients in the last televisit and, in line with recent guidelines, a TBR <70 mg/dL (<3.9 mmol/L) (level 1 hypoglycemia) of <4% and a TBR <54 mg/dL (<3.0 mmol/L) (level 2 hypoglycemia) of <1% were achieved in all patients. Seven patients achieved a CV of <36% at their last televisit. Telemedicine as an alternative follow-up tool during unusual circumstances such pandemics, even in countries where it is not routinely used, could be beneficial to achieve optimum glycemic control in patients with new-onset T1D.

Glycated hemoglobin variability and microvascular complications in patients with type 1 diabetes mellitus.

OBJECTIVES: Glycated hemoglobin (HbA1c) has proven to be indicative in the development of microvascular complications. In this study, the contribution of HbA1c variability to microvascular complications was evaluated. METHODS: Twenty-one cases with type 1 diabetes mellitus (T1DM) who developed microvascular complications and 39 cases without complications, that were similar in terms of gender, age of diagnosis, insulin treatment, insulin doses (U/kg), and mean HbA1c levels were included. RESULTS: Mean age of T1DM diagnosis was 5.87 ± 3.93 years in the complication group and 4.63 ± 3.33 years in the control group. Nephropathy was detected in 17 cases, neuropathy in 8 cases, and retinopathy in 1 case. Nephropathy occurred at a mean age of 11.52 ± 4.12 years and neuropathy at 14.13 ± 5.68 years. The mean HbA1c during follow-up was similar in the group with complications and the control group (8.60 ± 0.63 vs. 8.84 ± 1.32). Adjusted HbA1c-standard deviation (SD) and HbA1c-variation coefficient (CV) values were 1.30 ± 0.65 and 14.36 ± 6.23 in the group with complications (p=0.014), and 0.91 ± 0.37 and 10.59 ± 4.01 in the control group (p=0.013). In the Receiver Operating Characteristic (ROC)-analysis for microvascular complications, the limit value HbA1c-CV was 11.99 (sensitivity: 61.9%, specificity: 71.9%). This value for HbA1c-SD was 0.9699 (sensitivity: 71.43%, specificity: 66.67%). CONCLUSIONS: This study has shown that long-term fluctuations in HbA1c are associated with the development of microvascular complications in type 1 diabetes.