From islet transplantation to beta-cell regeneration: an update on beta-cell-based therapeutic approaches in type 1 diabetes.

INTRODUCTION: Type 1 diabetes (T1D) mellitus is an autoimmune disease in which immune cells, predominantly effector T cells, destroy insulin-secreting beta-cells. Beta-cell destruction led to various consequences ranging from retinopathy and nephropathy to neuropathy. Different strategies have been developed to achieve normoglycemia, including exogenous glucose compensation, whole pancreas transplantation, islet transplantation, and beta-cell replacement. AREAS COVERED: The last two decades of experience have shown that indigenous glucose compensation through beta-cell regeneration and protection is a peerless method for T1D therapy. Tremendous studies have tried to find an unlimited source for beta-cell regeneration, on the one hand, and beta-cell protection against immune attack, on the other hand. Recent advances in stem cell technology, gene editing methods, and immune modulation approaches provide a unique opportunity for both beta-cell regeneration and protection. EXPERT OPINION: Pluripotent stem cell differentiation into the beta-cell is considered an unlimited source for beta-cell regeneration. Devising engineered pancreas-specific regulatory T cells using Chimeric Antigen Receptor (CAR) technology potentiates an effective immune tolerance induction for beta-cell protection. Beta-cell regeneration using pluripotent stem cells and beta-cell protection using pancreas-specific engineered regulatory T cells promises to develop a curative protocol in T1D.

TRAIL induces proliferation in rodent pancreatic beta cells via AKT activation.

Strategies to increase functional pancreatic beta cell mass is of great interest in diabetes-related research. TNF-related apoptosis-inducing ligand (TRAIL) is well known to promote proliferation and survival in various cell types, including vascular smooth muscle and endothelial cells. Correlation between the protective nature of TRAIL on these cells and its proliferative effect is noteworthy. TRAIL's seemingly protective/therapeutic effect in diabetes prompted us to question whether it may act as an inducer of proliferation in pancreatic beta cells. We used rat primary islet cells and MIN6 mouse beta cell line to investigate TRAIL-induced proliferation. Cell viability and/or death was analyzed by MTT, WST-1, and Annexin-V/PI assays, while proliferation rates and pathways were assessed via immunocytochemical and Western blot analyses. Receptor neutralization antibodies identified the mediator receptors. Recombinant soluble TRAIL (sTRAIL) treatment led to 1.6-fold increased proliferation in insulin-positive cells in dispersed rat islets compared to the untreated group, while adenovirus-mediated overexpression of TRAIL increased the number of proliferating beta cells up to more than six-fold. sTRAIL or adenoviral vector-mediated TRAIL overexpression induced proliferation in MIN6 cells also. TRAIL's proliferative effect was mediated via AKT activation, which was suppressed upon specific inhibition. Neutralization of each TRAIL receptor reversed the proliferative effect to some degree, with the highest level of inhibition in death receptor 5 (DR5) blockage in MIN6 cells and in decoy receptor 1 (DcR1) blockage in primary rat beta cells. Thus, TRAIL induces proliferation in rodent pancreatic beta cells through activation of the AKT pathway.

Cytopathology-histopathology correlation and the effect of nodule diameter on diagnostic performance in patients undergoing thyroid fine-needle aspiration biopsy.

INTRODUCTION: Although thyroid fine-needle aspiration biopsy (FNAB) is established to have a good overall sensitivity and specificity, various outcomes have been reported on its performance in large nodules. The aim of the study was to evaluate the diagnostic performance of FNAB and the effect of the nodule diameter on its diagnostic performance. MATERIALS AND METHODS: The outcomes of a total of 7319 patients who underwent FNAB over the course of 5 years were analyzed retrospectively and 648 patients who had undergone post-FNAB thyroidectomy or lobectomy were included in the study. FNAB results were classified according to the Bethesda system. After evaluating the compatibility between cytology and pathology results, all-nodules and diameter-based (<4 cm and ≥4 cm) sensitivity, specificity, false positivity, false negativity, and accuracy rates of FNAB were calculated. RESULTS: Sensitivity of FNAB was 85.4% for all nodules, 88.3% for nodules <4 cm, and 75.8% for nodules >4 cm (P < 0.001). Specificity was 58.4% for all nodules, 49.3% for nodules <4 cm, and 75.1% for nodules >4cm (P < 0.001). While false positivity was 41.6% for all nodules, it was 50.7% for nodules smaller than 4 cm and was 24.9% for nodules larger than 4 cm (P < 0.001). False negativity was 14.6% for all nodules and was 11.7% for nodules smaller than 4 cm and 24.2% for nodules larger than 4 cm (P < 0.001). Finally, among the entire set of nodules, the accuracy was 64.4%, which was 59.2% in nodules smaller than 4 cm, and 75.2% in nodules larger than 4 cm (P < 0.001). CONCLUSION: Despite a higher rate of false negativity, FNAB has higher specificity and accuracy in large nodules than those in the small nodules. Nodule diameter should not be used alone as a criterion to recommend thyroidectomy to the patient.

Diagnostic value of the late-night salivary cortisol in the diagnosis of clinical and subclinical Cushing's syndrome: results of a single-center 7-year experience.

Late-night salivary cortisol (LNSaC) is an easy-to-use test reflecting the free cortisol level in the serum and does not require hospitalization. Controlled studies reported that LNSaC has a high sensitivity and specificity, but have not set a clearly defined cut-off value to be used in the diagnosis of Cushing's syndrome. In this study, we aimed to evaluate the diagnostic performance of LNSaC in patients with clinical Cushing's syndrome (CCS) and subclinical Cushing's syndrome (SCS). The data of 543 patients, whose LNSaC levels were assessed using electrochemiluminescence immunoassay method, were retrospectively evaluated. The study included a total of 324 patients: 58 patients with CCS, 53 patients with SCS, and 213 patients without Cushing's syndrome (NoCS). The cause of the Cushing's syndrome was hypophyseal in 26 patients (45%), adrenal in 24 patients (41%), and ectopic in 8 patients (14%) in the CCS group. Median LNSaC levels were 0.724 (0.107-33) µg/dL in CCS group, 0.398 (0.16-1.02) µg/dL in SCS group, and 0.18 (0.043-0.481) µg/dL in NoCS group (p=0.001). Accordingly, LNSaC had 89.6% sensitivity and 81.6% specificity at a cut-off value of 0.288 µg/dL in the diagnosis of CCS; and had 80.7% sensitivity and 85.1% specificity at a cut-off value of 0.273 µg/dL in the diagnosis of SCS. In the present study, a lower sensitivity and specificity than previously reported was found for LNSaC in the diagnosis of CCS. Moreover, the diagnostic performance of LNSaC in patients with SCS was close to its diagnostic performance in patients with CCS. Each center should determine its own cut-off value based on the method adopted for LNSaC measurement, and apply that cut-off value in the diagnosis of Cushing's syndrome.

Cystic lateral neck mass: Thyroid carcinoma metastasis to branchial cleft cyst.

Etiologies of lateral cervical masses are complex, most commonly these masses are branchial cleft cysts; however, metastatic thyroid carcinoma should be included in the differential. We report a case of lateral cystic neck mass in a 22-year-old female patient diagnosed as metastatic papillary thyroid carcinoma. The patient was diagnosed after she underwent surgery for branchial cleft cyst. The patient underwent thyroidectomy which revealed multifocal micropapillary thyroid carcinoma with capsular invasion and lymph node metastases. Radioactive iodine treatment was planned. Congenital malformations of the lateral neck may present themselves in the second and third decades of life. Ectopic thyroid tissue within a branchial cleft cyst may give rise to primary papillary carcinoma, as well as branchial cleft cyst may harbor metastases of primary thyroid papillary carcinoma. We classified our patient as a metastasis to the branchial cleft cyst rather than primary papillary carcinoma of the branchial cleft cyst.

Bone metastases without primary tumor: A well-differentiated follicular thyroid carcinoma case.

Metastases to the bone are the most common malignant bone tumors. Prostate, breast, and lung carcinomas are the most common primaries of bone metastases. Bone metastases show poor prognosis in means of median survival; however, some patients with highly curable tumors such as thyroid carcinoma may benefit from treatment. We report and discuss a unique case of a 70-year-old female patient presenting with arm pain, diagnosed with metastatic well-differentiated follicular carcinoma without a primary tumor in the thyroid.

Therapeutic Potential of Lentivirus-Mediated Glucagon-Like Peptide-1 Gene Therapy for Diabetes.

Postprandial glucose-induced insulin secretion from the islets of Langerhans is facilitated by glucagon-like peptide-1 (GLP-1)-a metabolic hormone with insulinotropic properties. Among the variety of effects it mediates, GLP-1 induces delta cell secretion of somatostatin, inhibits alpha cell release of glucagon, reduces gastric emptying, and slows food intake. These events collectively contribute to weight loss over time. During type 2 diabetes (T2DM), however, the incretin response to glucose is reduced and accompanied by a moderate reduction in GLP-1 secretion. To compensate for the reduced incretin effect, a human immunodeficiency virus-based lentiviral vector was generated to deliver DNA encoding human GLP-1 (LentiGLP-1), and the anti-diabetic efficacy of LentiGLP-1 was tested in a high-fat diet/streptozotocin-induced model of T2DM. Therapeutic administration of LentiGLP-1 reduced blood glucose levels in obese diabetic Sprague Dawley rats, along with improving insulin sensitivity and glucose tolerance. Normoglycemia was correlated with increased blood GLP-1 and pancreatic beta cell regeneration in LentiGLP-1-treated rats. Plasma triglyceride levels were also normalized after LentiGLP-1 injection. Collectively, these data suggest the clinical potential of GLP-1 gene transfer therapy for the treatment of T2DM.

Evaluation of Incidental Thyroid Nodules in Cancer Patients.

OBJECTIVE: Frequency of thyroid cancer in incidental thyroid nodules identified by imaging techniques in cancer patients is higher than that in the normal population. In the retrospective study, we have both investigated the incidence of thyroid cancer in incidentally identified nodules and compared the imaging techniques to determine whether there is any difference between them in detection of malign nodules. METHODS: A total of 7319 patients who underwent thyroid fine-needle aspiration biopsy (FNAB) were included in the study. The data of 174 patients who had previously been diagnosed with a hematologic or solid malignancy prior to the FNAB procedure and had incidentally identified thyroid nodules were evaluated retrospectively. RESULTS: Eighty-six (49.5%) of the incidental nodules were identified with ultrasonography (USG), 62 (35.6%) with positron emission tomography (PET) or PET/computed tomography (PET/CT), and 26 (14.9%) with CT. As a result of thyroidectomy, papillary carcinoma was identified in 8 (4.6%) patients, and metastasis to the thyroid of a primary cancer was found in 3 (1.7%) patients. While the papillary carcinoma proportion in the nodules identified by USG was 3.4%, PET/CT was 8.9%. A cut-off maximal standardized uptake value of 11.6 in PET/CT indicated malignancy achieving a sensitivity of 83.3% and a specificity of 91.1%. CONCLUSION: Whether the nodule in the incidental thyroid nodules of cancer patients is identified using USG or PET/CT, the risk of thyroid cancer is similar. However, cancer risk is higher in the event of a higher focal uptake in the nodules identified by PET/CT.

Which type 2 diabetes mellitus patients should be screened for subclinical Cushing's syndrome?

OBJECTIVE: Diabetes or impaired glucose tolerance are common in patients with Cushing's syndrome (CS). In this study we investigated the incidence of subclinical Cushing's syndrome (SCS) in type 2 diabetic patients with poor and good metabolic control and the relationship between blood glucose and cortisol levels. DESIGN: A total of 400 type 2 diabetes mellitus (T2DM) patients (Group A, HbA1c ≥8%, n=250; Group B, HbA1c ≤6.5%, n=150) were included in the study. Biochemical tests followed by the 1 mg dexamethasone suppression test (DST) were performed. If post-DST cortisol levels were above 1.8 µg/dl (non-suppressed group), the 2 mg DST test was performed for 2 days. Among the patients, post-DST cortisol of at least 1.8 µg/dL and midnight serum cortisol levels of at least 7.5 µg/dL confirmed the diagnosis of CS. RESULTS: While SCS was observed in 5 patients (2%) in Group A, no case was observed in group B. There was a statistically significant difference between groups for basal cortisol and post-DST cortisol results (p<0.001 and p<0.001, respectively). Microvascular complication rates were higher in the non-suppressed group (p=0.007). Post-DST cortisol levels had a positive correlation with the number of complications and HbA1c levels (respectively r=0.213, p<0.001, and r=0.191, p<0.001). Multivariate regression analysis revealed that durations of DM, HbA1c level, and post 1 mg DST cortisol levels were associated with the number of complications. CONCLUSIONS: This study shows the presence of SCS in a notable number in T2DM patients having poor metabolic control. Screening with 1 mg and 2 mg DST of T2DM patients with poor metabolic control who are also obese and hypertensive and have microvascular complications may be an appropriate method for detection of SCS.

ASSESSMENT OF DIASTOLIC DYSFUNCTION, ARTERIAL STIFFNESS, AND CAROTID INTIMA-MEDIA THICKNESS IN PATIENTS WITH ACROMEGALY.

OBJECTIVE: Early diagnosis and treatment of cardiovascular diseases, the most frequent cause of morbidity and mortality in acromegaly, may be an efficient approach to extending the lifespan of affected patients. Therefore, it is crucial to determine any cardiovascular diseases in the subclinical period. The study objectives were to determine markers of subclinical atherosclerosis and asses heart structure and function. METHODS: This was a cross-sectional, single-center study of 53 patients with acromegaly and 22 age- and sex-matched healthy individuals. Carotid intima-media thickness (CIMT), pulse-wave velocity (PWV), and echocardiographic data were compared between these groups. RESULTS: CIMT and PWV were higher in the acromegaly group than in the healthy group (P = .008 and P = .002, respectively). Echocardiography showed that left ventricular diastolic dysfunction was present in 11.3% of patients. Left ventricular mass index and left atrial volume index were higher in the patients (P = .016 and P<.001, respectively). No differences in the CIMT, PWV, or echocardiographic measurements were identified between the patients with biochemically controlled and uncontrolled acromegaly and the control group. CONCLUSION: Our results showed that subclinical atherosclerosis (i.e., CIMT and PWV markers) and heart structure and function were worse in patients with acromegaly than in healthy individuals. Because there were no differences in these parameters between patients with controlled and uncontrolled acromegaly, our results suggest that the structural and functional changes do not reverse with biochemical control. ABBREVIATIONS: AA = active acromegaly BSA = body surface area CA = biochemically controlled acromegaly CH = concentric hypertrophy CIMT = carotid intima-media thickness DBP = diastolic blood pressure DM = diabetes mellitus ECHO = echocardiography EDV = enddiastolic volume EF = ejection fraction ESV = endsystolic volume GH = growth hormone HC = healthy control HL = hyperlipidemia HT = hypertension IGF-1 = insulin-like growth factor 1 LA = left atrial LAV = left atrial volume LAVI = left atrial volume index LV = left ventricular LVDD = left ventricular diastolic dysfunction LVEF = left ventricular ejection fraction LVH = left ventricular hypertrophy LVMI = left ventricular mass index PWV = pulse-wave velocity RWT = relative wall thickness.

Parathyroidectomy in asymptomatic primary hyperparathyroidism reduces carotid intima-media thickness and arterial stiffness.

OBJECTIVE: Although an International Workshop has suggested that cardiovascular assessment in asymptomatic primary hyperparathyroidism (PHPT) patients is not necessary, improvements in risk factors of subclinical atherosclerosis have been shown following parathyroidectomy. The objectives of this study were to determine whether parathyroidectomy in asymptomatic PHPT patients causes any change in carotid intima-media thickness (CIMT), arterial stiffness [pulse wave velocity (PWV)] and soluble CD40 ligand (sCD40L) levels. DESIGN: Prospective study evaluating female patients diagnosed with asymptomatic PHPT in a single centre over a 6-month period. PATIENTS: A total of 48 subjects were included: 17 hypercalcaemic (HC, mean age: 51 ± 8 years, Ca: 2·73 ± 0·17 mmol/l) and 16 normocalcaemic (NC, mean age: 58 ± 7 years, Ca: 2·30 ± 0·10 mmol/l) PHPT patients, and 15 healthy controls (mean age: 52 ± 4 years, Ca: 2·27 ± 0·07 mmol/l). MEASUREMENTS: Biochemical tests, CIMT, PWV and sCD40L levels were compared at baseline and 6 months after parathyroidectomy (PTx). RESULTS: At baseline, CIMT and PWV values in the HC and NC patients were higher than in the control group. While there was a significant reduction in CIMT (601 ± 91 µm vs 541 ± 65 µm, P = 0·006) and PWV (9·6 ± 1·8 vs 8·4 ± 1·5 m/s, P = 0·000) in the hypercalcaemic group at the end of the 6th month after PTx, no change was observed in normocalcaemic group (P = 0·686 and P = 0·196 respectively). No differences were observed in sCD40L levels between patient and control groups or between baseline and 6 months in patients undergoing parathyroidectomy. CONCLUSION: Parathyroidectomy leads to an improvement in the structural and functional impairment associated with atherosclerosis in the vascular wall in asymptomatic hypercalcaemic PHPT patients.

Incretins: their physiology and application in the treatment of diabetes mellitus.

Therapies targeting the action of incretin hormones have been under close scrutiny in recent years. The incretin effect has been defined as postprandial enhancement of insulin secretion by gut-derived factors. Likewise, incretin mimetics and incretin effect amplifiers are the two different incretin-based treatment strategies developed for the treatment of diabetes. Although, incretin mimetics produce effects very similar to those of natural incretin hormones, incretin effect amplifiers act by inhibiting dipeptidyl peptidase-4 (DPP-4) enzyme to increase plasma concentration of incretins and their biologic effects. Because glucagon-like peptide-1 (GLP-1) is an incretin hormone with various anti-diabetic actions including stimulation of glucose-induced insulin secretion, inhibition of glucagon secretion, hepatic glucose production and gastric emptying, it has been evaluated as a novel therapeutic agent for the treatment of type 2 diabetes mellitus (T2DM). GLP-1 also manifests trophic effects on pancreas such as pancreatic beta cell growth and differentiation. Because DPP-4 is the enzyme responsible for the inactivation of GLP-1, DPP-4 inhibition represents another potential strategy to increase plasma concentration of GLP-1 to enhance the incretin effect. Thus, anti-diabetic properties of these two classes of drugs have stimulated substantial clinical interest in the potential of incretin-based therapeutic agents as a means to control glucose homeostasis in T2DM patients. Despite this fact, clinical use of GLP-1 mimetics and DPP-4 inhibitors have raised substantial concerns owing to possible side effects of the treatments involving increased risk for pancreatitis, and C-cell adenoma/carcinoma. Thus, controversial issues in incretin-based therapies under development are reviewed and discussed in this manuscript.

GLP-1-mediated gene therapy approaches for diabetes treatment.

Glucagon-like peptide (GLP)-1 is an incretin hormone with several antidiabetic functions including stimulation of glucose-dependent insulin secretion, increase in insulin gene expression and beta-cell survival. Despite the initial technical difficulties and profound inefficiency of direct gene transfer into the pancreas that seriously restricted in vivo gene transfer experiments with GLP-1, recent exploitation of various routes of gene delivery and alternative means of gene transfer has permitted the detailed assessment of the therapeutic efficacy of GLP-1 in animal models of type 2 diabetes (T2DM). As a result, many clinical benefits of GLP-1 peptide/analogues observed in clinical trials involving induction of glucose tolerance, reduction of hyperglycaemia, suppression of appetite and food intake linked to weight loss have been replicated in animal models using gene therapy. Furthermore, GLP-1-centered gene therapy not only improved insulin sensitivity, but also reduced abdominal and/or hepatic fat associated with obesity-induced T2DM with drastic alterations in adipokine profiles in treated subjects. Thus, a comprehensive assessment of recent GLP-1-mediated gene therapy approaches with detailed analysis of current hurdles and resolutions, is discussed.

Clinical utility of insulin and insulin analogs.

Diabetes is a pandemic disease characterized by autoimmune, genetic and metabolic abnormalities. While insulin deficiency manifested as hyperglycemia is a common sequel of both Type-1 and Type-2 diabetes (T1DM and T2DM), it does not result from a single genetic defect--rather insulin deficiency results from the functional loss of pancreatic ß cells due to multifactorial mechanisms. Since pancreatic ß cells of patients with T1DM are destroyed by autoimmune reaction, these patients require daily insulin injections. Insulin resistance followed by ß cell dysfunction and ß cell loss is the characteristics of T2DM. Therefore, most patients with T2DM will require insulin treatment due to eventual loss of insulin secretion. Despite the evidence of early insulin treatment lowering macrovascular (coronary artery disease, peripheral arterial disease and stroke) and microvascular (diabetic nephropathy, neuropathy and retinopathy) complications of T2DM, controversy exists among physicians on how to initiate and intensify insulin therapy. The slow acting nature of regular human insulin makes its use ineffective in counteracting postprandial hyperglycemia. Instead, recombinant insulin analogs have been generated with a variable degree of specificity and action. Due to the metabolic variability among individuals, optimum blood glucose management is a formidable task to accomplish despite the presence of novel insulin analogs. In this article, we present a recent update on insulin analog structure and function with an overview of the evidence on the various insulin regimens clinically used to treat diabetes.

Insulin gene therapy from design to beta cell generation.

Despite the fact that insulin injection can protect diabetic patients from developing diabetes-related complications, recent meta-analyses indicate that rapid and long-acting insulin analogues only provide a limited benefit compared with conventional insulin regarding glycemic control. As insulin deficiency is the main sequel of type-1 diabetes (T1D), transfer of the insulin gene-by-gene therapy is becoming an attractive treatment modality even though T1D is not caused by a single genetic defect. In contrast to human insulin and insulin analogues, insulin gene therapy targets to supplement patients not only with insulin but also with C-peptide. So far, insulin gene therapy has had limited success because of delayed and/or transient gene expression. Sustained insulin gene expression is now feasible using current gene-therapy vectors providing patients with basal insulin coverage, but management of postprandial hyperglycaemia is still difficult to accomplish because of the inability to properly control insulin secretion. Enteroendocrine cells of the gastrointestinal track (K cells and L cells) may be ideal targets for insulin gene therapy, but cell-targeting difficulties have limited practical implementation of insulin gene therapy for diabetes treatment. Therefore, recent gene transfer technologies developed to generate authentic beta cells through transdifferentiation are also highlighted in this review.

Molecular mechanisms of death ligand-mediated immune modulation: a gene therapy model to prolong islet survival in type 1 diabetes.

Type 1 diabetes results from the T cell-mediated destruction of pancreatic beta cells. Islet transplantation has recently become a potential therapeutic approach for patients with type 1 diabetes. However, islet-graft failure appears to be a challenging issue to overcome. Thus, complementary gene therapy strategies are needed to improve the islet-graft survival following transplantation. Immune modulation through gene therapy represents a novel way of attacking cytotoxic T cells targeting pancreatic islets. Various death ligands of the TNF family such as FasL, TNF, and TNF-Related Apoptosis-Inducing Ligand (TRAIL) have been studied for this purpose. The over-expression of TNF or FasL in pancreatic islets exacerbates the onset of type 1 diabetes generating lymphocyte infiltrates responsible for the inflammation. Conversely, the lack of TRAIL expression results in higher degree of islet inflammation in the pancreas. In addition, blocking of TRAIL function using soluble TRAIL receptors facilitates the onset of diabetes. These results suggested that contrary to what was observed with TNF or FasL, adenovirus mediated TRAIL gene delivery into pancreatic islets is expected to be therapeutically beneficial in the setting of experimental models of type 1 diabetes. In conclusion; this study mainly reveals the fundamental principles of death ligand-mediated immune evasion in diabetes mellitus.