Short fasting test as a reliable and effective tool to diagnose insulinoma.

PURPOSE: The diagnosis of insulinoma can be challenging, requiring documentation of hypoglycaemia associated with non-suppressed insulin and C-peptide, often achieved during a prolonged 72 h fast performed in inpatient setting. Our goal is to predict weather a shorter outpatient fasting test initiated overnight and prolonged up until 24 h could be a sensitive method for diagnosing insulinoma. METHODS: We conducted a retrospective monocentric study on subjects admitted to our Unit of Endocrinology from 2019 to 2022 for clinical suspicion of insulinoma and underwent the short fasting test. A comparison between the short test group and the group of subjects who underwent the standard prolonged fasting test (from 2003 to 2018) has also been performed. The short fasting test was initiated by the patient overnight at home and proceeded the following day in outpatient setting (Day Hospital). As in the standard protocol, symptoms and capillary blood glucose (CBG) were strictly monitored. Venous blood was drawn for glycaemia, insulin and C-peptide at admission and at established intervals, in case of symptoms of hypoglycaemia or if CBG ≤ 45 mg/dl, when the fast would be suspended. RESULTS: The final sample consisted of 37 patients, with mean age of 44.5 ± 12.6 years (17-74). Short and standard tests were performed in 15 and 22 subjects, respectively. Diagnostic values for insulinoma were observed in 12 patients: in 5/15 who underwent the short fasting test, in 6/22 who underwent the prolonged test and in 1 patient who was initially negative on the short test and subsequently showed diagnostic values during the prolonged test. The diagnosis of insulinoma was achieved in 11/12 cases within 24 h of the beginning of the fast (91.7%). CONCLUSIONS: A short fasting test could be a valid, sensitive and reliable first-line workup in diagnosing insulinoma.

The Effects of Human Immunodeficiency Virus Type 1 (HIV-1) Antigen-Expanded Specific T-Cell Therapy and Vorinostat on Persistent HIV-1 Infection in People With HIV on Antiretroviral Therapy.

BACKGROUND: The histone deacetylase inhibitor vorinostat (VOR) can reverse human immunodeficiency virus type 1 (HIV-1) latency in vivo and allow T cells to clear infected cells in vitro. HIV-specific T cells (HXTCs) can be expanded ex vivo and have been safely administered to people with HIV (PWH) on antiretroviral therapy. METHODS: Six PWH received infusions of 2 × 107 HXTCs/m² with VOR 400 mg, and 3 PWH received infusions of 10 × 107 HXTCs/m² with VOR. The frequency of persistent HIV by multiple assays including quantitative viral outgrowth assay (QVOA) of resting CD4+ T cells was measured before and after study therapy. RESULTS: VOR and HXTCs were safe, and biomarkers of serial VOR effect were detected, but enhanced antiviral activity in circulating cells was not evident. After 2 × 107 HXTCs/m² with VOR, 1 of 6 PWH exhibited a decrease in QVOA, and all 3 PWH exhibited such declines after 10 × 107 HXTCs/m² and VOR. However, most declines did not exceed the 6-fold threshold needed to definitively attribute decline to the study intervention. CONCLUSIONS: These modest effects provide support for the strategy of HIV latency reversal and reservoir clearance, but more effective interventions are needed to yield the profound depletion of persistent HIV likely to yield clinical benefit. Clinical Trials Registration. NCT03212989.

Calcium-stimulated calcitonin test for the diagnosis of medullary thyroid cancer: results of a multicenter study and comparison between different assays.

BACKGROUND: A basal serum calcitonin (Ct) increase >100 pg/mL in patients with a thyroid nodule is consistent with the diagnosis of medullary thyroid cancer (MTC). In cases where the CT test have a slight to moderate increase, the calcium gluconate stimulation test is helpful to increase diagnostic accuracy. However, reliable cut-offs for calcium-stimulated Ct are still lacking. The aim of this study was to evaluate the sex-specific calcium-stimulated Ct cutoffs for the diagnosis of MTC in a multicenter series. A comparison between different Ct assays has been also performed. METHODS: 90 subjects undergone calcium-stimulated Ct for a suspected MTC in 5 Endocrine Units between 2010-2021 were retrospectively analyzed. Serum Ct concentrations were assessed by immunoradiometric (IRMA) or chemiluminescence (CLIA) assays. RESULTS: MTC was diagnosed in 37 (41.1%) and excluded in 53 (58.9%) patients. The best calcium-stimulated Ct cut-off to identify MTC was 611 pg/mL in males (AUC =0.90, 95% CI (0.76;1) and 445 pg/mL in females (AUC=0.79, 95% CI (0.66;0.91). Logistic regression analysis showed that both basal (OR 1.01, P=0.003) and peak Ct after stimulation (OR 1.07, P=0.007) were significantly associated with MTC, together with sex (OR=0.06, P<0.001). The "Ct assay" variable was also considered in the logistic regression model, but it was not significantly associated with MTC (OR=0.93, P=0.919). CONCLUSIONS: This study indicates that calcium test could be helpful to identify patients with early-stage MTC and those without MTC. A Ct value of 611 pg/mL in males and 445 pg/mL in females are proposed as the optimal Ct cut-offs at the stimulation test.

Applications of virus-specific T cell therapies post-BMT.

Hematopoietic stem cell transplantation (HSCT) has been used as a curative standard of care for moderate to severe primary immunodeficiency disorders as well as relapsed hematologic malignancies for over 50 years [1,2]. However, chronic and refractory viral infections remain a leading cause of morbidity and mortality in the immune deficient period following HSCT, where use of available antiviral pharmacotherapies is limited by toxicity and emerging resistance [3]. Adoptive immunotherapy using virus-specific T cells (VSTs) has been explored for over 2 decades [4,5] in patients post-HSCT and has been shown prior phase I-II studies to be safe and effective for treatment or preventions of viral infections including cytomegalovirus, Epstein-Barr virus, BK virus, and adenovirus with minimal toxicity and low risk of graft vs host disease [6-9]. This review summarizes methodologies to generate VSTs the clinical results utilizing VST therapeutics and the challenges and future directions for the field.

Influence of Touch-Spun Nanofiber Diameter on Contact Guidance during Peripheral Nerve Repair.

Peripheral nerve regeneration across large gaps remains clinically challenging and scaffold design plays a key role in nerve tissue engineering. One strategy to encourage regeneration has utilized nanofibers or conduits to exploit contact guidance within the neural regenerative milieu. Herein, we report the effect of nanofiber topography on two key aspects of regeneration: Schwann cell migration and neurite extension. Substrates possessing distinct diameter distributions (300 ± 40 to 900 ± 70 nm) of highly aligned poly(ε-caprolactone) nanofibers were fabricated by touch-spinning. Cell migratory behavior and contact guidance were then evaluated both at the tissue level using dorsal root ganglion tissue explants and the cellular level using dissociated Schwann cells. Explant studies showed that Schwann cells emigrated significantly farther on fibers than control. However, both Schwann cells and neurites emigrated from the tissue explants directionally along the fibers regardless of their diameter, and the data were characterized by high variation. At the cellular level, dissociated Schwann cells demonstrated biased migration in the direction of fiber alignment and exhibited a significantly higher biased velocity (0.2790 ± 0.0959 µm·min-1) on 900 ± 70 nm fibers compared to other nanofiber groups and similar to the velocity found during explant emigration on 900 nm fibers. Therefore, aligned, nanofibrous scaffolds of larger diameters (900 ± 70 nm) may be promising materials to enhance various aspects of nerve regeneration via contact guidance alone. While cells track along with the fibers, this contact guidance is bidirectional along the fiber, moving in the plane of alignment. Therefore, the next critical step to direct regeneration is to uncover haptotactic cues that enhance directed migration.

Cortisol Deficiency in Lenvatinib Treatment of Thyroid Cancer: An Underestimated Common Adverse Event.

Background: Lenvatinib treatment has shown a significant improvement in progression-free survival in patients with metastatic, progressive, radioiodine-refractory differentiated thyroid cancer, although its use is associated with considerable toxicity. Fatigue is one of the most frequent adverse events (AEs). It has been reported that adrenal insufficiency (AI) may be involved in lenvatinib-related fatigue. In our study, we assessed the pituitary/adrenal axis before and during treatment, and the possible involvement of AI in lenvatinib-related fatigue. This was done to clarify the incidence, development, and time course of AI during lenvatinib treatment. Methods: We studied 13 patients who were selected for lenvatinib therapy. Adrenal function was evaluated by measuring cortisol and adrenocorticotropic hormone (ACTH) levels and through the ACTH (250 µg) stimulation test. Results: During treatment, seven patients (54%) developed AI. High levels of ACTH were observed in accordance with the diagnosis of primary AI (PAI). By evaluating the first ACTH test, before starting lenvatinib treatment, we found that patients with <646.6 nmol/L cortisol peak had an increased risk of developing PAI during lenvatinib treatment. Fatigue was observed in 11 patients (84.6%) during lenvatinib treatment. Cortisone acetate treatment induced an improvement in fatigue in six of seven patients (85.7%) in the PAI group, without the need to change the lenvatinib dosage. Conclusions: PAI may be considered one of the most common AEs associated with lenvatinib. Our data strongly suggest that PAI could be involved in lenvatinib-associated fatigue, particularly in patients with extreme fatigue. In this context, early diagnosis of PAI is essential, especially since glucocorticoid replacement therapy can induce a significant improvement in fatigue, without the need to reduce the dosage of lenvatinib. However, further studies are required to confirm these preliminary findings.

Enhancing Schwann cell migration using concentration gradients of laminin-derived peptides.

Neuroregeneration following peripheral nerve injury is largely mediated by Schwann cells (SC), the principal glial cell that supports neurons in the peripheral nervous system. Axonal regeneration in vivo is limited by the extent of SC migration into the gap between the proximal and distal nerve, however, little is known regarding the principal driving forces for SC migration. Engineered microenvironments, such as molecular and protein gradients, play a role in the migration of many cell types, including cancer cells and fibroblasts. However, haptotactic strategies have not been applied widely to SC. Herein, a series of tethered laminin-derived peptides were analyzed for their influence on SC adhesion, proliferation, and alignment. Concentration gradient substrates were fabricated using a controlled vapor deposition method, followed by covalent peptide attachment via a thiol-ene reaction, and characterized by X-ray photoelectron spectroscopy (XPS) and MALDI-MS imaging. While tethered RGD peptides supported SC adhesion and proliferation, concentration gradients of RGD had little influence on biased SC directional migration. In contrast, YIGSR promoted less SC attachment than RGD, yet YIGSR peptide gradients directed migration with a strong bias to the concentration profile. With YIGSR peptide, overall speed increased with the steepness of the peptide concentration profile. YIGSR gradients had no haptotactic effect on rat dermal fibroblast migration, in contrast to fibroblast migration on RGD gradients. The response of SC to these tethered peptide gradients will guide the development of translationally relevant constructs designed to facilitate endogenous SC infiltration into defects for nerve regeneration.

Synthesis and 3D Printing of PEG-Poly(propylene fumarate) Diblock and Triblock Copolymer Hydrogels.

PEG-based hydrogels are used widely in exploratory tissue engineering applications but in general lack chemical and structural diversity. Additive manufacturing offers pathways to otherwise unattainable scaffold morphologies but has been applied sparingly to cross-linked hydrogels. Herein, mono methyl ether poly(ethylene glycol) (PEG) and PEG-diol were used to initiate the ring-opening copolymerization (ROCOP) of maleic anhydride and propylene oxide to yield well defined diblock and triblock copolymers of PEG-poly(propylene maleate) (PPM) and ultimately poly(propylene fumarate) (PPF) with different molecular mass PEG macroinitiators and block length ratios. Using continuous digital light processing (cDLP) hydrogels were photochemically printed from an aqueous solution which resulted in a 10-fold increase in elongation at break compared to traditional diethyl fumarate (DEF) based printing. Furthermore, PPF-PEG-PPF triblock hydrogels were also found to be biocompatible in vitro across a number of engineered MC3T3, NIH3T3, and primary Schwann cells.

Combined Oral Contraception and Bicalutamide in Polycystic Ovary Syndrome and Severe Hirsutism: A Double-Blind Randomized Controlled Trial.

Context: Hirsutism often occurs in women with polycystic ovary syndrome (PCOS). The efficacy of oral contraceptive pill (OCP) plus antiandrogens in the treatment of its severe expression is controversial due to the lack of randomized, double-blind, long-term studies. Objective: The primary outcome was the reduction of hirsutism in PCOS women objectively measured by videodermoscopy on the androgen-sensitive skin areas assessed by the modified Ferriman and Gallwey (mF&G) total score, after 12 months of therapy with OCP + bicalutamide (BC) vs OCP plus placebo (P). The secondary outcomes were to evaluate tolerability of BC and body composition as well as the occurrence of adverse events. Design: An experimental, phase 3, prospective, multicenter, randomized, double-blind, P-controlled trial. Patients were evaluated at the baseline visit, at 6 and 12 months during treatment, and 6 months' posttreatment. Participants: Seventy women with classic PCOS (severe hirsutism, oligoanovulation, and ovarian polycystic ovarian morphology). Intervention: Patients received OCP + BC (50 mg/d) or OCP + P for 12 months. Results: The repeated measures analysis of variance showed that both treatments were effective in reducing hirsutism: The OCP + BC group had a higher reduction compared with the OCP + P group. No adverse effects were described during treatment except an increase in total cholesterol and low-density lipoprotein in the OCP + BC group. Conclusions: The association of OCP + BC is well tolerated and significantly more effective than OCP alone in treating severe hirsutism. We suggest a combined use of the videodermoscopic index and mF&G to evaluate the effects of androgen deprivation therapy for hirsutism.

IRIS: methodological assessment of psychopathological disease in a cohort of hirsute women.

BACKGROUND: Hirsutism in females can be a source of considerable psychological distress and a threat to female identity. The aim of our study was to evaluate a possible relationship between facial, total body hair involvement and physical, mental and social well-being during 12 months of follow-up and treatment. Both objective and subjective methods of evaluating hirsutism were used: the Ferriman-Gallwey (FG) scoring method and the questionnaires General Health Questionnaire (GHQ)-12, Polycystic Ovary Syndrome Questionnaire (PCOSQ) and SF-12. METHODS: The total of 469 female patients (mean age 27.61±7.63 years) was enrolled in 27 Italian centers participating in this study. Higher total body score was correlated to significant emotional discomfort. The correlation between the FG total body score, the facial score and physical/mental health was found to be significant in all the patients assessed by SF-12 questionnaire. The ongoing reduction of GHQ-12 score was found for the facial FG score at the first follow-up (T0-T1 period) and at the second one (T0-T2). No relationship was found between T1 and T2. At both 6 (T1) and 12 months (T2) follow-up an increase of PCOSQ Score (psychological improvement) was accompanied by a concomitant reduction of the FG Score (reduction of hirsutism). Physical health assessed by SF-12 questionnaire does not change at both 6- and 12-month follow-up, but mental health decreased at both T1 and T2. RESULTS: The clinical improvement was achieved at 6 months regardless on treatment used and it was maintained for the next six-month follow-up. The clinical outcome could be assessed both by FG Score both through questionnaires administrated to each patient with hirsutism. CONCLUSIONS: For the evaluation of psychopathological discomfort the most appropriate questionnaire was GHQ-12, because of it major sensitivity to identify the psychological discomfort in the hirsutism.

Ultrastructure of isolated mouse ovarian follicles cultured in vitro.

BACKGROUND: In vitro maturation of ovarian follicles, in combination with cryopreservation, might be a valuable method for preserving and/or restoring fertility in mammals with impaired reproductive function. Several culture systems capable of sustaining mammalian follicle growth in vitro have been developed and many studies exist on factors influencing the development of in vitro grown oocytes. However, a very few reports concern the ultrastructural morphology of in vitro grown follicles. METHODS: The present study was designed to evaluate, by transmission and scanning electron microscopy, the ultrastructural features of isolated mouse preantral follicles cultured in vitro for 6 days in a standard medium containing fetal calf serum (FCS). The culture was supplemented or not with FSH. RESULTS: The follicles cultured in FCS alone, without FSH supplementation (FCS follicles), did not form the antral cavity. They displayed low differentiation (juxta-nuclear aggregates of organelles in the ooplasm, a variable amount of microvilli on the oolemma, numerous granulosa cell-oolemma contacts, signs of degeneration in granulosa cell compartment). Eighty (80)% of FSH-treated follicles formed the antral cavity (FSH antral follicles). These follicles showed various ultrastructural markers of maturity (spreading of organelles in ooplasm, abundant microvilli on the oolemma, scarce granulosa cell-oolemma contacts, granulosa cell proliferation). Areas of detachment of the innermost granulosa cell layer from the oocyte were also found, along with a diffuse granulosa cell loosening compatible with the antral formation. Theca cells showed an immature morphology for the stage reached. Twenty (20)% of FSH-treated follicles did not develop the antral cavity (FSH non-antral follicles) and displayed morphological differentiation features intermediate between those shown by FCS and FSH antral follicles (spreading of organelles in the ooplasm, variable amount of microvilli, scattered granulosa cell-oolemma contacts, signs of degeneration in granulosa cell compartment). CONCLUSIONS: It is concluded that FSH supports the in vitro growth of follicles, but the presence of a diffuse structural granulosa cell-oocyte uncoupling and the absence of theca development unveil the incomplete efficiency of the system. The present study contributes to explain, from a morphological point of view, the effects of culture conditions on the development of mouse in vitro grown follicles and to highlight the necessity of maintaining efficient intercellular communications to obtain large numbers of fully-grown mature germ cells.

Reatividade do teste VDRL em bolsas de sangue da fundação de hematologia e hemoterapia do Amazonas-hemoam, os custos decorrentes de descarte e a estimativa de prevalência de sífilis em doadores de sangue do estado do amazonas / Reactivity of VDRL test in blood bags of the Amazon hematology and hemoterapy foundation-hemoam, the decurrent costs of discharge and estimative of syphilis prevalence in blood donnors of the amazon st

Introdução: a sífilis é uma doença infecto-contagiosa crônica que pode ser transmitida pelo sangue. No Brasil, a prevalência entre doadores de sanguevaria entre 1% a 4%. Atualmente é rotina a utilização de testes sorológicos sensíveis e específicos para eliminar a possibilidade de transmissão etais providências trazem vantagens em razão da qualidade do produto oferecido aos usuários, e desvantagens devido às taxas de descarte das bolsas, ocasionando prejuízos à instituição. Objetivos: determinar a freqüência de testes VDRL reativos em bolsas de sangue, a prevalência de sífilis em doadores atendidos no HEMOAM, a taxa de descarte de bolsas e custos estimados envolvidos no período entre 2000 a 2004. Métodos: estudo retrospectivo, transversal, descritivo com componentes analíticos no qual foram analisados todos os dados disponíveis provenientes da capital e do interior do estado do Amazonas submetidos aos testes de VDRL e FTA-Abs no período relativo aos anos 2000 e 2004. Resultados: das 237.643 amostras de bolsas de sangue analisadas 207.707 (87,40%) foram provenientes da capital e 30.016 (12,60%) do interior do estado. Do total de testes de VDRL reativos realizados na capital, 870 (37,9%) foram submetidos ao FTA-Abs, o mesmo acontecendo com as 283 (32,8%) amostras provenientes do interior, totalizando 1.153 testes realizados. O preço unitário de cada bolsa no período referente ao estudo era de R$ 31.69, totalizando um custo de descarte de R$ 100.679,30 com média anual de R$ 25.169,78. A prevalência de sífilis em doadores de sangue do interior e na capital do estado foi0,49%. Conclusão: os custos observados no período são significantes e requerem ações estratégicas técnico-educativas que possam levar à redução tanto da freqüência de VDRL reativos e prevalência de sífilis, quanto dos custos relacionados.

Introduction: syphilis is a chronic infectious disease that can be transmitted by blood. In Brazil, the prevalence among blood donors varies from 1 to4%. Sensitive and specific serological tests to eliminate the possibility of transmission are now routinely used result in better quality of the productoffered for transfusion. The amount of blood bags discarded after serology is significant, causing economical disadvantages for the Institution.Objective: to determine the frequency of reactive VRDL tests in blood bags, the prevalence of syphilis in blood donors of the HEMOAM, the discardedblood bags and the estimated costs involved in the period between 2000 - 2004. Methods: restrospective study, with analytical descriptive components. All disposal datas from city and interior of the Amazon state submitted for the VDRL tests and FTA-Abs in the period of 2000-2004 wereanalysed. Results: from 237.634 blood bags 207.707 (87,40%) were from the city of Manaus and 30.016 (12,60%) from the interior of the state.From the total of the reactive VDRL realized in the city of Manaus, 870 (37,9%) were submitted for the FTA-Abs test, as well as for the 283 (32,8%) from the interior of the state, totaling in 1.153 tests. The unit cost of each blood bags in the period of the study was R$ 31.69, totaling an discharge cust R$ 100.679,30, with annual media of the R$ 25.169,78. The prevalence of syphilis in blood donors at the interior and city of Manaus was 0.49%. Conclusion: The observed custs in the period of the study are significant and requery estrategical technical/educational actions to reduce as the reactive VDRL frequency and prevalence of syphilis as the related cost.

Occurrence of dopaminergic (D(2)) receptors within the rabbit pulmonary circulation.

The pharmacological characteristics and the microanatomical localization of dopamine D(2)-like receptors, or more correctly spiroperidol binding sites, in the rabbit pulmonary circulation were studied using combined marker binding and light microscopy autoradiography with [((3))H]-spiroperidol (spiperone) as marker. The marker was bound to the samples of the pulmonary artery in a manner consistent with the labelling of dopamine D(2)-like receptors with an equilibrium dissociation constant (K(d)) of about 2.4+/-0.07 nmol/l and a maximum density of binding sites of 65+/-4.5 fmol/mg tissue. Samples of bronchial artery show the same results as those of the pulmonary artery. In contrast, binding experiments made with samples of rabbit lung (capillary of the microcirculation), of pulmonary veins and/or of bronchial veins did not allow the evaluation of specific binding.Autoradiography, observed with light microscopy, showed the development of specific silver grains within the whole wall of extraparenchymal branches of the pulmonary artery and/or of the bronchial artery. Development of silver grains was inhibited by compounds active on the dopamine receptors. The greater sensitivity to displacement by domperidone, haloperidol, and bromocriptine than to displacement by N-propyl-nor-apomorphine, quinpirole and clozapine suggests that the binding sites observed in extraparenchymal, large and medium-sized branches of the rabbit pulmonary and bronchial arteries belong, likely, to the dopamine D(2) receptor subtype. Quantitative analysis of images let us count the amount of these receptors in many samples of the pulmonary and/or bronchial arteries.