Publications

In vitro growth systems of preantral follicles allow studying the effect of various endocrine, paracrine, and autocrine factors on follicular growth and oocyte maturation. CRH is a 41-amino-acid neuropeptide responsible for endocrine, autonomic, immunological, and behavioral responses of mammals to stress and has two receptors, CRH receptor type 1 (CRH-R1) and CRH-R2. Antalarmin, a CRH-R1 antagonist, has been used to elucidate the role of CRH in stress, inflammation, and reproduction. The present study describes in vitro growth of mouse preantral follicles, early embryo development, and steroidogenesis in the presence of CRH and its antagonist antalarmin. We cultured 732 follicles in control media, 1306 in CRH 10(-7) mol/liter, and 1202 in CRH 10(-7) plus antalarmin 10(-6) mol/liter. The culture medium was assayed on alternate days for 17β-estradiol, progesterone, and β-human chorionic gonadotropin. Total RNA was extracted from preantral follicles as well as early preimplantation embryos and was assessed by real-time RT-PCR for the expression of CRH-R1 and CRH-R2 mRNAs. Hormone analysis showed that the CRH group had lower levels of 17β-estradiol, progesterone, and β-human chorionic gonadotropin as the culture progressed, in comparison with the other two groups. RT-PCR demonstrated the presence of CRH-R1 and CRH-R2 in all stages of preantral follicle culture. Morula/blastocyst-stage embryos expressed only CRH-R1. In conclusion, CRH has an inhibitory effect on in vitro fertilized oocytes, resulting from cultured preantral follicles at all stages of preimplantation embryo development. Furthermore, the presence of CRH in the culture medium inhibits steroidogenesis by preantral mouse follicles cultured in vitro.

The identification of immunodominant B cell epitopes within surface pneumococcal virulence proteins in pediatric patients with invasive pneumococcal disease (IPD) is a valuable approach to define novel vaccine candidates. To this aim, we evaluated sera from children with IPD and age-matched controls against 141 20-mer synthetic peptides covering the entire sequence of major antigenic fragments within pneumococcal virulence proteins; namely, choline-binding protein D (CbpD), pneumococcal histidine triad proteins (PhtD and PhtE), pneumococcal surface protein A (PspA), plasminogen and fibronectin binding protein B (PfbB), and zinc metalloproteinase B (ZmpB). Ten immunodominant B cell epitopes were identified: CbpD-pep4 (amino acids (aa) 291-310), PhtD-pep11 (aa 88-107), PhtD-pep17 (aa 172-191), PhtD-pep19 (aa 200-219), PhtE-pep32 (aa 300-319), PhtE-pep40 (aa 79-98), PfbB-pep76 (aa 180-199), PfbB-pep79 (aa 222-241), PfbB-pep90 (aa 484-503), and ZmpB-pep125 (aa 431-450). All epitopes were highly conserved among different pneumococcal serotypes, and four of them were located within the functional zinc-binding domain of the histidine triad proteins PhtD and PhtE. Peptides CbpD-pep4, PhtD-pep19, and PhtE-pep40 were broadly recognized by IPD patient sera with prevalences of 96.4%, 92.9%, and 71.4%, respectively, whereas control sera exhibited only minor reactivities (<10.7%). Their specificities for IPD were 93.3%, 95%, and 96.7%; their sensitivities were 96.4%, 92.9%, and 71.4% and their positivity likelihood ratios for IPD were 14.5, 18.6, and 21.4, respectively. Furthermore, purified antibodies against CbpD-pep4, PhtD-pep19, and PhtE-pep40 readily bound on the surfaces of different pneumococcal serotypes, as assessed by FACS and immunofluorescence analysis. The identified immunodominant B cell epitopes provide a better understanding of immune response in IPD and are worth evaluation in additional studies as potential vaccine candidates.

The purpose of the present study was to evaluate special features and probable correlations between symptoms, laboratory findings, treatment, and outcomes of COVID-19 in children and adolescents, through a systematic review and pooled analysis. Following database (Pubmed, Google Scholar, Scopus and Embase) search, forty articles were considered eligible identifying a total of 2,971 confirmed pediatric COVID-19 patients. Fever was reported in 55.1% of the cases, while 28.4% were asymptomatic. Radiological signs of pneumonia were observed in more than half of the cases and in 40.7% of asymptomatic patients. Fever showed the highest sensitivity (sensitivity: 60.3%, specificity: 48.8%), followed by cough (sensitivity: 47.4%, specificity: 76.7%), rhinorrhea (sensitivity: 21.1%, specificity: 88.4%) and diarrhea (sensitivity: 10.3%, specificity: 88.4%), in differentiating cases with positive radiological signs for pneumonia. Compared to school age children, preschoolers (adjusted OR=6.01, 95%CI: 1.73-20.91) were more prone to pneumonia findings. Various combinations of treatments were used across studies, without following any strict guidelines. Most children (&gt;90%) had full recovery and rarely presented complications. Fever seems to be the most frequent symptom in pediatric COVID-19, but pediatricians should additionally evaluate cough, rhinorrhea, and diarrhea as indicators of SARS-CoV-2 infection. Asymptomatic cases were common, but not the majority, and a significant percentage had developed radiologic findings of pneumonia. Thorough reassessment of treatment and management guidelines should be helpful.

The responses of plasma immunoreactive ACTH (IR-ACTH), cortisol, GH, PRL, and LH to a single iv injection of 0.01-5 μg/kg human corticotropin-releasing factor (hCRF) were investigated in healthy volunteers. The lowest effective dose of hCRF was 0.5 μg/kg. hCRF caused brief pulselike elevations of plasma IR-ACTH and cortisol without any effect on the plasma concentrations of GH, PRL, and LH. By contrast, ovine CRF (oCRF), given for comparison, produced long-lasting stimulation of the human pituitary-adrenal axis. The difference in duration of effect between hCRF and oCRF may be attributed to an approximately 3-4 times higher MCR of hCRF [7.9 ± 1.2 (±SE) ml/kg min; n = 14] than oCRF (1.9 ± 0.1 ml/kg min; n = 9) in man. No serious side effects occurred at any of the doses of hCRF tested. The highest dose (5 μg/kg) caused a slight increase of heart rate that was not associated with significant changes in arterial blood pressure. All subjects receiving 5 μg/kg and one third of the subjects receiving 1 μg/kg hCRF experienced a transient facial flush. We conclude that hCRF causes brief plasma ACTH and cortisol secretory episodes in man, similar to the physiological plasma ACTH and cortisol secretory episodes. This is in contrast to oCRF, which causes prolonged ACTH and cortisol secretion. These differences between the two peptides may be explained by the higher MCR of hCRF than oCRF.

In primates, ovine and human CRH cause ACTH- and cortisol secretion in a dose-dependent fashion. Ovine CRH has a half-life, which is about three times longer than that of human CRH and results in long lasting ACTH- and cortisol responses. Plasma ACTH- and cortisol rises after administration of human CRH mimic the spontaneously occurring secretory episodes of these hormones in man. Pulsatile administration of human CRH restores the diurnal secretory pattern of ACTH and cortisol to normal in patients with apparent CRH deficiency. Facial and upper body flush occur in about 20% of patients receiving intravenously a 1 microgram/kg dose of CRH. Higher doses result in hypotension due to a decrease in peripheral vascular resistance. The latter is primarily due to dilatation of the superior mesenteric vessels. In in vitro models desensitization of the pituitary corticotroph by continuous, high-dose CRH administration occurs. No down-regulation of ACTH- and cortisol secretion has been shown in vivo, however. Pituitary-adrenal responsiveness to CRH can be modulated by many factors. These factors are involved in the basal regulation of the hypothalamic-pituitary-adrenal axis and in its activation during the stress response. Such factors include AVP, morphine, DAMME, and drugs modulating the endogenous serotoninergic and GABA/benzodiazepine system. Glucocorticoid feedback inhibition of the pituitary is one of the most important factors modulating ACTH- and cortisol responses to CRH. A negative correlation exists between the net ACTH- and cortisol response to exogenous CRH and the basal cortisol plasma concentration. Replacement doses of glucocorticoid result in a drastic decrease of basal plasma concentrations of cortisol and adrenal androgens.(ABSTRACT TRUNCATED AT 250 WORDS)

To examine the validity and the reliability of a novel measurement tool, the Newly Diagnosed Breast Cancer Stress Scale (NDBCSS) in the Greek population. The tool aimed to assess distress in patients recently diagnosed with breast cancer.We performed a principal component analysis (PCA) of the 17 items of the scale.The PCA resulted in 4 factors: 1. Personal life, 2. Procedural issues, 3. Facing challenges and 4. Psychological load. All subscales showed satisfactory internal consistency and variance, relative to theoretical score ranges. Subscale scores and total score were significantly correlated with perceived stress and hospital anxiety and depression scale, implying good criterion validity. Associations with social, demographic and disease related information were also found.The NDBCSS resulted in acceptable reliability and good validity, and was considered as a valuable tool for health-care workers and oncologists to measure psychological distress in early stage of breast cancer.

Diabetic nephropathy (DN), also named diabetic kidney disease (DKD), is a devastating complication in patients with both type 1 and 2 diabetes mellitus (T1D and T2D) and its diagnosis has been traditionally based on the presence of micro-albuminuria (MA). The aim of this article is to update, through review of the relevant medical literature, the most promising biomarkers for early DKD detection. MA has historically been employed as an early marker of microvascular complications, indicating risk for advanced CKD. However, due to the inability of MA to adequately predict DKD, especially in young patients or in non-albuminuric DKD, additional biomarkers of glomerular and/or tubular injury have been proposed to uncover early renal dysfunction and structural lesions, even before MA occurs. Defining new predictive biomarkers to use alongside urinary albumin excretion (UAE) during the initial stages of DKD would provide a window of opportunity for preventive and/or therapeutic interventions to prevent or delay the onset of irreversible long-term complications and to improve outcomes by minimizing the rates of severe cardio-renal morbidity and mortality in DKD patients.

Familial glucocorticoid resistance is characterized by increased cortisol secretion without clinical evidence of hypercortisolism, but with manifestations of androgen and mineralocorticoid excess. This condition is mainly caused by mutations of the GR gene that cause inadequate transduction of the glucocorticoid signal in glucocorticoid target tissues. The clinical features of glucocorticoid resistance in females include hirsutism, acne, male pattern baldness, oligomenorrhea, and oligoanovulation. We describe here a new phenotype, female pseudohermaphroditism and severe hypokalemia, caused by a homozygous inactivating mutation of the GR gene. The proband was born with ambiguous genitalia from consanguineous parents and was mistreated as a 21-hydroxylase deficiency case since the age of 5 yr. She had very high levels of plasma ACTH (759 pg/ml or 167 pmol/liter) and high levels of cortisol (28–54 μg/dl or 772-1490 nmol/liter), androstenedione (5–14 ng/ml or 17–48 nmol/liter), T (174–235 ng/dl or 7–8 nmol/liter), and 17-hydroxyprogesterone (8–12 ng/ml or 24–36 nmol/liter). Her cortisol and 17-hydroxyprogesterone levels were not compatible with the diagnosis of classic congenital adrenal hyperplasia; furthermore, cortisol was not properly suppressed after dexamethasone administration (28 μg/d or 772 nmol/liter). Her laboratory evaluation indicated a diagnosis of glucocorticoid resistance. To investigate this puzzling clinical and biochemical picture, we analyzed both GR and CYP21 genes. Indeed, a homozygous T to C substitution at nucleotide 1844 in exon 5 of the GR gene was identified in the patient that caused a valine to alanine substitution at amino acid 571 in the ligand domain of the receptor. Her parents and an older sister were heterozygous for this mutation. A whole Epstein-Barr virus-transformed cell dexamethasone-binding assay revealed that this Ala571 mutant had a 6-fold reduction in binding affinity compared with the wild-type receptor. In a functional assay using mouse mammary tumor virus promoter-driven luciferase reporter gene, the mutant receptor displayed 10- to 50-fold less trans-activation activity than the wild-type receptor. In addition, a large heterozygous CYP21 conversion was identified in the patient and her father. In conclusion, we described the first case of female pseudohermaphroditism caused by a novel homozygous GR gene mutation. This phenotype indicates that pre- and postnatal virilization can occur in females with the glucocorticoid resistance syndrome.

Background: Osteodysplastic syndromes compose a very diverse group of clinically and genetically heterogeneous disorders characterized by defects in bone and connective tissue development, as well as in bone density. Here, we report the case of a 48-year-old female with a complex medical history characterized by bone dysplasia, hyperostosis, and partial tooth agenesis. Methods: Genetic testing was performed using WES analysis and Sanger sequencing. Molecular modeling analysis and dynamics simulation explored the impact of detected pathogenic variants. Results: The genetic analysis detected multiple pathogenic variants in genes CREB3L1, SLCO2A1, SFRP4, LRP5 and LRP6, which are associated with rare osteodysplastic syndromes. The patient was homozygous for the same rare alleles associated with three of the identified autosomal recessive disorders, such as osteogenesis imperfecta type XVI, primary hypertrophic osteoarthropathy, and metaphyseal dysplasia Pyle type. She also had a variant linked to autosomal dominant endosteal hyperostosis and a variant previously associated with increased risk of osteoporosis and fractures. Two of the detected variants are predicted to cause abnormal splicing, while molecular modelling and dynamics simulations analysis suggest that the other three variants probably confer altered local secondary structure and flexibility that may be functionally devastating consequences. Conclusions: Our case highlights the rare coexistence of multiple osteodysplastic syndromes in a single patient that may complicate differential diagnosis. Furthermore, this case emphasizes the necessity for early genetic investigation of such complex cases with overlying phenotypic traits, followed by genetic counseling, in order to orchestrate clinical interventions and prevent or promptly manage manifestations.