Methylome analysis in girls with idiopathic central precocious puberty.

BACKGROUND: Genetic and environmental factors are implicated in many developmental processes. Recent evidence, however, has suggested that epigenetic changes may also influence the onset of puberty or the susceptibility to a wide range of diseases later in life. The present study aims to investigate changes in genomic DNA methylation profiles associated with pubertal onset analyzing human peripheral blood leukocytes from three different groups of subjects: 19 girls with central precocious puberty (CPP), 14 healthy prepubertal girls matched by age and 13 healthy pubertal girls matched by pubertal stage. For this purpose, the comparisons were performed between pre- and pubertal controls to identify changes in normal pubertal transition and CPP versus pre- and pubertal controls. RESULTS: Analysis of methylation changes associated with normal pubertal transition identified 1006 differentially methylated CpG sites, 86% of them were found to be hypermethylated in prepubertal controls. Some of these CpG sites reside in genes associated with the age of menarche or transcription factors involved in the process of pubertal development. Analysis of methylome profiles in CPP patients showed 65% and 55% hypomethylated CpG sites compared with prepubertal and pubertal controls, respectively. In addition, interestingly, our results revealed the presence of 43 differentially methylated genes coding for zinc finger (ZNF) proteins. Gene ontology and IPA analysis performed in the three groups studied revealed significant enrichment of them in some pathways related to neuronal communication (semaphorin and gustation pathways), estrogens action, some cancers (particularly breast and ovarian) or metabolism (particularly sirtuin). CONCLUSIONS: The different methylation profiles of girls with normal and precocious puberty indicate that regulation of the pubertal process in humans is associated with specific epigenetic changes. Differentially methylated genes include ZNF genes that may play a role in developmental control. In addition, our data highlight changes in the methylation status of genes involved in signaling pathways that determine the migration and function of GnRH neurons and the onset of metabolic and neoplastic diseases that may be associated with CPP in later life.

Bimodal buckling governs human fingers' luxation.

Equilibrium bifurcation in natural systems can sometimes be explained as a route to stress shielding for preventing failure. Although compressive buckling has been known for a long time, its less-intuitive tensile counterpart was only recently discovered and yet never identified in living structures or organisms. Through the analysis of an unprecedented all-in-one paradigm of elastic instability, it is theoretically and experimentally shown that coexistence of two curvatures in human finger joints is the result of an optimal design by nature that exploits both compressive and tensile buckling for inducing luxation in case of traumas, so realizing a unique mechanism for protecting tissues and preventing more severe damage under extreme loads. Our findings might pave the way to conceive complex architectured and bio-inspired materials, as well as next generation artificial joint prostheses and robotic arms for bio-engineering and healthcare applications.

Circulating levels of DLK1 and glucose homeostasis in girls with obesity: A pilot study.

Introduction: DLK1 gene is considered a molecular gatekeeper of adipogenesis. DLK1 mutations have been reported as a cause of central precocious puberty associated with obesity and metabolic syndrome with undetectable DLK1 serum levels. We investigated the association between DLK1 circulating levels with clinical and biochemical parameters in obese adolescents and healthy controls. Methods: Sixty-five obese adolescents and 40 controls were enrolled and underwent a complete clinical examination and biochemical assessment for glucose homeostasis and DLK1 plasma levels. Results: We observed lower DLK1 levels in cases compared to controls. Moreover, we found a negative correlation between DLK1 and HOMA-IR and a direct correlation with insulin-sensitivity index. Discussion: Our findings suggest that DLK1 might be involved in metabolic derangement in obese children.

MKRN3 role in regulating pubertal onset: the state of art of functional studies.

Puberty is a critical process characterized by several physical and psychological changes that culminate in the achievement of sexual maturation and fertility. The onset of puberty depends on several incompletely understood mechanisms that certainly involve gonadotropin-releasing hormone (GnRH) and its effects on the pituitary gland. The role of makorin ring finger protein 3 (MKRN3) in the regulation of pubertal timing was revealed when loss-of-function mutations were identified in patients with central precocious puberty (CPP), which to date, represent the most commonly known genetic cause of this condition. The MKRN3 gene showed ubiquitous expression in tissues from a broad spectrum of species, suggesting an important cellular role. Its involvement in the initiation of puberty and endocrine functions has just begun to be studied. This review discusses some of the recent approaches developed to predict MKRN3 functions and its involvement in pubertal development.

Case report: Goiter and overt hypothyroidism in an iodine-deficient toddler on soy milk and hypoallergenic diet.

Soy-based infant formulas (SFs) are often consumed by cow's milk allergic children. However, some concerns have risen since soy intake may adversely affect thyroid function in iodine-deficient or subclinical hypothyroid individuals. We report the first Italian case of SF induced goiter and hypothyroidism registered in our country since National Iodine program has been instituted. Finally, we review cases previously reported in literature. A 22-month-old toddler with a previous diagnosis of cow's milk protein allergy came to clinical attention for important goiter and overt hypothyroidism. Detailed dietary anamnesis revealed that he was on a restrictive dietary regimen based on soymilk since 12 months of age. A temporary levothyroxine substitution was instituted to avoid hypothyroidism complications. Adequate iodine supplementation and diet diversification completely reversed SF-induced hypothyroidism and goiter, confirming the diagnostic suspicion of soymilk-induced thyroid dysfunction in a iodine-deficient toddler. This case report demonstrates the importance of careful dietary habits investigation and adequate micronutrients supplementation in children on a restrictive diet due to multiple food allergies in order to prevent nutritional deficits.

Potential Therapeutic Effects of Long-Term Stem Cell Administration: Impact on the Gene Profile and Kidney Function of PKD/Mhm (Cy/+) Rats.

Cystic kidney disease (CKD) is a heterogeneous group of genetic disorders and one of the most common causes of end-stage renal disease. Here, we investigate the potential effects of long-term human stem cell treatment on kidney function and the gene expression profile of PKD/Mhm (Cy/+) rats. Human adipose-derived stromal cells (ASC) and human skin-derived ABCB5+ stromal cells (2 × 106) were infused intravenously or intraperitoneally monthly, over 6 months. Additionally, ASC and ABCB5+-derived conditioned media were administrated intraperitoneally. The gene expression profile results showed a significant reprogramming of metabolism-related pathways along with downregulation of the cAMP, NF-kB and apoptosis pathways. During the experimental period, we measured the principal renal parameters as well as renal function using an innovative non-invasive transcutaneous device. All together, these analyses show a moderate amelioration of renal function in the ABCB5+ and ASC-treated groups. Additionally, ABCB5+ and ASC-derived conditioned media treatments lead to milder but still promising improvements. Even though further analyses have to be performed, the preliminary results obtained in this study can lay the foundations for a novel therapeutic approach with the application of cell-based therapy in CKD.

Ultrasound waves in tumors via needle irradiation for precise medicine.

Grounded in the interdisciplinary crosstalk among physics and biological sciences, precision medicine-based diagnosis and treatment strategies have recently gained great attention for the actual applicability of new engineered approaches in many medical fields, particularly in oncology. Within this framework, the use of ultrasounds employed to attack cancer cells in tumors to induce possible mechanical damage at different scales has received growing attention from scholars and scientists worldwide. With these considerations in mind, on the basis of ad hoc elastodynamic solutions and numerical simulations, we propose a pilot study for in silico modeling of the propagation of ultrasound waves inside tissues, with the aim of selecting proper frequencies and powers to be irradiated locally through a new teragnostic platform based on Lab-on-Fiber technology, baptized as a hospital in the needle and already the object of a patent. It is felt that the outcomes and the related biophysical insights gained from the analyses could pave the way for envisaging new integrated diagnostic and therapeutic approaches that might play a central role in future applications of precise medicine, starting from the growing synergy among physics, engineering and biology.

Growth Hormone Receptor (GHR) 6Ω Pseudoexon Activation: a Novel Cause of Severe Growth Hormone Insensitivity.

CONTEXT: Severe forms of growth hormone insensitivity (GHI) are characterized by extreme short stature, dysmorphism, and metabolic anomalies. OBJECTIVE: This work aims to identify the genetic cause of growth failure in 3 "classical" GHI individuals. METHODS: A novel intronic growth hormone receptor gene (GHR) variant was identified, and in vitro splicing assays confirmed aberrant splicing. A 6Ω pseudoexon GHR vector and patient fibroblast analysis assessed the consequences of the novel pseudoexon inclusion and the impact on GHR function. RESULTS: We identified a novel homozygous intronic GHR variant (g.5:42700940T > G, c.618+836T > G), 44 bp downstream of the previously recognized intronic 6Ψ GHR pseudoexon mutation in the index patient. Two siblings also harbored the novel intronic 6Ω pseudoexon GHR variant in compound heterozygosity with the known GHR c.181C > T (R43X) mutation. In vitro splicing analysis confirmed inclusion of a 151-bp mutant 6Ω pseudoexon not identified in wild-type constructs. Inclusion of the 6Ω pseudoexon causes a frameshift resulting in a nonfunctional truncated GHR lacking the transmembrane and intracellular domains. The truncated 6Ω pseudoexon protein demonstrated extracellular accumulation and diminished activation of STAT5B signaling following GH stimulation. CONCLUSION: Novel GHR 6Ω pseudoexon inclusion results in loss of GHR function consistent with a severe GHI phenotype. This represents a novel mechanism of Laron syndrome and is the first deep intronic variant identified causing severe postnatal growth failure. The 2 kindreds originate from the same town in Campania, Southern Italy, implying common ancestry. Our findings highlight the importance of studying variation in deep intronic regions as a cause of monogenic disorders.

Growth Hormone Receptor (Ghr) 6ω Pseudoexon Activation: A Novel Cause Of Severe Growth Hormone Insensitivity (Ghi).

CONTEXT: Severe forms of Growth Hormone Insensitivity (GHI) are characterized by extreme short stature, dysmorphism and metabolic anomalies. OBJECTIVE: Identification of the genetic cause of growth failure in 3 'classical' GHI subjects. DESIGN: A novel intronic GHR variant was identified, and in vitro splicing assays confirmed aberrant splicing. A 6Ω pseudoexon GHR vector and patient fibroblast analysis assessed the consequences of the novel pseudoexon inclusion and the impact on GHR function. RESULTS: We identified a novel homozygous intronic GHR variant (g.5:42700940T>G, c.618 + 836T> G), 44bp downstream of the previously recognized intronic 6Ψ GHR pseudoexon mutation in the index patient. Two siblings also harbored the novel intronic 6Ω pseudoexon GHR variant in compound heterozygosity with the known GHR c.181C>T (R43X) mutation. In vitro splicing analysis confirmed inclusion of a 151bp mutant 6Ω pseudoexon not identified in wild-type constructs. Inclusion of the 6Ω pseudoexon causes a frameshift resulting in a non-functional truncated GHR lacking the transmembrane and intracellular domains. The truncated 6Ω pseudoexon protein demonstrated extracellular accumulation and diminished activation of STAT5B signaling following growth hormone stimulation. CONCLUSION: Novel GHR 6Ω pseudoexon inclusion results in loss of GHR function consistent with a severe GHI phenotype. This represents a novel mechanism of Laron syndrome and is the first deep intronic variant identified causing severe postnatal growth failure. The 2 kindreds originate from the same town in Campania, Southern Italy, implying common ancestry. Our findings highlight the importance of studying variation in deep intronic regions as a cause of monogenic disorders.

A lesson from earthquake engineering for selectively damaging cancer cell structures.

The progressive falling of barriers among disciplines is opening unforeseen scenarios in diagnosis and treatment of cancer diseases. By sharing models and mature knowledge in physics, engineering, computer sciences and molecular biology, synergistic efforts have in fact contributed in the last years to re-think still unsolved problems, shedding light on key roles of mechanobiology in tumors and envisaging new effective strategies for a precise medicine. The use of ultrasounds for altering cancer cells' program is one of the most attracting grounds to be explored in oncophysics, although how to administer mechanical energy to impair selected cell structures and functions simultaneously overcoming the critical trade-off between the impact of the cure and the patient risk still remains an open issue. Within this framework, by starting from the theoretical possibility of selectively attacking malignant cells by exploiting the stiffness discrepancies between tumor and healthy single cells, first proposed by Fraldi et al. (2015), we here investigate the in-frequency response of an overall spherical close-packing of geometrically equal polyhedral cells to gain insights into how mechanical resonance and vibration-induced failure phenomena can be oriented to destroy specific target units when both the cell populations coexist, as it happens for in vivo cases. Inspired by the dynamic action of earthquakes - which fracture only selected elements among adjacent ones in the same structure or damage individual constructions in contiguous buildings - we study the harmonic response of hierarchically architectured cell agglomerates, inhabited by both tumor and healthy cells that interact mutually throughout the extra-cellular matrix and whose cytoskeleton is modeled as a nonlinear soft-tensegrity structure. Numerical Finite Element results show that, at frequencies compatible with low intensity therapeutic ultrasounds, mechanical resonance and possible fatigue cycles of the pre-stressed actin filaments and microtubules can be selectively induced in cancer cells as a function of the global volume fraction of the cell species, paving the way for future engineered treatment protocols.

Competition between delamination and tearing in multiple peeling problems.

Adhesive attachment systems consisting of multiple tapes or strands are commonly found in nature, for example in spider web anchorages or in mussel byssal threads, and their structure has been found to be ingeniously architected in order to optimize mechanical properties: in particular, to maximize dissipated energy before full detachment. These properties emerge from the complex interplay between mechanical and geometric parameters, including tape stiffness, adhesive energy, attached and detached lengths and peeling angles, which determine the occurrence of three main mechanisms: elastic deformation, interface delamination and tape fracture. In this paper, we introduce a formalism to evaluate the mechanical performance of multiple tape attachments in different parameter ranges, where an optimal (not maximal) adhesion energy emerges. We also introduce a numerical model to simulate the multiple peeling behaviour of complex structures, illustrating its predictions in the case of the staple-pin architecture. Finally, we present a proof-of-principle experiment to illustrate the predicted behaviour. We expect the presented formalism and the numerical model to provide important tools for the design of bioinspired adhesive systems with tuneable or optimized detachment properties.

A New APEH Cluster with Antioxidant Functions in the Antarctic Hemoglobinless Icefish Chionodraco hamatus.

Acylpeptide hydrolase (APEH) is a ubiquitous cytosolic protease that plays an important role in the detoxification of oxidised proteins. In this work, to further explore the physiological role of this enzyme, two apeh cDNAs were isolated from the Chionodraco hamatus icefish, which lives in the highly oxygenated Antarctic marine environment. The encoded proteins (APEH-1(Ch) and APEH-2(Ch)) were characterised in comparison with the uniquely expressed isoform from the temperate fish Dicentrarchus labrax (APEH-1Dl) and the two APEHs from the red-blooded Antarctic fish Trematomus bernacchii (APEH-1(Tb) and APEH-2(Tb)). Homology modelling and kinetic characterisation of the APEH isoforms provided new insights into their structure/function properties. APEH-2 isoforms were the only ones capable of hydrolysing oxidised proteins, with APEH-2(Ch) being more efficient than APEH-2(Tb) at this specific function. Therefore, this ability of APEH-2 isoforms is the result of an evolutionary adaptation due to the pressure of a richly oxygenated environment. The lack of expression of APEH-2 in the tissues of the temperate fish used as the controls further supported this hypothesis. In addition, analysis of gene expression showed a significant discrepancy between the levels of transcripts and those of proteins, especially for apeh-2 genes, which suggests the presence of post-transcriptional regulation mechanisms, triggered by cold-induced oxidative stress, that produce high basal levels of APEH-2 mRNA as a reserve that is ready to use in case of the accumulation of oxidised proteins.