Molecular profile of sensitization in subjects with short occupational exposure to latex

Objectives: We examined the prevalence of latex allergy in subjects with occupational exposure to latex allergens for less than 5 years, determining the disease spectrum in symptomatic workers. We identified the most frequent molecular allergens by Immuno- CAP (ICAP), correlating the findings with skin prick test (SPT) results. Material and Methods: Seven hundred twenty-three healthcare students using latex gloves on a regular basis were invited to participate in a baseline questionnaire screening. An ICAP serum test was performed only when a possible latex allergy was indicated by the questionnaire. Results: The total number of participants responding to the baseline survey was 619. Glove-related symptoms were indicated by 4% (N = 25) of the students. The most common symptom was contact dermatitis (N = 18, 72%). In 12 subjects, ICAP revealed a real sensitization to latex, with a recombinant latex allergen profile showing a high frequency for rHev b 6.01 specific immunoglobulin E (sIgE) (N = 9, 67%). In these individuals, skin symptoms were more prevalent than other types (88%). Conclusions: The combined positivity for rHev b 6.01, rHev 8 and rHev b 5 determined by ICAP identified 92% of latex-allergic subjects with short-term exposure to latex

Microstructural features of human bones and funerary practices in Mount Sirai (Sardinia)

In the attempt to set up a useful methodology for the investigation of burned human remains in archaeological, anthropological and forensic fields, we decided to compare the most common protocols for the study of bone bioapatites (Fourier Transform Infrared spectroscopy, FT-IR, and X-ray Diffraction, XRD) to those deriving from the application of X-ray scattering techniques using synchrotron light. In this way, we expect to take advantage of the wider and more dynamic qualities of such a valuable tool in order to examine a higher number of samples in a very short time compared to the “traditional” techniques, meanwhile assessing its applicability in the archaeological field

Inhibition of microglia overactivation restores neuronal survival in a mouse model of CDKL5 deficiency disorder

open9noFunding: This work was supported by the Telethon foundation (grant number GGP19045 to EC), by the Italian parent associations “CDKL5 insieme verso la cura” to EC and by the CARISBO foundation (grant number 2020.0400 to ST).CDKL5 deficiency disorder (CDD), a severe neurodevelopmental disorder characterized by early onset epilepsy, intellectual disability, and autistic features, is caused by mutations in the CDKL5 gene. Evidence in animal models of CDD showed that absence of CDKL5 negatively affects neuronal survival, as well as neuronal maturation and dendritic outgrowth; however, knowledge of the substrates underlying these alterations is still limited. Neuroinflammatory processes are known to contribute to neuronal dysfunction and death. Recent evidence shows a subclinical chronic inflammatory status in plasma from CDD patients. However, to date, it is unknown whether a similar inflammatory status is present in the brain of CDD patients and, if so, whether this plays a causative or exacerbating role in the pathophysiology of CDD.openGalvani, Giuseppe; Mottolese, Nicola; Gennaccaro, Laura; Loi, Manuela; Medici, Giorgio; Tassinari, Marianna; Fuchs, Claudia; Ciani, Elisabetta; Trazzi, StefaniaGalvani, Giuseppe; Mottolese, Nicola; Gennaccaro, Laura; Loi, Manuela; Medici, Giorgio; Tassinari, Marianna; Fuchs, Claudia; Ciani, Elisabetta; Trazzi, Stefani

Early-onset brain alterations during postnatal development in a mouse model of CDKL5 deficiency disorder

Mutations in the CDKL5 gene are the cause of CDKL5 deficiency disorder (CDD), a rare and severe neurodevelopmental condition characterized by early-onset epilepsy, motor impairment, intellectual disability, and autistic features. A mouse model of CDD, the Cdkl5 KO mouse, that recapitulates several aspects of CDD symptomology, has helped to highlight brain alterations leading to CDD neurological defects. Studies of brain morphogenesis in adult Cdkl5 KO mice showed defects in dendritic arborization of pyramidal neurons and in synaptic connectivity, a hypocellularity of the hippocampal dentate gyrus, and a generalized microglia over-activation. Nevertheless, no studies are available regarding the presence of these brain alterations in Cdkl5 KO pups, and their severity in early stages of life compared to adulthood. A deeper understanding of the CDKL5 deficient brain during an early phase of postnatal development would represent an important milestone for further validation of the CDD mouse model, and for the identification of the optimum time window for treatments that target defects in brain development. In sight of this, we comparatively evaluated the dendritic arborization and spines of cortical pyramidal neurons, cortical excitatory and inhibitory connectivity, microglia activation, and proliferation and survival of granule cells of the hippocampal dentate gyrus in hemizygous Cdkl5 KO male (-/Y) mice aged 7, 14, 21, and 60 days. We found that most of the structural alterations in Cdkl5 -/Y brains are already present in pups aged 7 days and do not worsen with age. In contrast, the difference in the density of excitatory and inhibitory terminals between Cdkl5 -/Y and wild-type mice changes with age, suggesting an age-dependent cortical excitatory/inhibitory synaptic imbalance. Confirming the precocious presence of brain defects, Cdkl5 -/Y pups are characterized by an impairment in neonatal sensory-motor reflexes

Luteolin Treatment Ameliorates Brain Development and Behavioral Performance in a Mouse Model of CDKL5 Deficiency Disorder

CDKL5 deficiency disorder (CDD), a rare and severe neurodevelopmental disease caused by mutations in the X-linked CDKL5 gene, is characterized by early-onset epilepsy, intellectual disability, and autistic features. Although pharmacotherapy has shown promise in the CDD mouse model, safe and effective clinical treatments are still far off. Recently, we found increased microglial activation in the brain of a mouse model of CDD, the Cdkl5 KO mouse, suggesting that a neuroinflammatory state, known to be involved in brain maturation and neuronal dysfunctions, may contribute to the pathophysiology of CDD. The present study aims to evaluate the possible beneficial effect of treatment with luteolin, a natural flavonoid known to have anti-inflammatory and neuroprotective activities, on brain development and behavior in a heterozygous Cdkl5 (+/-) female mouse, the mouse model of CDD that best resembles the genetic clinical condition. We found that inhibition of neuroinflammation by chronic luteolin treatment ameliorates motor stereotypies, hyperactive profile and memory ability in Cdkl5 +/- mice. Luteolin treatment also increases hippocampal neurogenesis and improves dendritic spine maturation and dendritic arborization of hippocampal and cortical neurons. These findings show that microglia overactivation exerts a harmful action in the Cdkl5 +/- brain, suggesting that treatments aimed at counteracting the neuroinflammatory process should be considered as a promising adjuvant therapy for CDD

Treatment with a GSK-3β/HDAC Dual Inhibitor Restores Neuronal Survival and Maturation in an In Vitro and In Vivo Model of CDKL5 Deficiency Disorder

open11noFunding: This research was funded by the Telethon foundation (grant number GGP19045, awarded to E.C.), and by the Italian parent association “CDKL5 insieme verso la cura” (to E.C.).Mutations in the X-linked cyclin-dependent kinase-like 5 (CDKL5) gene cause a rare neurodevelopmental disorder characterized by early-onset seizures and severe cognitive, motor, and visual impairments. To date there are no therapies for CDKL5 deficiency disorder (CDD). In view of the severity of the neurological phenotype of CDD patients it is widely assumed that CDKL5 may influence the activity of a variety of cellular pathways, suggesting that an approach aimed at targeting multiple cellular pathways simultaneously might be more effective for CDD. Previous findings showed that a single-target therapy aimed at normalizing impaired GSK-3β or histone deacetylase (HDAC) activity improved neurodevelopmental and cognitive alterations in a mouse model of CDD. Here we tested the ability of a first-in-class GSK-3β/HDAC dual inhibitor, Compound 11 (C11), to rescue CDD-related phenotypes. We found that C11, through inhibition of GSK-3β and HDAC6 activity, not only restored maturation, but also significantly improved survival of both human CDKL5-deficient cells and hippocampal neurons from Cdkl5 KO mice. Importantly, in vivo treatment with C11 restored synapse development, neuronal survival, and microglia over-activation, and improved motor and cognitive abilities of Cdkl5 KO mice, suggesting that dual GSK-3β/HDAC6 inhibitor therapy may have a wider therapeutic benefit in CDD patients.openLoi, Manuela; Gennaccaro, Laura; Fuchs, Claudia; Trazzi, Stefania; Medici, Giorgio; Galvani, Giuseppe; Mottolese, Nicola; Tassinari, Marianna; Giorgini, Roberto Rimondini; Milelli, Andrea; Ciani, ElisabettaLoi, Manuela; Gennaccaro, Laura; Fuchs, Claudia; Trazzi, Stefania; Medici, Giorgio; Galvani, Giuseppe; Mottolese, Nicola; Tassinari, Marianna; Giorgini, Roberto Rimondini; Milelli, Andrea; Ciani, Elisabett

Interaction of Vault Particles with Estrogen Receptor in the MCF-7 Breast Cancer Cell

A 104-kD protein was coimmunoprecipitated with the estrogen receptor from the flowtrough of a phosphocellulose chromatography of MCF-7 cell nuclear extract. mAbs to this protein identified several cDNA clones coding for the human 104-kD major vault protein. Vaults are large ribonucleoprotein particles of unknown function present in all eukaryotic cells. They have a complex morphology, including several small molecules of RNA, but a single protein species, the major vault protein, accounts for >70% of their mass. Their shape is reminiscent of the nucleopore central plug, but no proteins of known function have been described to interact with them. Western blot analysis of vaults purified on sucrose gradient showed the presence of estrogen receptor co-migrating with the vault peak. The AER317 antibody to estrogen receptor coimmunoprecipitated the major vault protein and the vault RNA also in the 20,000 g supernatant fraction. Reconstitution experiments of estrogen receptor fragments with the major vault protein mapped the site of the interaction between amino acids 241 and 280 of human estrogen receptor, where the nuclear localization signal sequences are located. Estradiol treatment of cells increased the amount of major vault protein present in the nuclear extract and coimmunoprecipitated with estrogen receptor, whereas the anti-estrogen ICI182,780 had no effect. The hormone-dependent interaction of vaults with estrogen receptor was reproducible in vitro and was prevented by sodium molybdate. Antibodies to progesterone and glucocorticoid receptors were able to coimmunoprecipitate the major vault protein. The association of nuclear receptors with vaults could be related to their intracellular traffic

Cardiac Functional and Structural Abnormalities in a Mouse Model of CDKL5 Deficiency Disorder

CDKL5 (cyclin-dependent kinase-like 5) deficiency disorder (CDD) is a severe neurodevelopmental disease that mostly affects girls, who are heterozygous for mutations in the X-linked CDKL5 gene. Mutations in the CDKL5 gene lead to a lack of CDKL5 protein expression or function and cause numerous clinical features, including early-onset seizures, marked hypotonia, autistic features, gastrointestinal problems, and severe neurodevelopmental impairment. Mouse models of CDD recapitulate several aspects of CDD symptomology, including cognitive impairments, motor deficits, and autistic-like features, and have been useful to dissect the role of CDKL5 in brain development and function. However, our current knowledge of the function of CDKL5 in other organs/tissues besides the brain is still quite limited, reducing the possibility of broad-spectrum interventions. Here, for the first time, we report the presence of cardiac function/structure alterations in heterozygous Cdkl5 +/- female mice. We found a prolonged QT interval (corrected for the heart rate, QTc) and increased heart rate in Cdkl5 +/- mice. These changes correlate with a marked decrease in parasympathetic activity to the heart and in the expression of the Scn5a and Hcn4 voltage-gated channels. Interestingly, Cdkl5 +/- hearts showed increased fibrosis, altered gap junction organization and connexin-43 expression, mitochondrial dysfunction, and increased ROS production. Together, these findings not only contribute to our understanding of the role of CDKL5 in heart structure/function but also document a novel preclinical phenotype for future therapeutic investigation

Highlighting chromosome loops in DNA-picked chromatin (DPC).

"Growing evidence supports the concept that dynamic intra-and inter-chromosomal links between specific loci contribute to the creation of cell type-specific gene expression profiles. Therefore, analysis of the establishment of peculiar functional correlations between sites, also distant on linear DNA, that govern the transcriptional process appears to be of fundamental relevance. We propose here an experimental approach showing that 17 beta-estradiol-induced transcription associates to formation of loops between the promoter and termination regions of hormone-responsive genes. This strategy reveals as a tool to be also suitably used, in conjunction with automated techniques, for an extensive analysis of sites shared by multiple genes for induced expression.

Treatment with FRAX486 rescues neurobehavioral and metabolic alterations in a female mouse model of CDKL5 deficiency disorder

Introduction: CDKL5 deficiency disorder (CDD) is a rare neurodevelopmental condition, primarily affecting girls for which no cure currently exists. Neuronal morphogenesis and plasticity impairments as well as metabolic dysfunctions occur in CDD patients. The present study explored the potential therapeutic value for CDD of FRAX486, a brain-penetrant molecule that was reported to selectively inhibit group I p21-activated kinases (PAKs), serine/threonine kinases critically involved in the regulation of neuronal morphology and glucose homeostasis.Methods: The effects of treatment with FRAX486 on CDD-related alterations were assessed in vitro (100 nM for 48h) on primary hippocampal cultures from Cdkl5-knockout male mice (Cdkl5-KO) and in vivo (20 mg/Kg, s.c. for 5 days) on Cdkl5-KO heterozygous females (Cdkl5-Het).Results: The in vitro treatment with FRAX486 completely rescued the abnormal neuronal maturation and the number of PSD95-positive puncta in Cdkl5-KO mouse neurons. In vivo, FRAX486 normalized the general health status, the hyperactive profile and the fear learning defects of fully symptomatic Cdkl5-Het mice. Systemically, FRAX486 treatment normalized the levels of reactive oxidizing species in the whole blood and the fasting-induced hypoglycemia displayed by CdklS-Het mice. In the hippocampus of Cdkl5-Het mice, treatment with FRAX486 rescued spine maturation and PSD95 expression and restored the abnormal PAKs phosphorylation at sites which are critical for their activation (P-PAK-Ser144/141/139) or for the control cytoskeleton remodeling (P-PAK1-Thr212).Conclusions: Present results provide evidence that PAKs may represent innovative therapeutic targets for CDD