Transcriptional regulation of RACK1 and modulation of its expression: Role of steroid hormones and significance in health and aging

The Receptor for Activated C Kinase 1 (RACK1) is a scaffold protein for different kinases and membrane receptors. RACK1 can shuttle proteins to their sites of action, facilitate cross- talk among distinct signaling pathways or recruit other signaling proteins into the complexes. Therefore, it is a key mediator of various pathways and is involved in various biological events including development, immune response, brain activity and cancer. Because of its importance, it is of extreme significance to understand the transcriptional mechanisms governing its expression. The identification of regulatory elements in the promoter of RACK1 shed some light on its transcriptional modulation in physiological and pathological context. Literature data support the existence of a complex hormonal balance, between glucocorticoids and androgens, in the control of RACK1 expression due to specific and complex interactions on the RACK1 promoter. These and other informations suggest that a better understanding of RACK1 transcriptional regulation is essential to unravel its role. Furthermore, the modulation of its expression in physiological or pathological conditions may be of interest in different context, such as aging and cancer

Cloning and knock-out of the mouse gene coding for the high mobility group 2 protein (HMG2)

High Mobility Group proteins 1 and 2 are highly conserved nuclear proteins ubiquitously expressed in higher eukaryotic cells. HMG1 and HMG2 each contain two similar DNA binding domains, called HMG-boxes, and an acidic tail. HMG1 and HMG2 are able to facilitate the binding to DNA of several transcription factors (Steroid Hormone Receptors, HOX and OCT proteins) and they are involved in V(D)J recombination. hmg1 -1- mice, produced in our lab by conventional knock-out, are bom and die within the first day of life. The cause of death is a severe hypoglycaemia: the level of glucose is low in the blood of mutant mice, while abundant glycogen is still present in their liver. Glucocorticoid-dependent gene expression is impaired in -/- mice. Surprisingly, HMG1 is not essential for the life of the cell, as it might be suggested by the abundance of the protein and its evolutionary conservation. We then suggested that another protein can substitute HMG1 in its function in the cell. The best candidate is HMG2 because of its remarkable similarity with HMG1. Therefore, I decided to investigate the function of HMG2 by gene targeting, and I planned the work so as to investigate similarities and differences between HMG1 and HMG2. The mouse hmg2 gene was cloned, characterised and mapped on the centromeric region of chromosome 8. Further, functional analysis were done. By transient transfection assays, I demonstrated that HMG2 is very similar to HMG1 in the transcriptional activation mediated by HOX proteins. Using the same assay, I demonstrated that differences in the length of the acidic tail of HMG1 and HMG2 determine no functional differences. Moreover, I showed that HMG2, as HMG1, is not stably associated to chromatin. These data, together with previously published data, suggest that HMG2 and HMG1 play very similar functions in the cell. Nevertheless, a difference between the two proteins was found in their expression pattern. HMG2 is absent in adult liver and brain, while is very abundant in adult testis, spleen, and thymus. On the contrary, HMG1 is ubiquitously expressed. In situ hybridisation revealed that in the testis HMG2 has a specific distribution: it is absent in spermatogonia and in late spermatids and spermatozoa, while is very abundant in spermatocytes and round spermatids. The peculiar protein distribution during spermatogenesis correlates with the phenotype we found in hmg2 -/- mice.hmg2 -/- mice had no obvious phenotypic differences from wild type. No immunological, homeotic, and chromatin defects were found. However, -/- male mice are partially sterile. After six months of continuous breeding studies some -/- mice gave no litters, some others gave litters of reduced size while others gave normal litters. Histological analysis of the testis in completely sterile mice revealed the presence of a significant number of dysmorphogenic seminiferous tubules, similar to that described in knock-out mice for estrogen receptor and in mice overexpressing ABP (Androgen Binding Protein). Degenerated tubules were also found in fertile -/- mice, but the number was drastically reduced. Moreover, electron microscope analysis revealed severe defects in elongating spermatids. Spermatozoa are produced, but are mostly immobile. Nevertheless, they are able to fertilise eggs. The testes of many knock-outs were found in the inguinal ring instead of in the scrotum. The correct position of testes is regulated by androgens. The correct function of androgen receptor was tested in mouse embryonic fibroblasts, but no large difference in the response to testosterone was found in this kind of cells. Thus, HMG1 and HMG2 play a very similar role in the cell and it is highly probable that they can substitute each other to a large extent. HMG1 expression is high and ubiquitous, whereas HMG2 expression is limited to a subset of tissues and organs. In the testes, the absence of HMG2 leads to a reduction in the number of viable spermatozoa, suggesting that during male meiosis a high level ofHMGs is required

Typography and dyslexia: A preliminary study on university students

Specific fonts for people with dyslexia are designed under the assumption that text readability can benefit from decreased letter confusability. We asked university students (18-33 yr), either diagnosed with developmental dyslexia (DD= 8) or normal (N= 31) to perform several tasks on materials printed in 7 different fonts. The subjective ranking of fonts was not a strong predictor of individual performance in objective task

Ozone Therapy for Early Childhood Caries (ECC) Treatment: An In Vivo Prospective Study

The present study aimed to evaluate the effectiveness of ozone therapy in (1) increasing the compactness of decayed dentin (main aim), (2) reducing dentin hypersensitivity, (3) decreasing the salivary bacterial count, and (4) improving the quality of life in ECC-affected children during a 3-month follow-up. A total of 20 subjects uncooperative with conventional restorative treatment were treated with a 60 s ozone application/week (OzoneDTA(R)) for 4 weeks. Patients were evaluated at T0 = baseline; T1 = after ozone cycle and T2, T3, T4 = 1, 2, 3-month follow-up, respectively. Dentin compactness was evaluated according to the Affected Dentine Scale (ADS) proposed by the authors; dentin hypersensitivity was recorded with the Wong-Baker Faces Pain Rating Scale (WBFPRS); salivary bacteria were measured with the Saliva Check Mutans Test and quality of life with the Early Childhood Oral Health Impact Scale (ECOHIS). ADS values and salivary bacteria count results were analyzed using non-parametric tests. Statistically significant differences were detected for both parameters between T0 and T1 (p < 0.01). At T1, dentin hypersensitivity was resolved and quality of life improved. Ozone therapy is a simple and painless approach, which might be useful for controlling caries' progression in uncooperative children until appropriate cooperation is achieved

Social distancing measures for COVID-19 are changing winter season

Health authorities worldwide have adopted measures of social distancing and movement restrictions, in addition to other public health measures to reduce exposure and to suppress interhuman SARS-CoV- 2 transmission. In Italy, a national lockdown with school closure was introduced from March to May 2020. From November 2020, Italy has been divided into zones according to regional epidemiological data, with primary schools reopened, associated with the mandatory use of face masks and different levels of social distance measures. For children with symptoms suggestive of COVID-19, the surveillance mechanism for the control of SARS-CoV- 2 infection is based on the performance of a real-time PCR on a nasopharyngeal swab. A diagnostic test has been introduced at the tertiary-level university hospital, Institute for Maternal and Child Health, IRCCS \u201cBurlo Garofolo\u201d of Trieste, consisting of a multiple nucleic acid amplification assay for 13 common viral respiratory pathogens on nasopharyngeal swab (Respiratory Flow Chip assay (Vitro, Sevilla, Spain), including SARS-CoV- 2, influenza A and B, adenovirus, other coronaviruses, parainfluenza virus 1\u20134, enteroviruses, bocavirus, metapneumovirus, respiratory syncytial virus (RSV), rhinoviruses, Bordetella pertussis, Bordetella parapertussis and Mycoplasma pneumoniae. Before routine utilisation, international standard quality control samples for each pathogen were used for test validation, and no cross-detection was found between the different pathogens. Criteria for testing referral did not change during the study period. Weekly variability of the number of total tests performed was due to the normal variations of acute illness. During the last winter season, from September 2020 (week 39) to February 2021 (week 7), 1138 nasopharyngeal swabs were tested for patients younger than 17 years old (figure 1). No influenza A or B nor RSV was detected during this period. The most common pathogen was rhinovirus (n=505), followed by adenoviruses (n=131), other coronaviruses (n=101) and SARS-CoV- 2 (n=57). Our data show that common winter pathogens circulation changed, and influenza virus and RSV did not produce a seasonal epidemic in the 2020\u20132021 winter season. These data suggest that social distancing measures and mask wearing profoundly changed the seasonality of winter paediatric respiratory infections that are mainly spread by respiratory droplets. The reasons why rhinovirus remains the main pathogen despite social distancing and face mask use are still a matter of debate. Similar data showing a decrease of common viral respiratory infections during the winter season have recently been reported in the southern hemisphere.1\u20134 Our data refer to a single institute, covering paediatric population of the Trieste Province (about 230 000 inhabitants), limiting the generalisation of our findings. However, our results highlight the need for continuing surveillance for the delayed spread of such viruses during spring and summer

Essential Infantile Esotropia: A Course of Treatment From Our Experience

Background: Essential infantile esotropia (EIE) is the most common type of childhood esotropia. Although its classical approach is surgical, less invasive techniques have been proposed as an adjunct or alternative to traditional surgery. Among them, chemodenervation with botulinum toxin (BT) has been investigated, showing variable and sometimes conflicting results.& nbsp;Objectives: To compare the outcomes of bilateral BT injection and traditional surgery in a pediatric population with EIE in order to optimize and standardize the therapeutic approach. Other purposes are to evaluate whether early intervention may prevent the onset of vertical ocular deviation (which is part of the clinical picture of EIE) and/or influence the development of fine stereopsis, and also to assess changes in refractive status over time among the enrolled population.& nbsp;Methods: A retrospective consecutive cohort study was conducted in 86 children aged 0-48 months who underwent correction of EIE. The primary intervention in naive subjects was either bilateral BT injection (36 subjects, "BT group ") or strabismus surgery (50 subjects, "surgery group ").& nbsp;Results: Overall, BT chemodenervation (one or two injections) was effective in 13 (36.1%) subjects. With regard to residual deviation angle, the outcomes at least 5 years after the last intervention were overlapping in children receiving initial treatment with either injection or surgery; however, the success rate of primary intervention in the surgery group was higher, and the average number of interventions necessary to achieve orthotropia was smaller. Both early treatment with chemodenervation and surgery at a later age were not found to prevent the onset of vertical ocular deviation, whereas, surprisingly, the percentage of subjects developing fine stereopsis was higher in the surgery group. Finally, with regard to the change in refractive status over time, most of the subjects increased their initial hyperopia, whereas 10% became myopic.& nbsp;Conclusions: Our data suggest that a single bilateral BT injection by age 2 years should be considered as the first-line treatment of EIE without vertical component; whereas, traditional surgery should be considered as the first-line treatment for all other cases and in subjects unresponsive to primary single BT injection

Cortisol-induced SRSF3 expression promotes GR splicing, RACK1 expression and breast cancer cells migration

Recent data have demonstrated that triple negative breast cancer (TNBC) with high glucocorticoid receptor (GR) expression are associated to therapy resistance and increased mortality. Given that GR alternative splicing generates mainly GRα, responsible of glucocorticoids action, we investigated its role in the regulation of RACK1 (Receptor for Activated C Kinase 1), a scaffolding protein with a GRE (Glucocorticoid Response Element) site on its promoter and involved in breast cancer cells migration and invasion. We provide the first evidence that GRα transcriptionally regulates RACK1 by a mechanism connected to SRSF3 splicing factor, which promotes GRα, essential for RACK1 transcriptional regulation and consequently for cells migration. We also establish that this mechanism can be positively regulated by cortisol. Hence, our data elucidate RACK1 transcriptional regulation and demonstrate that SRSF3 involvement in cells migration implies its role in controlling different pathways thus highlighting that new players have to be considered in GR-positive TNBC