Pediatric glaucoma: current perspectives

Giorgio Marchini, Marco Toscani, Francesca Chemello Eye Clinic, Department of Neurological and Movement Sciences, University of Verona, Verona, Italy Abstract: "Childhood glaucoma" is a heterogeneous group of severe pediatric conditions often associated with significant visual loss and characterized by elevated intraocular pressure (IOP) and optic-disk cupping. Successful IOP control is crucial but challenging and most often achieved surgically, with medical therapy playing a supportive role. There are many classifications of childhood glaucoma, but they can simply be divided into primary, in which a developmental abnormality of the anterior chamber angle only exists, and secondary, in which aqueous outflow is reduced due to independent mechanisms that secondarily impair the function of the filtration angle. The worldwide prevalence of childhood blindness ranges from 0.03% in high-income countries to 0.12% in undeveloped countries. The majority of cases do not have an identified genetic mutation and, where the mutation is known, the genes often account for only a small proportion of cases. Several pathogenetic mechanisms are known to contribute to the development of childhood glaucoma. Whatever the cause, it results in a reduced aqueous outflow at the level of the trabecular meshwork. Age of onset and magnitude of the elevated IOP largely determine the clinical manifestation the high variability of clinical manifestations. Glaucoma from any cause in a neonate and infant is characterized by the classic triad of epiphora, photophobia, and blepharospasm, and could be associated with eye enlargement (buphthalmos) and Haab striae. The eye examination, usually performed under general anesthesia, includes: tonometry, anterior-segment examination, gonioscopy, corneal diameter and axial length measurement, dilated fundoscopy with optic-nerve-head evaluation. Medical therapy, considering the high frequency of side effects, is generally used as temporizing IOP-lowering treatment before surgery or as adjuvant treatment postoperatively in case of partially successful procedures for refractory glaucomas. Surgery is the nodal point of the management of refractory childhood glaucoma, so it is crucial to opt for a forward-looking strategy to reach the target IOP and minimize the visual loss. Keywords: childhood glaucoma, congenital glaucoma, glaucoma surgical therapy, childhood visual los

Cholesterol dependent macropinocytosis and endosomal escape control the transfection efficiency of lipoplexes in CHO Living Cells

Here we investigate the cellular uptake mechanism and final intracellular fate of two cationic liposome formulations characterized by similar physicochemical properties but very different lipid composition and efficiency for intracellular delivery of DNA. The first formulation is made of cationic lipid 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) and the zwitterionic helper dioleoylphosphocholine (DOPC), while the second one is made of the cationic 3 beta-[N-(N,N-dimethylaminoethane)-carbamoyl] cholesterol (DC-Chol) and the zwitterionic lipid dioleoylphosphatidylethanolamine (DOPE). Combining pharmacological and imaging approaches we show that both DOTAP-DOPC/DNA and DC-Chol-DOPE/DNA lipoplexes are taken up in Chinese hamster ovary (CHO) living cells mainly through fluid-phase macropinocytosis. Our results also indicate that lipoplex macropinocytosis is a cholesterol-sensitive uptake mechanism. On the other side, both clathrin-mediated and caveolae-mediated endocytosis play a minor role, if any, in the cell uptake. Colocalization of fluorescently tagged lipoplexes and Lysosensor, a primary lysosome marker, reveals that poorly efficient DOTAP-DOPC/DNA lipoplexes are largely degraded in the lysosomes, while efficient DC-Chol-DOPE/DNA systems can efficiently escape from endosomal compartments

Short-term use of dexamethasone/netilmicin fixed combination in controlling ocular inflammation after uncomplicated cataract surgery

Purpose: To evaluate the short-term anti-inflammatory effect of dexamethasone/netilmicin fixed combination in the management of ocular inflammation after cataract surgery. Patients and Methods: Open-label, randomized, active-controlled, clinical study con-ducted in 6 sites in Italy; 238 patients were randomized 2:1 to dexamethasone/netilmicin (dexa/net, n=158) or betamethasone/chloramphenicol (beta/chl, n=80). Treatment started the day of surgery and continued 4 times daily for 7 days. The primary efficacy parameter was the anterior chamber (AC) flare. The percentage of patients displaying none or mild (ie, only barely detectable) AC flare was defined as “efficacy rate”, whereas the percentage of patients showing a decrease of AC flare score from baseline was defined as “percentage of responders”. Additional parameters evaluated were AC cells, conjunctival hyperaemia, corneal and lid oedema, symptoms of ocular discomfort, visual acuity, and intraocular pressure. Dexa/net was considered effective if the efficacy rate was not inferior (by means of 97.5% confidence interval) to that of beta/chl. Results: After 7 days of treatment, no AC flare was observed in 92.8% (dexa/net) and 92.3% (beta/chl) of patients, whereas no AC cells were observed in 91.5% (dexa/net) and 93.6% (beta/chl) of patients, respectively. The “efficacy rate” was 100% in both groups, whereas the “percentage of responders” was 94.1% in the dexa/net and 93.6% in the beta/chl group. The p-value to reject the null hypothesis of inferiority was <0.001. Other efficacy parameters confirmed both treatments as highly effective, despite their difference in steroid content (2 mg/mL for beta/chl vs 1 mg/mL for dexa/net). IOP and visual acuity at the end of the study were comparable. Two cases of allergic conjunctivitis were considered adverse events and were both related to dexa/net. Conclusion: Short-term use of dexa/net fixed combination is safe and effective in the control of post-operative inflammation following uncomplicated cataract surgery

Reproduction of the azooxanthellate coral Caryophyllia inornata is not affected by temperature along an 850 km gradient on the Western Italian coast

The Intergovernmental Panel on Climate Change (IPCC) predicted that ocean surface temperature will rise of 0.6–2.0◦C by 2100. Ocean warming is expected to produce strong impacts on marine ecosystems such as coral reefs, affecting their physiological events including reproductive processes. To date, relatively few studies have examined the effects of climate change on the reproductive success of temperate corals and even less in the azooxanthellate ones. This study examined the reproductive output of the azooxanthellate Mediterranean coral Caryophyllia inornata along a wide latitudinal gradient of seawater temperature and solar radiation. A total of 260 samples, collected from five populations along the Western Italian coast, have been analyzed through histological techniques. The intriguing aspects characterizing all populations of C. inornata along the latitudinal gradient are a strong male-biased sex ratio and the presence of embryos in all stages of development throughout the year in females, males, and sexually inactive individuals. This peculiarity could suggest a mixed strategy of sexual and asexual reproduction in this species as has been observed for some anemones of the genus Actinia. Fecundity and spermary abundance (i.e., the number of reproductive elements per body volume unit), gonadal index (i.e., the percentage of body volume occupied by the germ cells) and fertility (i.e., the number of embryos per body volume unit) in females, males and sexually inactive individuals were unrelated to solar radiation and temperature along the latitudinal gradient. These results suggest that the reproduction in C. inornata is not affected by increasing solar radiation and temperature. The lack of zooxanthellae could make this species less dependent on these environmental parameters, as previously hypothesized for another azooxanthellate species, Leptopsammia pruvoti, investigated along the same gradient

Surface area of lipid membranes regulates the DNA-binding capacity of cationic liposomes.

We have applied electrophoresis on agarose gels to investigate the DNA-binding capacity of cationic liposomes made of cationic DC-cholesterol and neutral dioleoylphosphatidylethanolamine as a function of membrane charge density and cationic lipid/DNA charge ratio. While each cationic liposome formulation exhibits a distinctive DNA-protection ability, here we show that such a capacity is universally regulated by surface area of lipid membranes available for binding in an aspecific manner. The relevance of DNA protection for gene transfection is also discussed

Biomimetic Electrospun Self-Assembling Peptide Scaffolds for Neural Stem Cell Transplantation in Neural Tissue Engineering

Spinal cord regeneration using stem cell transplantation is a promising strategy for regenerative therapy. Stem cells transplanted onto scaffolds that can mimic natural extracellular matrix (ECM) have the potential to significantly improve outcomes. In this study, we strived to develop a cell carrier by culturing neural stem cells (NSCs) onto electrospun 2D and 3D constructs made up of specific crosslinked functionalized self-assembling peptides (SAPs) featuring enhanced biomimetic and biomechanical properties. Morphology, architecture, and secondary structures of electrospun scaffolds in the solid-state and electrospinning solution were studied step by step. Morphological studies showed the benefit of mixed peptides and surfactants as additives to form thinner, uniform, and defect-free fibers. It has been observed that β-sheet conformation as evidence of self-assembling has been predominant throughout the process except for the electrospinning solution. In vitro NSCs seeded on electrospun SAP scaffolds in 2D and 3D conditions displayed desirable proliferation, viability, and differentiation in comparison to the gold standard. In vivo biocompatibility assay confirmed the permissibility of implanted fibrous channels by foreign body reaction. The results of this study demonstrated that fibrous 2D/3D electrospun SAP scaffolds, when shaped as micro-channels, can be suitable to support NSC transplantation for regeneration following spinal cord injury

Mitogen-activated protein kinase kinase 5 regulates proliferation and biosynthetic processes in procyclic forms of Trypanosoma brucei

The pathogenic protozoan T. brucei alternates into distinct developmental stages in the mammalian and insect hosts. The mitogen-activated protein kinase (MAPK) signaling pathways transduce extracellular stimuli into a range of cellular responses, which ultimately lead to the adaptation to the external environment. Here, we combined a loss of function approach with stable isotope labeling with amino acids in cell culture (SILAC)-based mass spectrometry (MS) to investigate the role of the mitogen-activated protein kinase kinase 5 (MKK5) in T. brucei. The silencing of MKK5 significantly decreased the proliferation of procyclic forms of T. brucei. To shed light on the molecular alterations associated with this phenotype, we measured the total proteome and phosphoproteome of cells silenced for MKK5. In the total proteome, we observed a general decrease in proteins related to ribosome and translation as well as down-regulation of several components of the fatty acids biosynthesis pathway. In addition, we observed alterations in the protein levels and phosphorylation of key metabolic enzymes, which point toward a suppression of the oxidative metabolism. Taken together, our findings show that the silencing of MKK5 alters cell growth, energy metabolism, protein and fatty acids biosynthesis in procyclic T. brucei