Drug-Loaded Polymeric Particulated Systems for Ophthalmic Drugs Release.

Drug delivery to the anterior or posterior segments of the eye is a major challenge due to the protection barriers and removal mechanisms associated with the unique anatomical and physiological nature of the ocular system. The paper presents the preparation and characterization of drug-loaded polymeric particulated systems based on pre-emulsion coated with biodegradable polymers. Low molecular weight biopolymers (chitosan, sodium hyaluronate and heparin sodium) were selected due to their ability to attach polymer chains to the surface of the growing system. The particulated systems with dimensions of 190-270 nm and a zeta potential varying from -37 mV to +24 mV depending on the biopolymer charges have been obtained. Current studies show that particles release drugs (dexamethasone/pilocarpine/bevacizumab) in a safe and effective manner, maintaining therapeutic concentration for a longer period of time. An extensive modeling study was performed in order to evaluate the drug release profile from the prepared systems. In a multifractal paradigm of motion, nonlinear behaviors of a drug delivery system are analyzed in the fractal theory of motion, in order to correlate the drug structure with polymer. Then, the functionality of a SL(2R) type "hidden symmetry" implies, through a Riccati type gauge, different "synchronization modes" (period doubling, damped oscillations, quasi-periodicity and intermittency) during the drug release process. Among these, a special mode of Kink type, better reflects the empirical data. The fractal study indicated more complex interactions between the angiogenesis inhibitor Bevacizumab and polymeric structure.

Apoptosis of pro-B lymphocytes induced by NR4A1 activation in the presence of gingival fibroblast exosomes and TNFα, caspase 8, STAT3, and Akt pathways modulators.

There is a lack of data in the mainstream literature regarding the interactions between gingival fibroblasts, as a component of the local niche, and tumor precursors of B-lymphocytes. Although it is known that the development of tumors and tumor precursors depends on the local environment's characteristics. In order to experimentally evaluate the apoptosis of pro-B type lymphocytes, induced as a result of the known activation of orphan nuclear receptor 4A1 (NR4A1), through Cytosporone B (Csn-B, 10 µM), in the presence or absence of exosomes derived from gingival fibroblasts, we administered as a treatment: 1 µM R-7050 [functional inhibitor of tumor necrosis factor alpha (TNFα)], 1 µM Z-IETD-FMK (functional inhibitor of caspase 8), 1 µM GSK690693 (functional inhibitor of Akt 1∕2∕3 pathways) and, last but not least, 1 µM scutellarin [functional inhibitor of receptor activator of nuclear factor-kappa B ligand (RANKL)] and therefore of the signal transducer and activator of transcription 3 (STAT3) pathway. Firstly, it is really clear that the presence of exosomes in the pro-B lymphocytes culture medium amplified the apoptotic effects of 10 µM Csn-B. The inhibition of tumoral precursors development, namely the pro-B type, might be highly dependent on the inhibition of Akt 1∕2∕3 pathways, the first and most important consequence being apoptosis induced by the activation of NR4A1 orphan nuclear receptors.

Current diagnosis and management strategies in pachychoroid spectrum of diseases (Review).

This review presents a systematic analysis of the literature regarding the pachychoroid disease spectrum nomenclature, diagnosis criteria and therapeutic options. Pachychoroid related diseases are a new concept, introduced into the ophthalmological nomenclature in 2013 and evolving both as concept and as classification since then. The six disorders included in this phenotype have some common characteristics (thick choroid, pachyvessels, attenuation of the choriocapillaris), but also show individual features. The classification of the pachychoroid spectrum was revised many times, with the recent addition of the focal choroidal excavation (FCE) and peripapillary pachychoroid syndrome (PPS). As the terminology is developing, so is the number of case reports and case series from the initial report in 2013 to 57 reports in 2019. This review takes into account both the current literature and the clinical experience of the authors, emphasizing the understanding of the pathogenesis and aiming to update the therapeutic options available.

Chitosan grafted-poly(ethylene glycol) methacrylate nanoparticles as carrier for controlled release of bevacizumab.

The aim of the present study is to obtain, for the first time, polymeric nanocarriers based on the chitosan grafted-poly(ethylene glycol) methacrylate derivative. The strategy involves the use of chitosan grafted-poly(ethylene glycol) methacrylate with high solubility in water, obtained via Michael addition, in order to prepare potentially non-toxic micro/nanoparticles (MNPs). By modifying chitosan, its solubility in aqueous media was improved. Micro/nanoparticles-based chitosan grafted-poly(ethylene glycol) methacrylate were obtained under mild condition, with good and controlled swelling properties in acetate buffer solution (ABS) and phosphate buffer solution (PBS). The technique selected for the preparation of the MNPs was a double crosslinking (ionic and covalent) process in reverse emulsion which provide the mechanical stability of the polymeric nanocarrier. The chitosan derivative and MNPs were thoroughly characterized by Fourier Transform Infrared Spectroscopy (FT-IR), Thermogravimetric Analysis (TGA), Differential Scanning Calorimetry (DSC), Scanning Electron Microscopy (SEM). The Scanning Electron Microscopy photographs revealed that prepared MNPs have different diameters depending on the used stirring rate and polymer concentration. Nanoparticles potential as drug delivery system was analyzed by loading bevacizumab (BEV) a full-length monoclonal antibody. Also, the prepared particles were found suitable from the cytotoxicity and hemocompatibility point of view enabling their potential use as delivery system for the treatment of posterior segment of the eye conditions.

Drug-induced gingival hyperplasia - experimental model.

Several causes of gingival hyperplasia are known, the most widely accepted being the drug-induced gingival augmentation, a side effect associated mainly with three classes of drugs: anticonvulsants (Phenytoin), immunosuppressants (Cyclosporine A), and various calcium channel blockers (Nifedipine, Verapamil, Diltiazem). We studied the effect of Cyclosporine A (CsA) and Nifedipine on gingival fibroblasts extracted from the rat gum. Gingival fibroblasts were isolated from 6-week-old male rats weighing 150-170 g, from gingival explants, and grown in a specific culture medium consisting of Dulbecco's Modified Eagle's Medium (DMEM) supplemented with antibiotic and 10% fetal bovine serum (FBS). The medium was also supplemented with CsA (1 µg÷mL) and Nifedipine (3 mM). We also used a control group that received no treatment. The cells were photographed at 7, 14 and 30 days, with a Nikon Eclipse TE300 phase contrast microscope. For cell viability evidence, we used immunofluorescence (flow cytometry) with a FACS (fluorescence-activated cell sorting) Calibur device and its software. We noticed that the proliferation of these cells increased with the period of drug administration, and the subsequent morphological changes that occurred were related to the presence of drug accumulations in the cell cytoplasm. Modern molecular techniques are needed to shed some light upon the pathogenesis of drug induced gingival overgrowth and, thereby, provide novel information for preventative and effective future therapeutic strategies.

Interactions between apelin and angiotensin II on rat portal vein.

Apelin (AP), an endogenous ligand of the APJ receptor, is involved in the regulation of cardiovascular homeostasis. Regardless the multiple similarities between AP and angiotensin II (Ang II), their roles seem to be different. We studied both the interactions between Apelin 13 (AP13) and Ang II and to what extent, if any, nitric oxide (NO) is involved. The experiments were performed in endothelium-denuded or endothelium-intact rat portal vein in the presence of 10 microM N(G)-nitro L-arginine methyl ester or 10 microM aminoguanidine. AP13 did not modify the isolated rat portal vein tone by itself, but inhibited the Ang II-induced contractions acting mainly by a NO-dependent mechanism. Our results sustain the hypothesis that AP13 could increase the activity of both constitutive and inducible NO synthase on either endothelium intact or endothelium denuded rat portal vein rings.

Are multiple angiotensin receptor types involved in angiotensin (1-7) actions on isolated rat portal vein.

Angiotensin (1-7) [Ang (1-7)] is a bioactive component of the renin angiotensin system. Ang (1-7) may interact with angiotensin type 1 (AT1) or type 2 (AT2) receptors and with Ang (1-7) - specific receptors. We examined the interactions between different doses of Ang (1-7) (1 nM-1 microM) and angiotensin II (Ang II) (10 and 100 nM) on isolated rat portal vein. In endothelium-denuded portal vein rings, Ang (1-7) inhibited contractile effects induced by Ang II. The effects of Ang (1-7) were modified by indomethacin, N(G)-nitro-L-arginine methyl ester (L-NAME), (D-Ala7)-Angiotensin (1-7) (H-2888) and losartan. Our results suggest that on rat isolated portal vein rings without endothelium, Ang (1-7) reduces Ang II-induced contractions by acting mostly on Ang (1-7) specific receptors, and this effect is mediated by vasodilatatory prostaglandins. At high concentrations, Ang (1-7) effects are mediated by AT1-receptors, though to a lesser extent than by Ang (1-7) specific receptors.

Formulation and evaluation of cefuroxim loaded submicron particles for ophthalmic delivery.

Chitosan gelatin particles could be the ideal candidate for intraocular drug delivery due to their desirable properties. Double crosslinking in double emulsion has been used as an original and reliable method for particles preparation and their morphology has been optimized considering the main synthesis parameters such as polymers ratio, crosslinker amount, stirring speed, tensioactive amount and ionic crosslinking time, respectively. The particles have been analyzed for their physical-chemical properties (swelling degree, drug loading and release capacity, surface characteristics, etc.), the enzymatic degradation properties along with in vivo ocular investigations (ocular biodistribution, in vivo drug release). In the present study cefuroxim was used as a model drug, which is generally used in the prophylaxis of postoperative endophthalmitis following cataract surgery after intraocular administration. The present study proved that the dimensions and the physical-chemical properties of the particles can be modulated (by varying the preparation parameters) to facilitate the administration, the biodistribution and the drug release in the specific segment of the eye. This experimental study demonstrated also the ability of fluorescent nanoparticles to penetrate ocular tissues close to the administration site (intravitreal injection) and especially their tendency to migrate deep in the retina at time intervals of 72 h.

Genetic polymorphisms of TNFA and IL-1A and generalized aggressive periodontitis.

Virulent bacteria could cause gingival fibroblasts apoptosis through lipopolysaccharide release during generalized aggressive periodontitis (GAgP) development and evolution. We showed that treatment with lipopolysaccharide (LPS, 1 µg/mL) for 30 days induced the decrease in the number of cultured rat gingival fibroblasts as compared to control group, which received no treatment. GAgP is considered to have also a genetic etiology, so the aim of our study was to evaluate if some polymorphisms of tumor necrosis factor-alpha (TNFA) and interleukin 1A (IL-1A) genes are associated with GAgP in a sample of Romanian population. We selected a group of 32 subjects (22 cases and 10 controls) for studying the TNFA (-857) polymorphism and 97 subjects (66 cases and 31 controls) for IL-1A (-889) polymorphism. The single nucleotide polymorphisms were genotyped by real-time polymerase chain reaction for all subjects. The genotype and allelic distribution tended to be equally between the cases and the controls group. Similar results were obtained for the dominant and recessive model. The difference between the two groups did not reach statistic significance for neither of the two studied polymorphisms [p=0.76 for TNFA (-857) and p=0.84 for IL-1A (-889)]. The data suggest that TNFA (-857) C/T and IL-1A (-889) C/T polymorphisms are not associated with susceptibility to GAgP in this Romanian population, potentially because of the small sample size. This is the first such study for Romanian northeastern population.

Intraocular biodistribution of intravitreal injected chitosan/gelatin nanoparticles.

The aim of this study was to evaluate intraocular biodistribution of a fluorescent polymeric nanosystem composed of chitosan and gelatin after intravitreal administration in rat eyes. The nanoparticles based on chitosan and gelatin were synthesized using a reverse emulsion-double cross-linking technique (ionic and covalent) and their structural characteristics are presented. Two units of 1% suspension of fluorescein-labeled nanoparticles in saline solution were injected intravitreal in rat eyes. The histological cross-sections obtained at 24 and 72 hours were analyzed by confocal microscopy and compared to a similar number of control cross-sections. The scanning electron microscopy of the nanoparticles obtained by double cross-linking in reverse emulsion technique revealed spherical, smooth, highly porous particles with no tendency to form aggregates. The chitosan-fluorescein conjugate was present in all the ocular tissues both at 24 and at 72 hours. The nanoparticles were present in the retina in a larger quantity and persisted longer than in the other ocular tissues. They were mainly fixed paravascular. The double cross-linking in reverse emulsion technique was efficient in synthesizing a biocompatible polymeric nanosystem. The in vivo study of intraocular biodistribution of fluorescein-labeled nanoparticles revealed their affinity for the retina after intravitreal administration.

Double crosslinked interpenetrated network in nanoparticle form for drug targeting--preparation, characterization and biodistribution studies.

The development of polymer nanosystems able to target and control/sustain the drug delivery is still considered an important desideratum in pharmaceutical research. The present study reports the preparation of nanoparticles based on chitosan and gelatin, using a reverse emulsion-double crosslinking (ionic followed by covalent one) technique. The nanoparticles structural and morphological characteristics (diameter and size distribution), their swelling capacity in aqueous media of different pH (4 and 7.4) and their ability to include and release poorly water-soluble drugs were seen to be influenced by the composition of the polymer mixture and by the surfactants concentration. Also, nanoparticles biodistribution after intraperitoneal or intravenous administration was evaluated by polymer marking with fluorescein. Particles ability to penetrate different organs (liver, heart, lungs, and less brain, gums, testicles) was increased when injected intravenously.

[Involvement of angiotensin on adenosine-induced bronchial hyperreactivity]. / Implicarea angiotensinei in modularea hiperreactivitatii bronsice determinate de catre adenozina.

Adenosine is an endogenously purine nucleoside which plays a significant role in regulation of airways tone and reactivity by multiple and incompletely known mechanisms, including the release of endogenously active peptides from mast cells via activation of the A3 receptors. Our previous results suggested that releasing of enzymes from activated mast cells could activate the intrapulmonary renin angiotensin system (RAS). In this study, we investigated the involvement of angiotensin II (Ang II) in adenosine-induced bronchoconstriction in an experimental model of allergic asthma. On bronchial rings from ovalbumin (OVA) sensitized rats, after in vitro challenge, adenosine induced small contractile effects which became significant after indomethacin pre-treatment. On the other hand adenosine pre-treatment amplified bronchoconstriction induced by the allergen (OVA) challenge and reduced bronchial relaxation of acetylcholine pre-contracted bronchial rings by cumulative doses of terbutaline. All these effects are significantly lower on rats treated with losartan (a blocker of Ang II type 1 specific receptors, AT1) in the last two weeks of sensitization protocol (50 mg/kg/day). Our data confirmed that adenosine induced bronchial hyperreactivity could be partially a result of RAS activation in abnormal conditions as antigen sensitization and challenge.

Janus kinases affect thrombopoietin receptor cell surface localization and stability.

The thrombopoietin receptor (TpoR) regulates hematopoietic stem cell renewal, megakaryocyte differentiation, and platelet formation. TpoR signals by activating Janus kinases JAK2 and Tyk2. Here we show that, in addition to signaling downstream from the activated TpoR, JAK2 and Tyk2 strongly promote cell surface localization and enhance total protein levels of the TpoR. This effect is caused by stabilization of the mature endoglycosidase H-resistant form of the receptor. Confocal microscopy indicates that TpoR colocalizes partially with recycling transferrin in Ba/F3 cells. The interaction with JAK2 or Tyk2 appears to protect the receptor from proteasome degradation. Sequences encompassing Box1 and Box2 regions of the receptor cytosolic domain and an intact JAK2 or Tyk2 FERM domain are required for these effects. We discuss the relevance of our results to the reported defects of TpoR processing in myeloproliferative diseases and to the mechanisms of Tpo signaling and clearance via the TpoR.