Customized Corneal Cross-Linking-A Mathematical Model

Purpose: To improve the safety, reproducibility, and depth of effect of corneal cross-linking with the ultraviolet A (UV-A) exposure time and fluence customized according to the corneal thickness. Methods: Twelve human corneas were used for the experimental protocol. They were soaked using a transepithelial (EPI-ON) technique using riboflavin with the permeation enhancer vitamin E-tocopheryl polyethylene glycol succinate. The corneas were then placed on microscope slides and irradiated at 3 mW/cm2 for 30 minutes. The UV-A output parameters were measured to build a new equation describing the time-dependent loss of endothelial protection induced by riboflavin during cross-linking, as well as a pachymetry-dependent and exposure time-dependent prescription for input UV-A fluence. The proposed equation was used to establish graphs prescribing the maximum UV-A fluence input versus exposure time that always maintains corneal endothelium exposure below toxicity limits. Results: Analysis modifying the Lambert-Beer law for riboflavin oxidation leads to graphs of the maximum safe level of UV-A radiation fluence versus the time applied and thickness of the treated cornea. These graphs prescribe UV-A fluence levels below 1.8 mW/cm2 for corneas of thickness 540 [mu]m down to 1.2 mW/cm2 for corneas of thickness 350 [mu]m. Irradiation times are typically below 15 minutes. Conclusions: The experimental and mathematical analyses establish the basis for graphs that prescribe maximum safe fluence and UV-A exposure time for corneas of different thicknesses. Because this clinically tested protocol specifies a corneal surface clear of shielding riboflavin on the corneal surface during UV-A irradiation, it allows for shorter UV-A irradiation time and lower fluence than in the Dresden protocol

Citrus sinensis and Vitis vinifera Protect Cardiomyocytes from Doxorubicin-Induced Oxidative Stress: Evaluation of Onconutraceutical Potential of Vegetable Smoothies

The interest towards nutraceuticals able to counteract drug side effects is continuously growing in current chemotherapeutic protocols. In the present study, we demonstrated that smoothies containing mixtures of Citrus sinensis and Vitis vinifera L. cv. Aglianico N, two typical fruits of the Mediterranean diet, possess bioactive polyphenols that protect cardiomyocytes against doxorubicin-induced oxidative stress. The polyphenolic extracts isolated from Citrus sinensis- and Vitis vinifera-based functional smoothies were deeply characterized by Liquid Chromatography-Mass Spectrometry methods. Subsequently, the functional smoothies and relative mixtures were tested to verify their ability to affect cellular viability and oxidative stress parameters in embryonic cardiomyocyte cells (H9c2), and human breast adenocarcinoma cell line (MCF-7) exposed to doxorubicin. Interestingly, we found that the mix resulting from Citrus sinensis and Vitis vinifera association in ratio 1:1 was able to reduce cardiomyocytes damage induced by anthracyclines, without significantly interfering with the pro-apoptotic activity of the drug on breast cancer cells. These results point out the potential use of vegetable smoothies as adjuvants functional foods for chemotherapeutic anticancer protocols

N-4 Alkyl Cytosine Derivatives Synthesis: A New Approach

The selective N-4 alkylation of cytosine plays a critical role in the synthesis of biologically active molecules. This work focuses on the development of practical reaction conditions toward a regioselective synthesis of N-4-alkyl cytosine derivatives. The sequence includes a direct and selective sulfonylation at the N-1 site of the cytosine, followed by the alkylation of the amino siteusing KHMDS in CH2Cl2/THF mixture, providing a fast and efficient approach consistent withpyrimidine-based drug design

Detailed peptide profiling of “Scotta”: from a dairy waste to a source of potential health-promoting compound

“Scotta” is a liquid waste deriving from Ricotta cheese production, which is wrongly considered only a dairy by-product. In this work, with the aim to elucidate the presence of valuable bioactive compounds in Buffalo’s Scotta, a peptide fraction under 3000 Da was isolated by ultra-filtration, purified by solid-phase extraction, and,subsequently, characterized in detail by liquid chromatography coupled to Orbitrap mass spectrometry. Analytical results revealed a complex profile, leading to the identification of 226 peptides, belonging to alpha, beta, and kappa caseins. A database-driven search approach was used to assess the biological effects of some of the identified peptides. A wide range of healthy properties was ascribed to the encrypted peptides, comprising antihypertensive, antimicrobial, immunomodulating, opioid, antioxidant, and antithrombotic. The peptidomic profile of Scotta was highlighted in depth for the first time, and the results revealed that this matrix should not be considered only a mere by-product, but a source of potential health-promoting peptides, which can be recovered and employed in nutraceuticals and functional foods

Therapeutic potential of TRPM8 antagonists in prostate cancer.

Transient receptor potential melastatin-8 (TRPM8) represents an emerging target in prostate cancer, although its mechanism of action remains unclear. Here, we have characterized and investigated the effects of TRPM8 modulators in prostate cancer aggressiveness disclosing the molecular mechanism underlying their biological activity. Patch-clamp and calcium fluorometric assays were used to characterize the synthesized compounds. Androgen-stimulated prostate cancer-derived cells were challenged with the compounds and the DNA synthesis was investigated in a preliminary screening. The most effective compounds were then employed to inhibit the pro-metastatic behavior of in various PC-derived cells, at different degree of malignancy. The effect of the compounds was then assayed in prostate cancer cell-derived 3D model and the molecular targets of selected compounds were lastly identified using transcriptional and non-transcriptional reporter assays. TRPM8 antagonists inhibit the androgen-dependent prostate cancer cell proliferation, migration and invasiveness. They are highly effective in reverting the androgen-induced increase in prostate cancer cell spheroid size. The compounds also revert the proliferation of castrate-resistant prostate cancer cells, provided they express the androgen receptor. In contrast, no effects were recorded in prostate cancer cells devoid of the receptor. Selected antagonists interfere in non-genomic androgen action and abolish the androgen-induced androgen receptor/TRPM8 complex assembly as well as the increase in intracellular calcium levels in prostate cancer cells. Our results shed light in the processes controlling prostate cancer progression and make the transient receptor potential melastatin-8 as a ‘druggable’ target in the androgen receptor-expressing prostate cancers

A Novel Vasoactive Peptide “PG1” from Buffalo Ice-Cream Protects from Angiotensin-Evoked High Blood Pressure

Arterial hypertension is the most important risk factor for cardiovascular diseases, myocardial infarction, heart failure, renal failure and peripheral vascular disease. In the last decade, milk-derived bioactive peptides have attracted attention for their beneficial cardiovascular properties. Methods: Here, we combined in vitro chemical assay such as LC-MS/MS analysis of buffalo ice cream, ex vivo vascular studies evaluating endothelial and smooth muscle responses using pressure myograph, and translational assay testing in vivo the vascular actions of PG1 administration in murine models. Results: We demonstrate that a novel buffalo ice-cream-derived pentapeptide “QKEPM”, namely PG1, is a stable peptide that can be obtained at higher concentration after gastro-intestinal digestions (GID) of buffalo ice-cream (BIC). It owns potent vascular effect in counteract the effects of angiotensin II-evoked vasoconstriction and high blood pressure levels. Its effects are mediated by the inhibitory effect on AT1 receptor leading to a downregulation of p-ERK½/Rac1-GTP and consequent reduction of oxidative stress. Conclusions: These results strongly candidate PG1, as a novel bioactive peptide for the prevention and management of hypertension, thus expanding the armamentarium of preventive strategies aimed at reducing the incidence and progression of hypertension and its related cardiovascular complication

Cross-linking del collagene corneale e biomeccanica delle cornee

L'obiettivo principale del progetto consiste nell'ultilizzare il cross-linking del collagene corneale per un uovo approccio alle patologie che provocano una ridotta resitenza biomeccanica della cornea.Una varietà di condizioni primarie, quali cheratocono e degenerazione marginale pellucida, o secondarie, quali ectasia iatrogena e post-infettiva, infatti, determinano disturbi ectastici che portano ad una ridotta resistenza biomeccanica della cornea. Il cross-linking del collagene corneale rappresenta un approccio innovativo a tali patologie in grado di evitare, in molti casi, il ricorrere a procedure di trapianto corneale (cheratoplastica perforante o lamellare). La tecnica standard viene eseguita in sala operatoria e consiste nella rimozione dell'epitelio corneale, al fine di favorire la penetrazione della sostanza fotosensibile, seguita dall'irraggiamento mediante ultravioletti a specifiche lunghezze d'onda. Il trattamento proposto dal presente progetto di ricerca prevede, al contrario, la permeazione della sostanza fotosensibile attraverso l'epitelio corneale, evitando l'intervento chirugico e le complicazioni ad esso potenzialmente associate (infezioni, cheratiti, edema, cicatrici), in grado di comportare un ulteriore abbassamento del visus ed una scarsa compliance da parte del paziente.Tale obiettivo sarà perseguito attraverso la formulazione di medical devices atti a favorire una maggiore permeazione della sostanza fotosensibile in presenza di epitelio corneale intatto attraverso l'uso di enhancer di penetrazione e/o sistemi di rilascio micro e nanoparticellari. Allo stesso tempo, data la potenziale tossicità corneale dell'irraggiamento UV-A, il progetto prevede la creazione di un protocollo di cross-linking corneale personalizzata, identificando i parametri fondamentali nella riduzione della fototossicità (intesità della radiazione, spessore della cornea, tempo di irraggiamento) e creando una nuova apparecchiatura che, sulla base dell'incidenza dei suddetti parametri sia in grado di variare potenza, intensità e durata per garantire una maggiore sicurezza ed efficacia di trattamento al paziente

Corneal Cross-Linking: The Science Beyond the Myths and Misconceptions

PURPOSE: There has been a recent explosion in the variety of techniques used to accomplish corneal cross-linking (CXL) for the treatment of ectatic corneal diseases. To understand the success or failure of various techniques, we review the physicochemical basis of corneal CXL and re-evaluate the current principles and long-standing conventional wisdom in the light of recent, compelling, and sometimes contradictory research. METHODS: Two clinicians and a medicinal chemist developed a list of current key topics, controversies, and questions in the field of corneal CXL based on information from current literature, medical conferences, and discussions with international practitioners of CXL. RESULTS: Standard corneal CXL with removal of the corneal epithelium is a safe and efficacious procedure for the treatment of corneal ectasias. However, the necessity of epithelium removal is painful for patients, involves risk and requires significant recovery time. Attempts to move to transepithelial corneal CXL have been hindered by the lack of a coherent understanding of the physicochemistry of corneal CXL. Misconceptions about the applicability of the Bunsen-Roscoe law of reciprocity and the Lambert-Beer law in CXL hamper the ability to predict the effect of ultraviolet A energy during CXL. Improved understanding of CXL may also expand the treatment group for corneal ectasia to those with thinner corneas. Finally, it is essential to understand the role of oxygen in successful CXL. CONCLUSIONS: Improved understanding of the complex interactions of riboflavin, ultraviolet A energy and oxygen in corneal CXL may provide a successful route to transepithelial corneal CXL