Less is more: Self-amplifying mRNA becomes self-killing upon dose escalation in immune-competent retinal cells.

In the last few years, mRNA therapeutics experienced a new wave of interest as therapy for retinal diseases. Nevertheless, despite the widespread use of mRNA vaccines in the COVID-19 pandemic, mRNA delivery to the eye is still in its infancy. Recently, our research group has demonstrated that after subretinal and intravitreal delivery of modified mRNA, the number of transfected retinal cells and protein expression per cell remains limited.

Every nano-step counts: a critical reflection on do's and don'ts in researching nanomedicines for retinal gene therapy.

Introduction: Retinal disease affects millions of people worldwide, generating a massive social and economic burden. Current clinical trials for retinal diseases are dominated by gene augmentation therapies delivered with recombinant viruses as key players. As an alternative, nanoparticles hold great promise for the delivery of nucleic acid therapeutics as well.

ICG-mediated photodisruption of the inner limiting membrane enhances retinal drug delivery.

Many groundbreaking therapies for the treatment of blindness require delivery of biologics or cells to the inner retina by intravitreal injection. Unfortunately, the advancement of these therapies is greatly hampered by delivery difficulties where obstruction of the therapeutics at the inner limiting membrane (ILM) represents the dominant bottleneck. In this proof-of-principle study, we explore an innovative light-based approach to locally ablate the ILM in a minimally invasive and highly controlled manner, thus making the ILM more permeable for therapeutics.

Laser-induced nanobubbles safely ablate vitreous opacities in vivo.

In myopia, diabetes and ageing, fibrous vitreous liquefaction and degeneration is associated with the formation of opacities inside the vitreous body that cast shadows on the retina, appearing as 'floaters' to the patient. Vitreous opacities degrade contrast sensitivity function and can cause notable impairment in vision-related quality of life. Here we introduce 'nanobubble ablation' for safe destruction of vitreous opacities.

Vaccinia Virus Protein B18R: Influence on mRNA Immunogenicity and Translation upon Non-Viral Delivery in Different Ocular Cell Types.

In the last few years, interest has grown in the use of nucleic acids as an ocular therapy for retinal genetic diseases. Recently, our research group has demonstrated that mRNA delivery could result in effective protein expression in ocular cells following subretinal injection. Yet, although mRNA therapy comes with many advantages, its immunogenicity resulting in hampered mRNA translation delays development to the clinic.

Ocular barriers to retinal delivery of intravitreal liposomes: Impact of vitreoretinal interface.

Drug delivery to the posterior segment of the eye is challenging due to several anatomical and physiological barriers. Thus, there is a need for prolonged action and targeted drug delivery to treat retinal diseases. Intravitreal injections avoid anterior eye barriers, but the vitreoretinal interface and inner limiting membrane (ILM) may prevent access of drug delivery systems to the retina.

Influence of pathogenic stimuli on Müller cell transfection by lipoplexes.

Neuroprotection is a mutation-independent therapeutic strategy that seeks to enhance the survival of neuronal cell types through delivery of neuroprotective factors. The Müller cell, a retinal glial cell type appreciated for its unique morphology and neuroprotective functions, could be regarded as an ideal target for this strategy by functioning as a secretion platform within the retina following uptake of a transgene of our choice. In this in vitro study we aimed to investigate the capability of Müller cells to take up a standard liposomal vector (i.

Photoablation of Human Vitreous Opacities by Light-Induced Vapor Nanobubbles.

Myopia, diabetes, and aging are the main causes of progressive vitreous collagen aggregation, resulting in vitreous opacities, which can significantly disturb vision. As vitreous opacities, which induce the visual phenomenon of "floaters", are accessible with nanomaterials and light, we propose a nanotechnology-based approach to locally ablate them with highly reduced light energy compared to the more traditional YAG laser therapy. Our strategy relies on the plasmon properties of gold nanoparticles that generate vapor nanobubbles upon pulsed-laser illumination whose mechanical force can ablate vitreous opacities.

Non-viral delivery of chemically modified mRNA to the retina: Subretinal versus intravitreal administration.

mRNA therapeutics have recently experienced a new wave of interest, mainly due to the discovery that chemical modifications to mRNA's molecular structure could drastically reduce its inherent immunogenicity and perceived instability. On this basis, we aimed to explore the potential of chemically stabilized mRNA for ocular applications. More specifically, we investigated the behavior of mRNA-loaded lipid-based carriers in human retinal cells (in vitro), in bovine retinal explants (ex vivo) and in mouse retinas (in vivo).

The obstacle course to the inner retina: Hyaluronic acid-coated lipoplexes cross the vitreous but fail to overcome the inner limiting membrane.

Considerable research over the last few years has revealed dysregulation of growth factors in various retinal diseases, such as glaucoma, diabetic retinopathy and photoreceptor degenerations. The use of messengerRNA (mRNA) to transiently overexpress a specific factor could compensate for this imbalance. However, a critical challenge of this approach lies in the ability to efficiently deliver mRNA molecules to the retinal target cells.

Müller cells as a target for retinal therapy.

Müller cells are specialized glial cells that span the entire retina from the vitreous cavity to the subretinal space. Their functional diversity and unique radial morphology render them particularly interesting targets for new therapeutic approaches. In this review, we reflect on various possibilities for selective Müller cell targeting and describe how some of their cellular mechanisms can be used for retinal neuroprotection.

Morphology and Composition of the Inner Limiting Membrane: Species-Specific Variations and Relevance toward Drug Delivery Research.

The inner limiting membrane (ILM) represents the structural boundary between the vitreous and the retina, and is suggested to act as a barrier for a wide range of retinal therapies. While it is widely acknowledged that the morphology of the human ILM exhibits regional variations and undergoes age-related changes, insight into its structure in laboratory animals is very limited. Besides presenting a detailed overview of the morphology and composition of the human ILM, this review specifically reflects on the species-specific differences in ILM structure.

In vitro and ex vivo models to study drug delivery barriers in the posterior segment of the eye.

Many ocular disorders leading to blindness could benefit from efficient delivery of therapeutics to the retina. However, despite extensive research into drug delivery vehicles and administration techniques, efficacy remains limited because of the many static and dynamic barriers present in the eye. Comprehension of the various barriers and especially how to overcome them can improve our ability to estimate the potential of existent drug delivery vectors and support the design of new ones.

Toward smart design of retinal drug carriers: a novel bovine retinal explant model to study the barrier role of the vitreoretinal interface.

Retinal gene delivery via intravitreal injection is hampered by various physiological barriers present in the eye of which the vitreoretinal (VR) interface represents the most serious hurdle. In this study, we present a retinal explant model especially designed to study the role of this interface as a barrier for the penetration of vectors into the retina. In contrast to all existing explant models, the developed model is bovine-derived and more importantly, keeps the vitreous attached to the retina at all times to guarantee an intact VR interface.

Ovarian tissue cryopreservation in female-to-male transgender people: insights into ovarian histology and physiology after prolonged androgen treatment.

Female-to-male transgender people (trans men) are faced with the risk of losing their reproductive potential owing to gender-affirming hormone treatment and genital reconstructive surgery. This observational, prospective cohort study investigates the effect of prolonged androgen therapy on their ovarian histology and fertility preservation perspectives. Hormone serum levels, ovarian histology and cumulus-oocyte complexes (COC) of 40 trans men were analysed at the moment of hysterectomy with bilateral oophorectomy in the context of genital reconstructive surgery after testosterone treatment (58.

Effect of hyaluronic acid-binding to lipoplexes on intravitreal drug delivery for retinal gene therapy.

Intravitreal administration of nanomedicines could be valuable for retinal gene therapy, if their mobility in the vitreous and therapeutic efficacy in the target cells can be guaranteed. Hyaluronic acid (HA) as an electrostatic coating of polymeric gene nanomedicines has proven to be beneficial on both accounts. While electrostatic coating provides an easy way of coating cationic nanoparticles, the stability of electrostatic complexes in vivo is uncertain.

Coating of Quantum Dots strongly defines their effect on lysosomal health and autophagy.

Unlabelled: In the last decade the interest in autophagy got an incredible boost and the phenomenon quickly turned into an extensive research field. Interestingly, dysfunction of this cytoplasmic clearance system has been proposed to lie at the root of multiple diseases including cancer. We therefore consider it crucial from a toxicological point of view to investigate if nanomaterials that are developed for biomedical applications interfere with this cellular process.

Comparison of gold nanoparticle mediated photoporation: vapor nanobubbles outperform direct heating for delivering macromolecules in live cells.

There is a great interest in delivering macromolecular agents into living cells for therapeutic purposes, such as siRNA for gene silencing. Although substantial effort has gone into designing nonviral nanocarriers for delivering macromolecules into cells, translocation of the therapeutic molecules from the endosomes after endocytosis into the cytoplasm remains a major bottleneck. Laser-induced photoporation, especially in combination with gold nanoparticles, is an alternative physical method that is receiving increasing attention for delivering macromolecules in cells.

Assessing nanoparticle toxicity in cell-based assays: influence of cell culture parameters and optimized models for bridging the in vitro-in vivo gap.

The number of newly engineered nanomaterials is vastly increasing along with their applications. Despite the fact that there is a lot of interest and effort is being put into the development of nano-based biomedical applications, the level of translational clinical output remains limited due to uncertainty in the toxicological profiles of the nanoparticles (NPs). As NPs used in biomedicines are likely to directly interact with cells and biomolecules, it is imperative to rule out any adverse effect before they can be safely applied.