Retinal explant culture: A platform to investigate human neuro-retina

The retina is the tissue responsible for light detection, in which retinal neurons convert light energy into electrical signals to be transported towards the visual cortex. Damage of retinal neurons leads to neuronal cell death and retinal pathologies, compromising visual acuity and eventually leading to irreversible blindness. Models of retinal neurodegeneration include 2D systems like cell lines, disassociated cultures and co-cultures, and 3D models like organoids, organotypic retinal cultures and animal models. Of these, ex-vivo human retinal cultures are arguably the most suitable models for translational research as they retain complex inter-cellular interactions of the retina and precisely mimic in-situ responses. In this review, we summarise the distinguishing features of the human retina which are important to preserve in experimental culture, the historical development of human retinal culture systems, the factors affecting ex-vivo human retinal culture and the applications and challenges associated with current methods of human retinal explant culture

Gene therapy and the adeno-associated virus in the treatment of genetic and acquired ophthalmic diseases in humans: trials, future directions and safety considerations

Voretigene neparvovec‐rzyl was recently approved for the treatment of Leber Congenital Amaurosis, and the use of gene therapy for eye disease is attracting even greater interest. The eye has immune privileged status, is easily accessible, requires a reduced dosage of therapy due to its size, and is highly compartmentalized, significantly reducing systemic spread. Adeno‐associated virus (AAV), with its low pathogenicity, prolonged expression profile and ability to transduce multiple cell types, has become the leading gene therapy vector. Target diseases have moved beyond currently untreatable inherited dystrophies to common, partially treatable acquired conditions such as exudative AMD and glaucoma, but use of the technology in these conditions imposes added obligations for caution in vector design. This review discusses the current status of AAV gene therapy trials in genetic and acquired ocular diseases, and explores new scientific developments which could help ensure effective and safe use of the therapy in the future

Evaluation of the Glypican 3 promoter for transcriptional targeting of hepatocellular carcinoma

© 2018 Macmillan Publishers Ltd., part of Springer Nature. Hepatocellular carcinoma (HCC) is a major health problem as evidenced by its increasing incidence and high morbidity and mortality rates. Most patients with HCC have underlying liver disease and dysfunction which limits the current therapeutic options. Treatments that spare the liver and destroy the HCC are needed. Targeting transcriptional differences between HCC and liver cells may provide this therapeutic window. In this study, we examine the potential of the Glypican 3 (GPC3) promoter as a targeting strategy. GPC3 is an oncofetal protein belonging to the proteoglycan family which is normally only expressed during fetal development. However, in HCC, the expression of this protein is reactivated. Here, we show that GPC3 is expressed primarily in HCC and not in normal liver lines. We show that the GPC3 promoter can be used to drive expression of significantly more luciferase and eYFP in HCC cell lines compared to normal liver cells. Further, we show that vectors containing cytosine deaminase (CD) under GPC3 promotor control induced significantly more killing of HCC cell lines after treatment with 5-FC compared to normal liver cell lines. These data suggest that transcriptionally targeted delivery of transgene in HCC cells can be achieved using the GPC3 promoter and this targeting strategy produces limited toxicity to normal liver cells

Adeno-associated virus neutralising antibodies in type 1 diabetes mellitus

Recombinant Adeno-associated viruses (AAVs) are an attractive vector for gene therapy delivery which may be blocked by AAV neutralising antibodies (NAbs). As Type 1 Diabetes (T1DM) is an endocrine disease of immunological origin, it is likely that NAb profiles are altered in the disease. In this study NAb to AAV2, AAV5, AAV6, and AAV8 in 72 subjects with T1DM and 45 non-diabetic patients were measured over a 4-year follow-up period. AAV2 NAb titres were significantly lower in non-diabetic subjects (P = 0.036). The T1DM group had more AAV8 NAb activity at baseline (P = 0.019), whilst after 4 years follow-up the T1DM group displayed developed increased AAV 5 (P = 0.03), 6 (P = 0.03) and 8 (P = 0.002) activity relative to the control group, however, overall AAV5 and 8 NAb levels were very low in patients <40. AAV NAb titre activity and prevalence generally appears higher in T1DM, however, low levels of AAV 5 and 8, particular in younger adult age groups at which T1DM can be targeted, could make these attractive vectors to target the disease