Transplantation of Human Embryonic Stem Cell-Derived Retinal Pigment Epithelial Cells in Macular Degeneration

PURPOSE: Transplantation of human embryonic stem cell (hESC)-derived retinal pigment epithelial (RPE) cells offers the potential for benefit in macular degeneration. Previous trials have reported improved visual acuity (VA), but lacked detailed analysis of retinal structure and function in the treated area. DESIGN: Phase 1/2 open-label dose-escalation trial to evaluate safety and potential efficacy (clinicaltrials.gov identifier, NCT01469832). PARTICIPANTS: Twelve participants with advanced Stargardt disease (STGD1), the most common cause of macular degeneration in children and young adults. METHODS: Subretinal transplantation of up to 200 000 hESC-derived RPE cells with systemic immunosuppressive therapy for 13 weeks. MAIN OUTCOME MEASURES: The primary end points were the safety and tolerability of hESC-derived RPE cell administration. We also investigated evidence of the survival of transplanted cells and measured retinal structure and function using microperimetry and spectral-domain OCT. RESULTS: Focal areas of subretinal hyperpigmentation developed in all participants in a dose-dependent manner in the recipient retina and persisted after withdrawal of systemic immunosuppression. We found no evidence of uncontrolled proliferation or inflammatory responses. Borderline improvements in best-corrected VA in 4 participants either were unsustained or were matched by a similar improvement in the untreated contralateral eye. Microperimetry demonstrated no evidence of benefit at 12 months in the 12 participants. In one instance at the highest dose, localized retinal thinning and reduced sensitivity in the area of hyperpigmentation suggested the potential for harm. Participant-reported quality of life using the 25-item National Eye Institute Visual Function Questionnaire indicated no significant change. CONCLUSIONS: Subretinal hyperpigmentation is consistent with the survival of viable transplanted hESC-derived RPE cells, but may reflect released pigment in their absence. The findings demonstrate the value of detailed analysis of spatial correlation of retinal structure and function in determining with appropriate sensitivity the impact of cell transplantation and suggest that intervention in early stage of disease should be approached with caution. Given the slow rate of progressive degeneration at this advanced stage of disease, any protection against further deterioration may be evident only after a more extended period of observation

Assessment of AAV vector tropisms for mouse and human pluripotent stem cell-derived RPE and photoreceptor cells

Adeno-associated viral vectors are showing great promise as gene therapy vectors for a wide range of retinal disorders. To date, evaluation of therapeutic approaches has depended almost exclusively on the use of animal models. With recent advances in human stem cell technology, stem-cell derived retina now offers the possibility to assess efficacy in human organoids in vitro. Here we test 6 AAV serotypes (AAV2/2, AAV2/9, AAV2/8, AAV2/8T(Y733F), AAV2/5 and ShH10) to determine their efficiency in transducing mouse and human pluripotent stem cell (PSC)-derived RPE and photoreceptor cells in vitro. All the serotypes tested were capable of transducing RPE and photoreceptor cells in vitro. AAV ShH10 and AAV2/5 are the most efficient vectors at transducing both mouse and human RPE, while AAV2/8 and ShH10 achieved similarly robust transduction of human ESC-derived cone photoreceptors. Furthermore, we show that hESC-derived photoreceptors can be used to establish promoter specificity in human cells in vitro. The results of this study will aid capsid selection and vector design for pre-clinical evaluation of gene therapy approaches, such as gene editing, that require the use of human cells and tissues

VIDIIA Hunter: a low-cost, smartphone connected, artificial intelligence-assisted COVID-19 rapid diagnostic platform approved for medical use in the UK

Data availability statement: The original contributions presented in the study are included in the article/Supplementary Material, further inquiries can be directed to the corresponding authors.The Supplementary Material for this article can be found online at: https://www.frontiersin.org/articles/10.3389/fmolb.2023.1144001/full#supplementary-material SUPPLEMENTARY TABLE S1 | Full data set obtained from the 400 clinical samples, as it was submitted to the CTDA for approval.A correction has been applied to this article in: Corrigendum: VIDIIA Hunter: a low-cost, smartphone connected, artificial intelligence-assisted COVID-19 rapid diagnostic platform approved for medical use in the UK - correction: In the published article, there was an error in the Article title. Instead of “VIDIIA Hunter diagnostic platform: a low-cost, smartphone connected, artificial intelligence-assisted COVID-19 rapid diagnostics approved for medical use in the UK,” it should be “VIDIIA Hunter: a low-cost, smartphone connected, artificial intelligence-assisted COVID-19 rapid diagnostic platform approved for medical use in the UK”. The authors apologize for this error and state that this does not change the scientific conclusions of the article in any way. The original article has been updated.Copyright . Introduction: Accurate and rapid diagnostics paired with effective tracking and tracing systems are key to halting the spread of infectious diseases, limiting the emergence of new variants and to monitor vaccine efficacy. The current gold standard test (RT-qPCR) for COVID-19 is highly accurate and sensitive, but is time-consuming, and requires expensive specialised, lab-based equipment. Methods: Herein, we report on the development of a SARS-CoV-2 (COVID-19) rapid and inexpensive diagnostic platform that relies on a reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay and a portable smart diagnostic device. Automated image acquisition and an Artificial Intelligence (AI) deep learning model embedded in the Virus Hunter 6 (VH6) device allow to remove any subjectivity in the interpretation of results. The VH6 device is also linked to a smartphone companion application that registers patients for swab collection and manages the entire process, thus ensuring tests are traced and data securely stored. Results: Our designed AI-implemented diagnostic platform recognises the nucleocapsid protein gene of SARS-CoV-2 with high analytical sensitivity and specificity. A total of 752 NHS patient samples, 367 confirmed positives for coronavirus disease (COVID-19) and 385 negatives, were used for the development and validation of the test and the AI-assisted platform. The smart diagnostic platform was then used to test 150 positive clinical samples covering a dynamic range of clinically meaningful viral loads and 250 negative samples. When compared to RT-qPCR, our AI-assisted diagnostics platform was shown to be reliable, highly specific (100%) and sensitive (98–100% depending on viral load) with a limit of detection of 1.4 copies of RNA per µL in 30 min. Using this data, our CE-IVD and MHRA approved test and associated diagnostic platform has been approved for medical use in the United Kingdom under the UK Health Security Agency’s Medical Devices (Coronavirus Test Device Approvals, CTDA) Regulations 2022. Laboratory and in-silico data presented here also indicates that the VIDIIA diagnostic platform is able to detect the main variants of concern in the United Kingdom (September 2023). Discussion: This system could provide an efficient, time and cost-effective platform to diagnose SARS-CoV-2 and other infectious diseases in resource-limited settings.The study was funded by a Biotechnology and Biological Sciences Research Council grant (BB/R012695/1), awarded to Brunel University, University of Surrey, and Lancaster University. The study also received funding from the Higher Education Innovation Funding (HEIF–KN9190A) and the UK Research and Innovation (UKRI) EPSRC Impact Acceleration Accounts (IAA–RN0441R, RN0461E, and EP/R511560/1), which were awarded to the University of Surrey and Lancaster University in collaboration with Brunel University, United Kingdom

Proteomic evaluation of coffee zygotic embryos in two different stages of seed development.

Coffee seed development is accompanied by severe modifications in water soluble proteins, several of these being associated to a specific developmental stage. For this reason, a proteomic approach has been used to describe spatial-temporal proteome modifications in zygotic embryos at different stages of seed development. Embryos from Coffea arabica seeds were harvested in two different developmental stages: stage 1 at 210 days after anthesis and stage 2 at 255 days. Total proteins were extracted and submitted to 2-DE. From these gels, several spots were identified by mass spectrometry including kinases, MYB transcription factor and enzymes involved in metabolic pathways. All proteins identified seem to affect coffee development in different ways, being directly involved in plant growth or used as an intermediate in some metabolic pathway that, indirectly, will influence coffee development. This is the first work using two-dimensional electrophoresis followed by mass spectrometry analyses that evaluates the expression of proteins during coffee zygotic embryos development. Data here reported supply some light over coffee development and could be used in a near future to improve coffee plants? growth and development by molecular strategies.Made available in DSpace on 2011-04-09T22:27:53Z (GMT). No. of bitstreams: 1 Proteomicevaluation.pdf: 268577 bytes, checksum: fdc832eb548eb7752486660f4600f571 (MD5) Previous issue date: 2011-03-10200

Mitragynine (Kratom)-induced cognitive impairments in mice resemble Δ9-THC and morphine effects: reversal by cannabinoid CB1 receptor antagonism

Kratom is a widely abused plant-based drug preparation with a global interest in recent years, well beyond its native grounds in Southeast Asia. Mitragynine, its major psychoactive constituent is known to exhibit opioid-like behavioral effects with resultant neuroplasticity in the brain reward system. Its chronic administration is associated with cognitive impairments in animal studies. However, the underlying molecular mechanism for such a deficit remains elusive. In this study, the involvement of cannabinoid type-1 (CB1) receptors in cognitive deficits after chronic mitragynine exposures was investigated for 28 days (with incremental dose sensitization from 1 to 25 mg/kg) in adult male Swiss albino mice using the IntelliCage® system. Chronic high-dose mitragynine exposure (5–25 mg/kg, intraperitoneal [i.p.]), but not low-dose exposure (1–4 mg/kg, i.p.), induced hyperlocomotion, potentiated the preference for sucrose reward, increased resistance to punishment, and impaired place learning and its reversal. Comparable deficits were also observed after chronic treatments with Δ-9-tetrahydrocannabinol (THC, 2 mg/kg, i.p.) or morphine (5 mg/kg, subcutaneous). Mitragynine-, morphine-, and THC-induced learning and memory deficits were reversed by co-treatment with the CB1 receptor antagonist, NIDA-41020 (10 mg/kg, i.p.). A significant upregulation of CB1 receptor expression was found in the hippocampal CA1 region and ventral tegmental area after chronic high-dose mitragynine and morphine, whereas a downregulation was observed after chronic THC. In conclusion, the present study suggests a plausible role of the CB1 receptor in mediating the dose-dependent cognitive deficits after chronic high-dose mitragynine exposure. This also highlights the potential of CB1 receptor antagonism in ameliorating the cognitive deficits associated with long-term kratom/mitragynine consumption in humans

Development of a loop-mediated isothermal amplification (LAMP)-based electrochemical test for rapid detection of SARS-CoV-2

Summary: Rapid, reliable, sensitive, portable, and accurate diagnostics are required to control disease outbreaks such as COVID-19 that pose an immense burden on human health and the global economy. Here we developed a loop-mediated isothermal amplification (LAMP)-based electrochemical test for the detection of SARS-CoV-2 that causes COVID-19. The test is based on the oxidation-reduction reaction between pyrophosphates (generated from positive LAMP reaction) and molybdate that is detected by cyclic voltammetry using inexpensive and disposable carbon screen printed electrodes. Our test showed higher sensitivity (detecting as low as 5.29 RNA copies/μL) compared to the conventional fluorescent reverse transcriptase (RT)-LAMP. We validated our tests using human serum and saliva spiked with SARS-CoV-2 RNA and clinical (saliva and nasal-pharyngeal) swab samples demonstrating 100% specificity and 93.33% sensitivity. Our assay provides a rapid, specific, and sensitive test with an electrochemical readout in less than 45 min that could be adapted for point-of-care settings