An improved formulation of human neural stem cells for the treatment of geographic atrophy

Development of a Targeted Choroidal Injury Model for the Study of Retinal Degenerations and Therapeutic Cell Replacement

Purpose Loss of the choroid is an important pathophysiological step in retinal diseases ranging from age-related macular degeneration (AMD) to choroideremia. However, current models of choroidal injury, such as laser photocoagulation-induced injury, induce acute tissue damage that does not parallel the gradual loss seen in most retinal degenerations. As such, there remains a need for models of targeted choroidal injury to study mechanisms of disease and develop potential treatments. Methods We report a targeted choroidal injury model using bioconjugated saporins (immunotoxins) against cluster of differentiation (CD) domains CD38 and CD105. To induce injury, 10 µL of bioconjugate solution or control (phosphate-buffered saline) was delivered via suprachoroidal injection in wild-type Sprague-Dawley (SD) and Sprague-Dawley Rag2/Il2rg (SRG; double-knockout) rats. The extent and severity of choroidal injury were compared with animals treated with sodium iodate administered via tail vein injections at 1, 2, and 3 weeks post-treatment using clinical imaging and immunohistochemistry. Results Choroidal injury induced by either anti-CD38 or anti-CD105 immunotoxin was targeted to the choroid and localized to the sector of the injection, unlike global destruction induced by sodium iodate. Immunotoxin-induced vascular damage remained constant over 3 weeks, without choroidal neovascularization seen, whereas sodium iodate caused progressive and severe injury of the choroid and overlying retina. Conclusions Suprachoroidal injection of anti-CD38 and anti-CD105 immunotoxins can induce localized and durable choroidal injury in both wild-type and immunosuppressed rats. Translational Relevance Immunotoxin-based models of targeted choroidal injury may be useful for understanding pathophysiologic mechanisms and developing therapies in conditions involving choroidal cell death.

Human Stem Cell Transplantation, Immunosuppression, and Noninvasive In vivo Cell Tracking in the Mouse Brain

Abstract Stem cell therapies have emerged as potential therapeutics of interest for many central nervous system diseases, including stroke, epilepsy, neurodegenerative diseases, and traumatic brain injury. Preclinical models are required to evaluate the migration of transplanted stem cells to various areas of the brain, monitor graft survival to improve immunosuppression regimens, and assess stem cell safety and their efficacy on disease outcomes. Understanding all these aspects of stem cells as therapeutics can help optimize parameters for translation to humans. To facilitate this overarching goal, we report a protocol for human stem cell transplantation into mouse fimbria fornix, the outflow white matter tract of the hippocampus, which facilitates stem cell dissemination to regions distant to the injection site. Although our protocol involves injecting human neural stem cells expressing insulin‐like growth factor 1 into an Alzheimer's disease mouse model, it can be altered for application to diverse stem cells and cellular therapeutics, secreting various trophic factors, at multiple dosing schemes, and in different central nervous system targets and disorder models. Our approach is robust and highly reproducible, with demonstrated success in improving memory performance in an Alzheimer's disease mouse model and achieving long‐term survival of transplanted cells (up to 32 weeks) in immunocompetent mice, facilitating stem cell studies in the context of neuroimmune crosstalk. Our primary protocol involves administering an anti‐CD4/anti‐CD40L monoclonal antibody cocktail to prevent graft rejection, with bioluminescence imaging to track transplanted stem cells in vivo. We also present an alternate immunosuppression protocol that relies on tacrolimus and mycophenolate mofetil, which is less costly, albeit less effective (especially in the longer term), than the CD4‐CD40L antibody regimen. © 2026 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol: Human stem cell transplantation into mouse hippocampus, antibody‐based immunosuppression, and in vivo tracking by bioluminescence imaging Alternate Protocol: Immunosuppression by tacrolimus and mycophenolate mofetil

Preclinical evaluation of AL-001, a gene therapy for wet age-related macular degeneration

Frequent intravitreal administration of antivascular endothelial growth factor Vascular endothelial growth factor agents remains a major limitation in the management of wet age-related macular degeneration (wAMD). This study evaluated whether suprachoroidal delivery of an engineered recombinant adeno-associated viral (rAAV)-aflibercept vector could achieve sustained, targeted expression with improved efficacy and safety compared with intravitreal administration. AL-001, an engineered rAAV vector expressing aflibercept, was developed and characterized. Its expression profile was first assessed in New Zealand white rabbits following suprachoroidal space (SCS) injection. Efficacy, pharmacokinetics, and safety were then evaluated in a nonhuman primate model of laser-induced choroidal neovascularization (CNV), comparing SCS and intravitreal (IVT) administration routes. AL-001 efficiently expressed aflibercept in relevant ocular cells in vitro . In rabbits, SCS administration produced sustained aflibercept levels in ocular tissues. In the nonhuman primate CNV model, a single SCS injection of AL-001 showed favorable efficacy to IVT injection and a notable mild inflammatory response. At week 4, grade IV lesion incidence was 0% (0/48) after SCS administration versus 14.3% (6/42) after IVT administration (absolute difference, −14.3 percentage points; 95% CI, 3.7%–27.8%; P = 0.0258). Throughout follow-up, mean leakage area and grade IV lesion incidence remained 0 with SCS, versus IVT peaks of approximately 0.3 mm 2 and 33.0%, respectively, declining to 0.03 mm 2 and 2.0% by day 100. Both the medium and high doses decreased pathological vascular leakage and subretinal hyperreflective material. Vector administration preceded laser-induced CNV modeling, demonstrating that sustained intraocular aflibercept expression in the retina and choroid provided durable antiangiogenic protection. Pharmacokinetic analysis confirmed distinct ocular exposure profiles between routes, with viral genomes confined predominantly to the injected eye and no significant systemic accumulation. AL-001 was well tolerated, without sustained intraocular pressure elevation or severe ocular inflammation, and only mild-to-moderate treatment-emergent adverse events. Low pre-existing anti-AAV2 immunity and time-dependent neutralizing antibody responses postdosing, informing a translational model for patient stratification and redosing feasibility. Suprachoroidal administration of AL-001 is well tolerated and provides durable, targeted aflibercept expression with pronounced antiangiogenic efficacy. These results support AL-001 as a promising, long-acting therapeutic candidate for wAMD.

endo-Regulation of functional corneal endothelial cells isolated from sphere colonies by Rho-associated protein kinase inhibitor

Intrathecal injection of 3D-mesenchymal stem cells attenuates disseminated neuroinflammation and improves cognitive function in controlled cortical impact rats

Background Persistent cognitive impairment following traumatic brain injury (TBI) represents a significant unmet clinical need. Neuroinflammation has been implicated as one of the pathological processes that may contribute to this outcome. Given the limitations of current therapies, this study explored the therapeutic potential of three-dimensional cultured mesenchymal stem cells (3D-MSCs) for TBI, with a focus on long-term cognitive outcome and their immunomodulatory properties. Methods A controlled cortical impact (CCI) model was established in male Sprague-Dawley rats. Twenty-four hours post-injury, rats received an intrathecal injection of 1x106 3D-MSCs or vehicle. Cell migration, apoptosis, lesion volume, sensorimotor and cognitive functions, and neuroinflammation were subsequently evaluated. Results Intrathecally administered 3D-MSCs exhibited significant homing to the perilesional region within 24 h. Subsequently, the treatment reduced perilesional apoptosis by 59.0% at 48 h and lesion volume by 28.8% at 7 days post-injury (dpi). 3D-MSCs significantly enhanced sensorimotor recovery from day 3 and improved spatial learning and memory at 30 dpi. Importantly, it also attenuated CCI-induced widespread Iba1-associated neuroinflammatory responses across multiple brain regions at 7 dpi. Conclusion Focal TBI can induce disseminated neuroinflammation. A single intrathecal injection of 3D-MSCs within 24 h after TBI enables rapid migration to the injury site and exerts early neuroprotective effects. In addition, 3D-MSC administration improved long-term cognitive function and attenuated disseminated neuroinflammatory responses, suggesting that immunomodulation may be one of the factors contributing to its therapeutic benefit.

Neuroprotective effect of intraperitoneal Humanin-G in retinal degeneration of Royal College of Surgeons rats.

This study aimed to examine whether Humanin-G (HNG), a mitochondrial derived peptide with cytoprotective properties, could improve the retinal function and gene expression in Royal College of Surgeons (RCS) rats with retinal pigment epithelium (RPE) dysfunction and retinal degeneration. Starting at postnatal day 21, RCS rats received twice a week intraperitoneal injection of either Low Dose HNG (0.4 mg/kg), High Dose HNG (4 mg/kg), or sham-saline for 1 or 4 weeks. Visual function was tested with electroretinography (ERG) and optokinetic testing (OKT). Then the rats were euthanized for RNA, cDNA and Quantitative Real-time PCR (qRT-PCR) analysis. The results showed that high dose HNG at 4 weeks after first injection (WAFI) was associated with the largest change in gene expression in the RPE and retina of treated animals, altering expression of genes involved in apoptosis, oxidative stress, inflammation and retinal/RPE function. At 4 WAFI, ERG showed no difference between either low or high dose of HNG and sham injection, while the visual acuity tested by OKT in rats treated with high dose HNG showed significant improvement. Our findings suggested that HNG can modulate gene expression and improve vision. Further studies are warranted to show whether HNG may be a potential treatment for retinal degeneration diseases.