Cell Delivery Strategies Research Articles

A review on recent advances in polymeric microneedle loading cells: Design strategies, fabrication technologies, transdermal application and challenges.

Microneedle systems (MNs) loading living cells are a powerful platform to treat various previously incurable diseases in the era of precision medicine. Herein, an overview of recent advances in MN-based strategies for cell delivery is summarized, including material selection, design of morphological structures, and processing methods. We also systematically outlined the law of microstructural design relative to the structure-effective/function relationship in transdermal delivery or precision medicine and the design principles of cell microneedle (CMN). Furthermore, the representative works of precision treatments focusing on inflammatory skin diseases were tracked and discussed using CMN. Indeed, it highlights a practical path to solving the dilemma of cell therapy and raising the hope of precision medicine. However, there are still some challenges in developing CMN since they need multi-dimensional comprehensive properties, including mechanical properties, cell viability preservation, release, therapeutic effect, etc. The manuscript could provide insights into developing an innovative fit-to-purpose vehicle in cell therapy for interested researchers.

Gelatin microcarriers as an effective adipose-derived stem cells delivery strategy in osteoarthritis treatment

Despite their clinical success, stem cells (SCs) in the treatment of osteoarthritis (OA) are limited by lower retention efficiency and restricted paracrine function. The development of effective SCs culture and delivery systems to maintain or promote SCs paracrine function is urgently needed. In this study, we focused on gelatin microcarriers with different sizes as SCs culture scaffold for OA therapy. The effect of culturing adipose-derived stem cells (ADSCs) on different size microcarriers (180 μm and 320 μm) to promote paracrine function was evaluated. The secretome of ADSCs cultured on microcarriers more effectively regulated macrophages and chondrocytes in a direction favorable to OA healing compared to culture plates. In particular, microcarriers with ADSCs effectively reduced the coefficient of friction at the cartilage interface. Injection low-dose ADSCs without and with different size microcarriers into the knee joints in rats' OA model was achieved. Even better OA therapeutic effects were achieved by using smaller size (higher curvature) microcarriers to deliver ADSCs at low doses. Microcarrier-based cell delivery strategies offer potential solution method for OA therapy.

Small heat shock protein B8: from cell functions to its involvement in diseases and potential therapeutic applications.

Heat shock protein family B (small) member 8 (HSPB8) is a 22 kDa ubiquitously expressed protein belonging to the family of small heat shock proteins. HSPB8 is involved in various cellular mechanisms mainly related to proteotoxic stress response and in other processes such as inflammation, cell division, and migration. HSPB8 binds misfolded clients to prevent their aggregation by assisting protein refolding or degradation through chaperone-assisted selective autophagy. In line with this function, the pro-degradative activity of HSPB8 has been found protective in several neurodegenerative and neuromuscular diseases characterized by protein misfolding and aggregation. In cancer, HSPB8 has a dual role being capable of exerting either a pro- or an anti-tumoral activity depending on the pathways and factors expressed by the model of cancer under investigation. Moreover, HSPB8 exerts a protective function in different diseases by modulating the inflammatory response, which characterizes not only neurodegenerative diseases, but also other chronic or acute conditions affecting the nervous system, such as multiple sclerosis and intracerebellar hemorrhage. Of note, HSPB8 modulation may represent a therapeutic approach in other neurological conditions that develop as a secondary consequence of other diseases. This is the case of cognitive impairment related to diabetes mellitus, in which HSPB8 exerts a protective activity by assuring mitochondrial homeostasis. This review aims to summarize the diverse and multiple functions of HSPB8 in different pathological conditions, focusing on the beneficial effects of its modulation. Drug-based and alternative therapeutic approaches targeting HSPB8 and its regulated pathways will be discussed, emphasizing how new strategies for cell and tissue-specific delivery represent an avenue to advance in disease treatments.

Dental pulp stem cells-loaded kartogenin-modified hydrogel microspheres with chondrocyte differentiation property for cartilage repair

Stem cell-based regimens have evolved as a promising approach to cartilage reparations. Research efforts in this realm aim to enhance the therapeutic capabilities of stem cells and develop more effective cell delivery strategies. Herein, we present novel dental pulp stem cells (DPSCs)-loaded kartogenin-modified hydrogel microspheres with chondrocyte differentiation property for temporomandibular joint (TMJ) osteoarthritis treatment. Leveraging microfluidic technology, DPSCs were successfully encapsulated in hydrogel microspheres, serving as the pivotal contributors to cartilage repair. By modifying the hydrogel with kartogenin, these microspheres facilitated sustained stimulation of DPSCs’ chondrogenic differentiation. In vitro experimental results revealed that the DPSCs exhibited advanced cartilage-forming differentiation, as evidenced by the up-regulation of relative gene expressions and protein markers. Additionally, the DPSCs-loaded microspheres system displayed a capacity to regenerate and repair tissue at the cartilage defects in a TMJ osteoarthritis animal model. Thus, the DPSCs-loaded kartogenin-modified hydrogel microspheres exhibit significant promise for cartilage repair, showcasing the substantial potential for future clinical applications.

Dexamethasone-Integrated Mesenchymal Stem Cells for Systemic Lupus Erythematosus Treatment via Multiple Immunomodulatory Mechanisms.

The therapeutic application of mesenchymal stem cells (MSCs) has good potential as a treatment strategy for systemic lupus erythematosus (SLE), but traditional MSC therapy still has limitations in effectively modulating immune cells. Herein, we present a promising strategy based on dexamethasone liposome-integrated MSCs (Dexlip-MSCs) for treating SLE via multiple immunomodulatory pathways. This therapeutic strategy prolonged the circulation time of dexamethasone liposomes in vivo, restrained CD4+T-cell proliferation, and inhibited the release of proinflammatory mediators (IFN-γ and TNF-α) by CD4+T cells. In addition, Dexlip-MSCs initiated cellular reprogramming by activating the glucocorticoid receptor (GR) signaling pathway to upregulate the expression of anti-inflammatory factors such as cysteine-rich secretory protein LCCL-containing domain 2 (CRISPLD2) and downregulate the expression of proinflammatory factors. In addition, Dexlip-MSCs synergistically increased the anti-inflammatory inhibitory effect of CD4+T cells through the release of dexamethasone liposomes or Dex-integrated MSC-derived exosomes (Dex-MSC-EXOs). Based on these synergistic biological effects, we demonstrated that Dexlip-MSCs alleviated disease progression in MRL/lpr mice more effectively than Dexlip or MSCs alone. These features indicate that our stem cell delivery strategy is a promising therapeutic approach for clinical SLE treatment.

Lyophilized lymph nodes for improved delivery of chimeric antigen receptor T cells.

Lymph nodes are crucial organs of the adaptive immune system, orchestrating T cell priming, activation and tolerance. T cell activity and function are highly regulated by lymph nodes, which have a unique structure harbouring distinct cells that work together to detect and respond to pathogen-derived antigens. Here we show that implanted patient-derived freeze-dried lymph nodes loaded with chimeric antigen receptor T cells improve delivery to solid tumours and inhibit tumour recurrence after surgery. Chimeric antigen receptor T cells can be effectively loaded into lyophilized lymph nodes, whose unaltered meshwork and cytokine and chemokine contents promote chimeric antigen receptor T cell viability and activation. In mouse models of cell-line-derived human cervical cancer and patient-derived pancreatic cancer, delivery of chimeric antigen receptor T cells targeting mesothelin via the freeze-dried lymph nodes is more effective in preventing tumour recurrence when compared to hydrogels containing T-cell-supporting cytokines. This tissue-mediated cell delivery strategy holds promise for controlled release of various cells and therapeutics with long-term activity and augmented function.

Rationally designed approaches to augment CAR-T therapy for solid tumor treatment

Chimeric antigen receptor T cell denoted as CAR-T therapy has realized incredible therapeutic advancements for B cell malignancy treatment. However, its therapeutic validity has yet to be successfully achieved in solid tumors. Different from hematological cancers, solid tumors are characterized by dysregulated blood vessels, dense extracellular matrix, and filled with immunosuppressive signals, which together result in CAR-T cells’ insufficient infiltration and rapid dysfunction. The insufficient recognition of tumor cells and tumor heterogeneity eventually causes cancer reoccurrences. In addition, CAR-T therapy also raises safety concerns, including potential cytokine release storm, on-target/off-tumor toxicities, and neuro-system side effects. Here we comprehensively review various targeting aspects, including CAR-T cell design, tumor modulation, and delivery strategy. We believe it is essential to rationally design a combinatory CAR-T therapy via constructing optimized CAR-T cells, directly manipulating tumor tissue microenvironments, and selecting the most suitable delivery strategy to achieve the optimal outcome in both safety and efficacy.

The Osteogenic Role of Biomaterials Combined with Human-Derived Dental Stem Cells in Bone Tissue Regeneration.

The use of stem cells in regenerative medicine had great potential for clinical applications. However, cell delivery strategies have critical importance in stimulating the differentiation of stem cells and enhancing their potential to regenerate damaged tissues. Different strategies have been used to investigate the osteogenic potential of dental stem cells in conjunction with biomaterials through in vitro and in vivo studies. Osteogenesis has a broad implication in regenerative medicine, particularly for maxillofacial defects. This review summarizes some of the most recent developments in the field of tissue engineering using dental stem cells.

Fabrication, characterization and in vivo assessment of cardiogel loaded chitosan patch for myocardial regeneration

Cell therapy is one of the promising approaches for cardiac repair, subsequently after infarction or injury. However, contemporary mesenchymal stromal/stem cell (MSCs) delivery strategies result in low retention and poor engraftment of donor cells, thus limiting the therapeutic efficacy. Here, we developed an engineered biomimetic cardiogel patch (EBCP) comprising of the native decellularized cardiac extracellular matrix (ECM) “cardiogel” and chitosan, leading to the efficient regeneration of injured myocardium. We also developed novel bio-adhesive that is capable of suture-free epicardial placement of EBCP to injured myocardium. We have illustrated the potential of the mussels-inspired bioadhesive system, which comprises gelatin catechol and partially oxidized chitosan, which relies on self-crosslinking capability, to promote wet adhesion. In vitro studies with isolated cardiogel promoted cell proliferation, adhesion, and migration while aiding cardiomyogenic differentiation. The EBCP's ability to protect cells from abrasion due to surrounding tissues in the myocardial infarction (MI) rat model makes it more desirable.Furthermore, the epicardial implantation of the EBCP loaded with MSCs improves the initial retention of cells and subsequent functional cardiac recovery with enhanced myocardial tissue restoration. Histological examination showed the presence of EBCP and infiltration of cells to the infarcted heart tissue. The fast and facile synthesis of bioadhesive and major therapeutic benefits of EBCP make it a potential candidate for recuperating the ailing heart.

Biological properties of human periodontal ligament cell spheroids cultivated on chitosan and polyvinyl alcohol membranes

Multicellular spheroid cultures have attracted increasing attention in the field of periodontal regeneration. However, very few studies have reported the periodontal ligament (PDL) cell spheroid formation via biomaterials-induced processes. This study investigated the biological characteristics of human PDL cell spheroids formed on two hydrophilic polymer-based biomaterials, namely chitosan and polyvinyl alcohol. The expressions of periostin, paxillin, hypoxia-inducible factor 1-α (HIF-1α), and vascular endothelial growth factor (VEGF) were analyzed. Cell migration ability was assessed using a scratch assay. Furthermore, PDL cell spheroids were cultured in 3D-printed polylactic acid scaffolds to evaluate mineralizing capability. Western blot analysis revealed increased expressions of periostin, HIF-1α, and VEGF in the 3D spheroids. After the spheroids were reseeded, the cells gradually migrated outward from the spheroids and time-dependent distribution of paxillin was observed. The cells migrating outward from the 3D spheroids demonstrated greater migration ability than that of 2D monolayer cells. Compared to the dissociated cells from a monolayer culture, the cell spheroids formed on the chitosan membrane exhibited elevated alkaline phosphatase activity and an increase in mineralized matrix deposition. The biomaterial-induced formation of PDL cell spheroids suggests a novel strategy for cell delivery in research and clinical applications of periodontal regeneration.

Adaptive Gelatin Microspheres Enhanced Stem Cell Delivery and Integration With Diabetic Wounds to Activate Skin Tissue Regeneration.

The delayed and complicated diabetic wound healing raises clinical and social concerns. The application of stem cells along with hydrogels is an attractive therapeutic approach. However, low cell retention and integration hindered the performance. Herein, gelatin microspheres were fabricated for local delivery of adipose-derived stem cells (from rats, rADSCs), and the effect of rADSCs with microspheres on diabetic wound healing was examined. Uniform, well-dispersed microspheres were fabricated using the microfluidic technique. Due to geometry differences, the proteinase degradation rate for microspheres was four times that of the bulk hydrogel. The obtained gelatin microspheres supported cell's adhesion and proliferation and provided a suitable microenvironment for rADSC survival. For in vivo animal tests, rADSCs were labeled with CM-Dil for tracking purposes. Microspheres were well embedded in the regenerated tissue and demonstrated good biocompatibility and an adaptive biodegradation rate. Histological examination revealed rADSC-loaded gelatin microspheres that significantly accelerated wound healing via promoting M2 macrophage polarization, collagen deposition, angiogenesis associated with peripheral nerve recovery, and hair follicle formation. Notably, the relative fluorescence intensity around the hair follicle was 17-fold higher than that of the blank group, indicating rADSC participated in the healing process via exosomes. Taken together, the rADSC-laden gelatin microspheres provided a promising strategy for local stem cell delivery to improve diabetic wound healing.

Cell therapy and delivery strategies for spinal cord injury.

Spinal cord injury (SCI) is a complex neuropathological condition that represents a major challenge for clinicians and scientists due to patient's functional dysfunction and paralysis. Several treatments have been proposed including biological factors, drugs and cells administered in various ways. Stem cells arise as good candidates to treat SCI since they are known to secrete neurotrophic factors, improving neuroregeneration, but also due to their role in modulating the inflammatory process, favoring a pro-regenerative status. There are several types of cells that have been tested to treat SCI in experimental and clinical studies, but we still face many unanswered questions; one of them is the type of cells that can offer the best benefits and, also the ideal dose and administration routes. This review aimed to summarize recent research on cell treatment, focusing on current delivery strategies for SCI therapy and their effects in tissue repair and regeneration.

(Invited) Non-Blinking Long Lifetime Quantum Dots and Passive Cell Delivery Strategies

Over the past ~20 years there has been considerable effort towards the optimization of the photophysical properties of emissive nanomaterials to engender efficacy for biological imaging and sensing. The development of core-shell CdSe/ZnS and CdSe/CdS quantum dots (QDs) was instrumental due to those materials’ resistance to photobleaching and high quantum yields. However, these particles experience fluorescence intermittency (“blinking”), which is detrimental for single particle tracking studies. Furthermore, ~10 nanosecond lifetimes are not compatible with time-gating visualization, a technique that reduces background noise. Last, cytosolic delivery of QDs is a well-known problem due to the propensity for QDs to become sequestered in the endosomes of live cells.Presented are developments that address these issues. Long lifetimes, >100 ns, and non-blinking behavior have been realized in “giant” quantum dots with a highly alloyed core/shell interface. This results in type-II behavior that separates the electron and hole, which slows emissive recombination. Also presented is a novel cell-penetrating peptide that can be conjugated to surfactants that are commonly employed for QD water solubilization. These materials can be passively delivered to the cytosol of live T-cell lymphocytes. Single particle tracking studies reveal fast diffusion of single dots that are not consistent with endosomal capture, which is observed in control nanomaterials. These developments create new inroads for chemical and biological sensing studies inside of live cells for basic research and diagnostic purposes. Figure 1

Nanomaterials toward the treatment of Alzheimer’s disease: Recent advances and future trends

Alzheimer’s disease (AD) is a progressive and fatal neurodegenerative condition and the most prevalent cause of dementia. This disease is characterized by progressive cognitive impairment. The prevalence of AD is currently affecting more than 35 million people and is rising worldwide. No efficient therapy is currently available due to low drug potency and a number of various obstacles to delivery. Recent nanotechnological advancements have the potential to offer promising therapeutic options. Progress on nanomaterials as well as their applications in biomedicine is receiving increasing attention, especially the advantages of nanomaterial-based drug delivery systems. The aim of this review is to comprehensively summarize the latest developments in nanomaterial-based strategies for AD treatment, including nanoparticles, liposomes and other options for the delivery of therapeutic compounds and scaffolds for cell delivery strategies. Future research directions are also proposed. We hope this review can provide important information to guide the future development of nanomaterials in AD treatment.

Enhanced Endothelial Cell Delivery for Repairing Injured Endothelium via Pretargeting Approach and Bioorthogonal Chemistry.

Arterial wall injury often leads to endothelium cell activation, endothelial detachment, and atherosclerosis plaque formation. While abundant research efforts have been placed on treating the end stages of the disease, no cure has been developed to repair injured and denude endothelium often occurred at an early stage of atherosclerosis. Here, a pretargeting cell delivery strategy using combined injured endothelial targeting nanoparticles and bioorthogonal click chemistry approach was developed to deliver endothelial cells to replenish the injured endothelium via a two-step process. First, nanoparticles bearing glycoprotein 1b α (Gp1bα) proteins and tetrazine (Tz) were fabricated to provide a homogeneous nanoparticle coating on an injured arterial wall via the interactions between Gp1bα and von Willebrand factor (vWF), a ligand that is present on denuded endothelium. Second, transplanted endothelium cells bearing transcyclooctene (TCO) would be quickly immobilized on the surfaces of nanoparticles via TCO:Tz reactions. In vitro binding studies under both static and flow conditions confirmed that our novel Tz-labeled Gp1bα-conjugated poly(lactic-co-glycolic acid) (PLGA) nanoparticles can successfully pretargeted toward the injured site and support rapid adhesion of endothelial cells from the circulation. Ex vivo results also confirm that such an approach is highly efficient in mediating the local delivery of endothelial cells at the sites of arterial injury. The results support that this pretargeting cell delivery approach may be used for repairing injured endothelium in situ at its early stage.

A Quantitative Description for Designing the Extrudability of Shear-Thinning Physical Hydrogels.

Physically associated hydrogels (PHs) capable of reversible transitions between solid and liquid-like states have enabled novel strategies for 3D printing, therapeutic drug and cell delivery, and regenerative medicine. Among the many design criteria (e.g., viscoelasticity, cargo diffusivity, biocompatibility) for these applications, engineering PHs for extrudability is a necessary and critical design criterion for the successful application of these materials. As the development of many distinct PH material systems continues, a strategy to determine the extrudability of PHs a priori will be exceedingly useful for reducing costly and time-consuming trial-and-error experimentation. Here, a strategy to determine the property-function relationships for PHs in injectable drug delivery applications at clinically relevant flow rates is presented. This strategy-validated with two chemically and physically distinct PHs-reveals material design spaces in the form of Ashby-style plots that highlight acceptable, application-specific material properties. It is shown that the flow behavior of PHs does not obey a single shear-thinning power law and the implications for injectable drug delivery are discussed. This approach for generating design criteria has potential for streamlining the screening of PHs and their utility in applications with varying geometrical (i.e., needle diameter) and process (i.e., flow rate) constraints.

Cardiomyocyte Induction and Regeneration for Myocardial Infarction Treatment: Cell Sources and Administration Strategies.

Occlusion of coronary artery and subsequent damage or death of myocardium can lead to myocardial infarction (MI) and even heart failure-one of the leading causes of deaths world wide. Notably, myocardium has extremely limited regeneration potential due to the loss or death of cardiomyocytes (i.e., the cells of which the myocardium is comprised) upon MI. A variety of stem cells and stem cell-derived cardiovascular cells, in situ cardiac fibroblasts and endogenous proliferative epicardium, have been exploited to provide renewable cellular sources to treat injured myocardium. Also, different strategies, including direct injection of cell suspensions, bioactive molecules, or cell-incorporated biomaterials, and implantation of artificial cardiac scaffolds (e.g., cell sheets and cardiac patches), have been developed to deliver renewable cells and/or bioactive molecules to the MI site for the myocardium regeneration. This article briefly surveys cell sources and delivery strategies, along with biomaterials and their processing techniques, developed for MI treatment. Key issues and challenges, as well as recommendations for future research, are also discussed.

Abstract 4482: Monitoring T cell reactivity to allogeneic placental cell products using multiple lymphocyte reaction and TCR sequencing approaches

Abstract Introduction: Allogeneic stem cell-derived therapies offer patients a readily accessible source of off-the-shelf product. However, allogeneic cells may be recognized by the recipient's immune system thus negatively affecting persistence and efficacy. Among people, allelic differences in human leukocyte antigen (HLA) complex genes are the major source of alloantigen which can elicit immunity. While there are established assays for measuring alloantibodies, monitoring T cell alloreactivity is more challenging. Unique clonal populations vary between individuals and functional assays are difficult to scale and standardize if alloreactive populations are rare/infrequent. Here, we evaluate how alloreactive T cells from patients may be identified using multiple lymphocyte reaction (MLR). Next generation sequencing of the TCR locus permits the subsequent identification and monitoring of allogeneic T cell clones in a patient over time. Methods: Placental leukocytes depleted of CD34 cells or peripheral blood mononuclear cells (PBMC) were enriched for monocytes using magnetic-bead isolation followed by dendritic cell (DC) maturation in the presence of GM-CSF and IL-4 for 5 days. DC were activated with lipopolysaccharide for 16 hours then cultured in the presence of CD3+ T cells isolated from healthy 3rd party PBMC for 5 days in the presence of IL-2. Proliferating alloantigen-specific T cells were observed by flow cytometry. TCR locus sequencing was performed on DNA libraries generated from unstimulated and post-MLR cultures using the Illumina platform. Results: In contrast to monocyte derived DCs from PBMC, monocyte derived DCs from placenta required higher levels of cytokine and lipopolysaccharide stimulation to achieve similar level of maturation. Co-culture of placental-derived DCs with unrelated T cells over 5 days resulted in the outgrowth of T cell colonies visible by brightfield microscopy. Flow cytometry confirmed clonal expansion of CD4 and CD8 T cell populations based upon the expression of Ki-67, CD25, and CD69 on cells with a higher forward scatter. To assess feasibility of performing TCR sequencing in unstimulated and post-MLR T cells, one T cell responder was evaluated in response to 3 different DC donors. For each DC donor, a single T cell clone representing 0.35% of the starting repertoire expanded to become the dominant clone at 44%, 61%, and 66% following MLR with the next most frequent clones represented 4%, 3%, and 2% among total T cells, respectively. Our data demonstrate the feasibility of identifying alloreactive T cell clones for translational research monitoring in patients. This approach could inform correlations with cell persistence and help optimize allogeneic cell delivery and dosage strategies in the context of disease state and lymphodepletion. Citation Format: Bhavani Stout, Tanel Mahlakoiv, Robert Hariri, Xiaokui Zhang, William van der Touw. Monitoring T cell reactivity to allogeneic placental cell products using multiple lymphocyte reaction and TCR sequencing approaches [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4482.

Fabrication of homotypic neural ribbons as a multiplex platform optimized for spinal cord delivery

Cell therapy for the injured spinal cord will rely on combined advances in human stem cell technologies and delivery strategies. Here we encapsulate homotypic spinal cord neural stem cells (scNSCs) in an alginate-based neural ribbon delivery platform. We perform a comprehensive in vitro analysis and qualitatively demonstrate graft survival and injury site retention using a rat C4 hemi-contusion model. Pre-configured neural ribbons are transport-stable modules that enable site-ready injection, and can support scNSC survival and retention in vivo. Neural ribbons offer multifunctionality in vitro including co-encapsulation of the injury site extracellular matrix modifier chondroitinase ABC (chABC), tested here in glial scar models, and ability of cervically-patterned scNSCs to differentiate within neural ribbons and project axons for integration with 3-D external matrices. This is the first extensive in vitro characterization of neural ribbon technology, and constitutes a plausible method for reproducible delivery, placement, and retention of viable neural cells in vivo.

HLA-Matched Allogeneic iPS Cells-Derived RPE Transplantation for Macular Degeneration.

Immune attacks are key issues for cell transplantation. To assess the safety and the immune reactions after iPS cells-derived retinal pigment epithelium (iPS-RPE) transplantation, we transplanted HLA homozygote iPS-RPE cells established at an iPS bank in HLA-matched patients with exudative age-related macular degeneration. In addition, local steroids without immunosuppressive medications were administered. We monitored immune rejections by routine ocular examinations as well as by lymphocytes-graft cells immune reaction (LGIR) tests using graft RPE and the patient’s blood cells. In all five of the cases that underwent iPS-RPE transplantation, the presence of graft cells was indicated by clumps or an area of increased pigmentation at 6 months, which became stable with no further abnormal growth in the graft during the 1-year observation period. Adverse events observed included corneal erosion, epiretinal membrane, retinal edema due to epiretinal membrane, elevated intraocular pressure, endophthalmitis, and mild immune rejection in the eye. In the one case exhibiting positive LGIR tests along with a slight fluid recurrence, we administrated local steroid therapy that subsequently resolved the suspected immune attacks. Although the cell delivery strategy must be further optimized, the present results suggest that it is possible to achieve stable survival and safety of iPS-RPE cell transplantation for a year.