Ziming Luo

All Publications

• Kif5a Regulates Mitochondrial Transport in Developing Retinal Ganglion Cells In Vitro. Investigative ophthalmology & visual science Yokota, S., Shah, S. H., Huie, E. L., Wen, R. R., Luo, Z., Goldberg, J. L. 2023; 64 (3): 4

  Abstract

  Axon transport of organelles and neurotrophic factors is necessary for maintaining cellular function and survival of retinal ganglion cells (RGCs). However, it is not clear how trafficking of mitochondria, essential for RGC growth and maturation, changes during RGC development. The purpose of this study was to understand the dynamics and regulation of mitochondrial transport during RGC maturation using acutely purified RGCs as a model system.Primary RGCs were immunopanned from rats of either sex during three stages of development. MitoTracker dye and live-cell imaging were used to quantify mitochondrial motility. Analysis of single-cell RNA sequencing was used to identify Kinesin family member 5A (Kif5a) as a relevant motor candidate for mitochondrial transport. Kif5a expression was manipulated with either short hairpin RNA (shRNA) or exogenous expression adeno-associated virus viral vectors.Anterograde and retrograde mitochondrial trafficking and motility decreased through RGC development. Similarly, the expression of Kif5a, a motor protein that transports mitochondria, also decreased during development. Kif5a knockdown decreased anterograde mitochondrial transport, while Kif5a expression increased general mitochondrial motility and anterograde mitochondrial transport.Our results suggested that Kif5a directly regulates mitochondrial axonal transport in developing RGCs. Future work exploring the role of Kif5a in vivo in RGCs is indicated.

  View details for DOI 10.1167/iovs.64.3.4

  View details for PubMedID 36862119

• Characterization of Primary Cilia Formation in Human ESC-Derived Retinal Organoids. Stem cells international Ning, K., Luo, Z., Kowal, T. J., Tran, M., Majumder, R., Jarin, T. M., Wu, A. Y., Goldberg, J. L., Sun, Y. 2023; 2023: 6494486

  Abstract

  Objectives: Primary cilia are conserved organelles found in polarized mammalian cells that regulate neuronal growth, migration, and differentiation. Proper cilia formation is essential during eye development. Our previous reports found that both amacrine and retinal ganglion cells (RGCs) contain primary cilia in primate and rodent retinas. However, whether primary cilia are present in the inner retina of human retinal organoids remains unknown. The purpose of this study is to characterize the primary cilia distribution in human embryonic stem cell (hESC-derived retinal organoid development.Materials and Methods: Retinal organoids were differentiated from a hESC line, harvested at various developmental timepoints (day 44-day 266), and immunostained with antibodies for primary cilia, including Arl13b (for the axoneme), AC3, and Centrin3 (for the basal body). AP2alpha, Prox1, GAD67, Calretinin, GFAP, PKCalpha, and Chx10 antibodies as well as Brn3b-promoted tdTomato expression were used to visualize retinal cell types.Results: A group of ciliated cells were present in the inner aspects of retinal organoids from day 44 to day 266 in culture. Ciliated Chx10-positive retinal progenitor cells, GFAP-positive astrocytes, and PKCalpha-positive rod-bipolar cells were detected later during development (day 176 to day 266). Ciliation persisted during all stages of retinal developmental in AP2alpha-positive amacrine cells, but it was decreased in Brn3b-positive retinal ganglion cells (RGCs) at later time points. Additionally, AC3-positive astrocytes significantly decreased during the later stages of organoid formation.Conclusions: Amacrine cells in retinal organoids retain cilia throughout development, whereas RGC ciliation gradually and progressively decreases with organoid maturation.

  View details for DOI 10.1155/2023/6494486

  View details for PubMedID 36684387

• Directly induced human retinal ganglion cells mimic fetal RGCs and are neuroprotective after transplantation invivo. Stem cell reports Luo, Z., Chang, K., Wu, S., Sun, C., Xia, X., Nahmou, M., Bian, M., Wen, R. R., Zhu, Y., Shah, S., Tanasa, B., Wernig, M., Goldberg, J. L. 2022

  Abstract

  Retinal ganglion cell (RGC) replacement therapy could restore vision in glaucoma and other optic neuropathies. We developed a rapid protocol for directly induced RGC (iRGC) differentiation from human stem cells, leveraging overexpression of NGN2. Neuronal morphology and neurite growth were observed within 1week of induction; characteristic RGC-specific gene expression confirmed identity. Calcium imaging demonstrated gamma-aminobutyric acid (GABA)-induced excitation characteristic of immature RGCs. Single-cell RNA sequencing showed more similarities between iRGCs and early-stage fetal human RGCs than retinal organoid-derived RGCs. Intravitreally transplanted iRGCs survived and migrated into host retinas independent of prior optic nerve trauma, but iRGCs protected host RGCs from neurodegeneration. These data demonstrate rapid iRGC generation invitro into an immature cell with high similarity to human fetal RGCs and capacity for retinal integration after transplantation and neuroprotective function after optic nerve injury. The simplicity of this system may benefit translational studies on human RGCs.

  View details for DOI 10.1016/j.stemcr.2022.10.011

  View details for PubMedID 36368332

• Understanding RGC differentiation and development in retinal organoids by scRNA-seq Luo, Z., Chang, K., Tanasa, B., Goldberg, J. L. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2022

  View details for Web of Science ID 000844437006261

• Impact of Neurofibromatosis type 1 (NF1) heterozygosity on RGC death after optic nerve injury Xia, X., Sun, C., Luo, Z., Shyamsundar, S., Russano, K., Goldberg, J. L. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2022

  View details for Web of Science ID 000844401302314

• Distribution Of Primary Cilia In hESC-Derived Retinal Organoid Jarin, T., Ning, K., Luo, Z., Kowal, T., Li, B., Hu, Y., Wu, A. Y., Goldberg, J. L., Sun, Y. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2022

  View details for Web of Science ID 000844437004144

• Carbon Nanotube Polymer Scaffolds as a Conductive Alternative for the Construction of Retinal Sheet Tissue. ACS chemical neuroscience Yang, R., Yang, S., Li, K., Luo, Z., Xian, B., Tang, J., Ye, M., Lu, S., Zhang, H., Ge, J. 2021

  Abstract

  With the great success of graphene in the biomedical field, carbon nanotubes have attracted increasing attention for different applications in ophthalmology. Here, we report a novel retinal sheet composed of carbon nanotubes (CNTs) and poly(lactic-co-glycolic acid) (PLGA) that can enhance retinal cell therapy. By tuning our CNTs to regulate the mechanical characteristics of retina sheets, we were able to improve the in vitro viability of retinal ganglion cells derived from human-induced pluripotent stem cells incorporated into CNTs. Engrafted retinal ganglion cells displayed signs of regenerating processes along the optic nerve. Compared with PLGA scaffolds, CNT-PLGA retinal sheet tissue has excellent electrical conductivity, biocompatibility, and biodegradation. This new biomaterial offers new insight into retinal injury, repair, and regeneration.

  View details for DOI 10.1021/acschemneuro.1c00242

  View details for PubMedID 34375091

• Scaffolds Facilitate Epiretinal Transplantation of hiPSC-Derived Retinal Neurons in Nonhuman Primates. Acta biomaterialia Luo, Z., Xian, B., Li, K., Li, K., Yang, R., Chen, M., Xu, C., Tang, M., Rong, H., Hu, D., Ye, M., Yang, S., Lu, S., Zhang, H., Ge, J. 2021

  Abstract

  Transplantation of stem cell-derived retinal neurons is a promising regenerative therapy for optic neuropathy. However, significant anatomic differences compromise its efficacy in large animal models. The present study describes the procedure and outcomes of human-induced pluripotent stem cell (hiPSC)-derived retinal sheet transplantation in primate models using biodegradable materials. Stem cell-derived retinal organoids were seeded on polylactic-coglycolic acid (PLGA) scaffolds and directed toward a retinal ganglion cell (RGC) fate. The seeded tissues showed active proliferation, typical neuronal morphology, and electrical excitability. The cellular scaffolds were then epiretinally transplanted onto the inner surface of rhesus monkey retinas. With sufficient graft-host contact provided by the scaffold, the transplanted tissues survived for up to 1 year without tumorigenesis. Histological examinations indicated survival, further maturation, and migration. Moreover, green fluorescent protein-labeled axonal projections toward the host optic nerve were observed. Cryopreserved organoids were also able to survive and migrate after transplantation. Our results suggest the potential efficacy of RGC replacement therapy in the repair of optic neuropathy for the restoration of visual function.

  View details for DOI 10.1016/j.actbio.2021.07.040

  View details for PubMedID 34314890

• Rapid protocol for induced retinal ganglion cell differentiation from human stem cells Luo, Z., Chang, K., Tanasa, B., Wernig, M., Goldberg, J. L. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2021

  View details for Web of Science ID 000690761600554

• Retinal organoid differentiation and transplantatio Chang, K., Luo, Z., Xia, X., Knasel, C., Goldberg, J. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2020

  View details for Web of Science ID 000554495705088

• Islet1 and Brn3 Expression Pattern Study in Human Retina and hiPSC-Derived Retinal Organoid. Stem cells international Luo, Z., Xu, C., Li, K., Xian, B., Liu, Y., Li, K., Liu, Y., Rong, H., Tang, M., Hu, D., Yang, S., Ye, M., Zhong, X., Ge, J. 2019; 2019: 8786396

  Abstract

  This study was conducted to determine the dynamic Islet1 and Brn3 (POU4F) expression pattern in the human fetal retina and human-induced pluripotent stem cell- (hiPSC-) derived retinal organoid. Human fetal eyes from 8 to 27 fetal weeks (Fwks), human adult retina, hiPSC-derived retinal organoid from 7 to 31 differentiation weeks (Dwks), and rhesus adult retina were collected for cyrosectioning. Immunofluorescence analysis showed that Islet1 was expressed in retinal ganglion cells in the fetal retina, human adult retina, and retinal organoids. Unexpectedly, after Fwk 20, Brn3 expression gradually decreased in the fetal retina. In the midstage of development, Islet1 was detected in bipolar and developing horizontal cells. As the photoreceptor developed, the Islet1-positive cone precursors gradually became Islet1-negative/S-opsin-positive cones. This study highlights the distinguishing characteristics of Islet1 dynamic expression in human fetal retina development and proposes more concerns which should be taken regarding Brn3 as a cell-identifying marker in mature primate retina.

  View details for DOI 10.1155/2019/8786396

  View details for PubMedID 31885629

• Dexamethasone Provides Effective Immunosuppression for Improved Survival of Retinal Organoids after Epiretinal Transplantation. Stem cells international Xian, B. n., Luo, Z. n., Li, K. n., Li, K. n., Tang, M. n., Yang, R. n., Lu, S. n., Zhang, H. n., Ge, J. n. 2019; 2019: 7148032

  Abstract

  We investigated the efficacy of the immunosuppressants rapamycin (RAP) and dexamethasone (DEX) in improving the survival of retinal organoids after epiretinal transplantation. We first compared the immunosuppressive abilities of DEX and RAP in activated microglia in an in vitro setting. Following this, we used immunofluorescence, real-time polymerase chain reaction, and flow cytometry to investigate the effects of DEX and RAP on cells in the retinal organoids. Retinal organoids were then seeded onto poly(lactic-co-glycolic) acid (PLGA) scaffolds and implanted into rhesus monkey eyes (including a healthy individual and three monkeys with chronic ocular hypertension (OHT) induction) and subjected to different post-operative immunosuppressant treatments; 8 weeks after the experiment, histological examinations were carried out to assess the success of the different treatments. Our in vitro experiments indicated that both DEX and RAP treatments were equally effective in suppressing microglial activity. Although both immunosuppressants altered the morphologies of cells in the retinal organoids and caused a slight decrease in the differentiation of cells into retinal ganglion cells, the organoid cells retained their capacity to grow and differentiate into retinal tissues. Our in vivo experiments indicate that the retinal organoid can survive and differentiate into retinal tissues in a healthy rhesus monkey eye without immunosuppressive treatment. However, the survival and differentiation of these organoids in OHT eyes was successful only with the DEX treatment. RAP treatment was ineffective in preventing immunological rejection, and the retinal organoid failed to survive until the end of 8 weeks. DEX is likely a promising immunosuppressant to enhance the survival of epiretinal implants.

  View details for DOI 10.1155/2019/7148032

  View details for PubMedID 31428159

  View details for PubMedCentralID PMC6683795

• BAM15 attenuates transportation-induced apoptosis in iPS-differentiated retinal tissue. Stem cell research & therapy Tang, M. n., Luo, Z. n., Wu, Y. n., Zhuang, J. n., Li, K. n., Hu, D. n., Rong, H. n., Xian, B. n., Ge, J. n. 2019; 10 (1): 64

  Abstract

  BAM15 is a novel mitochondrial protonophore uncoupler capable of protecting mammals from acute renal ischemic-reperfusion injury and cold-induced microtubule damage. The purpose of our study was to investigate the effect of BAM15 on apoptosis during 5-day transportation of human-induced pluripotent stem (hiPS)-differentiated retinal tissue.Retinal tissues of 30 days and 60 days were transported with or without BAM15 for 5 days in the laboratory or by real express. Immunofluorescence staining of apoptosis marker cleaved caspase3, proliferation marker Ki67, and neural axon marker NEFL was performed. And expression of apoptotic-related factors p53, NFkappaB, and TNF-a was detected by real-time PCR. Also, location of ganglion cells, photoreceptor cells, amacrine cells, and precursors of neuronal cell types in retinal tissue was stained by immunofluorescence after transportation. Furthermore, cell viability was assessed by CCK8 assay.Results showed transportation remarkably intensified expression of apoptotic factor cleaved caspase3, p53, NFkappaB, and TNF-a, which could be reduced by supplement of BAM15. In addition, neurons were severely injured after transportation, with axons manifesting disrupted and tortuous by staining NEFL. And the addition of BAM15 in transportation was able to protect neuronal structure and increase cell viability without affecting subtypes cells location of retinal tissue.BAM15 might be used as a protective reagent on apoptosis during transporting retinal tissues, holding great potential in research and clinical applications.

  View details for DOI 10.1186/s13287-019-1151-y

  View details for PubMedID 30795805

  View details for PubMedCentralID PMC6387563

• Alpha 1-antitrypsin inhibits microglia activation and facilitates the survival of iPSC grafts in hypertension mouse model. Cellular immunology Yang, S., Xian, B., Li, K., Luo, Z., Liu, Y., Hu, D., Ge, J. 2018; 328: 49-57

  Abstract

  This study was conducted to investigate the use of Alpha 1-antitrypsin (AAT) to inhibit microglia activation in chronic hypertension model and provide a permissive environment for stem cell transplantation. Chronic ocular hypertension of C57BL/6 mice using magnetic microbead injection was induced 3 weeks prior to iPSCs transplantation. The ocular hypertension model was assessed histologically and intraocular pressure was measured. Survival of grafted cells and microglia activation were examined by flow cytometry and immunofluorescence in AAT and PBS treated hosts. Retinal cytokines expression was also detected by real-time PCR. Chronic ocular hypertension resulted in persistent microglia activation and stem cell grafts loss. AAT treatment significantly inhibited microglia activation and facilitated the survival of transplant iPSCs 4w post transplantation compared to PBS treatment. AAT holds tremendous potential for the clinical application to control neuroinflammation factor in glaucoma and improve the stem cell replacement therapy of retinal neurodegenerative disease.

  View details for DOI 10.1016/j.cellimm.2018.03.006

  View details for PubMedID 29573789

• Establishing a Surgical Procedure for Rhesus Epiretinal Scaffold Implantation with HiPSC-Derived Retinal Progenitors. Stem cells international Luo, Z. n., Li, K. n., Li, K. n., Xian, B. n., Liu, Y. n., Yang, S. n., Xu, C. n., Fan, Z. n., Lu, S. n., Zhang, H. n., Ge, J. n. 2018; 2018: 9437041

  Abstract

  To develop an effective surgical procedure for cellular scaffold epiretinal implantation in rhesus, facilitating subsequent epiretinal stem cell transplantation.Retinal progenitors were seeded onto a poly(lactic-co-glycolic) acid (PLGA) scaffold. First, the cellular scaffolds were delivered by 18G catheter or retinal forceps into rabbit epiretinal space (n = 50). Then, the cell survival rate was evaluated by Cell Counting Kit-8 (CCK-8). Second, three methods of scaffold fixation, including adhesion after gas-liquid exchange (n = 1), tamponade by hydrogel (n = 1), and fixation by retinal tacks (n = 4), were performed in rhesus monkeys. After one month, fundus photography and SD-OCT were performed to assess the outcomes, and histological examination was performed to evaluate proliferation.The cell survival rate was significantly higher in the catheter group. Follow-up examination showed that retinal tack fixation was the only method to maintain the scaffolds attached to host retina for at least 3 weeks, which is the minimal time required for cell integration. Histological staining demonstrated slight glial fibrillary acidic protein (GFAP) accumulation in the retinal tack insertion area.The established surgical procedure offers a new insight into research of epiretinal cell replacement therapy in rhesus eyes. The successful delivery and long-term fixation provide a prerequisite for cell migration and integration.

  View details for DOI 10.1155/2018/9437041

  View details for PubMedID 29760741

  View details for PubMedCentralID PMC5924980

• An Optimized System for Effective Derivation of Three-Dimensional Retinal Tissue via Wnt Signaling Regulation. Stem cells (Dayton, Ohio) Luo, Z. n., Zhong, X. n., Li, K. n., Xie, B. n., Liu, Y. n., Ye, M. n., Li, K. n., Xu, C. n., Ge, J. n. 2018; 36 (11): 1709–22

  Abstract

  Effective derivation of three-dimensional (3D) retinal tissue from human-induced pluripotent stem cells (hiPSCs) could provide models for drug screening and facilitate patient-specific retinal cell replacement therapy. However, some hiPSC lines cannot undergo 3D self-organization and show inadequate differentiation efficiency to meet clinical demand. In this study, we developed an optimized system for derivation of 3D retinal tissue. We found that the Wnt signaling pathway antagonist Dickkopf-related protein 1 (DKK-1) rescued the inability of differentiated retinal progenitors to self-organize. By evaluating DKK-1 expression and supplying DKK-1 if necessary, retinal organoids were differentiated from six hiPSC lines, which were reprogramed from three common initiating cell types. Retinal tissues derived from the optimized system were well organized and capable of surviving for further maturation. Thus, using this system, we generated retinal tissues from various hiPSC lines with high efficiency. This novel system has many potential applications in regenerative therapy and precision medicine. Stem Cells 2018;36:1709-1722.

  View details for DOI 10.1002/stem.2890

  View details for PubMedID 29999566

• HiPSC-derived retinal ganglion cells grow dendritic arbors and functional axons on a tissue-engineered scaffold. Acta biomaterialia Li, K., Zhong, X., Yang, S., Luo, Z., Li, K., Liu, Y., Cai, S., Gu, H., Lu, S., Zhang, H., Wei, Y., Zhuang, J., Zhuo, Y., Fan, Z., Ge, J. 2017; 54: 117-127

  Abstract

  Numerous therapeutic procedures in modern medical research rely on the use of tissue engineering for the treatment of retinal diseases. However, the cell source and the transplantation method are still a limitation. Previously, it was reported that a self-organizing three-dimensional neural retina can be induced from human-induced pluripotent stem cells (hiPSCs). In this study, we disclose the generation of retinal ganglion cells (RGCs) from the neural retina and their seeding on a biodegradable poly (lactic-co-glycolic acid) (PLGA) scaffold to create an engineered RGC-scaffold biomaterial. Moreover, we explored the dendritic arbor, branching point, functional axon and action potential of the biomaterial. Finally, the cell-scaffold was transplanted into the intraocular environment of rabbits and rhesus monkeys.As a part of the mammalian central nervous system (CNS), the retinal ganglion cell (RGC) shows little regenerative capacity. With the use of medical biomaterial for cells seeding and deliver, a new domain is now emerging that uses tissue engineering therapy for retinal disease. However, previous studies utilized RGCs from rodent model, which has limitations for human disease treatment. In the present study, we generated RGCs from hiPSCs-3D neural retina and then seeded these RGCs on PLGA scaffold to create an engineered RGC-scaffold biomaterial. Moreover, we assessed the transplantation method for biomaterial in vivo. Our study provides a technique to produce the engineered human RGC-scaffold biomaterial.

  View details for DOI 10.1016/j.actbio.2017.02.032

  View details for PubMedID 28216299