Masayo Takahashi MD, PhD | Vision Care Inc. & Kobe City Eye Hospital, Japan

Background

Fourteen years have passed since the publication of the human iPS cell paper in 2007 by Prof. Yamanaka’s group. During that time, we conducted the first clinical study of iPS cells, that was autologous iPS cell-derived retinal pigment epithelial cell (iPSC-RPE) sheet transplantation, from 2013 to 2015 and the second clinical study of HLA-matched allogeneic iPSC-RPE suspension transplantation from 2017 to 2019. More than 10 different clinical studies using iPS cells in Japan, including a clinical trial for Parkinson’s disease (dopamine cells), corneal epithelial cells, cardiac muscle sheets, spinal cord transplantation, and platelets has been approved. The fact that clinical applications are progressing at such a rapid pace is largely due to the environment and legal framework that is suitable for regenerative therapies, that is completely different from small molecule drugs.

The 1st Clinical study – autologous iPSC-RPE sheet transplantation (1 case)

In 2012 we were the first in the world to submit a clinical research protocol using human iPS cells to the “Review Committee on Human Stem Cell Clinical Research”. For autologous transplantation, it was necessary to be certain that (1) iPSCs of clinically usable quality could be generated from any case and (2) safe, mature iPSC-RPE could always be produced with a purity of almost 100% (effectively 99% or more = no more than 500 non-purpose cells mixed in the final product). Based on many years of experience in stem cell research, we thought that the special characteristics of RPE would be advantageous to make the cells absolutely safe nevertheless, given that stem cells are always a heterogeneous population even if they look the same and that cell culture is always subject to genetic mutation at each pass. Conversely, if the above two conditions can be achieved, RPEs can be used safely even if the slightest genetic mutation occurs, as far as RPEs with no reports of primary metastatic tumors are concerned. During the five years since the announcement of human iPS cells, we repeatedly verified the safety of the final product by modifying the differentiation method that had been established for human ES cells. Once we were confident of the safety of the final product, we applied for a protocol. In parallel with the scientific verification, we decided on the direction of the clinical study while keeping abreast of the direction and trends of the new regenerative medicine law that was being drafted. Finally, the cell risk was reduced to almost zero, and the cells went into clinical trials with only a few percent surgical risk, just as in ordinary retinal surgery.

Accumulation of information for risk

The reason why we can draw such a clear line of safety is because we performed a long period of stem cell research and also knew well about RPE cells’ behavior in the eye through ophthalmic surgery. In other words, we knew the risks from basic research to clinical surgery, so that we could consider the risk matrix (significance and frequency of the risks) and apply the clinical study. The risks associated with RPE transplantation and the degree of efficacy in each case can be predicted to some extent based on our experience in treating age-related macular degeneration (AMD).

The RPE, which has been working almost irreplaceably in the eye for its entire life, has, as expected, survived in the same sheet shape even 5 years after transplantation, maintaining the photoreceptor cells only in the area where the transplanted sheet exists. Retinal photoreceptor cells will always degenerate without pigment epithelium, so it is clear that the iPSC-RPE is still functioning in this case. Before the surgery, her visual acuity deteriorated although she got 13 times anti-VEGF injection. After the surgery her visual acuity is still stable.

We planned two cases for the first clinical study, but different from the first case, the patient’s visual acuity in the second case had stabilized at 0.3 during the 10 months of cell preparation, so that the risk of surgery (visual acuity loss) was greater than in the first case, so we decided to stop RPE transplantation, although the safety was confirmed by intensive tumorigenicity tests.

Thus, only one case, but we showed the safe way to use the iPS cells, that many people thought dangerous to use for humans at that time.

The 2nd Clinical Study – HLA-matched allogeneic iPSC-RPE suspension transplantation (5 cases)

The reason why we first started with autologous sheet transplants, which are technically difficult, was to do the scientifically best treatment for the first iPS application. Even though it is costly and time-consuming, autologous transplantation is the best treatment from the viewpoint of immunity. We think the autologous transplantation will eventually become an option, but for the time being, allogeneic transplantation, which can be applied to many people by preparing several types of cells, is desired as the actual standard treatments.

CiRA in Kyoto University produced HLA6 homozygous iPS cells, which covered 17% of the Japanese population, so that we conducted clinical research on HLA-matched allogeneic transplants using this iPS cell line. The reason for using iPS cells in the development of a treatment for age-related macular degeneration, which had originally been conducted using ES cells, was because for the elder patients to use immunosuppressive drugs in the treatment of age-related macular degeneration, in which more than 70% of patients are over 70 years old, was not desirable. In fact, many of the side effects of retinal cell transplantation that have been reported in papers are caused by immunosuppressive drugs.³

The manufacturing method of RPE is the same, but in the second clinical study, the target was milder cases than the first clinical study and a suspension transplant was used. In those cases, RPE atrophy is often sparse and a large sheet is not necessary. It is less invasive and safer to inject the suspension through a small hole in the retina into the back of the retina (subretinal) to form a sheet within the eye to control the neovascularization.

Immune response – LGIR test

The primary endpoint of the second clinical study, as in the first study, was safety, but the key point was whether the rejection of HLA-matched allogeneic transplants could be suppressed by local treatment alone without systemic immune suppression. For this purpose, we created a test that can examine the patient’s immune response in real time by examining the proliferation of mononuclear cells in mixed culture of patient peripheral blood mononuclear cells and transplanted RPE cells, which we call the LGIR (Lymph Graft Immune Reaction) test⁴ (Figure 4). This test was very useful in clinical studies, and in the first case, we were able to confirm that the appearance of a small amount of subretinal fluid (a few µl) at 5 weeks after transplantation indicated the beginning of rejection, not the recurrence of the disease, which was suppressed by three injections of steroid outside the eye. In the other cases, there was no sign of rejection in this test or in the clinical findings during the one-year follow-up, and all five cases showed successful implantation of the transplanted RPE. In the case with the best control of the implantation site, polarized OCT showed that the transplanted cells were covering the neovascularization in a sheet-like pattern.⁵

In other cases, unlike in the animal model, it proved difficult to control the location of grafted suspension, and in all cases, epiretinal membrane formation was observed due to backflow of cells from the injection site. In one of the cases, retinal edema appeared, and surgery was performed to remove the epiretinal membrane, a procedure that is frequently performed in ophthalmology, with no serious effects on visual function. These clinical experiences were immediately returned to the laboratory to improve the RPE formulation and surgical technique. We now reached to the RPE strip, that can be transplanted from a small hole in the retina and expand to form small sheets beneath the retina several weeks after the surgery. We think this is the safest and most effective formulation of RPE.

Learning from Clinical Research – Effectiveness

We have learned a great deal from our experience with these six cases in total. There are good reasons for the RPE to be the first case of iPS application to humans. In most of the cell types, it is difficult to induce differentiation of pluripotent stem cells to functionally mature cells in a culture dish, but RPE is one of the few cells that can be differentiated to mature cells and transplanted with almost the same properties as in vivo. Furthermore, the inner layer of the eye cup during the development in the fetus becomes the neural retina and the outer layer becomes the RPE, and the neural retina and RPE have no cell adhesive apparatus but just contact with each other. Thus, monolayer of RPE cells with a basement membrane is already a tissue, and the RPE sheet is a complete tissue graft. In other words, it is perfect tissue equivalent to the native RPE in the body.

Furthermore, the safety was obtained relatively easily because of the RPE that does not form tumors even with any gene mutations. These are the unique features of RPE that distinguish it from other tissues.

Thus, we proved the safety of autologous and allogeneic iPS-RPE. Now we move to the next step to evaluate the effectiveness of the RPE transplantation. For this, it is important to select the appropriate cases even in the same diseases while various conditions are referred to by a single disease name. The effect of the treatment is not determined by RPE cells that already have perfect function, but by patients’ retinal environment, where the morphology and functions of cells change drastically depending on the microenvironment. That is the most different point of cell therapy compared to the current treatment. In regenerative medicine, the end products do not immediately make treatments like small molecule drugs.

Retinal Organoids Transplantation – photoreceptor replacement

In 2011, Dr. Yoshiki Sasai’s group invented a method called “organoids” to create a three-dimensional retina from ES or iPS cells. Since then, organoid research is flourishing for various parts of the body in the world. Again, in the case of photoreceptor cells, the world’s first organoid transplantation was done in the field of retina.

Dr. Michiko Mandai in our group worked diligently to create the POC. It took about 7 years to prove 1) whether the grafted immature retinal organoid sheets would mature and become a photoreceptor cell, 2) whether it would re-create synapses with the retina in the body, 3) whether it would function electrophysiologically, and 4) whether the photoreceptor degenerated model animals would be able to see the light after transplantation. After obtaining these POC in the animal, the Kobe City Eye Hospital performed the retinal sheet (photoreceptor) transplantation clinical study with two patients. The hospital reported the results at the Japanese Society of Clinical Ophthalmology meeting last month. In both cases, photoreceptor cells were successfully transplanted into the patient without any abnormal proliferation and survived for more than one year with some hint of functional recovery.

In ophthalmology, microstructures such as OCT are observed to make decisions on pathological conditions and treatment. In replacement therapy in ophthalmology, anatomical effects (tissue reconstruction) are used as an indicator of effectiveness. We hope that we will be able to show the effect in the near future.

Conclusion

Surgical treatment has gone through a process of trial and error in its history, and at one point, through the ingenuity of many doctors, it was perfected. Having witnessed the development of cataract surgery these past 30 years, I think that regenerative medicine will trace the same course. Since safe and functional cells have been created, many doctors should be involved to accomplish the effective new treatment. A suitable path for regenerative medicine is necessary to deliver effective treatments to patients quickly and inexpensively.

References

1. Takahashi M, Palmer TD, Takahashi J, Gage FH. Widespread integration and survival of adult-derived neural progenitor cells in the developing optic retina. Mol. Cell. Neurosci. 12:340-348, 1998.
2. Kawasaki H, Suemori H, Mizuseki K, Watanabe K, Urano F, Ichinose H, Haruta M, Takahashi M, Yoshikawa K, Nishikawa S, Nakatsuji N, Sasai Y. Generation of dopaminergic neurons and pigmented epithelia from primate ES cells by stromal cell-derived inducing activity. Proc Natl Acad Sci U S A. 2002 Feb 5;99(3):1580-5. doi: 10.1073/pnas.032662199. Epub 2002 Jan 29.
3. Haruta M, Sasai Y, Kawasaki H, Amemiya K, Ooto S, Kitada M, Suemori H, Nakatsuji N, Ide C, Honda Y, Takahashi M. In vitro and in vivo characterization of pigment epithelial cells differentiated from primate embryonic stem cells. Invest Ophthalmol Vis Sci. 2004 Mar;45(3):1020-5. doi: 10.1167/iovs.03-1034.
4. Ikeda H, Osakada F, Watanabe K, Mizuseki K, Haraguchi T, Miyoshi H, Kamiya D, Honda Y, Sasai N, Yoshimura N, Takahashi M, Sasai Y. Generation of Rx+/Pax6+ neural retinal precursors from embryonic stem cells. Proc Natl Acad Sci U S A. 2005 Aug 9;102(32):11331-6. doi: 10.1073/pnas.0500010102. Epub 2005 Aug 2.
5. Osakada F, Ikeda H, Mandai M, Wataya T, Watanabe K, Yoshimura N, Akaike A, Sasai Y, Takahashi M. Toward the generation of rod and cone photoreceptors from mouse, monkey and human embryonic stem cells. Nat Biotechnol. 2008 Feb;26(2):215-24. doi: 10.1038/nbt1384. Epub 2008 Feb 3.
6. Hirami Y, Osakada F, Takahashi K, Okita K, Yamanaka S, Ikeda H, Yoshimura N, Takahashi M. Generation of retinal cells from mouse and human induced pluripotent stem cells. Neurosci Lett. 2009 Jul 24;458(3):126-31. doi: 10.1016/j.neulet.2009.04.035. Epub 2009 Apr 18.
7. Osakada F, Ikeda H, Sasai Y, Takahashi M. Stepwise differentiation of pluripotent stem cells into retinal cells. Nat Protoc. 2009;4(6):811-24. doi: 10.1038/nprot.2009.51. Epub 2009 May 7.
8. Jin ZB, Okamoto S, Osakada F, Homma K, Assawachananont J, Hirami Y, Iwata T, Takahashi M. Modeling retinal degeneration using patient-specific induced pluripotent stem cells. PLoS One. 2011 Feb 10;6(2):e17084. doi: 10.1371/journal.pone.0017084.
9. Mandai M, Homma K, Okamoto S, Yamada C, Nomori A, Takahashi M. Adequate Time Window and Environmental Factors Supporting Retinal Graft Cell Survival in rd Mice. Cell Med. 2012 Apr 20;4(1):45-54. doi: 10.3727/215517912X639315. eCollection 2012 Jan.
10. Kanemura H, Go MJ, Nishishita N, Sakai N, Kamao H, Sato Y, Takahashi M, Kawamata S. Pigment epithelium-derived factor secreted from retinal pigment epithelium facilitates apoptotic cell death of iPSC. Sci Rep. 2013;3:2334. doi: 10.1038/srep02334.
11. Maeda T, Lee MJ, Palczewska G, Marsili S, Tesar PJ, Palczewski K, Takahashi M, Maeda A. Retinal pigmented epithelial cells obtained from human induced pluripotent stem cells possess functional visual cycle enzymes in vitro and in vivo. J Biol Chem. 2013 Nov 29;288(48):34484-93. doi: 10.1074/jbc.M113.518571. Epub 2013 Oct 15.
12. Kanemura H, Go MJ, Shikamura M, Nishishita N, Sakai N, Kamao H, Mandai M, Morinaga C, Takahashi M, Kawamata S. Tumorigenicity studies of induced pluripotent stem cell (iPSC)-derived retinal pigment epithelium (RPE) for the treatment of age-related macular degeneration. PLoS One. 2014 Jan 14;9(1):e85336. doi: 10.1371/journal.pone.0085336. eCollection 2014.
13. Kamao H, Mandai M, Okamoto S, Sakai N, Suga A, Sugita S, Kiryu J, Takahashi M. Characterization of human induced pluripotent stem cell-derived retinal pigment epithelium cell sheets aiming for clinical application. Stem Cell Reports. 2014 Jan 23;2(2):205-18. doi: 10.1016/j.stemcr.2013.12.007. eCollection 2014 Feb 11.
14. Assawachananont J, Mandai M, Okamoto S, Yamada C, Eiraku M, Yonemura S, Sasai Y, Takahashi M. Transplantation of embryonic and induced pluripotent stem cell-derived 3D retinal sheets into retinal degenerative mice. Stem Cell Reports. 2014 Apr 24;2(5):662-74. doi: 10.1016/j.stemcr.2014.03.011. eCollection 2014 May 6.
15. Sugita S, Kamao H, Iwasaki Y, Okamoto S, Hashiguchi T, Iseki K, Hayashi N, Mandai M, Takahashi M. Inhibition of T-cell activation by retinal pigment epithelial cells derived from induced pluripotent stem cells. Invest Ophthalmol Vis Sci. 2015 Jan 20;56(2):1051-62. doi: 10.1167/iovs.14-15619.
16. Sun J, Mandai M, Kamao H, Hashiguchi T, Shikamura M, Kawamata S, Sugita S, Takahashi M. Protective Effects of Human iPS-Derived Retinal Pigmented Epithelial Cells in Comparison with Human Mesenchymal Stromal Cells and Human Neural Stem Cells on the Degenerating Retina in rd1 mice. Stem Cells. 2015 May;33(5):1543-53. doi: 10.1002/stem.1960.
17. Kawamata S, Kanemura H, Sakai N, Takahashi M, Go MJ. Design of a Tumorigenicity Test for Induced Pluripotent Stem Cell (iPSC)-Derived Cell Products. J Clin Med. 2015 Jan 14;4(1):159-71. doi: 10.3390/jcm4010159.
18. Shirai H, Mandai M, Matsushita K, Kuwahara A, Yonemura S, Nakano T, Assawachananont J, Kimura T, Saito K, Terasaki H, Eiraku M, Sasai Y, Takahashi M. Transplantation of human embryonic stem cell-derived retinal tissue in two primate models of retinal degeneration. Proc Natl Acad Sci U S A. 2016 Jan 5;113(1):E81-90. doi: 10.1073/pnas.1512590113. Epub 2015 Dec 22.
19. Daley GQ, Hyun I, Apperley JF, Barker RA, Benvenisty N, Bredenoord AL, Breuer CK, Caulfield T, Cedars MI, Frey-Vasconcells J, Heslop HE, Jin Y, Lee RT, McCabe C, Munsie M, Murry CE, Piantadosi S, Rao M, Rooke HM, Sipp D, Studer L, Sugarman J, Takahashi M, Zimmerman M, Kimmelman J. Setting Global Standards for Stem Cell Research and Clinical Translation: The 2016 ISSCR Guidelines. Stem Cell Reports. 2016 Jun 14;6(6):787-797. doi: 10.1016/j.stemcr.2016.05.001. Epub 2016 May 12.
20. Fujii M, Sunagawa GA, Kondo M, Takahashi M, Mandai M. Evaluation of Micro Electroretinograms Recorded with Multiple Electrode Array to Assess Focal Retinal Function. Sci Rep. 2016 Aug 2;6:30719. doi: 10.1038/srep30719.
21. Sugita S, Iwasaki Y, Makabe K, Kimura T, Futagami T, Suegami S, Takahashi M. Lack of T Cell Response to iPSC-Derived Retinal Pigment Epithelial Cells from HLA Homozygous Donors. Stem Cell Reports. 2016 Oct 11;7(4):619-634. doi: 10.1016/j.stemcr.2016.08.011. Epub 2016 Sep 15.
22. Sugita S, Iwasaki Y, Makabe K, Kamao H, Mandai M, Shiina T, Ogasawara K, Hirami Y, Kurimoto Y, Takahashi M. Successful Transplantation of Retinal Pigment Epithelial Cells from MHC Homozygote iPSCs in MHC-Matched Models. Stem Cell Reports. 2016 Oct 11;7(4):635-648. doi: 10.1016/j.stemcr.2016.08.010. Epub 2016 Sep 15.
23. Mandai M, Fujii M, Hashiguchi T, Sunagawa GA, Ito SI, Sun J, Kaneko J, Sho J, Yamada C, Takahashi M. iPSC-Derived Retina Transplants Improve Vision in rd1 End-Stage Retinal-Degeneration Mice. Stem Cell Reports. 2017 Feb 14;8(2):489. doi: 10.1016/j.stemcr.2017.01.018.
24. Mandai M, Watanabe A, Kurimoto Y, Hirami Y, Morinaga C, Daimon T, Fujihara M, Akimaru H, Sakai N, Shibata Y, Terada M, Nomiya Y, Tanishima S, Nakamura M, Kamao H, Sugita S, Onishi A, Ito T, Fujita K, Kawamata S, Go MJ, Shinohara C, Hata KI, Sawada M, Yamamoto M, Ohta S, Ohara Y, Yoshida K, Kuwahara J, Kitano Y, Amano N, Umekage M, Kitaoka F, Tanaka A, Okada C, Takasu N, Ogawa S, Yamanaka S, Takahashi M. Autologous Induced Stem-Cell-Derived Retinal Cells for Macular Degeneration. N Engl J Med. 2017 Mar 16;376(11):1038-1046. doi: 10.1056/NEJMoa1608368.
25. Mandai M, Fujii M, Hashiguchi T, Sunagawa GA, Ito SI, Sun J, Kaneko J, Sho J, Yamada C, Takahashi M. iPSC-Derived Retina Transplants Improve Vision in rd1 End-Stage Retinal-Degeneration Mice. Stem Cell Reports. 2017 Apr 11;8(4):1112-1113. doi: 10.1016/j.stemcr.2017.03.024.
26. Iraha S, Tu HY, Yamasaki S, Kagawa T, Goto M, Takahashi R, Watanabe T, Sugita S, Yonemura S, Sunagawa GA, Matsuyama T, Fujii M, Kuwahara A, Kishino A, Koide N, Eiraku M, Tanihara H, Takahashi M, Mandai M. Establishment of Immunodeficient Retinal Degeneration Model Mice and Functional Maturation of Human ESC-Derived Retinal Sheets after Transplantation. Stem Cell Reports. 2018 Mar 13;10(3):1059-1074. doi: 10.1016/j.stemcr.2018.01.032. Epub 2018 Mar 1.
27. Sugita S, Makabe K, Fujii S, Takahashi M. Detection of Complement Activators in Immune Attack Eyes After iPS-Derived Retinal Pigment Epithelial Cell Transplantation. Invest Ophthalmol Vis Sci. 2018 Aug 1;59(10):4198-4209. doi: 10.1167/iovs.18-24769.
28. Tu HY, Watanabe T, Shirai H, Yamasaki S, Kinoshita M, Matsushita K, Hashiguchi T, Onoe H, Matsuyama T, Kuwahara A, Kishino A, Kimura T, Eiraku M, Suzuma K, Kitaoka T, Takahashi M, Mandai M. Medium- to long-term survival and functional examination of human iPSC-derived retinas in rat and primate models of retinal degeneration. EBioMedicine. 2019 Jan;39:562-574. doi: 10.1016/j.ebiom.2018.11.028. Epub 2018 Nov 28.
29. Akiba R, Matsuyama T, Tu HY, Hashiguchi T, Sho J, Yamamoto S, Takahashi M, Mandai M. Quantitative and Qualitative Evaluation of Photoreceptor Synapses in Developing, Degenerating and Regenerating Retinas. Front Cell Neurosci. 2019 Feb 11;13:16. doi: 10.3389/fncel.2019.00016. eCollection 2019.
30. Takagi S, Mandai M, Gocho K, Hirami Y, Yamamoto M, Fujihara M, Sugita S, Kurimoto Y, Takahashi M. Evaluation of Transplanted Autologous Induced Pluripotent Stem Cell-Derived Retinal Pigment Epithelium in Exudative Age-Related Macular Degeneration. Ophthalmol Retina. 2019 Oct;3(10):850-859. doi: 10.1016/j.oret.2019.04.021. Epub 2019 Apr 26.
31. Kuwahara A, Yamasaki S, Mandai M, Watari K, Matsushita K, Fujiwara M, Hori Y, Hiramine Y, Nukaya D, Iwata M, Kishino A, Takahashi M, Sasai Y, Kimura T. Preconditioning the Initial State of Feeder-free Human Pluripotent Stem Cells Promotes Self-formation of Three-dimensional Retinal Tissue. Sci Rep. 2019 Dec 12;9(1):18936. doi: 10.1038/s41598-019-55130-w.
32. Kuwahara A, Yamasaki S, Mandai M, Watari K, Matsushita K, Fujiwara M, Hori Y, Hiramine Y, Nukaya D, Iwata M, Kishino A, Takahashi M, Sasai Y, Kimura T. Publisher Correction: Preconditioning the Initial State of Feeder-free Human Pluripotent Stem Cells Promotes Self-formation of Three-dimensional Retinal Tissue. Sci Rep. 2020 Feb 5;10(1):2237. doi: 10.1038/s41598-020-58892-w.
33. Sugita S, Mandai M, Hirami Y, Takagi S, Maeda T, Fujihara M, Matsuzaki M, Yamamoto M, Iseki K, Hayashi N, Hono A, Fujino S, Koide N, Sakai N, Shibata Y, Terada M, Nishida M, Dohi H, Nomura M, Amano N, Sakaguchi H, Hara C, Maruyama K, Daimon T, Igeta M, Oda T, Shirono U, Tozaki M, Totani K, Sugiyama S, Nishida K, Kurimoto Y, Takahashi M.HLA-Matched Allogeneic iPS Cells-Derived RPE Transplantation for Macular Degeneration. J Clin Med. 2020 Jul 13;9(7):2217. doi: 10.3390/jcm9072217.
34. Sugita S, Futatsugi Y, Ishida M, Edo A, Takahashi M. Retinal Pigment Epithelial Cells Derived from Induced Pluripotent Stem (iPS) Cells Suppress or Activate T Cells via Costimulatory Signals. Int J Mol Sci. 2020 Sep 5;21(18):6507. doi: 10.3390/ijms21186507.
35. Yamanari M, Mase M, Obata R, Matsuzaki M, Minami T, Takagi S, Yamamoto M, Miyamoto N, Ueda K, Koide N, Maeda T, Totani K, Aoki N, Hirami Y, Sugiyama S, Mandai M, Aihara M, Takahashi M, Kato S, Kurimoto Y. Melanin concentration and depolarization metrics measurement by polarization-sensitive optical coherence tomography. Sci Rep. 2020 Nov 11;10(1):19513. doi: 10.1038/s41598-020-76397-4.
36. Sugita S, Mandai M, Kamao H, Takahashi M. Immunological aspects of RPE cell transplantation. Prog Retin Eye Res. 2021 Sep;84:100950. doi: 10.1016/j.preteyeres.2021.100950. Epub 2021 Jan 19.
37. Matsuyama T, Tu HY, Sun J, Hashiguchi T, Akiba R, Sho J, Fujii M, Onishi A, Takahashi M, Mandai M. Genetically engineered stem cell-derived retinal grafts for improved retinal reconstruction after transplantation. iScience. 2021 Jul 16;24(8):102866. doi: 10.1016/j.isci.2021.102866. eCollection 2021 Aug 20.
38. Nishida M, Tanaka Y, Tanaka Y, Amaya S, Tanaka N, Uyama H, Masuda T, Onishi A, Sho J, Yokota S, Takahashi M, Mandai M. Human iPS cell derived RPE strips for secure delivery of graft cells at a target place with minimal surgical invasion. Sci Rep. 2021 Nov 2;11(1):21421. doi: 10.1038/s41598-021-00703-x.
39. Motozawa N, Miura T, Ochiai K, Yamamoto M, Horinouchi T, Tsuzuki T, Kanda GN, Ozawa Y, Tsujikawa A, Takahashi K, Takahashi M, Kurimoto Y, Maeda T, Mandai M. Automated evaluation of retinal pigment epithelium disease area in eyes with age-related macular degeneration. Sci Rep. 2022 Jan 18;12(1):892. doi: 10.1038/s41598-022-05006-3.
40. Kanda GN, Tsuzuki T, Terada M, Sakai N, Motozawa N, Masuda T, Nishida M, Watanabe CT, Higashi T, Horiguchi SA, Kudo T, Kamei M, Sunagawa GA, Matsukuma K, Sakurada T, Ozawa Y, Takahashi M, Takahashi K, Natsume T. Robotic search for optimal cell culture in regenerative medicine. Elife. 2022 Jun 28;11:e77007. doi: 10.7554/eLife.77007.