Invest Ophthalmol Vis Sci. 2018 May 1;59(6):2470-2477. doi: 10.1167/iovs.18-24096.
RGC Neuroprotection Following Optic Nerve Trauma Mediated By Intranasal Delivery of Amnion Cell Secretome.
Grinblat GA1, Khan RS2, Dine K2, Wessel H3, Brown L3, Shindler KS2.
Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, United States.
Scheie Eye Institute and F.M. Kirby Center for Molecular Ophthalmology, University of Pennsylvania, Philadelphia, Pennsylvania, United States.
Noveome Biotherapeutics, Inc., Pittsburgh, Pennsylvania, United States.
Intranasally delivered ST266, the biological, proteinaceous secretome of amnion-derived multipotent progenitor cells, reduces retinal ganglion cell (RGC) loss, optic nerve inflammation, and demyelination in experimental optic neuritis. This unique therapy and novel administration route delivers numerous cytokines and growth factors to the eye and optic nerve, suggesting a potential to also treat other optic neuropathies. Thus, ST266-mediated neuroprotection was examined following traumatic optic nerve injury.
Optic nerve crush injury was surgically induced in C57BL/6J mice. Mice were treated daily with intranasal PBS or ST266. RGC function was assessed by optokinetic responses (OKRs), RGCs were counted, and optic nerve sections were stained with luxol fast blue and anti-neurofilament antibodies to assess myelin and RGC axon damage.
Intranasal ST266 administered daily for 5 days, beginning at the time that a 1-second optic nerve crush was performed, significantly attenuated OKR decreases. Furthermore, ST266 treatment reduced damage to RGC axons and myelin within optic nerves, and blocked RGC loss. Following a 4-second optic nerve crush, intranasal ST266 increased RGC survival and showed a trend toward reduced RGC axon and myelin damage. Ten days following optic nerve crush, ST266 prevented myelin damage, while also inducing a trend toward increased RGC survival and visual function.
ST266 significantly attenuates traumatic optic neuropathy. Neuroprotective effects of this unique combination of biologic molecules observed here and previously in optic neuritis suggest potential broad application for preventing neuronal damage in multiple optic nerve disorders. Furthermore, results support intranasal delivery as a novel, noninvasive therapeutic modality for eyes and optic nerves.
ST266 preserves visual function following ONC. The right optic nerves of C57BL/6J mice were surgically exposed and crushed for 1 second with fine-tip forceps. Left eyes served as controls. Mice (n = 6 mice/treatment group) were treated daily with one intranasal drop of ST266 or sham-treated with PBS. OKR responses were measured before ONC injury (day 0) and repeated through 5 days post-ONC. ONC induced significant decreases in OKR responses as compared with normal OKR responses in control eyes (***P < 0.001). Treatment with daily ST266 led to significantly higher OKR responses in ONC eyes as compared with ONC eyes from PBS-treated mice (@P < 0.05). Data from one of two representative experiments shown.
Invest Ophthalmol Vis Sci. 2018 May;59(6):2470-2477.
ST266 reduces RGC axon loss. Mice (n = 6 mice/treatment group) were treated daily with one intranasal drop of ST266 or sham-treated with PBS beginning immediately after 1-second ONC was performed on the right eye only. Retinas and optic nerves were removed 5 days post-ONC. (A) ONC induced significant RGC loss, measured by Brn3a labeling, as compared with control eyes in retinas of PBS-treated mice (*P < 0.05). In contrast, RGC numbers were not significantly decreased compared with control eyes in ST266-treated mice, but showed only a trend toward increased RGC numbers compared with ONC eyes from PBS-treated mice. (B) Representative images (original magnification ×40) of RGCs from each group are shown. (C) ONC induced a significant decrease in RGC axon immunostaining with neurofilament antibodies as compared with control eyes in optic nerves of PBS-treated mice (***P < 0.001), and RGC axon staining was significantly higher in ONC eyes of ST266-treated mice compared with ONC eyes from PBS-treated mice (@@@P < 0.001). (D) A collage of images (original magnification ×40) of optic nerves from each group show variable neurofilament staining in ONC eyes reflective of focal areas of axon loss. Data from one of two experiments shown.
ST266 reduces demyelination. Optic nerve sections from mice (n = 6 mice/treatment group) treated daily with one intranasal drop of ST266 or sham-treated with PBS following 1-second ONC performed on the right eye only were stained with LFB to assess myelin. Optic nerves were removed 5 days post-ONC. (A) ONC induced significant demyelination as compared with control eyes in PBS-treated mice (***P < 0.001), and ST266 treatment significantly reduced the level of demyelination induced by ONC (@P < 0.05 versus PBS-treated ONC eyes). (B) Representative images of myelin staining from each group show normal myelin in control eyes, focal myelin loss in an ONC eye from a PBS-treated mouse, and less myelin loss in the ONC eye of an ST266-treated mouse (original magnification ×20). Data from one of two experiments shown.
ST266 effects following 4-second ONC. The right optic nerves of C57BL/6J mice were surgically exposed and crushed for 4 seconds with fine-tip forceps. Left eyes served as controls. Mice (n = 5 mice/treatment group) were treated daily with one intranasal drop of ST266 or sham-treated with PBS. (A) ONC induced significant decreases in OKR responses as compared with normal OKR responses in control eyes (***P < 0.001), with or without ST266 treatment. (B) ONC also induced a significant decrease in neurofilament staining of RGC axons with (**P < 0.01) or without (***P < 0.001) ST266 treatment. (C) ONC induced significant RGC loss compared with control eyes (**P < 0.01), and ST266 treatment significantly increased RGC survival (@P < 0.05) after ONC compared with ONC eyes of PBS-treated mice. (D) ONC induced significant demyelination as compared with control eyes in PBS-treated mice (**P < 0.01), but not in ST266-treated mice. Data from one of two representative experiments shown.
ST266 effects 10 days post-ONC. The right optic nerves of C57BL/6J mice were surgically exposed and crushed for 1 second with fine-tip forceps. Left eyes served as controls. Mice (n = 5 mice/group; ONC eye from one mouse in the PBS-treated cohort was not used in analyses, as the optic nerve was not well exposed and surgical crush could not be confirmed) were treated daily with one intranasal drop of ST266 or sham-treated with PBS for 10 days. (A) ONC induced significant decreases in OKR responses as compared with normal OKR responses in control eyes (***P < 0.001), with or without ST266 treatment, and ST266 treatment led to a trend toward increased OKR responses following ONC. (B) ONC induced a significant decrease in neurofilament staining of RGC axons 10 days post-ONC (*P < 0.05) that was not present with ST266 treatment. (C) ONC induced significant RGC loss compared with control eyes, with (**P < 0.01) or without (***P < 0.001) ST266 treatment, with ST266 inducing a trend toward increased RGC survival. (D) ONC induced significant demyelination as compared with control eyes in PBS-treated mice (**P < 0.01), and ST266 treatment significantly reduced the level of demyelination induced by ONC (@P < 0.05 versus PBS-treated ONC eyes).