Reprogramming Müller glia via in vivo cell fusion regenerates murine photoreceptors.

Vision impairments and blindness caused by retinitis pigmentosa result from severe neurodegeneration that leads to a loss of photoreceptors, the specialized light-sensitive neurons that enable vision. Although the mammalian nervous system is unable to replace neurons lost due to degeneration, therapeutic approaches to reprogram resident glial cells to replace retinal neurons have been proposed. Here, we demonstrate that retinal Müller glia can be reprogrammed in vivo into retinal precursors that then differentiate into photoreceptors. We transplanted hematopoietic stem and progenitor cells (HSPCs) into retinas affected by photoreceptor degeneration and observed spontaneous cell fusion events between Müller glia and the transplanted cells. Activation of Wnt signaling in the transplanted HSPCs enhanced survival and proliferation of Müller-HSPC hybrids as well as their reprogramming into intermediate photoreceptor precursors. This suggests that Wnt signaling drives the reprogrammed cells toward a photoreceptor progenitor fate. Finally, Müller-HSPC hybrids differentiated into photoreceptors. Transplantation of HSPCs with activated Wnt functionally rescued the retinal degeneration phenotype in rd10 mice, a model for inherited retinitis pigmentosa. Together, these results suggest that photoreceptors can be generated by reprogramming Müller glia and that this approach may have potential as a strategy for reversing retinal degeneration.

The NIMA-family kinase Nek6 phosphorylates the kinesin Eg5 at a novel site necessary for mitotic spindle formation.

Nek6 and Nercc1 (also known as Nek9) belong to the NIMA family of protein kinases. Nercc1 is activated in mitosis, whereupon it binds, phosphorylates and activates Nek6. Interference with Nek6 or Nercc1 in mammalian cells causes prometaphase-metaphase arrest, and depletion of Nercc1 from Xenopus egg extracts prevents normal spindle assembly. Herein we show that Nek6 is constitutively associated with Eg5 (also known as Kinesin-5 and Kif11), a kinesin that is necessary for spindle bipolarity. Nek6 phosphorylated Eg5 at several sites in vitro and one of these sites, Ser1033, is phosphorylated in vivo during mitosis. Whereas CDK1 phosphorylates nearly all Eg5 at Thr926 during mitosis, Nek6 phosphorylates approximately 3% of Eg5, primarily at the spindle poles. Eg5 depletion caused mitotic arrest, resulting in cells with a monopolar spindle. This arrest could be rescued by wild-type Eg5 but not by Eg5[Thr926Ala]. Despite substantial overexpression, Eg5[Ser1033Ala] rescued 50% of cells compared with wild-type Eg5, whereas an Eg5[Ser1033Asp] mutant was nearly as effective as wild type. Thus, during mitosis Nek6 phosphorylates a subset of Eg5 polypeptides at a conserved site, the phosphorylation of which is crucial for the mitotic function of Eg5.

Assessment of the protective effects of amifostine against cisplatin-induced toxicity.

OBJECTIVE: The goals of this study were to determine whether the toxicity of cisplatin, a chemotherapeutic alkylating agent, could be reduced by the use of amifostine and to determine whether amifostine alone could cause ototoxicity. STUDY DESIGN: Prospective, animal study. METHODS: Auditory brainstem response click threshold, latencies, and blood work were used to measure ototoxicity, nephrotoxicity, and myelotoxicity before and 4 weeks after treatment. Groups of guinea pigs received either cisplatin alone (30 mg/kg), amifostine (1000 mg/kg), cisplatin plus amifostine (1000 mg/kg), or no agent. RESULTS: Amifostine reduced the hearing loss caused by cisplatin for many animals. Amifostine partially protected against cisplatin-induced ototoxicity and renal toxicity. We did not find evidence for myelotoxicity owing to cisplatin in this sample. CONCLUSION: Amifostine may have a role in reducing toxicity or permitting larger doses of cisplatin to be given, but toxicity can still be significant even with protection.