Purchase this article with an account.
Mohor Sengupta, Naoki Nakaya, Christo Kole, Stanislav I Tomarev; Brorin, a Chordin-related protein, facilitates neuroprotection of injured retinal ganglion cells. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4843.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Brorin (Vwc2), a member of the Chordin family of proteins, is a component of the AMPA receptor complex and is expressed in neuronal tissues, including the retina. The objective of this study was to evaluate whether Vwc2 can protect retinal ganglion cells (RGCs) after injury and elucidate the molecular mechanisms of its action.
Mouse Vwc2 or LacZ were cloned into the AAV2.9/2yF viral vector. Retinal wholemounts from adult C57/BL6 mice after recombinant AAV intravitreal injection and optic nerve crush were stained for RBPMS and osteopontin and imaged. Mixed retinal cultures were prepared from postnatal day 20 (P20) mouse, infected on day 1 with AAV2.9/2yF-Vwc2 or AAV2.9/2yF-LacZ and harvested on day 14 for counting of Brn3a+ surviving RGCs. Mixed P7 retinal cultures from wild-type (WT) or Vwc2 knockout (KO) mice were maintained for 2 weeks, after which they were either treated with Vwc2 protein (10 nM - 100nM) or an equivalent volume of PBS control. The culture was harvested 48 hours later and imaged. A different set of similar cultures was maintained for two weeks and analyzed for pre- and post-synaptic markers, synaptophysin and PSD95, respectively.
Intravitreal injections of recombinant AAV2.9/2yF-Vwc2 vector (109 viral particles) into adult mouse eyes increased Brn3a+ RGC survival one week after crush compared with recombinant AAV2.9/2yF-LacZ vector injection (543±93 versus 360±61 RGCs/mm2, respectively; p=0.0001). Infection of mixed cultures from P20 retinae with AAV2.9/2yF-Vwc2 showed increased survival of Brn3a+ RGCs compared to AAV2.9/2yF-LacZ infection (440±46 versus 200±29 RGCs/mm2, respectively; p=0.0021). In retinal mixed cultures, 100 nM purified Vwc2 significantly increased phospho-P70S6K level compared to PBS (normalized average intensity values 11156±742 and 8702±551, respectively; p=0.02), suggesting mTOR activation. In 2-week P7 retinal mixed cultures, Vwc2 KO mice showed increased PSD95 (four times) compared to synaptophysin, whereas, the WT mice showed similar levels of both proteins.
Our data suggest that Vwc2 has a neuroprotective effect on injured RGCs, which might be due to mTOR activation. Higher number of PSD95 in Vwc2 KO mice compared with WT mice suggests altered synaptic composition and/or maturation, which may be attributed to the absence of Vwc2.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
This PDF is available to Subscribers Only