Pro-survival redox signalling in progesterone-mediated retinal neuroprotection

Retinitis pigmentosa (RP) is a group of hereditary retinal diseases, characterised by photoreceptor cell loss. Despite a substantial understanding of the mechanisms leading to cell death, an effective therapeutic strategy is sought. Our laboratory has previously demonstrated the neuroprotective properties of Norgestrel, a progesterone analogue, in the degenerating retina, mediated in part by the neurotrophic factor basic fibroblast growth factor (bFGF). In other retinal studies, we have also presented a pro-survival role for reactive oxygen species (ROS), downstream of bFGF. Thus, we hypothesized that Norgestrel utilises bFGF-driven ROS production to promote photoreceptor survival. Using the 661W photoreceptor-like cell line, we now show that Norgestrel, working through progesterone receptor membrane complex 1 (PGRMC1); generates an early burst of pro-survival bFGF-induced ROS. Using the rd10 mouse model of RP, we confirm that Norgestrel induces a similar early pro-survival increase in retinal ROS. Norgestrel-driven protection in the rd10 retina was attenuated in the presence of antioxidants. This study therefore presents an essential role for ROS signalling in Norgestrel-mediated neuroprotection in vitro and demonstrates that Norgestrel employs a similar pro-survival mechanism in the degenerating retina

Progesterone analogue protects stressed photoreceptors via bFGF-mediated calcium influx.

Retinitis pigmentosa (RP) is a degenerative retinal disease leading to photoreceptor cell loss. In 2011, our group identified the synthetic progesterone ‘Norgestrel’ as a potential treatment for RP. Subsequent research showed Norgestrel to work through progesterone receptor membrane component 1 (PGRMC1) activation and upregulation of neuroprotective basic fibroblast growth factor (bFGF). Using trophic factor deprivation of 661W photoreceptor-like cells, we aimed to further elucidate the mechanism leading to Norgestrel-induced neuroprotection. In the present manuscript, we show by flow cytometry and live-cell immunofluorescence that Norgestrel induces an increase in cytosolic calcium in both healthy and stressed 661Ws over 24h. Specific PGRMC1 inhibition by AG205 (1 μM) showed this rise to be PGRMC1-dependent, primarily utilising calcium from extracellular sources, for blockade of L-type calcium channels by verapamil (50 μM) prevented a Norgestrel-induced calcium influx in stressed cells. Calcium influx was also shown to be bFGF-dependent, for siRNA knock down of bFGF prevented Norgestrel-PGRMC1 induced changes in cytosolic calcium. Notably, we demonstrate PGRMC1-activation is necessary for Norgestrel-induced bFGF upregulation. We propose that Norgestrel protects through the following pathway: binding to and activating PGRMC1 expressed on the surface of photoreceptor cells, PGRMC1 activation drives bFGF upregulation and subsequent calcium influx. Importantly, raised intracellular calcium is critical to Norgestrel's protective efficacy, for extracellular calcium chelation by EGTA abrogates the protective effects of Norgestrel on stressed 661W cells in vitro

The effect of oral contraceptives on orthodontic tooth movement in rat

Introduction: The purpose of this study was to investigate the effect of ethinyl estradiol/norgestrel ? used in some oral contraceptives- on orthodontic tooth movement in Wistar rats. Material and Methods: Forty eight female three-month old Wistar rats with an average weight of 250?25gr were divided into two experimental and control groups. One week prior to appliance insertion and during the appliance therapy period, 100 mcg/kg/day of ethinyl estradiol and 1mg/kg/days of norgestrel were administered to the experimental group by gavage; meanwhile the control group received an equivalent volume of Sodium Chloride 0.9 % (Saline). Maxillary central incisors were tipped distally by insertion of springs exerting 30g force. Two, seven and fourteen days after spring insertion animals were sacrificed. The mesioincisal distance between maxillary incisors were measured. Subsequently, histological sections were prepared for histomorphometric studies. Results: 14 days after force application the orthodontic tooth movement was significantly lower in the experimental group (p<0.05). The number of osteoclasts were significantly lower in the experimental group 2, 7 and 14 days after spring insertion (p<0.05). Conclusion: Ethinyl estradiol/norgestrel (oral contraceptives) can significantly decrease the amount of tooth movement in the linear phase