Abstract
Purpose :
Many conjunctival inflammatory diseases differ between the sexes, and altered conjunctival goblet cells(GC) response is involved. Inflammation starts with the releasing of pro-inflammatory mediators and is terminated by biosynthesis of specialized pro-resolution mediators(SPMs). Herein, we will determine the sex-based difference in the responses of conjunctival goblet cells to inflammatory stimuli or pro-resolution lipid SPMs and their interaction with sex hormones.
Methods :
GCs were cultured from pieces of human conjunctiva. 24h before experiments the cells were transferred into phenol red-free and FBS-free media to minimize the effect of serum hormones. RT-PCR and immunofluorescence microscopy(IFM) were performed to determine the presence of sex hormone receptors. Cellular response to stimuli or SPMs was studied by measuring the increase in intracellular [Ca2+]([Ca2+]i)using fura 2/AM microscopy.
Results :
1) Androgen receptors were identified in 4/4 male and 3/3 female GCs by RT-PCR and IFM(Fig 1). Estrogen receptor(ER) α was found in 3/5 males and 3/3 females and ERβ was found in 2/3 females and 2/5 males by RT-PCR.
2) Carbachol(10-5-10-3M), anaphylactic mediator histamine(10-8-10-5M) and pro-inflammatory mediator LTB4(10-11-10-8M) all caused a concentration-dependent increase in [Ca2+]i in GC. Significant different Ca2+ response between sexes was found with carbachol 10-3M(p<0.05), but not in histamine or LTB4.
3) The SPM lipoxin A4(LXA4)(10-9-10-8M), but not in the SPM resolvin(Rv) D1 or RvD2, induced a [Ca2+]i increase that was significantly higher in males compared to females(p<0.008).
4) We incubated GC with dihydrotestosterone(DHT), estrone(E1) or estradiol(E2). Incubation with DHT, E1 and E2 10-7M for 30 min significantly inhibited the LTB4-stimulated [Ca2+]i increase in males(p<0.05, 0.04, 0.04, respectively); DHT and E1 significantly inhibited the LTB4 response in females (p=0.04, 0.04 respectively). Incubation with DHT, E1 and E2 overnight significantly inhibited LTB4 response in females(p=0.02, 0.02, 0.03 respectively); DHT and E2 significantly inhibited the LTB4 response in males(p<0.05, 0.04 respectively).
Conclusions :
We concluded that male and female GCs were different in the expression of sex hormone receptors, response to the SPM LXA4 and pro-inflammatory mediator LTB4-induced response to sex hormone treatment.
This is a 2020 ARVO Annual Meeting abstract.