212 EXPRESSION OF FIBROBLAST GROWTH FACTOR 18 (FGF18) AND COGNATE RECEPTORS (FGFR3C AND FGFR4) DURING LUTEAL DEVELOPMENT AND INDUCED LUTEOLYSIS IN CATTLE

Fibroblast growth factor receptor 2 (FGFR2) has been shown to induce luteinization in granulosa cells, luteal angiogenesis, and luteal growth. Alternative splicing of 4 genes give rise to 7 subtypes of fibroblast growth factor receptors (FGFR) with varying affinity for different fibroblast growth factors (FGF). Fibroblast growth factor receptor 2 and FGF18 efficiently activate FGFR3C and FGFR4 and may act in cooperation in tissues expressing these receptors. We aimed to determine mRNA expression patterns for FGF18, FGFR3C, and FGFR4 during bovine luteal development and following induced luteolysis. In addition, we assessed FGF18 localization in the bovine CL. Bovine CL were obtained from abattoir ovaries and classed into 4 stages of development: stage 1 =corpus hemorragicum; stage 2 = developing CL; stage 3 = mature or early functional luteolysis CL; and stage 4 = structural luteolysis. To assess FGF18 and FGFR mRNA expression during induced luteolysis, adult cows (Bos taurus Holstein-Friesians) were injected with the PGF2 analogue cloprostenol (500 mg i.m. Intervet, Unterschleissheim, Germany) during the mid-luteal phase of the cycle (Days 8-12). Corpus luteum were collected by transvaginal ovariectomy at 0, 0.5, 2, 4, 12, 24, 48, and 64 hr (n = 5/time point) after PGF2 injection. Tissue samples were submitted to total RNA extraction. Expression of FGF18, FGFR3C, and 4 mRNA during the bovine CL lifespan and induced luteolysis were measured by real-time RT-PCR with oligo-dT in the RT and bovine-specific primers in the PCR. Expression of cyclophilin was used as internal control. The effect of developmental stage and time post-PGF2 on gene expression was tested by ANOVA, followed by Tukey- Kramer HSD test. Immunohistochemical analysis was performed with a commercial human antibody (anti-FGF18; Santa Cruz Biotechnology, Santa Cruz, CA, USA). Fibroblast growth factor 18, FGFR3C, and FGFR4 mRNA was detected in all 4 developmental stages; FGF18 mRNA abundance was higher in stage 3 (2.89 ± 0.05; mean ± SEM) compared with stages 1 (0.3 ± 0.27), 2 (0.56 ± 1.27), and 4 (0.99 ± 0.32). Fibroblast growth factor 18 and FGFR4 mRNA expression did not significantly change during induced luteolysis. Fibroblast growth factor receptor 3C mRNA abundance peaked 4 h after PGF2 injection and significantly decreased at 24 h post-treatment in comparison with peak levels. Immunohistochemical analysis revealed the presence of FGF18 in small and large luteal cells and in blood vessels. In conclusion, the mRNA expression patterns of FGF18 and its receptors suggest their participation in the control of luteal differentiation, particularly during functional luteolysis. The localization of FGF18 protein to blood vessels suggests it may play a role in the control of angiogenesis in the bovine CL. Supported by CAPES/FAPESP.