GnRH analogs induce a LH peak and increase pregnancy per timed-AI in ewes.

To date, there have been no studies testing the capacity of GnRH analogs and respective doses to induce a LH peak in sheep. In this sense, the present study aimed to evaluate the capacity of different synthetic forms and doses of GnRH in inducing LH release in sheep, and the effect of GnRH administration at timed artificial insemination (TAI) on pregnancy per timed-AI. In experiment 1, ewes (n = 40) received an intravaginal device (IVD) of medroxyprogesterone acetate (MPA; 60 mg) for 7 d and prostaglandin F2α analog on Day 5. On Day 7, the ewes were allocated randomly into one of eight groups (n = 5/group), which received a GnRH analog at a specific dose, as follows: lecirelin (12.5 or 25 µg), gonadorelin (50 or 100 µg), buserelin acetate (4.2 or 8.4 µg), or deslorelin (375 or 750 µg). Blood samples for LH determination were obtained at 0, 2, 4, and 6 h after GnRH and the IVDs were removed after the last blood collection. The maximal LH concentration induced by gonadorelin at doses of 50 µg and 100 µg (12.0 ± 2.4 ng/mL and 28.6 ± 7.1 ng/mL, respectively) was lower (P < 0.05) than serum LH induced by 8.4 µg of buserelin (78.9 ± 12.9 ng/mL), 375 µg and 750 µg of deslorelin (75.6 ± 7.4 ng/mL and 72.1 ± 10.6 ng/mL, respectively) and 12.5 µg and 25 µg of lecirelin (73.3 ± 17.8 ng/mL and 61.6 ± 5.9 ng/mL, respectively). However, the maximal LH concentration induced by 4.2 µg of buserelin (49.4 ± 5.9 ng/mL) was similar (P > 0.05) to the 100 µg of gonadorelin. The total release of LH (area under the curve - AUC) after treatment with 50 µg of gonadorelin (31.7 ± 5.9 ng h/mL) was lower (P < 0.05) than after other agonists. In a second experiment, 330 ewes were treated with IVD containing MPA for 7 d. Simultaneously with IVD removal, 250 µg of cloprostenol and 200 IU of eCG were administered. Then, ewes were assigned randomly to either no further treatment (control); or to receive 4.2 µg of buserelin acetate (GnRH group) at cervical TAI, which was performed with fresh semen 54 h after IVD withdrawal in all the animals. Higher pregnancy per timed-AI was observed for GnRH (50.3 %) compared to control (40.7 %). We conclude that buserelin acetate (8.4 µg), lecirelin (12.5 and 25 µg) and deslorelin (375 and 750 µg) induced a greater stimulatory effect on LH secretion than gonadorelin treatment. Furthermore, buserelin acetate treatment at TAI increased pregnancy per timed-AI in ewes previously treated with MPA and eCG.

The evaluation of the effect of tafluprost on the intraocular pressure of healthy male guinea pigs under different light-and-darkness regimes.

BACKGROUND: Ocular hypertension is one of the most underdiagnosed ocular abnormalities among guinea pigs around the world. OBJECTIVES: The current study investigates the effect of 0.0015% preservative-free tafluprost ophthalmic solution (Zioptan) on the intraocular pressure of 16 healthy male guinea pigs (Cavia porcellus) under different light/darkness regimes. METHODS: All guinea pigs received a single drop of tafluprost at 5:30 in the right eye, whereas the contralateral eyes served as control to receive a placebo. Then, the animals were randomly divided into two groups; group A was exposed to light, whereas group B was placed in darkness from 5:30 to 18:00. Rebound tonometry (TonoVet) was instrumented to measure IOP values at 5:30 (baseline), 6:00, 7:00, 8:00, 9:00 and then every 3 h until 18:00. RESULTS: The maximum IOP reduction associated with tafluprost was observed at 6:00 by -1.4 ± 1.1 mmHg (p-value = 0.026) and -2.5 ± 1.2 mmHg (p-value = 0.011) in group A and B, respectively (repeated measure ANOVA test). There was a significant difference between the mean right and left eye IOP values in both groups at 5:30, 6:00, 7:00 and 8:00 (p-value <0.05), which was greater in amount in group B compared to group A due to the effect of darkness on IOP reduction. CONCLUSIONS: It is suggested that the variations of IOP in different light/dark conditions be taken into consideration when applying ocular hypotensive agents on guinea pigs' eyes.

Effect of metritis on in-vitro uterine contractility in cows during the puerperium.

Aim of this study was to determine the effect of metritis on in-vitro uterine contractility. Uteri obtained from 16 euthanized Holstein-Friesian cows were divided into two groups depending on whether metritis was absent (M-, n = 6) or present (M+, n = 10). Four longitudinal and four circular myometrial strips of all uteri were incubated in an organ bath. Spontaneous contractility was recorded in five consecutive 30-minute periods (T1-T5). This was followed by stimulation of one longitudinal and one circular strip with increasing concentrations of oxytocin, prostaglandinF2α (PGF2α), and calcium chloride (each during four 30-minute periods [T6-T9]). Strips in group M+ had higher minimum amplitude (minA) values at T1 and higher minA, mean amplitude (meanA), and area under the curve (AUC) values at T2 than strips in group M- (P ≤ 0.05). In the M+ group, the maximum amplitude (maxA), meanA, and AUC values of circular strips were higher than those of longitudinal strips during spontaneous contractility (T1, T4, and T5; P ≤ 0.05). The minA, meanA, and AUC values for strips in group M+ were higher than those in group M- when exposed to the highest concentration of PGF2α (P ≤ 0.05). During stimulation with PGF2α (T9), longitudinal strips had higher maxA values than the circular strips in group M+ (P ≤ 0.05). Spontaneous and stimulated contractility were temporarily increased in uteri with metritis compared to healthy uteri. Both myometrial layers, especially in uteri with metritis, reacted differently during spontaneous contractility and to stimulation with PGF2α.

Effects of 0.0015% preservative-free tafluprost on the equine eye.

OBJECTIVE: The purpose of this study was to determine the effects and potential side effects of topical preservative-free (PF) tafluprost 0.0015% in ophthalmologically normal horses. ANIMALS: Five adult grade horses. PROCEDURES: One of the eyes of each horse was randomly chosen as the "treatment" eye, and consequently, the contralateral eye served as the "control." A single dose of PF tafluprost 0.0015% (0.2 mL) was instilled in the treated eye of each horse. Intraocular pressure (IOP), Schirmer's tear test (STT) levels of each eye, and an ophthalmic examination were performed at T0 (baseline), T30, T120, T24 h, and T48 h. RESULTS: The mean IOP values of the treated eyes at baseline (T0), T30, T120, T24 h, and T48 h were 25.4 ± 4.8 mmHg, 21.2 ± 1.92 mmHg, 15.20 ± 2.48 mmHg, 18.40 ± 1.51 mmHg, and 24.60 ± 1.94 mmHg, respectively. Significant differences were observed between the mean baseline IOP level and the T120 and T24 h time points (p = .001 and p = .009). The mean STT levels at each time point showed insignificant fluctuations during the study (p = .140). Adverse effects such as chemosis and episcleral injection were observed 30 min after the instillation of tafluprost 0.0015% (T30). Blepharospasm and conjunctival hyperemia were observed 120 min (T120) after the administration of the medication. CONCLUSION AND CLINICAL RELEVANCE: Tafluprost 0.0015% showed potential in reducing IOP, but due to its local side effects, it is not a good candidate for management of glaucoma in horses. Tafluprost did not notably affect STT.

Pre-synchronization of ovulation timing and delayed fixed-time artificial insemination increases pregnancy rates when sex-sorted semen is used for insemination of heifers.

This study was conducted to determine effects of pre-synchronization of ovulation timing among heifers and delayed fixed-time artificial insemination (TAI) with sex-sorted semen on proportion of heifers pregnant after TAI (PR/AI). Heifers were assigned to one of eight treatments: 1 and 2), 7-d CO-Synch + CIDR treatment regimen with administration of gonadotropin-releasing hormone and a CIDR insert on Day 0, prostaglandin F2α (PGF) at CIDR removal on Day 7, and TAI occurring 54 h later with conventionally processed (CTRL54-CNV) or sex-sorted semen (CTRL54-SEX); 3 and 4), same as CTRL54 but TAI delayed to 72 h with conventionally processed (CTRL72-CNV) or sex-sorted semen (CTRL72-SEX); 5 and 6), same as CTRL54 but additional administration of PGF on Day -7 and TAI with conventionally processed (PRE54-CNV) or sex-sorted semen (PRE54-SEX); 7 and 8), same as PRE54 treatments but TAI delayed to 72 h with conventionally processed (PRE72-CNV) or sex-sorted semen (PRE72-SEX). Proportion of heifers pregnant after TAI was greater (P ≤  0.02) with conventionally processed semen compared with sex-sorted semen, yet PR/AI did not differ (P =  0.14) between heifers in PRE72-CNV and PRE72-SEX groups. There were greater PR/AI in the PRE72-SEX (P =  0.03) than CTRL54-SEX group (46.1 % and 36.9 %) and there was no difference (P =  0.31) in PR/AI between CTRL54-CNV and PRE72-SEX groups (50.4 % and 46.1 %). In conclusion, pre-synchronization of ovulation timing among heifers combined with delayed TAI resulted in increased PR/AI with sex-sorted semen compared with the 7-d CO-Synch+CIDR treatment regimen.

Biomechanical Effects of Two Forms of PGF2α on Ex-vivo Rabbit Cornea.

PURPOSE: To investigate the biomechanical effects of two synthetic prostaglandin F2α analogues (PGF2α), namely Travoprost and Tafluprost, on the ex-vivo rabbit cornea. MATERIALS AND METHODS: Ninety-six eyes of 48 Japanese white rabbits were divided into 3 equal groups randomly; the Travoprost treated group (Tra), the Tafluprost treated group (Taf) and the control group (Co). Eyes in Tra and Taf groups were preserved in storage medium for 10 days with 1:10 Travoprost and Tafluprost diluents, respectively; while the Co eyes were preserved in a similar but PGF2α-free medium. Twenty-four corneas of each group were tested under inflation conditions with up to 30 mmHg posterior pressure. The pressure-deformation data obtained experimentally were used in an inverse analysis process to derive the stress-strain behavior of the tissue, using which the tangent modulus, a direct measure of the tissue's material stiffness, was calculated. The remaining eight specimens of each group were analyzed using electron microscopy for fibril diameter and interfibrillar spacing. RESULTS: Although the central corneal thickness increased significantly in the three groups after storage (p < .01), it was similar in all groups both before (p = .598) and after storage (p = .181). After treatment with Travoprost and Tafluprost, the corneas exhibited lower tangent modulus (by 29.2% and 29.8%, respectively, at 6 kPa stress) and larger stromal interfibril spacing (by 21.9% and 23.6%) compared with the control group. There was no significant change in fibril diameter with either Travoprost or Tafluprost treatment (p = .769). CONCLUSIONS: The results demonstrated significant reductions in tangent modulus and increases in interfibrillar spacing, which were of similar magnitudes, with the application of two different forms of PGF2α.

Effects of pupil size on canine visual evoked potential with pattern stimulation.

The purpose of this study was to investigate the effects of pupil diameter on canine visual evoked potentials with pattern stimulation (P-VEP). Atropine eye drop (1.0%) was applied to both eyes as a cycloplegic drug, and tafluprost eye drop (0.015%) was applied to one eye that was selected randomly for miosis (miosis group). The other eye did not receive tafluprost (mydriasis group). P-VEP was recorded at three pattern sizes. The P100 implicit time at a small pattern size in the mydriasis group was significantly prolonged compared to the miosis group. We hypothesized that the prolonged P100 implicit time under mydriatic conditions was due to increased spherical aberrations and concluded that mydriatic conditions affected P100 implicit time in canine P-VEP recordings.

Effect of Topical Prostaglandin F2α Analogs on Selected Oxidative Stress Parameters in the Tear Film.

Background and Objectives: Topically administered antiglaucoma medications, especially those containing benzalkonium chloride (BAC), may cause local adverse effects and compromise ocular surface. The aim of the study was to assess the effect of topical prostaglandin F2α analogs (PGAs): preservative-free latanoprost, BAC-preserved latanoprost, preservative-free tafluprost, and BAC-preserved bimatoprost, on selected oxidative stress parameters in the tear film. Materials and Methods: The patients were divided into five groups: group C (n = 25) control group-subjects who did not use topical antiglaucoma medications, group L (n = 22)-patients using topical preservative-free latanoprost, group L+BAC (n = 25)-patients using topical BAC-preserved latanoprost, group T (n = 19)-patients using topical preservative-free tafluprost, and group B+BAC (n = 17)-patients using topical BAC-preserved bimatoprost. The oxidative stress markers in the tear film samples were evaluated: total protein (TP) concentration, advanced oxidation protein products (AOPP) content, total sulfhydryl (-SH) groups content, the activity of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx), as well as Total Oxidant Status (TOS), Total Antioxidant Response (TAR), and Oxidative Stress Index (OSI). Results: The TP concentrations in the groups L, L+BAC, and B+BAC were statistically significantly higher in comparison with group C. The SOD and CAT activities in the groups L+BAC and B+BAC were statistically significantly higher when compared to group C. As compared to group C, AOPP and TOS were statistically significantly higher in all the study groups. OSI was found to be statistically significantly higher in the groups L+BAC, T, and B+BAC in comparison with group C. Conclusion: Use of topical PGAs by the patients with ocular hypertension or primary open-angle glaucoma is associated with increased oxidative stress in the tear film which is additionally exacerbated by the presence of BAC in the formulation.

Meibomian gland dropout rate as a method to assess meibomian gland morphologic changes during use of preservative-containing or preservative-free topical prostaglandin analogues.

PURPOSE: To investigate the usefulness of meibomian gland (MG) dropout rate in the evaluation of MG morphological change associated with the use of prostaglandin for glaucoma treatment through the association between MG and the ocular surface parameters and medication duration and presence of preservative. METHODS: This cross-sectional study was conducted on 88 eyes of 88 patients who were diagnosed with glaucoma and used only Tafluprost as treatment. The patients were divided into four "user" groups: 1) 23 patients used preservative-free (PF) Tafluprost for 6 months; 2) 21 patients used preservative-containing (PC) Tafluprost for 6 months; 3) 23 patients used PF-Tafluprost for 24 months; 4) 21 patients used PC-Tafluprost for 24 months. Ocular surface parameters and the MG condition, including MG dropout rate and meiboscale, were evaluated. Multiple regression was used to identify associations. RESULTS: There were significant differences in age (p = 0.003), tear breakup time (p = 0.016), lid margin abnormality (p = 0.016), expressibility (p = 0.039), meiboscale (p<0.001), and MG dropout rate (p<0.001) among the 4 groups. MG dropout rate and meiboscale showed significant differences in all post hoc analyses, except for the comparison between the PF-Tafluprost and PC-Tafluprost 6-month user groups. Medication duration, preservative status, and meiboscale were significantly correlated with MG dropout rate (p<0.001, p = 0.024, p<0.001, respectively). In the 6-month user group, preservative status significantly correlated with MG dropout rate (p = 0.015). However, in the 24-month user group, meiboscale was the only parameter significantly associated with MG dropout rate (p<0.001). CONCLUSION: MG dropout rate in patients using Tafluprost showed a significant correlation with medication duration and preservative status. This result indicates MG dropout rate reflects MG morphologic change associated with prostaglandin.

Efficacy and safety of the fixed combinations of tafluprost/timolol and latanoprost/carteolol.

In this study, we made a comparative efficacy and safety assessment of two different fixed combinations of drugs, viz., tafluprost/timolol (TAF/TIM) and latanoprost/carteolol (LAT/CAR), by determining their effects on intraocular pressure (IOP) in ocular normotensive monkeys and examining their toxic effects on ocular surface using human corneal epithelial cells. TAF/TIM was found to be more effective in lowering IOP for a longer duration compared to LAT/CAR. We found that the difference in the intensity of IOP-lowering effect was because of the differences in the strength of timolol compared with that of carteolol as a beta-adrenergic antagonist and strength of tafluprost compared with that of latanoprost as a prostaglandin analogue. In addition, TAF/TIM showed much less cytotoxic effects compared to LAT/CAR on the human corneal epithelial cells. Our findings showed that TAF/TIM is better than LAT/CAR with regard to the IOP-lowering effect in monkeys and toxicity on ocular surface.

Effects of Anti-Glaucoma Prostaglandin Ophthalmic Solutions on Cultured Human Corneal Epithelial Cells.

Purpose: We compare the cytotoxicity of anti-glaucoma prostaglandin ophthalmic solutions on human corneal epithelial cells and elucidate mechanisms of toxicity. Methods: Cell viability was examined using MTS assay, and morphological changes of the cells were observed. Induction of necrosis/apoptosis was measured by colorimetric caspase assay. The production of Reactive oxygen species (ROS) and release of cytokines were analyzed using 2', 7'-dichlorodihydrofluorescein diacetate and bead-based indirect immunofluorescent assay, respectively. Results: Xalatan, Lumigan 0.01%, and Lumigan 0.03% decreased cell viability and induced morphological changes. Xalatan and Lumigan 0.01% induced necrosis. Xalatan, Lumigan 0.01%, Lumigan 0.03%, and Taflotan stimulated ROS production. Travatan and Lumigan 0.03% increased concentrations of Interleukin (IL)-6 and IL-8 in culture media. Conclusions: Xalatan and Lumigan 0.01% ophthalmic solutions demonstrated potent cytotoxicity compared with Lumigan 0.03%, Travatan, Taflotan, and Taflotan UD. Taflotan UD, compared to Taflotan 0.0015%, induced less oxidative stress and apoptotic signalling. The cytotoxicity might be partly associated with benzalkonium chloride.

Effect of the administration of alfaprostol 3 or 6 days after ovulation in jennies: ultrasonographic characteristic of corpora lutea and serum progesterone concentration.

Donkey jenny's corpus luteum (CL) response to PGF2α analogues has not been investigated in depth. Aim of this study was to evaluate the donkey jenny's corpus luteum (CL) ultrasonographic characteristics (diameter, area and vascularized area) by B-Mode, Color Doppler and serum progesterone concentration ([P4]) after treatment with the prostaglandins F2α analogue alfaprostol at day 3 or day 6 after ovulation (groups PG3 and PG6, respectively). [P4] was positively correlated (P < 0.0001) with CL diameter: r2 = 0.17; area: r2 = 0.21 and vascularized area: r2 = 0.54. The interovulatory interval was significantly reduced in the PG6 group (15 ±â€¯1.8 days), compared to the control group (24.5 ±â€¯2.9 days; P < 0,05), while there were no significant differences in interovulatory interval between PG3 (21.7 ±â€¯7.9 days) and control or PG6 group. [P4], in the 6 jennies of the PG6 group, dropped under 1 ng/mL within 2 days after treatment, remaining under this concentration until [P4] raised again to levels comparable with those of the control group until spontaneous luteolysis. After alfaprostol administration, one of the 2 remaining PG3 group jennies showed a complete luteolysis, and the other one underwent a partial luteolysis and ovulated in diestrus.

The involvement of NR4A1 and NR4A2 in the regulation of the luteal function in rats.

The nuclear receptor 4A (NR4A) members play important roles in cellular proliferation, differentiation and apoptosis. The current study first evaluate the expression of ovarian NR4A1 during different luteal stages in rats. Immature rats aged 28 days were treated with sequential Pregnant mare serum gonadotropin (PMSG) (D -2) / human chorionic gonadotropin (hCG) (D 0) to induce pseudopregnancy. Serum progesterone (P4) and ovarian expression of NR4A1 were detected by RIA and WB, respectively, at follicle stage (D 0), early (D 2), middle (D 7) and late (D 14 and D 20) luteal stages. To confirm the role of NR4A1 during the luteal regression, rats were treated with prostaglandin F2α analog (PGF) for 0-8 h on D 7 to detect the expressions of NR4A1 and NR4A2. RIA result showed that serum P4 reached highest level on D 7 and then declined. WB results showed that there were two types of NR4A1 (NR4A1-L and NR4A1-S) expressed in the ovary. The ovarian NR4A1-L decreased at the late luteal stage (D 20). However, the NR4A1-S increased at the late luteal stage (D 14). After PGF treatment on D 7, the expression of NR4A1-S increased which peaked at 0.5-1 h and then declined; while NR4A1-L expression did not change within 8 h. Real-time PCR results showed that the ovarian NR4A1 mRNA increased within 0.5 h, maintained high at 1 h and then declined. The NR4A2 mRNA expression exhibited a similar pattern to that of NR4A1 mRNA, though its abundance was not as high as NR4A1. IHC results revealed that NR4A1-L was expressed mainly in the cytoplasm of luteal steroidogenic cells, faintly expressed in the follicle theca cells, oocytes and the pericytes; while NR4A2 was primarily localized in the cytoplasm of luteal steroidogenic cells. In conclusion, all these results demonstrate that NR4A2 as well as NR4A1 might be involved in the luteal development and luteolysis in rats.

Prostaglandin (PTG) E and F receptors in the porcine corpus luteum; effect of tumor necrosis factor-α.

The porcine corpus luteum (CL) is NOT sensitive to the luteolytic effects of PGF-2α until days 12-13 of cycle. The control of "luteolytic sensitivity" (LS) of the pig CL to PGF-2α is unknown, but it is temporally associated with macrophage infiltration into the CL. Since macrophages are the predominant source of TNF-α in the porcine CL, in other studies we examined the effects of TNF-α on porcine luteal cells in culture and showed that TNF-α induces LS in vitro. In Experiment 1 of this study possible mechanisms involved in the control of LS were examined, and involved measurement of the protein levels of PTGER2/EP-2, and PTGER3/EP-3 in porcine CL collected before (days 7-10), versus after (day 13), the onset of the LS. In Experiment 2, an examination of potential mechanisms involved in the control of LS by TNF-α, was carried out in which the effects of TNF-α on mRNA and protein expression of EP-2, EP-3 and FP in cultured luteal cells, were examined. The results of Experiment 1 showed that PTGER-3/EP-3 (but not PTGER-2/EP-2) levels decreased in porcine CLs after (day 13) compared to before (day 7-10) LS. In Experiment 2, the data obtained showed that TNF-α decreased PTGER-3/EP-3 and increased PTGFR/FP protein (in EARLY stage CL). In conclusion, these studies suggest a role for PTGER-3/EP-3 in the acquisition of LS, and support the hypothesis that TNF-α from CL macrophages plays a critical role in the control of LS in the porcine CL, by increasing PTGFR/FP, and decreasing PTGER-3/EP-3 protein.

Expression patterns of claudin-5 and its related signals during luteal regression in pseudopregnant rats: The enhanced effect of additional PGF treatment.

To study the expression patterns of claudin-5 and its related signals during luteal regression in rats, a sequential PMSG/hCG treatment paradigm was used to obtain a single, well-defined generation of corpus luteum (CL). A total of 35 rats were treated with one PGF or two PGF at an interval of 24 h from day 7 of pseudopregnancy to induce CL regression. Serum and ovaries were collected at 0, 2, 4, 8 or 24 h after one PGF injection (1 PGF), 2 or 24 h after two PGF injections (2 PGF). The serum progesterone level was detected by RIA; the ovarian expression of claudin-5, the phosphorylations of STAT3 (p-STAT3), Akt (p-Akt), ERK1/2 (p-ERK) and p38 MAPK (p-p38) were detected by western blot, real-time PCR and IHC. Results showed that serum progesterone (P4) decreased after PGF treatment. Claudin-5 mRNA decreased at 4 h and 8 h after 1 PGF and 2 h after 2 PGF, and claudin-5 protein decreased at 4 h after 1 PGF. p-STAT3 increased at 4 h after 1 PGF and 2 h after 2 PGF. p-ERK increased at 2 h after 2 PGF. The level of p-Akt decreased at 4 h after 1 PGF. PGF treatment did not alter the phosphorylation of p38 MAPK at any time points in this study. IHC results revealed that claudin-5 was expressed in the nuclei and cytoplasm of steroidogenic cells and in the vessels, while PGF induced-p-STAT3 was expressed uniformly in the cytoplasm of luteal steroidogenic cells. In conclusion, PGF treatment decreased the expression of claudin-5 and the additional PGF treatment enhanced the decrease in claudin-5 mRNA expression and the increases in ERK1/2 and STAT3 phosphorylation in the corpus luteum of pseudopregnant rats, which will contribute new information to the further study of molecular mechanism of luteal regression.

Preperimetric Glaucoma Prospective Observational Study (PPGPS): Design, baseline characteristics, and therapeutic effect of tafluprost in preperimetric glaucoma eye.

PURPOSE: There is no consensus on the diagnosis or treatment policy for Preperimetric Glaucoma (PPG) because the pathogenesis of PPG is not clear at this time. Preperimetric Glaucoma Prospective Observational Study (PPGPS) is a first multicenter, prospective, observational study to clarify the pathogenesis of PPG. This article indicates study design, patient baseline characteristics, and analysis focused on optic nerve head (ONH) blood flow in PPG, as well as the intraocular pressure (IOP) -lowering effect and ONH blood flow-improving effects of Tafluprost. METHOD: In this study, 122 eyes from 122 subjects (mean age: 53.1 ± 14.3) newly diagnosed as PPG were enrolled. The circumpapillary retinal nerve fiber layer thickness (cpRNFLT) was evaluated with optical coherence tomography (OCT). The ONH blood flow was measured with laser speckle flowgraphy. The therapeutic effect of Tafluprost was evaluated at Month 0 (ONH blood flow-improving effect) and Month 4 (IOP-lowering effect). RESULTS: The untreated IOP, cpRNFLT, and baseline Mean deviation (MD) value was 16.4 ± 2.5 mmHg, 80.4 ± 8.2 µm, and -0.48 ± 1.29 dB, respectively. In the site-specific visual field evaluation using the sector map, there was no appreciable site-specific visual field defect in the eye with PPG. The inferior region of cpRNFLT in 4-quadrant OCT sector analysis and 6 o'clock region in 12-o'clock OCT sector analysis was the highest rate of abnormality in PPG eyes. Topical administration of Tafluprost significantly reduced IOP from 16.4 ± 2.5 mmHg at baseline to 14.5 ± 2.3 mmHg at Month 4 (P < 0.001, paired t-test). In the linear regression analysis, there was a significant relationship between the increase of ONH blood flow and baseline value. CONCLUSION: PPGPS is a first prospective study focusing on the pathology of PPG. This study is expected to elucidate the pathology of PPG, with evidence useful for determining a treatment strategy for PPG.

A Novel Convergent Synthesis of the Potent Antiglaucoma Agent Tafluprost.

Tafluprost (AFP-168, 5) is a unique 15-deoxy-15,15-difluoro-16-phenoxy prostaglandin F2α (PGF2α) analog used as an efficacious ocular hypotensive agent in the treatment of glaucoma and ocular hypertension, as monotherapy, or as adjunctive therapy to ß-blockers. A novel convergent synthesis of 5 was developed employing Julia-Lythgoe olefination of the structurally advanced prostaglandin phenylsulfone 16, also successfully applied for manufacturing of pharmaceutical grade latanoprost (2), travoprost (3) and bimatoprost (4), with an aldehyde ω-chain synthon 17. The use of the same prostaglandin phenylsulfone 16, as a starting material in parallel syntheses of all commercially available antiglaucoma PGF2α analogs 2-5, significantly reduces manufacturing costs resulting from its synthesis on an industrial scale and development of technological documentation. Another key aspect of the route developed is deoxydifluorination of a trans-13,14-en-15-one 30 with Deoxo-Fluor. Subsequent hydrolysis of protecting groups and final esterification of acid 6 yielded tafluprost (5). The main advantages are the preparation of high purity tafluprost (5) and the application of comparatively cheap reagents. The preparation and identification of two other tafluprost acid derivatives, tafluprost methyl ester (32) and tafluprost ethyl amide (33), are also described.

Effect of preservative-free tafluprost on intraocular pressure, pupil diameter, and anterior segment structures in normal canine eyes.

OBJECTIVE: This study was designed to evaluate the changes in intraocular pressure (IOP), pupil diameter (PD), and anterior segment parameters using ultrasound biomicroscopy (UBM) after instillation of preservative-free (PF) tafluprost in normal dogs. PROCEDURES: Six beagle dogs were used. PF tafluprost was instilled in one randomly selected eye, and PF artificial tear was instilled in the other eye (control). IOP and PD were measured every 15 min for the first hour, every 2 h for the next 17 h, and at 24 h and 36 h postinstillation (PI). Anterior segment parameters including geometric iridocorneal angle (ICA), width of the entry of the ciliary cleft (CCW), length of the ciliary cleft, area of the ciliary cleft, and depth of the anterior chamber were measured with UBM before and after PF tafluprost instillation. RESULTS: Compared with the control group, IOP was significantly lower from 4 h PI to 24 h PI and PD was significantly smaller from 30 min PI to 18 h PI (P < 0.05). Among UBM parameters, ICA and CCW significantly decreased and increased after PF tafluprost instillation, respectively (P < 0.05). Other parameters showed no significant changes. CONCLUSIONS: Instillation of PF tafluprost lowered IOP and induced miosis in normal canine eyes. Alterations in ICA and CCW occurred simultaneously, which probably affected the outflow of aqueous humor. PF tafluprost could be considered an alternative prostaglandin analog in the treatment of canine glaucoma.

Impact of prostaglandin glaucoma drops on platelet-activating factor action: an in vitro study.

AIM: The aim of this study was to investigate the effect of different prostaglandin analogs on platelet-activating factor (PAF) levels. METHODS: Three prostaglandin analogs were selected: bimatoprost 0.3 mg/mL, latanoprost 50 µg/mL, and tafluprost 15 µg/mL. Each drug sample was tested for its ability to cause platelet aggregation, which was measured as PAF-induced aggregation, before and after the addition of various concentrations of the examined sample, creating a linear curve of percentage inhibition (ranging from 0% to 100%) versus different concentrations of the sample. The concentration of the sample that inhibited 50% PAF-induced aggregation was calculated based on this curve, and this value was defined as IC50. In addition, the effect of eye drops on PAF metabolism was examined, through an in vitro analysis on PAF basic metabolic enzymes (PAF-cholinephosphotransferase, PAF-acetyl-CoA:1-O-alkyl-sn-glycero-3-phosphocholine acetyltransferase, and PAF-acetylhydrolase). RESULTS: The IC50 values for Lumigan UD® (bimatoprost 0.3 mg/mL), Monoprost® (latanoprost 50 µg/mL), and Saflutan (tafluprost 15 µg/mL) were 8.7, 0.28, and 1.4 µg/mL, respectively. DISCUSSION: All three prostaglandin analogs suspended PAF, but bimatoprost induced the most potent inhibition, compared to tafluprost and to the weak effect of latanoprost.

Advantages of Efficacy and Safety of Fixed-Dose Tafluprost/Timolol Combination Over Fixed-Dose Latanoprost/Timolol Combination.

PURPOSE: To compare the safety and efficacy of fixed-dose tafluprost/timolol combination (Taf/T-FDC) with those of fixed-dose latanoprost/timolol combination (Lat/T-FDC) by measuring the intraocular pressure (IOP)-lowering effect, ocular pharmacokinetics, and ocular surface toxicity. METHODS: The IOP-lowering effect of Taf/T-FDC and Lat/T-FDC in ocular normotensive monkeys was evaluated at 4 and 8 h after instillation in study A, at 12, 14, 16, and 18 h after instillation in study B, and at 24, 26, 28, and 30 h after instillation in study C. Drug penetration into the eye was evaluated by measuring the concentrations of timolol, tafluprost acid (active metabolic form of tafluprost), and latanoprost acid (active metabolic form of latanoprost) using liquid chromatography coupled with tandem mass spectrometry after single instillation of Taf/T-FDC or Lat/T-FDC to Sprague Dawley rats. Cytotoxicity following 1-30 min exposure of SV40-transformed human corneal epithelial cells to Taf/T-FDC or Lat/T-FDC was analyzed using 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assays. Undiluted and 10-fold diluted solutions of each FDC were evaluated. RESULTS: The IOP-lowering effect of Taf/T-FDC was almost equivalent to that of Lat/T-FDC at 4-8 h after instillation. The peak IOP reduction of Taf/T-FDC and Lat/T-FDC was observed at 8 h after instillation, and there is no difference between the two. The difference between them was observed at 24-30 h after instillation, and Taf/T-FDC demonstrated a significantly greater IOP-lowering effect than Lat/T-FDC at 24-30 h after instillation. The IOP-lowering effect of Taf/T-FDC was sustained up to 30 h after instillation, while that of Lat/T-FDC had almost disappeared at 28 h after instillation. Timolol concentrations in aqueous humor after Taf/T-FDC instillation were higher than those after Lat/T-FDC instillation (Cmax, 3870 ng/mL vs 1330 ng/mL; AUCinf, 3970 ng·h/mL vs 1250 ng·h/mL). The concentrations of tafluprost acid and latanoprost acid in aqueous humor after instillation of Taf/T-FDC and Lat/T-FDC, respectively, were similar to those after instillation of mono-preparations of tafluprost and latanoprost, respectively. The cytotoxic effect of Taf/T-FDC to the human corneal epithelial cells was significantly lower than that of Lat/T-FDC at all evaluated time points in both undiluted and 10-fold diluted FDCs. CONCLUSION: Taf/T-FDC provides increased IOP-lowering effect duration and lower potential ocular surface toxicity than Lat/T-FDC.