去卵巢食蟹猴眼表状态的动态观察

Dynamic observation of ocular surface in ovariectomized cynomolgus monkey

目的探讨去卵巢食蟹猴作为绝经期干眼研究动物模型的可行性。方法选择4～6岁健康成年雌性食蟹猴6只，采用随机数字表法将食蟹猴分为卵巢摘除组和假手术组，每组各3只。卵巢摘除组食蟹猴手术摘除双侧卵巢，假手术组以相同的手术方式进行操作但不摘除卵巢。分别于术前及术后1、3、6个月收集各组食蟹猴下肢静脉血各3 ml，采用放射免疫法检测猴血清雌二醇和睾酮质量浓度。分别于上述时间点采用泪液渗透压仪5600检测各组食蟹猴泪液渗透压，并进行干眼相关检查，包括基础泪液分泌试验（SⅠt）、角膜荧光素染色及结膜印迹细胞学检查。结果卵巢摘除组食蟹猴手术前后血清雌二醇、睾酮质量浓度均低于假手术组，但各组间差异均无统计学意义（F分组＝5.16，P＝0.09；F分组＝1.46，P＝0.29）；卵巢摘除组和假手术组食蟹猴组间泪液渗透压的比较，差异无统计学意义（F分组＝0.83，P＝0.37），卵巢摘除组食蟹猴术后3个月泪液渗透压较术后1个月明显升高，差异有统计学意义（t＝5.12，P〈0.05）；卵巢摘除组与假手术组食蟹猴SⅠt检测值比较，差异无统计学意义（F分组＝0.06，P＝0.82），卵巢摘除组食蟹猴术后3个月SⅠt检测值较术后1个月明显降低，差异有统计学意义（t＝1.13，P〈0.05）；各组食蟹猴手术前后各时间点均未见角膜荧光素染色。结膜印迹细胞学检查表明，卵巢摘除组食蟹猴术前及术后1个月结膜上皮细胞形态正常，结膜杯状细胞数量多，过碘酸希夫染色呈紫色，与假手术组食蟹猴结膜上皮细胞和杯状细胞形态相似；卵巢摘除组食蟹猴术后3个月结膜上皮细胞体积较大，形态多样，结膜杯状细胞体积缩小；术后6个月结膜上皮细胞形态多样且不易分辨，杯状细胞数量明显减少。结论去卵巢食蟹猴眼表结膜细胞形态发生异常变化，泪液渗透压升高，其眼表动态变化与血清雌激素水平下降趋势一致，因此去卵巢食蟹猴可作为性激素相关干眼研究的动物模型。

ObjectiveTo investigate the feasibility of ovariectomized cynomolgus monkey as an animal model of postmenopausal dry eye.MethodsSix healthy adult female cynomolgus monkeys （aged 4-6 years old） were selected.Cynomolgus monkeys were divided into ovariectomized group and sham operation group by random number table.Bilateral ovariectomy was performed in each cynomolgus of the ovariectomy group, and the same operation was operated in the sham group but the ovaries were not removed.The venous blood of the lower extremities of each cynomolgus monkey was collected before operation and at 1 month, 3 and 6 months after operation respectively.The concentrations of estradiol and testosterone were measured by radioimmunoassay.At the above time points, tear-liquid osmotic pressure detector 5600 was used to detect the tear osmotic pressure of cynomolgus monkeys in each group, and dry eye related examinations, including SchirmerⅠtest （SⅠt）, corneal fluorescein staining and conjunctive impression cytology were performed.ResultsPreoperative and postoperative serum estradiol, testosterone concentrations in the ovariectomy group were lower than those in the sham operation group, but there were no significant differences between the two groups （Fgroup=5.16, P=0.09; Fgroup=1.46, P=0.29）. There was no significant difference in the osmotic pressure between the two groups （Fgroup=0.83, P=0.37）. The osmotic pressure in the 3 months after ovariectomy was higher than that in the 1 month after ovariectomy in the ovariectomy group, with a significant difference between them （t=5.12, P〈0.05）. There was no significant difference in SⅠt between ovariectomized group and sham operation group （Fgroup=0.06, P=0.82）, the SⅠt in the 3 months after ovariectomy was lower than that in the 1 month after ovariectomy in the ovariectomy group, with a significant difference between them （t=1.13, P〈0.05）. There was no corneal fluorescein staining at each time points before and after ovariectomy in each group.The conjunctive impression cytology examination showed that, in the ovariectomy group, the morphology of conjunctival epithelial cells was normal, the number of conjunctival goblet cells was large, the periodic acid Schiff staining was purple before and 1 month after ovariectomy, which were similar to those in the sham operation group; three months after ovariectomy, the conjunctiva epithelial cell body was larger and had diverse forms, goblet cells were shrinked; six months after ovariectomy, the shapes of conjunctival epithelial cells were diverse and not easily been distinguished, the number of goblet cells was distinctly decreased.ConclusionsThe ocular conjunctival cells in the ovariectomized cynomolgus monkey have abnormal changes in morphology, and tear osmotic pressure increase, the ocular surface dynamic changes are consistent with the decrease of serum estrogen level, so ovariectomized cynomolgus monkeys can be used as sex hormone-related dry eye animal models.