Koh S W, Yeh T H, Morris S M, Leffler M, Higginbotham E J, Brenneman D E, Yue B Y
Department of Ophthalmology, University of Maryland at Baltimore 21201, USA.
Invest Ophthalmol Vis Sci. 1997 Dec;38(13):2781-9.
To demonstrate that vasoactive intestinal peptide (VIP), a 28-amino acid neuropeptide, is a growth factor of human trabecular meshwork (TM) cells in culture and in a corneoscleral explant organ culture treated with laser trabeculoplasty (LTP).
Proliferating human TM cells in cell cultures were incubated with VIP for 20 hours, followed by total cell number determination, using a Coulter counter. The percentage of proliferating TM cells was assessed, using an antibody against the proliferating cell nuclear antigen (PCNA). To test the growth effect of VIP on TM cells in situ, corneoscleral explants in organ cultures were first treated with argon LTP to initiate TM-cell proliferation and then were exposed to VIP for 48 hours. The mitotic TM cells were demonstrated immunocytochemically, using anti-PCNA in paraffin sections of the explants; and the total number of TM cells was determined after paraffin sections were counterstained by hematoxylin.
Vasoactive intestinal peptide dose-dependently stimulated the proliferation of TM cells in cell culture. Treatment with 5 x 10(-10) M VIP resulted in a maximal increase of 40% in cell number. The effect of VIP was blocked by a VIP antagonist. The number of PCNA-stained TM cells and the total cell number in the TM in LTP-treated corneoscleral explants were increased by VIP.
Exogenously applied VIP stimulated the proliferation of human TM cells in subconfluent cultures and in LTP-treated corneoscleral explants. In that LTP has been shown to increase the number of TM cells in situ, the growth stimulatory effect of VIP may help enhance this therapy.
证明血管活性肠肽(VIP),一种由28个氨基酸组成的神经肽,是培养的人小梁网(TM)细胞以及经激光小梁成形术(LTP)处理的角膜巩膜外植体器官培养中的生长因子。
将细胞培养中增殖的人TM细胞与VIP孵育20小时,然后使用库尔特计数器测定总细胞数。使用抗增殖细胞核抗原(PCNA)抗体评估增殖的TM细胞百分比。为了测试VIP对原位TM细胞的生长作用,首先对器官培养中的角膜巩膜外植体进行氩激光小梁成形术以启动TM细胞增殖,然后将其暴露于VIP 48小时。使用抗PCNA对植入物石蜡切片进行免疫细胞化学染色以显示有丝分裂的TM细胞;在用苏木精对石蜡切片进行复染后测定TM细胞的总数。
血管活性肠肽剂量依赖性地刺激细胞培养中TM细胞的增殖。用5×10⁻¹⁰ M VIP处理导致细胞数量最大增加40%。VIP拮抗剂可阻断VIP的作用。VIP增加了LTP处理的角膜巩膜外植体中PCNA染色的TM细胞数量和TM中的总细胞数量。
外源性应用的VIP刺激了亚汇合培养物中和LTP处理的角膜巩膜外植体中人类TM细胞的增殖。鉴于LTP已被证明可增加原位TM细胞数量,VIP的生长刺激作用可能有助于增强这种治疗方法。