Buch S K, Khurdayan V K, Lutz S E, Knapp P E, El-Hage N, Hauser K F
Department of Anatomy and Neurobiology, University of Kentucky, College of Medicine, 800 Rose Street, Lexington, KY 40536-0298, USA.
Neuroscience. 2007 Jun 8;146(4):1546-54. doi: 10.1016/j.neuroscience.2007.03.006. Epub 2007 May 2.
Recent evidence suggests that human immunodeficiency virus (HIV)-induced pathogenesis is exacerbated by opioid abuse and that the synergistic toxicity may result from direct actions of opioids in immature glia or glial precursors. To assess whether opioids and HIV proteins are directly toxic to glial-restricted precursors (GRPs), we isolated neural stem cells from the incipient spinal cord of embryonic day 10.5 ICR mice. GRPs were characterized immunocytochemically and by reverse transcriptase-polymerase chain reaction (RT-PCR). At 1 day in vitro (DIV), GRPs failed to express mu opioid receptors (MOR or MOP) or kappa-opioid receptors (KOR or KOP); however, at 5 DIV, most GRPs expressed MOR and KOR. The effects of morphine (500 nM) and/or Tat (100 nM) on GRP viability were assessed in GRPs at 5 DIV by examining the apoptotic effector caspase-3 and cell viability (ethidium monoazide exclusion) at 96 h following continuous exposure. Tat or morphine alone or in combination caused significant increases in GRP cell death at 96 h, but not at 24 h, following exposure. Although morphine or Tat caused increases in caspase-3 activity at 4 h, this was not accompanied with increased cleaved caspase-3 immunoreactive or ethidium monoazide-positive dying cells at 24 h. The results indicate that prolonged morphine or Tat exposure is intrinsically toxic to isolated GRPs and/or their progeny in vitro. Moreover, MOR and KOR are widely expressed by Sox2 and/or Nkx2.2-positive GRPs in vitro and the pattern of receptor expression appears to be developmentally regulated. The temporal requirement for prolonged morphine and HIV-1 Tat exposure to evoke toxicity in glia may coincide with the attainment of a particular stage of maturation and/or the development of particular apoptotic effector pathways and may be unique to spinal cord GRPs. Should similar patterns occur in vivo then we predict that immature astroglia and oligodendroglia may be preferentially vulnerable to HIV-1 infection or chronic opiate exposure.
最近的证据表明，阿片类药物滥用会加剧人类免疫缺陷病毒（HIV）引发的发病机制，且这种协同毒性可能源于阿片类药物对未成熟神经胶质细胞或神经胶质前体细胞的直接作用。为了评估阿片类药物和HIV蛋白是否对神经胶质限制前体细胞（GRP）具有直接毒性，我们从胚胎第10.5天的ICR小鼠的初始脊髓中分离出神经干细胞。通过免疫细胞化学和逆转录聚合酶链反应（RT-PCR）对GRP进行了表征。在体外培养1天（DIV）时，GRP未能表达μ阿片受体（MOR或MOP）或κ阿片受体（KOR或KOP）；然而，在5 DIV时，大多数GRP表达了MOR和KOR。通过在连续暴露96小时后检测凋亡效应因子半胱天冬酶-3和细胞活力（单叠氮溴化乙锭排除法），评估了吗啡（500 nM）和/或Tat（100 nM）对5 DIV时GRP活力的影响。单独或联合使用Tat或吗啡在暴露后96小时导致GRP细胞死亡显著增加，但在24小时时未出现这种情况。尽管吗啡或Tat在4小时时导致半胱天冬酶-3活性增加，但在24小时时并未伴随裂解的半胱天冬酶-3免疫反应性增加或单叠氮溴化乙锭阳性的死亡细胞增加。结果表明，长期暴露于吗啡或Tat对体外分离的GRP及其后代具有内在毒性。此外，MOR和KOR在体外被Sox2和/或Nkx2.2阳性的GRP广泛表达，且受体表达模式似乎受发育调控。长期暴露于吗啡和HIV-1 Tat以诱发神经胶质细胞毒性的时间要求可能与达到特定成熟阶段和/或特定凋亡效应途径的发展相吻合，并且可能是脊髓GRP所特有的。如果体内出现类似模式，那么我们预测未成熟的星形胶质细胞和少突胶质细胞可能更容易受到HIV-1感染或慢性阿片类药物暴露的影响。
