Shambaugh G E, Haines G K, Koch A, Lee E J, Zhou J n, Pestell R
Department of Medicine, Northwestern University Medical School and Veterans Affairs Chicago Health Care System, Lakeside Division, 333 East Huron St., Chicago, IL 60611-3008, USA.
Brain Res. 2000 Feb 7;855(1):11-22. doi: 10.1016/s0006-8993(99)02028-4.
Expression of the cyclin-dependent kinase inhibitors (CKIs) has been linked to the inhibition of cellular proliferation and the induction of differentiation. Based on structure function analysis, two distinct families of CDKIs, the INK4 and the Cip/Kip family have been identified. The INK4 family member p16(Ink4), and the Cip/Kip protein p27(Kip1) have been implicated in normal development of the CNS and cerebellum. Recent studies have suggested a functional inter-dependence between the CKI and the abundance of cyclin D1 in orchestrating growth factor-induced cellular proliferation. The neonatal rat cerebellum undergoes proliferative growth and differentiation, localized to distinct topographical regions and cell types. The cell type and the temporal profile of CKI expression during postnatal cerebellar development had not been described. The current studies determined the specific cerebellar cell types in which the CKIs were expressed during post natal development by co-staining for cell-type specific markers. p16(Ink4a) and p27(Kip1) immunostaining was identified in both neurons and glial cells, increasing progressively between postnatal days 6 to 13 into adulthood. By contrast, neuronal and glial cell p21(Cip1) staining was prominent at days 6-11 and decreased thereafter. Cyclin D1 was expressed in the proliferating external granular cells, with occassional staining in the molecular, and internal granular layers. Dual immunostaining demonstrated cyclin D1 within cells expressing CKI (p16(Ink4a), p21(Cip1),p27(Kip1)). Cerebellar cellular growth arrest, induced by protein-calorie malnutrition, inhibited cyclin D1 protein levels without affecting CKI immunostaining suggesting CKI do not mediate the developmental arrest. These results demonstrate that the CKIs are induced by differentiation cues in specific cell types with distinct kinetics in the developing cerebellum in vivo.
细胞周期蛋白依赖性激酶抑制剂（CKIs）的表达与细胞增殖的抑制和分化的诱导有关。基于结构功能分析，已鉴定出两个不同的CDKIs家族，即INK4和Cip/Kip家族。INK4家族成员p16(Ink4)和Cip/Kip蛋白p27(Kip1)与中枢神经系统和小脑的正常发育有关。最近的研究表明，在协调生长因子诱导的细胞增殖过程中，CKI与细胞周期蛋白D1的丰度之间存在功能上的相互依赖关系。新生大鼠小脑经历增殖性生长和分化，定位于不同的地形区域和细胞类型。出生后小脑发育过程中CKI表达的细胞类型和时间模式尚未描述。目前的研究通过对细胞类型特异性标记物进行共染色，确定了出生后发育过程中表达CKIs的特定小脑细胞类型。在神经元和神经胶质细胞中均检测到p16(Ink4a)和p27(Kip1)免疫染色，从出生后第6天到第13天逐渐增加直至成年。相比之下，神经元和神经胶质细胞的p21(Cip1)染色在第6 - 11天很突出，此后减少。细胞周期蛋白D1在增殖的外颗粒细胞中表达，在分子层和内颗粒层偶尔有染色。双重免疫染色显示细胞周期蛋白D1在表达CKI（p16(Ink4a)、p21(Cip1)、p27(Kip1)）的细胞内。蛋白质-热量营养不良诱导的小脑细胞生长停滞抑制了细胞周期蛋白D1蛋白水平，但不影响CKI免疫染色，表明CKI不介导发育停滞。这些结果表明，在体内发育中的小脑中，CKIs是由特定细胞类型中的分化信号以不同的动力学诱导产生的。
