Schmidt A J, Woerthwein K F
Differentiation. 1975 Aug 11;3(1-3):29-42. doi: 10.1111/j.1432-0436.1975.tb00843.x.
The regenerating forelimb of the adult newt, Notophthalmus viridescens was investigated for 5'-nucleotidase (5' ribonucleotide phosphohydrolase, 3.1.3.5) acitivity. The newt's humeri were surgically removed, and after a twenty-one-day recovery period, the forelimbs amputated above the elbows. Regenerates were sampled at predetermined times for specific phases in the progress of regeneration, frozen, sectioned in a cryostat, and the sections fixed in 10% cold formol calcium. The Wachstein and Meisel [25] lead procedure at neutral pH was used predominately in these experiments, although tests were also conducted with Gomori's [14] calcium, Allen's [21] highly alkaline procedures. The substrates used to obtain specific enzyme reactions were adenine, cytosine, guanine, uracil and inosine 5'-monophosphate nucleotides. Sodium beta-glycerophosphate served as a non-specific phosphomonoesterase substrate, distilled water replaced substrate, and inhibitors such as zinc and cyanide ions were used as control measures to assist in increasing the precision in interpreting the results obtained. The most reactive 5'-nucleotidase (5'-Nase) loci were in the walls of the blood vascular system, mysial and neural sheaths, dermis, and periosteum: the principal cells involved were macrophages, endothelium of blood vessels, and fibrocytes of connective tissues. A moderate enzyme response was elicited from secretory cells of some of the subcutaneous glands, hypertrophied chondrocytes and osteogenic centers, chondrocytes in the articular regions and within red blood cells and leucocytes. Normal, injured and degenerating, or regenerating striated muscle and nerve fibers were judged unreactive for 5'-Nase. The epidermis and wound epithelium displayed negative responses for 5'-Nase. Cells forming the regeneration blastema were 5'-Nase reactive during the early formative phase, but with growth and development of the blastema into bulb and conic forms, these cells did not respond for this enzyme-activity. One suggestion offered is that the absence of 5'-Nase in cells of the blastema may be related to the lack of an adequate blood-vascular supply. Several functions of 5'-Nase in normal and regenerating tissues are discussed. A basic conclusion reached is that 5'-nucleotidase hydrolyses may be more involved in fundamental anabolic than in catabolic metabolism.
对成年绿红东美螈再生的前肢进行了5'-核苷酸酶（5'-核糖核苷酸磷酸水解酶，3.1.3.5）活性研究。通过手术切除蝾螈的肱骨，在21天的恢复期后，于肘部上方截断前肢。在再生过程的特定阶段，于预定时间采集再生组织样本，冷冻后在低温恒温器中切片，切片用10%冷甲醛钙固定。在这些实验中，主要采用Wachstein和Meisel [25]在中性pH条件下的铅染色法，不过也用Gomori [14]的钙染色法、Allen [21]的高碱性染色法进行了测试。用于获得特定酶反应的底物有腺嘌呤、胞嘧啶、鸟嘌呤、尿嘧啶和肌苷5'-单磷酸核苷酸。β-甘油磷酸钠用作非特异性磷酸单酯酶底物，用蒸馏水替代底物，并使用锌离子和氰离子等抑制剂作为对照措施，以提高解释所得结果的准确性。反应活性最强的5'-核苷酸酶（5'-Nase）位点位于血管系统壁、肌鞘和神经鞘、真皮和骨膜中：涉及的主要细胞为巨噬细胞、血管内皮细胞和结缔组织的纤维细胞。一些皮下腺的分泌细胞、肥大的软骨细胞和成骨中心、关节区域的软骨细胞以及红细胞和白细胞内引发了中等程度的酶反应。正常、受伤、退化或再生的横纹肌和神经纤维被判定对5'-Nase无反应。表皮和伤口上皮对5'-Nase呈阴性反应。形成再生芽基的细胞在早期形成阶段对5'-Nase有反应，但随着芽基生长发育成球茎状和圆锥状，这些细胞对该酶活性无反应。有人提出一种观点，即芽基细胞中5'-Nase的缺失可能与缺乏充足的血管供应有关。讨论了5'-Nase在正常组织和再生组织中的几种功能。得出的一个基本结论是，5'-核苷酸酶水解可能更多地参与基本的合成代谢而非分解代谢。
