Barbul A, Damewood R B, Wasserkrug H L, Penberthy L T, Efron G
J Surg Res. 1983 Jun;34(6):505-9. doi: 10.1016/0022-4804(83)90102-6.
It has been shown previously that fluid obtained from 7-day-old wounds noncytotoxically inhibits normal thymic lymphocyte blastogenesis and that mononuclear cells (MNC) from the same wounds lack mitogenic responsiveness. The present series of experiments studies whether wound MNC are the source of the wound inhibitory factor(s) and the effect of adult thymectomy (ATDX) on their generation. Adult male Sprague-Dawley rats (300-350 g), intact or ATDX (performed at 8-10 weeks of age), underwent dorsal wounding (7 cm) and subcutaneous implantation of sterile Ivalon sponges. Seven days later sponges were harvested, wound fluid was obtained, and the cell pellet was purified to 90% MNC. Normal rat thymocyte blastogenesis (stimulation index) to Con A and PHA evaluated in a microculture system (10 separate experiments) was 169.9 +/- 10.0 and 30.1 +/- 3.7. Addition of 10% wound fluid markedly inhibited thymocyte mitogenesis--6.3 +/- 1.0 and 2.7 +/- 0.6, respectively (P less than 0.001). Heat-inactivated wound fluid (56 degrees C, 30 min) had similar inhibitory activity--3.4 +/- 0.9 and 2.7 +/- 0.6 (P less than 0.001). Normal thymic blastogenesis could also be inhibited by the addition of 5 X 10(4) wound MNC to the microculture system--4.4 +/- 1.1 and 1.9 +/- 0.3 (P less than 0.001). Wound fluid from ATDX rats had much less inhibitory activity (77.1 +/- 22.4 and 7.2 +/- 2.1, P less than 0.01) vs control wound fluid. In addition wound MNC from ADTX animals were also less immune suppressive (30.7 +/- 4.9 and 13.5 +/- 3.7, P less than 0.001) than control MNC. Forty-eight-hour supernatants of wound MNC from intact rats, added in 25% concentration to normal thymocyte cultures, demonstrated inhibition similar to that of the wound fluid from the same animals: 4.4 +/- 0.7 and 3.9 +/- 0.6, while ATDX MNC supernatants had minimal inhibitory activity (110.1 +/- 18.2 and 25.7 +/- 6.5, P less than 0.005). No cytotoxicity could be demonstrated in any of these experiments by trypan blue exclusion. It is concluded that 7-day-old wound fluid noncytotoxically inhibits thymocyte blastogenesis; this effect is also demonstrated by wound MNC and their supernatants, suggesting immune "suppressor" lymphocytes are present in wounds; ATDX, which abrogates suppressor cell induction, leads to marked diminution of wound inhibitory activity. The data suggest that important immune events occur at the wound site; their relation to normal wound healing remains to be elucidated.
先前已经表明，从7日龄伤口获得的液体可无细胞毒性地抑制正常胸腺淋巴细胞的母细胞生成，并且来自相同伤口的单核细胞（MNC）缺乏有丝分裂反应性。本系列实验研究伤口MNC是否为伤口抑制因子的来源以及成年胸腺切除术（ATDX）对其产生的影响。成年雄性Sprague-Dawley大鼠（300 - 350克），完整或接受ATDX（在8 - 10周龄时进行），进行背部创伤（7厘米）并皮下植入无菌Ivalon海绵。7天后收获海绵，获得伤口液体，并将细胞沉淀纯化至90% MNC。在微培养系统中评估的正常大鼠胸腺细胞对Con A和PHA的母细胞生成（刺激指数）（10个独立实验）分别为169.9±10.0和30.1±3.7。添加10%伤口液体可显著抑制胸腺细胞有丝分裂——分别为6.3±1.0和2.7±0.6（P < 0.001）。热灭活的伤口液体（56℃，30分钟）具有相似的抑制活性——3.4±0.9和2.7±0.6（P < 0.001）。向微培养系统中添加5×10⁴个伤口MNC也可抑制正常胸腺母细胞生成——4.4±1.1和1.9±0.3（P < 0.001）。与对照伤口液体相比，ATDX大鼠的伤口液体抑制活性低得多（77.1±22.4和7.2±2.1，P < 0.01）。此外，来自ATDX动物的伤口MNC的免疫抑制作用也比对照MNC弱（30.7±4.9和13.5±3.7，P < 0.001）。来自完整大鼠的伤口MNC的48小时上清液，以25%的浓度添加到正常胸腺细胞培养物中，显示出与相同动物的伤口液体相似的抑制作用：4.4±0.7和3.9±0.6，而ATDX MNC上清液的抑制活性最小（110.1±18.2和25.7±6.5，P < 0.005）。在任何这些实验中，通过台盼蓝排斥法均未显示出细胞毒性。结论是，7日龄伤口液体可无细胞毒性地抑制胸腺细胞母细胞生成；伤口MNC及其上清液也显示出这种作用，提示伤口中存在免疫“抑制”淋巴细胞；ATDX消除了抑制细胞的诱导，导致伤口抑制活性显著降低。数据表明伤口部位发生重要的免疫事件；它们与正常伤口愈合的关系仍有待阐明。
