Laforest Sofia, Pelletier Mélissa, Michaud Andréanne, Daris Marleen, Descamps Justine, Soulet Denis, Jensen Michael D, Tchernof André
Endocrinology and Nephrology, CHU de Quebec-Laval University, 2705 Laurier Blvd. (R-4779), Quebec, QC, G1V 4G2, Canada.
Quebec Heart and Lung Institute, Quebec, Canada.
Histochem Cell Biol. 2018 Mar;149(3):209-218. doi: 10.1007/s00418-018-1635-3. Epub 2018 Jan 22.
Histomorphometric analyses of adipose tissue usually require formalin fixation of fresh samples. Our objective was to determine if intact, flash-frozen whole adipose tissue samples stored at - 80 °C could be used for measurements developed for fresh-fixed adipose tissues. Portions of adipose tissue samples were either formalin-fixed immediately upon sampling or flash-frozen and stored at - 80 °C and then formalin-fixed during the thawing process. Mean adipocyte diameter was measured. Immunohistochemistry was performed on additional samples to identify macrophage subtypes (M1, CD14 + and M2, CD206 +) and total (CD68 +) number. All slides were counterstained using haematoxylin and eosin (H&E). Visual inspection of H&E-stained adipose tissue slides performed in a blinded fashion showed little or no sign of cell breakage in 74% of frozen-fixed samples and in 68% of fresh-fixed samples (p > 0.5). There was no difference in the distribution frequencies of adipocyte sizes in fresh-fixed vs. frozen-fixed tissues in both depots (p > 0.9). Mean adipocyte size from frozen-fixed samples correlated significantly and positively with adipocyte size from fresh-fixed samples (r = 0.74, p < 0.0001, for both depots). The quality of staining/immunostaining and appearance of tissue architecture were comparable in fresh-fixed vs. frozen-fixed samples. In conclusion, intact flash-frozen adipose tissue samples stored at - 80 °C can be used to perform techniques conventionally applied to fresh-fixed samples. This approach allows for retrospective studies with frozen human adipose tissue samples.
脂肪组织的组织形态计量学分析通常需要对新鲜样本进行福尔马林固定。我们的目的是确定保存在-80°C的完整、速冻的全脂肪组织样本是否可用于针对新鲜固定脂肪组织开发的测量方法。脂肪组织样本的一部分在采样后立即进行福尔马林固定，或速冻并保存在-80°C，然后在解冻过程中进行福尔马林固定。测量平均脂肪细胞直径。对额外的样本进行免疫组织化学分析，以识别巨噬细胞亚型（M1，CD14+和M2，CD206+）以及总数（CD68+）。所有玻片均用苏木精和伊红（H&E）复染。以盲法对H&E染色的脂肪组织玻片进行目视检查，结果显示，74%的冷冻固定样本和68%的新鲜固定样本几乎没有或没有细胞破裂迹象（p>0.5）。在两个储存库中，新鲜固定组织与冷冻固定组织中脂肪细胞大小的分布频率没有差异（p>0.9）。冷冻固定样本的平均脂肪细胞大小与新鲜固定样本的脂肪细胞大小显著正相关（两个储存库的r=0.74，p<0.0001）。新鲜固定样本与冷冻固定样本的染色/免疫染色质量和组织结构外观具有可比性。总之，保存在-80°C的完整速冻脂肪组织样本可用于执行常规应用于新鲜固定样本的技术。这种方法允许对冷冻的人类脂肪组织样本进行回顾性研究。
