Department of Biochemistry and Molecular Biology, Harbin Medical University, Harbin, 150086, China.
Heilongjiang Academy of Medical Sciences, Harbin, 150086, China.
Skelet Muscle. 2021 Mar 30;11(1):9. doi: 10.1186/s13395-021-00263-8.
ALAS2 (delta-aminolevulinate synthase 2) is one of the two isoenzymes catalyzing the synthesis of delta-aminolevulinic acid (ALA), which is the first precursor of heme synthesis. ALAS2-overexpressing transgenic mice (Tg mice) showed syndrome of porphyria, a series of diseases related to the heme anabolism deficiency. Tg mice showed an obvious decrease in muscle size. Muscle atrophy results from a decrease in protein synthesis and an increase in protein degradation, which ultimately leads to a decrease in myofiber size due to loss of contractile proteins, organelles, nuclei, and cytoplasm.
The forelimb muscle grip strength of age-matched ALAS-2 transgenic mice (Tg mice) and wild-type mice (WT mice) were measured with an automated grip strength meter. The activities of serum LDH and CK-MB were measured by Modular DPP. The histology of skeletal muscle (quadriceps femoris and gastrocnemius) was observed by hematoxylin and eosin (HE) staining, immunohistochemistry, and transmission electron microscope. Real-time PCR was used to detect mtDNA content and UCP3 mRNA expression. Evans blue dye staining was used to detect the membrane damage of the muscle fiber. Single skeletal muscle fiber diameter was measured by single-fiber analyses. Muscle adenosine triphosphate (ATP) levels were detected by a luminometric assay with an ATP assay kit.
Compared with WT mice, the strength of forelimb muscle and mass of gastrocnemius were decreased in Tg mice. The activities of serum CK-MB and LDH, the number of central nuclei fibers, and Evans blue positive fibers were more than those in WT mice, while the diameter of single fibers was smaller, which were associated with suppressed expression levels of MHC, myoD1, dystrophin, atrogin1, and MuRF1. Re-expression of eMyHC was only showed in the quadriceps of Tg mice, but not in WT mice. Muscle mitochondria in Tg mice showed dysfunction with descented ATP production and mtDNA content, downregulated UCP3 mRNA expression, and swelling of mitochondria.
ALAS2 overexpressing-transgenic mice (Tg mice) showed muscle dystrophy, which was associated with decreased atrogin-1 and MuRF-1, and closely related to mitochondrial dysfunction.
ALAS2（δ-氨基酮戊酸合酶 2）是催化δ-氨基酮戊酸（ALA）合成的两种同工酶之一，ALA 是血红素合成的第一个前体。ALAS2 过表达转基因小鼠（Tg 小鼠）表现出卟啉症综合征，这是一系列与血红素生物合成缺陷相关的疾病。Tg 小鼠的肌肉大小明显减小。肌肉萎缩是由于蛋白质合成减少和蛋白质降解增加，最终导致由于收缩蛋白、细胞器、核和细胞质丢失而使肌纤维大小减小。
使用自动握力计测量年龄匹配的 ALAS-2 转基因小鼠（Tg 小鼠）和野生型小鼠（WT 小鼠）的前肢肌肉握力。通过 Modular DPP 测量血清 LDH 和 CK-MB 的活性。通过苏木精和伊红（HE）染色、免疫组织化学和透射电镜观察骨骼肌（股四头肌和腓肠肌）的组织学。实时 PCR 用于检测 mtDNA 含量和 UCP3 mRNA 表达。伊文思蓝染色用于检测肌肉纤维的膜损伤。通过单纤维分析测量单骨骼肌纤维直径。用发光测定法和 ATP 测定试剂盒检测肌肉三磷酸腺苷（ATP）水平。
与 WT 小鼠相比，Tg 小鼠的前肢肌肉力量和腓肠肌质量下降。血清 CK-MB 和 LDH 的活性、中央核纤维的数量和伊文思蓝阳性纤维增多，而单纤维直径较小，这与 MHC、myoD1、dystrophin、atrogin1 和 MuRF1 的表达水平降低有关。eMyHC 的重新表达仅在 Tg 小鼠的股四头肌中显示，而在 WT 小鼠中未显示。Tg 小鼠的肌肉线粒体功能障碍，表现为 ATP 生成和 mtDNA 含量下降、UCP3 mRNA 表达下调以及线粒体肿胀。
ALAS2 过表达转基因小鼠（Tg 小鼠）表现出肌肉萎缩，这与 atrogin-1 和 MuRF-1 减少有关，与线粒体功能障碍密切相关。
