フジタ　キヨウヘイ   FUJITA Kiyouhei   藤田　恭平    所属   医学部 医学科    職種   助教
言語種別	英語
発表タイトル	Studying markers of metabolism, apoptosis, and necrosis in cullin-3 skeletal muscle knockouts
会議名	The 29th Annual Congress of the World Muscle Society
学会区分	国際学会及び海外の学会
発表形式	ポスター掲示
講演区分	一般
発表者・共同発表者	J. Herskind, K. Fujita, V. Dato, V. Fochi, J. Blondelle, B. Medelyte, J. Seto, Y. Jones, G. Castillon, M. Ghassemian, J. Singer, S. Lange
発表年月日	2024/10
開催地 （都市, 国名）	Prague, Czechia
開催期間	2024/10/08～2024/10/12
概要	Loss of cullin-3 in skeletal muscle (Cul3-skmKO) leads to postnatal death in mice caused by respiratory distress. Muscles appeared underdeveloped, and muscle mass was reduced by approximately 50% compared to controls owing to reduced hypertrophy of developing muscles. Our histological analyses using Gomori trichrome stained diaphragms found aggregates in Cul3-skmKO that are reminiscent of nemaline bodies, characteristic for nemaline myopathy. We also performed proteome and enrichment analyses of diaphragm and tongue muscles. These results indicated a reduction of mitochondrial metabolism and energy generation as well as sarcomerogenesis in Cul3-skmKO, while proteins related to reactive oxygen species metabolic processes are deregulated. The latter finding suggests that Cul3-skmKO may undergo metabolic stress, leading to ROS production and induction of cell death. We used immune-blot analyses to study changes to mitochondrial and sarcomeric proteins, and markers for apoptosis and necrosis in Cul3-skmKO and control diaphragms. We also performed ultrastructural analyses using electron microscopy to identify the nature of the pathological protein aggregates, quantify mitochondrial content and size, and correlate changes to the biochemical analyses. Our ultrastructural analyses revealed unchanged mitochondrial content and sizes between Cul3-skmKO and controls. We observed electron dense material in Cul3-skmKO diaphragms that could stem from cells undergoing cell death, giving rise to Gomori trichrome positive aggregates. However, levels of apoptotic and necrotic marker proteins remained unchanged when normalized to sarcomeric myosin or decreased when normalized to β-actin. Biochemical analysis of electron transport chain and sarcomeric proteins confirmed data from our proteome analyses when normalized to β-actin. Conversely, when normalized to sarcomeric myosin, no differences could be identified. This suggests a decreased contribution of muscle tissues to the total tissue sample analysed by immunoblots in skmKO, perhaps reflective of the earlier identified loss in overall muscle mass. Further studies are required to identify if metabolic stress leading to production of reactive oxygen species may be involved the origin and development of the observed cell death in Cul3-skmKO muscles.