Chen S-E, Jin B, Li Y-P. TNF-␣ regulates myogenesis and muscle regeneration by activating p38 MAPK. Am J Physiol Cell Physiol 292: C1660 -C1671, 2007. First published December 6, 2006; doi:10.1152/ajpcell.00486.2006.-Although p38 MAPK activation is essential for myogenesis, the upstream signaling mechanism that activates p38 during myogenesis remains undefined. We recently reported that p38 activation, myogenesis, and regeneration in cardiotoxininjured soleus muscle are impaired in TNF-␣ receptor double-knockout (p55) mice. To fully evaluate the role of TNF-␣ in myogenic activation of p38, we tried to determine whether p38 activation in differentiating myoblasts requires autocrine TNF-␣, and whether forced activation of p38 rescues impaired myogenesis and regeneration in the p55Ϫ/Ϫ soleus. We observed an increase of TNF-␣ release from C2C12 or mouse primary myoblasts placed in low-serum differentiation medium. A TNF-␣-neutralizing antibody added to differentiation medium blocked p38 activation and suppressed differentiation markers myocyte enhancer factor (MEF)-2C, myogenin, p21, and myosin heavy chain in C2C12 myoblasts. Conversely, recombinant TNF-␣ added to differentiation medium stimulated myogenesis at 0.05 ng/ml while inhibited it at 0.5 and 5 ng/ml. In addition, differentiation medium-induced p38 activation and myogenesis were compromised in primary myoblasts prepared from p55Ϫ/Ϫ mice. Increased TNF-␣ release was also seen in cardiotoxin-injured soleus over the course of regeneration. Forced activation of p38 via the constitutive activator of p38, MKK6bE, rescued impaired myogenesis and regeneration in the cardiotoxin-injured p55Ϫ/Ϫ soleus. These results indicate that TNF-␣ regulates myogenesis and muscle regeneration as a key activator of p38. myocyte enhancer factor-2C; myogenin; p21; myosin heavy chain; Akt; tumor necrosis factor-␣; mitogen-activated protein kinase SKELETAL MUSCLE HAS A REMARKABLE ability to regenerate itself. However, muscle regeneration is a complex process comprising many highly coordinated events in a sequence of satellite cell activation, proliferation, and differentiation (myogenesis) that repairs damaged myofibers or forms new ones. The activation of satellite cells is characterized by the expression of myogenic regulatory factors (MRFs) Myf 5 and MyoD. After the proliferation phase, satellite cells express myogenin and MRF4 to initiate myogenic differentiation. This is followed by expression of the Cdk inhibitor p21 and a permanent exit from the cell cycle (reviewed in Ref. 11). The regenerative process requires a complicated array of intrinsic and extrinsic signals that regulate various satellite cell activities. Thus, despite many past efforts, our understanding of the intrinsic and extrinsic signals that regulate muscle regeneration remains limited.Initiation of the myogenic program in adult muscle is a critical step in skeletal muscle regeneration, which requires chromatin remodeling in myogenic cells that allows transcriptional activation of myogenic target genes. The act...

Heme oxygenase (HO)-1 is known to metabolize heme into biliverdin/bilirubin, carbon monoxide, and ferrous iron, and it has been suggested to demonstrate cytoprotective effects against various stress-related conditions. HO-1 is commonly regarded as a survival molecule, exerting an important role in cancer progression and its inhibition is considered beneficial in a number of cancers. However, increasing studies have shown a dark side of HO-1, in which HO-1 acts as a critical mediator in ferroptosis induction and plays a causative factor for the progression of several diseases. Ferroptosis is a newly identified iron- and lipid peroxidation-dependent cell death. The critical role of HO-1 in heme metabolism makes it an important candidate to mediate protective or detrimental effects via ferroptosis induction. This review summarizes the current understanding on the regulatory mechanisms of HO-1 in ferroptosis. The amount of cellular iron and reactive oxygen species (ROS) is the determinative momentum for the role of HO-1, in which excessive cellular iron and ROS tend to enforce HO-1 from a protective role to a perpetrator. Despite the dark side that is related to cell death, there is a prospective application of HO-1 to mediate ferroptosis for cancer therapy as a chemotherapeutic strategy against tumors.

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Recent data suggest a physiological role for the proinflammatory cytokine TNF-alpha in skeletal muscle regeneration. However, the underlying mechanism is not understood. In the present study, we analyzed TNF-alpha-activated signaling pathways involved in myogenesis in soleus muscle injured by cardiotoxin (CTX) in TNF-alpha receptor double-knockout mice (p55(-/-)p75(-/-)). We found that activation of p38MAPK, which is critical for myogenesis, was blocked in CTX-injured p55(-/-)p75(-/-) soleus on day 3 postinjury when myogenic differentiation was being initiated, while activation of ERK1/2 and JNK MAPK, as well as transcription factor NF-kappaB, was not reduced. Consequently, the phosphorylation of transcription factor myocyte enhancer factor-2C, which is catalyzed by p38 and crucial for the expression of muscle-specific genes, was blunted. Meanwhile, expression of p38-dependent differentiation marker myogenin and p21 were suppressed. In addition, expression of cyclin D1 was fivefold that in wild-type (WT) soleus. These results suggest that myogenic differentiation is blocked or delayed in the absence of TNF-alpha signaling. Histological studies revealed abnormalities in regenerating p55(-/-)p75(-/-) soleus. On day 5 postinjury, new myofiber formation was clearly observed in WT soleus but not in p55(-/-)p75(-/-) soleus. To the contrary, p55(-/-)p75(-/-) soleus displayed renewed inflammation and dystrophic calcification. On day 12 postinjury, the muscle architecture of WT soleus was largely restored. Yet, in p55(-/-)p75(-/-) soleus, multifocal areas of inflammation, myofiber death, and myofibers with smaller cross-sectional area were observed. Functional studies demonstrated an attenuated recovery of contractile force in injured p55(-/-)p75(-/-) soleus. These data suggest that TNF-alpha signaling plays a critical regulatory role in muscle regeneration.

manner. Stretch stimulated the cleavage activity of TACE. Conversely, TACE inhibitor TAPI or TACE siRNA abolished stretch activation of p38. In addition, conditioned medium from stretched myoblast cultures activated p38 in unstretched myoblasts, which required TACE activity in the donor myoblasts, and TNF-␣ receptors in the recipient myoblasts. These results indicate that posttranscriptional activation of TACE mediates the mechanotransduction that activates p38-dependent myogenesis via the release of TNF-␣.

In mammals, triacylglycerol (TAG) accumulation in nonadipose tissue, termed lipotoxicity, develops with obesity and can provoke insulin resistance, overt diabetes, and ovarian dysfunction. Leptin, an adipose tissue hormone, may mediate these effects. Feed-satiated broiler breeder hens manifest lipotoxicity-like symptoms. Changes in body and organ weights, hepatic and plasma TAG, nonesterified fatty acids (NEFA), ovarian morphology, and egg production in response to acute voluntary increases of feed intake were measured in 2 studies with Cobb 500 broiler breeder hens provided with either 145 or > or = 290 g of feed/d per hen for 10 d. In both studies, no hen fed 145 g of feed/d exhibited ovarian abnormalities, whereas approximately 50% of feed-satiated hens did. Egg production in feed-satiated hens was reduced from 73.3 to 55.8% (P = 0.001). Morphology indicated that apoptosis-induced atresia occurred in the hierarchical follicles. Fractional weight of yolk increased from 29.3 to 30.6% (P = 0.016) and no longer correlated to egg weight. Body, liver, and abdominal adipose weights were significantly greater (P < 0.05) in feed-satiated hens, as were plasma concentrations of glucose, NEFA, TAG, insulin, and leptin (P < 0.05). Feed-satiated hens with abnormal ovaries had significantly more liver and abdominal fat, greater plasma leptin and TAG concentrations, and more saturated fatty acids in plasma NEFA than did feed-satiated hens with normal ovaries. Differences in severity of lipotoxic metabolic and hormonal responses among feed-satiated hens were closely linked to the incidence of ovarian abnormalities and granulosa cell susceptibility to apoptosis and necrosis.

Skeletal muscle satellite cells can sense various forms of environmental cues and initiate coordinated signaling that activates myogenesis. Although this process involves cellular membrane receptor integrins, the role of integrins in myogenesis is not well defined. Here, we report a regulatory role of β3-integrin, which was previously thought not expressed in muscle, in the initiation of satellite cell differentiation. Undetected in normal muscle, β3-integrin expression in mouse hindlimb muscles is induced dramatically from 1 to 3 d after injury by cardiotoxin. The source of β3-integrin expression is found to be activated satellite cells. Proliferating C2C12 myoblasts also express β3-integrin, which is further up-regulated transiently on differentiation. Knockdown of β3-integrin expression attenuates Rac1 activity, impairs myogenic gene expression, and disrupts focal adhesion formation and actin organization, resulting in impaired myoblast migration and myotube formation. Conversely, overexpression of constitutively active Rac1 rescues myotube formation. In addition, a β3-integrin-neutralizing antibody similarly blocked myotube formation. Comparing with wild-type littermates, myogenic gene expression and muscle regeneration in cardiotoxin-injured β3-integrin-null mice are impaired, as indicated by depressed expression of myogenic markers and morphological disparities. Thus, β3-integrin is a mediator of satellite cell differentiation in regenerating muscle.

SummaryMyogenic differentiation in adult muscle is normally suppressed and can be activated by myogenic cues in a subset of activated satellite cells. The switch mechanism that turns myogenesis on and off is not defined. In the present study, we demonstrate that tissue inhibitor of metalloproteinase 3 (TIMP3), the endogenous inhibitor of TNFa-converting enzyme (TACE), acts as an on-off switch for myogenic differentiation by regulating autocrine TNFa release. We observed that constitutively expressed TIMP3 is transiently downregulated in the satellite cells of regenerating mouse hindlimb muscles and differentiating C2C12 myoblasts. In C2C12 myoblasts, perturbing TIMP3 downregulation by overexpressing TIMP3 blocks TNFa release, p38 MAPK activation, myogenic gene expression and myotube formation. TNFa supplementation at a physiological concentration rescues myoblast differentiation. Similarly, in the regenerating soleus, overexpression of TIMP3 impairs release of TNFa and myogenic gene expression, and delays the formation of new fibers. In addition, downregulation of TIMP3 is mediated by the myogenesis-promoting microRNA miR-206. Thus, TIMP3 is a physiological regulator of myogenic differentiation.