Long term culture of rat soleus muscle in vitro

Stace, Pamela B.; Marchington, David R.; Kerbey, A L; Randle, P. J.

doi:10.1016/0014-5793(90)81058-v

Cited by 17 publications

(4 citation statements)

References 24 publications

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“…Previously reported culture methods of isolated skeletal muscles were based on the incubation in a shaking water bath at 35°C with O 2 /CO 2 insufflation: the muscles were maintained under such conditions until 18 h and then submitted to biochemical analysis. 3 , 14-17 The bioreactor used for the present study seems to be more efficient as it may be directly placed inside the incubator, thus allowing to maintain steady levels of temperature, humidity, O 2 and CO 2 , which are essential factors to improve cell survival under in vitro conditions. In addition, this fluid dynamic system may ensure a culture environment more similar to the physiological one than other previously proposed incubation methods: in our experiment, the flow rate in the bioreactor was set at 300 μL/min, to reproduce the blood flow values in skeletal muscle, 18 but the fluid flow may be finely modulated, to mimic metabolic changes for functional studies.…”

Section: Resultsmentioning

confidence: 99%

Incubation under fluid dynamic conditions markedly improves the structural preservation in vitro of explanted skeletal muscles

2017

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Explanted organs and tissues represent suitable experimental systems mimicking the functional and structural complexity of the living organism, with positive ethical and economic impact on research activities. However, their preservation in culture is generally limited, thus hindering their application as experimental models for biomedical research. In the present study, we investigated the potential of an innovative fluid dynamic culture system to improve the structural preservation in vitro of explanted mouse skeletal muscles (soleus). We used light and transmission electron microscopy to compare the morphological features of muscles maintained either in multiwell plates under conventional conditions or in a bioreactor mimicking the flow of physiological fluids. Our results demonstrate that fluid dynamic conditions markedly slowed the progressive structural deterioration of the muscle tissue occurring during the permanence in the culture medium, prolonging the preservation of some organelles such as mitochondria up to 48 h.

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Section: Resultsmentioning

confidence: 99%

Incubation under fluid dynamic conditions markedly improves the structural preservation in vitro of explanted skeletal muscles

2017

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“…The protocol used was quite similar to that described by Arias et al [25] and modified from Stace et al [26]. Freshly excised muscles were incubated for 12 hours in dish plates containing 2 ml of DMEM supplemented to bring final concentrations of 5.5 mM glucose, 2.54 mM CaCl 2 , 25 mM NaHCO 3 , 0.6 nM insulin, 0.1% BSA, 100 µU/ml penicillin and 100 µg/ml streptomycin, the media being replaced with fresh media every 4 hours.…”

Section: Methodsmentioning

confidence: 99%

Increasing O-GlcNAcylation Level on Organ Culture of Soleus Modulates the Calcium Activation Parameters of Muscle Fibers

et al. 2012

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O-N-acetylglucosaminylation is a reversible post-translational modification which presents a dynamic and highly regulated interplay with phosphorylation. New insights suggest that O-GlcNAcylation might be involved in striated muscle physiology, in particular in contractile properties such as the calcium activation parameters. By the inhibition of O-GlcNAcase, we investigated the effect of the increase of soleus O-GlcNAcylation level on the contractile properties by establishing T/pCa relationships. We increased the O-GlcNAcylation level on soleus biopsies performing an organ culture of soleus treated or not with PUGNAc or Thiamet-G, two O-GlcNAcase inhibitors. The enhancement of O-GlcNAcylation pattern was associated with an increase of calcium affinity on slow soleus skinned fibers. Analysis of the glycoproteins pattern showed that this effect is solely due to O-GlcNAcylation of proteins extracted from skinned biopsies. We also characterized the O-GlcNAcylated contractile proteins using a proteomic approach, and identified among others troponin T and I as being O-GlcNAc modified. We quantified the variation of O-GlcNAc level on all these identified proteins, and showed that several regulatory contractile proteins, predominantly fast isoforms, presented a drastic increase in their O-GlcNAc level. Since the only slow isoform of contractile protein presenting an increase of O-GlcNAc level was MLC2, the effect of enhanced O-GlcNAcylation pattern on calcium activation parameters could involve the O-GlcNAcylation of sMLC2, without excluding that an unidentified O-GlcNAc proteins, such as TnC, could be potentially involved in this mechanism. All these data strongly linked O-GlcNAcylation to the modulation of contractile activity of skeletal muscle.

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“…Thus, after 2 h of chow re-feeding, fasttwitch skeletal-muscle PDHa activities in late-pregnant rats remained substantially lower than corresponding values for the 2 h-re-fed virgin controls ( Table 2). The attenuated responses of fast-twitch skeletal-muscle PDHa activities to acute re-feeding after 24 h starvation in late pregnancy could not be attributed to changes in the activities of citrate synthase (Table 2), a mitochondrial marker enzyme which is suitable for use as a reference enzyme to assess the efficacy of mitochondrial extraction (Stace et al, 1990(Stace et al, , 1992Denyer et al, 1989).…”

Section: % Gsamentioning

confidence: 99%

Changes in rates of glucose utilization and regulation of glucose disposal by fast-twitch skeletal muscles in late pregnancy

Holness

Sugden

1993

Biochemical Journal

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Glucose utilization indices (GUI) were measured in vivo in conjunction with active pyruvate dehydrogenase complex (PDH(a) and glycogen synthase (GS) activities in fast-twitch skeletal muscles [extensor digitorum longus (EDL), tibialis anterior and gastrocnemius] of late-pregnant rats and age-matched virgin control rats in the fed state, after 24 h starvation and at 2 h after re-feeding with standard laboratory chow ad libitum after 24 h starvation. As demonstrated previously [Holness and Sugden (1990) Biochem. J 277, 429-433], GUI values of fast-twitch skeletal muscles of virgin rats were low in the fed ad libitum and the 24 h-starved states, but dramatically increased after subsequent chow re-feeding. GUI values of fast-twitch skeletal muscles of late-pregnant rats were also low in the fed and starved states and were increased by re-feeding, but the increase in GUI values elicited by re-feeding was greatly attenuated. PDHa activities in EDL, tibialis anterior and gastrocnemius in the fed state were unaffected by late pregnancy, and skeletal-muscle PDHa activities were decreased after 24 h of starvation in both groups. Whereas re-feeding of virgin rats with standard diet for 2 h restored PDHa activities in fast-twitch skeletal muscles to values for rats continuously fed ad libitum, PDHa activities in fast-twitch skeletal muscles of late-pregnant rats, although increased in response to re-feeding, remained considerably less than the corresponding fed ad libitum values after 2 h of re-feeding. In contrast, neither skeletal-muscle GS re-activation nor rates of skeletal-muscle glycogen deposition after re-feeding were markedly affected by late pregnancy. The results are discussed in relation to the specific targeting of individual pathways of glucose disposal in fast-twitch skeletal muscles during re-feeding in late pregnancy.

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Long term culture of rat soleus muscle in vitro

Cited by 17 publications

References 24 publications

Incubation under fluid dynamic conditions markedly improves the structural preservation in vitro of explanted skeletal muscles

Incubation under fluid dynamic conditions markedly improves the structural preservation in vitro of explanted skeletal muscles

Increasing O-GlcNAcylation Level on Organ Culture of Soleus Modulates the Calcium Activation Parameters of Muscle Fibers

Changes in rates of glucose utilization and regulation of glucose disposal by fast-twitch skeletal muscles in late pregnancy

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