Phosphodiesterase 4 inhibition restrains muscle proteolysis in diabetic rats by activating PKA and EPAC/Akt effectors and inhibiting FoxO factors.

AIM: There is growing evidence about the ability of cyclic adenosine monophosphate (cAMP) signaling and nonselective phosphodiesterase (PDE) inhibitors on mitigate muscle atrophy. PDE4 accounts for the major cAMP hydrolyzing activity in skeletal muscles, therefore advances are necessary about the consequences of treatment with PDE4 inhibitors on protein breakdown in atrophied muscles. We postulated that rolipram (selective PDE4 inhibitor) may activate cAMP downstream effectors, inhibiting proteolytic systems in skeletal muscles of diabetic rats. MAIN METHODS: Streptozotocin-induced diabetic rats were treated with 2 mg/kg rolipram for 3 days. Changes in the levels of components belonging to the proteolytic machineries in soleus and extensor digitorum longus (EDL) muscles were investigated, as well as cAMP effectors. KEY FINDINGS: Treatment of diabetic rats with rolipram decreased the levels of atrogin-1 and MuRF-1 in soleus and EDL, and reduced the activities of calpains and caspase-3; these findings partially explains the low ubiquitin conjugates levels and the decreased proteasome activity. The inhibition of muscle proteolysis may be occurring due to phosphorylation and inhibition of forkhead box O (FoxO) factors, probably as a consequence of the increased cAMP levels, followed by the activation of PKA and Akt effectors. Akt activation may be associated with the increased levels of exchange protein directly activated by cAMP (EPAC). As a result, rolipram treatment spared muscle mass in diabetic rats. SIGNIFICANCE: The antiproteolytic responses associated with PDE4 inhibition may be helpful to motivate future investigations about the repositioning of PDE4 inhibitors for the treatment of muscle wasting conditions.

Involvement of cAMP/EPAC/Akt signaling in the antiproteolytic effects of pentoxifylline on skeletal muscles of diabetic rats.

Advances in the knowledge of the mechanisms controlling protein breakdown in skeletal muscles have allowed the exploration of new options for treating muscle-wasting conditions. Pentoxifylline (PTX), a nonselective phosphodiesterase (PDE) inhibitor, attenuates the loss of muscle mass during catabolic conditions, mainly via inhibiting protein breakdown. The aim of this study was to explore the mechanisms by which PTX inhibits proteolysis in the soleus and extensor digitorum longus (EDL) muscles of streptozotocin-induced diabetic rats. The levels of atrogin-1 and muscle RING finger-1 were decreased, as were the activities of caspase-3 (EDL) and calpains (soleus and EDL), in diabetic rats treated with PTX, which at least partly explains the drop in the ubiquitin conjugate (EDL) levels and in proteasome activity (soleus and EDL). Treatment with PTX decreased PDE activity and increased cAMP content in muscles of diabetic rats; moreover, it also increased both the protein levels of exchange protein directly activated by cAMP (EPAC, a cAMP effector) and the phosphorylation of Akt. The loss of muscle mass was practically prevented in diabetic rats treated with PTX. These findings advance our understanding of the mechanisms underlying the antiproteolytic effects of PTX and suggest the use of PDE inhibitors as a strategy to activate cAMP signaling, which is emerging as a promising target for treating muscle mass loss during atrophic conditions. NEW & NOTEWORTHY cAMP signaling has been explored as a strategy to attenuate skeletal muscle atrophies. Therefore, in addition to ß2AR agonists, phosphodiesterase inhibitors such as pentoxifylline (PTX) can be an interesting option. This study advances the understanding of the mechanisms related to the antiproteolytic effects of PTX on skeletal muscles of diabetic rats, which involve the activation of both exchange protein directly activated by cAMP and Akt effectors, inhibiting the expression of atrogenes and calpain/caspase-3-proteolytic machinery.

Increased glyceroneogenesis in adipose tissue from rats adapted to a high-protein, carbohydrate-free diet: role of dietary fatty acids.

We have previously shown in in vivo experiments that adipose tissue glyceroneogenesis is increased in rats adapted to a high-protein, carbohydrate-free (HP) diet. The objectives of the present study were (1) to verify if the increased glyceroneogenic activity is also observed in isolated adipocytes and (2) to investigate the role of preformed fatty acids in the production of the increased adipose tissue glyceroneogenesis. Control rats received a balanced diet, with the same lipid content of the HP diet. Glyceroneogenic activity was found to be higher in adipocytes from HP rats than in controls, as evidenced by increased rates of conversion of pyruvate and lactate to triacylglycerol (TAG)-glycerol. Administration of Triton WR 1339, which blocks the removal of TAG incorporated into circulating lipoproteins, to HP diet-adapted rats caused a significant reduction in the incorporation of 14C-pyruvate into TAG-glycerol by adipose tissue, which was accompanied by a marked inhibition of phosphoenolpyruvate carboxykinase activity, the key enzyme of glyceroneogenesis. The inhibitory effect of Triton on TAG-glycerol synthesis by adipose tissue was also observed in vivo, after administration of 3H2O. Adaptation to the HP diet induced a marked increase in the activity of retroperitoneal and epididymal fat LPL, which was restored to control values 24 hours after replacement of the HP diet by the balanced diet. The data suggest that in rats adapted to a carbohydrate-free diet, adipose tissue glyceroneogenesis is activated by an increased use of diet-derived fatty acids.

Curcumin Pharmacokinetic and Pharmacodynamic Evidences in Streptozotocin-Diabetic Rats Support the Antidiabetic Activity to Be via Metabolite(s).

This study measures the curcumin concentration in rat plasma by liquid chromatography and investigates the changes in the glucose tolerance and insulin sensitivity of streptozotocin-diabetic rats treated with curcumin-enriched yoghurt. The analytical method for curcumin detection was linear from 10 to 500 ng/mL. The C max⁡ and the time to reach C max⁡ (t max⁡) of curcumin in plasma were 3.14 ± 0.9 µg/mL and 5 minutes (10 mg/kg, i.v.) and 0.06 ± 0.01 µg/mL and 14 minutes (500 mg/kg, p.o.). The elimination half-time was 8.64 ± 2.31 (i.v.) and 32.70 ± 12.92 (p.o.) minutes. The oral bioavailability was about 0.47%. Changes in the glucose tolerance and insulin sensitivity were investigated in four groups: normal and diabetic rats treated with yoghurt (NYOG and DYOG, resp.) and treated with 90 mg/kg/day curcumin incorporated in yoghurt (NC90 and DC90, resp.). After 15 days of treatment, the glucose tolerance and the insulin sensitivity were significantly improved in DC90 rats in comparison with DYOG, which can be associated with an increase in the AKT phosphorylation levels and GLUT4 translocation in skeletal muscles. These findings can explain, at least in part, the benefits of curcumin-enriched yoghurt to diabetes and substantiate evidences for the curcumin metabolite(s) as being responsible for the antidiabetic activity.

Involvement of cAMP/Epac/PI3K-dependent pathway in the antiproteolytic effect of epinephrine on rat skeletal muscle.

Very little is known about the signaling pathways by which catecholamines exert anabolic effects on muscle protein metabolism, stimulating protein synthesis and suppressing proteolysis. The present work tested the hypothesis that epinephrine-induced inhibition of muscle proteolysis is mediated through the cAMP/Epac/PI3K-dependent pathway with the involvement of AKT and Foxo. The incubation of extensor digitorum longus (EDL) muscles from rats with epinephrine and/or insulin increased the phosphorylation of AKT and its downstream target Foxo3a, a well-known effect that prevents Foxo translocation to the nucleus and the activation of proteolysis. Similar effects on AKT/Foxo signaling were observed in muscles incubated with DBcAMP (cAMP analog). The stimulatory effect of epinephrine on AKT phosphorylation was completely blocked by wortmannin (selective PI3K inhibitor), suggesting that the epinephrine-induced activation of AKT is mediated through PI3K. As for epinephrine and DBcAMP, the incubation of muscles with 8CPT-2Me-cAMP (selective Epac agonist) reduced rates of proteolysis and increased phosphorylation levels of AKT and Foxo3a. The specific PKA agonist (N6BZ-cAMP) inhibited proteolysis and abolished the epinephrine-induced AKT and Foxo3a phosphorylation. On the other hand, inhibition of PKA by H89 further increased the phosphorylation levels of AKT and Foxo3a induced by epinephrine, DBcAMP or 8CPT-2Me-cAMP. These findings suggest that the antiproteolytic effect of the epinephrine on isolated skeletal muscle may occur through a cAMP/Epac/PI3K-dependent pathway, which leads to the phosphorylation of AKT and Foxo3a. The parallel activation of PKA-dependent pathway also inhibits proteolysis and seems to limit the stimulatory effect of cAMP on AKT/Foxo3a signaling.