Functional deficits in skeletal muscle grafts

Individual skeletal muscle fibers degenerate and regenerate with minimal functional deficits. When whole skeletal muscles are grafted in rats or cats by standard grafting techniques, revascularization and reinnervation must occur spontaneously. Under these circumstances, contraction times and maximum velocities of shortening eventually return to control values, but a significant deficit is observed in maximum tetanic tension. Grafts made with anastomosis of nerves or with nerves left intact have smaller deficits in tension development than do standard grafts made without nerve repair. The measurement of contractile properties of single motor units in extensor digitorum longus (EDL) muscles and in EDL grafts in rats indicates that the decreased maximum tetanic tension of whole grafts is due to a 10-20% decrease in the maximum tetanic tension of individual motor units, whereas standard grafts also show a 40-45% decrease in the number of motor units. Compared with control values, the fatigability of 100-mg grafts in rats is decreased, whereas larger 3-g grafts in cats show an increased fatigability. The deficits observed in large grafts can be reduced, but not eliminated, by grafting with neurovascular anastomoses.     

Properties of standard avian slow muscle grafts following long-term regeneration

The tonic anterior latissimus dorsi (ALD) of adult pigeons was orthotopically homografted and evaluated after 11 months of regeneration for histological, histochemical, electromyographic (EMG), and mechanical properties. The resting EMG activity of the grafts was lower in amplitude than that of the controls, but showed the tonic pattern typical for these tonic muscles. The control and grafted muscles had a histochemically homogeneous population of fibers with moderate myofibrillar adenosine triphosphatase activity. Succinic dehydrogenase activity was moderate for the control muscles, but low for the grafts. The regenerated muscles had fewer and smaller fibers and had much larger intersynaptic distances. Both the regenerated and the contralateral control muscles were slow contracting and maintained tetanic tension for prolonged periods with direct electrical stimulation. The relaxation was slower in the grafted muscle than in the control. The grafts produced 40% of the maximum tension of the control muscles, but the rate of tension development was similar between the two groups. The results indicate that the tonic properties were regenerated, but the innervation pattern was altered and the grafted muscles did not have normal mature fibers even after long-term regeneration.     

Muscle grafts overloaded by ablation of synergistic muscles

We hypothesized that the mass and maximum tetanic tension (Po) of nerve-intact grafts overloaded by ablation of synergistic muscles would be greater than that of standard nerve-intact grafts or of control soleus muscles. Soleus muscles were grafted orthotopically and bilaterally in 35 female rats. Control soleus muscles were obtained from 30 age-matched cohorts. Twenty-eight days following grafting, gastrocnemius muscles were ablated bilaterally in half of the animals. Comparisons were made between 28 and 112 days following grafting. By 112 days the wet mass of the overload nerve-intact grafts was 138% of the standard grafts and 152% of the control soleus muscles, whereas the Po was 161% and 107%, respectively. Specific tension stabilized at approximately 19 +/- 1 N/cm2 for both types of grafts, significantly lower than the value of 24 +/- 1 N/cm2 for control soleus muscles. Ablation of synergistic muscles resulted in a significant and sustained increase in mass and Po in regenerating skeletal muscle autografts. We conclude that provided the appropriate conditioning stimulus small grafts (100-200 mg) are capable of achieving the values for the mass and Po of control muscles.     

Force deficit of vascularized skeletal muscle grafts in rabbits

Through autografting experiments on 9-g rectus femoris (RFM) muscles in rabbits, we substantiated a previous observation that the maximum isometric tetanic force (Po) and specific Po (N/cm2) of neurovascular-intact grafts are not different from grafts made with neurovascular repair. We then tested the hypotheses that the specific Po of vascularized grafts is significantly less than that of control RFM muscles and the deficit in the specific Po is associated with increases in connective tissue and interstitial space. The specific Po of the grafts was 65% of the value for control RFM muscles. Connective tissue protein concentration of grafts was 3.8 times greater than the control value of 16.6 +/- 3 micrograms/mg wet mass, but this only accounted for a 5% correction in specific Po. The volume of interstitial space did not differ between grafts and control muscles. We conclude that the deficit of 35% in specific Po of vascularized grafts compared with control values is partially explained by an increase in connective tissue, but a 30% unresolved deficit remains.     

Glucocorticoid-induced glycogen increases in extensor digitorum longus and soleus grafts in the rat

The purpose of the present study was to compare dexamethasone-induced glycogen increases in normal EDL and SOL muscles with that in free muscle grafts. Glycogen in mature EDL and SOL grafts in the rat equalled control concentrations irrespective of whether the graft was a nerve-intact (NI), nerve-crushed (NC), reimplanted, or cross-transplanted graft. The grafts also possessed the glycogen-regulatory mechanisms to respond to the glucocorticoid dexamethasone (DEX), which increases muscle glycogen. The increase in glycogen induced by DEX in the EDL and SOL grafts resembled that of the EDL and SOL muscles, respectively, whether the grafted muscle was originally an EDL or SOL. DEX induced an approximate twofold increase in glycogen concentration in control muscles and nerve-intact SOL grafts, and a smaller but significant increase in all other free grafts. Nerve crushing prior to grafting resulted in no significant change in muscle weight, glycogen concentration, or DEX-induced glycogen increase in these grafts. The data suggest that skeletal muscle grafts are qualitatively similar to normal muscles in terms of metabolic responsiveness to hormones. Leaving the nerve intact during grafting quantitatively enhances the graft's hormonal sensitivity but the technique of nerve crushing prior to grafting has no such effect.     

Innervation of free muscle grafts in the rat in the absence of mechanical trauma to surrounding muscles or nerves.

Soleus muscles in the rat were freely grafted alongside a normal soleus muscle in the absence of mechanical trauma to any of the surrounding muscles or motor nerves. The object of this experiment was to determine whether or not the muscle grafts would become reinnervated under these circumstances. Contractile and histochemical properties of the grafts were compared with those of the contralateral denervated soleus as well as normal muscles. Innervation of the grafts did occur, and it was concluded that the innervation of the grafts arose primarily from sprouts from nerves supplying neighboring muscles. The grafts were studied with specific nerve stains, histochemical techniques and by analysis of their contractile properties.     

Synergist muscle ablation and recovery from nerve-repair grafting: contractile and metabolic function.

After nerve-repair grafting of medial gastrocnemius muscle, there is incomplete recovery of specific force and sustainable power, perhaps due to overcompensation by synergistic muscles. We hypothesized that increased workload due to synergist ablation would enhance graft recovery. Contractile and metabolic properties of control and nerve-repair grafted muscles, with and without synergist ablation, were determined after 120 days recovery. Specific force (N/cm(2)) and normalized power (W/kg) were less in the experimental groups compared with controls. Sustained power (W/kg) in the synergist-ablated nerve-repair grafted muscle was higher than nerve-repair grafted muscle, returning to control values. GLUT-4 protein was higher and glycogen content was diminished in both synergist-ablated groups. In summary, synergist ablation did not enhance the recovery of specific force or normalized power, but sustained power did recover, suggesting that metabolic and not mechanical parameters were responsible for this recovery. The enhanced endurance after synergist ablation was accompanied by increased GLUT-4 protein, suggesting a role for increased uptake of circulating glucose during contraction.     

Parallel-fibered muscles transplanted with neurovascular repair into bipennate muscle sites in rabbits.

Experiments were performed on 20 New Zealand White male rabbits. Our hypotheses were that (1) latissimus dorsi (LTD) muscles transplanted into the site of a bipennate rectus femoris (RFM) muscle with neurovascular repair would retain their parallel-fibered structure and (2) the parallel-fibered structure of latissimus dorsi grafts would reduce their total fiber cross-sectional area and adversely affect force development relative to that of bipennate rectus femoris grafts and muscles. Compared with their respective donor muscles, 120 to 150 days after grafting, latissimus dorsi and rectus femoris grafts showed no change in the number of fibers and a decrease in the mean single-fiber cross-sectional area to approximately 70 percent. The latissimus dorsi grafts, which remained parallel-fibered, developed maximum forces 34 and 23 percent of the values for fully activated rectus femoris grafts and muscles, respectively. The deficit in the maximum force of the latissimus dorsi grafts resulted primarily from the smaller total-fiber cross-sectional area as a result of the parallel-fibered structure.     

Effect of exercise training on passive stiffness in locomotor skeletal muscle: role of extracellular matrix

The purpose ofthis study was to evaluate the effect of endurance exercise training onboth locomotor skeletal muscle collagen characteristics and passivestiffness properties in the young adult and old rat. Young(3-mo-old) and senescent (23-mo-old) male Fischer 344 rats wererandomly assigned to either a control or exercise training group[young control (YC), old control (OC), young trained (YT), oldtrained (OT)]. Exercise training consisted of treadmill runningat ~70% of maximal oxygen consumption (45 min/day, 5 days/wk, for 10 wk). Passive stiffness (stress/strain) of the soleus (Sol) muscle fromall four groups was subsequently measured in vitro at 26°C.Stiffness was significantly greater for Sol muscles in OC rats comparedwith YC rats, but in OT rats exercise training resulted in muscles withstiffness characteristics not different from those in YC rats. Solmuscle collagen concentration and the level of the nonreduciblecollagen cross-link hydroxylysylpyridinoline (HP) significantlyincreased from young adulthood to senescence. Although training had noeffect on Sol muscle collagen concentration in either age group, itresulted in a significant reduction in the level of Sol muscle HP in OTrats. In contrast, exercise had no effect on HP in the YT animals.These findings indicate that 10 wk of endurance exercise significantlyalter the passive viscoelastic properties of Sol muscle in old but notin young adult rats. The coincidental reduction in the principalcollagen cross-link HP also observed in response to training in OTmuscle highlights the potential role of collagen in influencing passivemuscle viscoelastic properties.

     

Short-term immobilization and recovery affect skeletal muscle but not collagen tissue turnover in humans

Not much is known about the effects of immobilization and subsequent recovery on tendon connective tissue. In the present study, healthy young men had their nondominant leg immobilized for a 2-wk period, followed by a recovery period of the same length. Immobilization resulted in a mean decrease of 6% (5,413 to 5,077 mm(2)) in cross-sectional area (CSA) of the triceps surae muscles and a mean decrease of 9% (261 to 238 N.m) in strength of the immobilized calf muscles. Two weeks of recovery resulted in a 6% increased in CSA (to 5,367 mm(2)), whereas strength remained suppressed (240 N.m). No difference in Achilles tendon CSA was detected between the two legs at any time point. Local tendon collagen synthesis, measured as the peritendinous concentrations of PINP (NH(2)-terminal propeptide of type I collagen; indirect marker for collagen synthesis), was unchanged after 2 wk of immobilization. However, peritendinous levels of PINP were significantly elevated in the immobilized leg (15 to 139 ng/ml) following 2 wk of remobilization compared with preimmobilization levels. In contradiction hereto, systemic concentrations of PINP remained unchanged throughout the study. Immobilization reduced muscle size and strength, while tendon size and collagen turnover were unchanged. While recovery resulted in an increase in muscle size, strength was unchanged. No significant difference in tendon size could be detected between the two legs after 2 wk of recovery, although collagen synthesis was increased in the previously immobilized leg. Thus 2 wk of immobilization are sufficient to induce significant changes in muscle tissue, whereas tendon tissue seems to be more resistant to short-term immobilization.     

Contractile properties,structure and fiber phenotype of intact and regenerating slow-twitch muscles of mice treated with cyclosporin A

We have studied the contractile properties, structure, fiber-type composition, and myosin heavy chain (MyHC) expression pattern of regenerating and intact soleus muscles of adult CBA/J mice treated with cyclosporin A (CsA) or vehicle solutions (Cremophor, saline). A comparison of muscles after 4-7 weeks drug application with those receiving vehicle showed that the isometric contractile force of intact drug-treated muscles was reduced (tetanus, -21%; twitch, -34%) despite normal mass and muscle cross-sectional area. The frequency of fast-twitch fibers was increased, whereas no innervation deficits, histopathological alterations, or changes in fiber numbers were observed. Regeneration after cryolesion of the contralateral soleus proceeded more slowly in CsA-treated than in vehicle-treated animals. Despite this, when muscle properties reached mature levels (4-7 weeks), muscle mass recovery was better in CsA-treated animals (30% higher weight, 50% more fiber profiles in cross-sections). The force production per unit cross-sectional area was deficient, but not the maximum tension. Twitch time-to-peak and half-relaxation time were shorter than controls correlating with a predominance of fast-twitch fibers (98% Type II fibers versus 16%-18% in control muscles) and fast MyHC isoforms. Partial reversal of this fast phenotype and an increase in muscle force were observed when the animals were left to recover without treatment for 5-8 weeks after CsA application over 7 weeks. The high numbers of fiber profiles in CsA-treated regenerated muscles and increased mass remained unchanged after withdrawal. Thus, CsA treatment has a hyperplastic effect on regenerating muscles, and drug-induced phenotype alterations are much more prominent in regenerated muscles.     

Regeneration and transplantation of muscles in old rats and between young and old rats.

In order to compare the regenerative ability of skeletal muscle between young (5 month) and old (26 month) rats, sliced or intact extensor digitorum longus muscles were freely autografted into young and old rats and also reciprocally grafted from young to old inbred animals and vice versa. Sixty days after grafting, the transplants were analyzed for contractile and histochemical properties. There was a relative similarity between the contraction times of both normal control muscles and of all groups of transplants, although the contraction time tended to be prolonged and histochemical fiber pattern was more often found to be uniform in grafts of senescent animals. All groups of transplants possessed histochemically heterogeneous fiber types at 60 days. The experiments demonstrate that skeletal muscle in old rats possesses a substantial degree of regenerative ability and that the free tranpllantation of entire muscles in old animals is feasible.     

The development of dopamine overflow from foetal nigral grafts in the intact rat striatum and their influence on contralateral striatal dopamine overflow.

In this study, cell suspensions of foetal rat ventral mesencephalic dopaminergic tissue were grafted to the intact (non-lesioned) striatum of adult rats. Differential pulse voltammetry at carbon-fibre micro electrodes (12 microm diameter) was employed to first, monitor the development of dopamine overflow over a 20 week period within the grafts and secondly, their influence on contralateral striatal dopamine overflow. At 8 and 20 weeks, animals were pre-treated with pargyline and both striata were monitored for dopamine overflow for 90 min following d-amphetamine administration. Amphetamine led to a significant increase in dopamine overflow in both the grafted striatum and the contralateral striatum. The time course of dopamine overflow in both the grafted striatum and the striatum contralateral to the graft was similar in all groups of animals. Although the actual concentration of dopamine measured in 20 week old grafts was more (approximately 21%) than that measured in 8 week old grafts, there was no significant difference between the two time points. The concentration of dopamine measured in the striatum contralateral to 8 week old grafts was significantly lower (approximately 43%) than that measured in the striatum of a normal control rats. There was no significant difference between the concentration of dopamine measured in the striatum contralateral to 20 week old grafts and normal control rats. In conclusion, dopamine overflow from a ventral mesencephalic graft does not change significantly between 8 and 20 weeks following grafting. However, the grafted tissue causes a decrease of d-amphetamine-induced dopamine overflow in the contralateral side 8 weeks following grafting, which is restored 12 weeks later.     

Spontaneous regeneration of older dystrophic muscle does not reflect its regenerative capacity

Young dystrophic (dy) murine muscle is capable of "spontaneous" regeneration (i.e., regeneration in the absence of external trauma); however, by the time the mice are 8 weeks old, this regeneration ceases. It has been suggested that the cessation of regeneration in dystrophic muscle may be due to exhaustion of the mitotic capability of myosatellite cells during the early stages of the disease. To test this hypothesis, orthotopic transplantation of bupivacaine treated, whole extensor digitorum longus muscles has been performed on 14 to 16-week-old 129 ReJ/++ and 129 ReJ/dydy mice. The grafted dystrophic muscle is able to produce and maintain for 100 days post-transplantation 356 +/- 22 myofibers, a number similar to that found in age-matched dystrophic muscle. The ability of old dystrophic muscle to regenerate subsequent to extreme trauma indicates that the cessation of "spontaneous" regeneration is due to factor(s) other than the exhaustion of mitotic capability of myosatellite cells. Moreover, there is no significant difference in myosatellite cell frequencies between grafted normal and dystrophic muscles (100 days post-transplantation). Myosatellite cell frequencies in grafted muscles are similar to those in age-matched, untraumatized muscles. While grafting of young dystrophic muscle modifies the phenotypic expression of histopathological changes usually associated with murine dystrophy, grafts of older dystrophic muscle show extensive connective-tissue infiltration and significantly fewer myofibers than do grafts of age-matched normal muscle. As early as 14 days post-transplantation, it is possible to distinguish between grafts of old, normal and dystrophic muscles. It is suggested that the connective tissue stroma, present in the dystrophic muscle at the time of transplantation, may survive the grafting procedure.     

The effects of dynamic tension and reduced graft size on muscle regeneration in rabbit free muscle grafts.

In a study of 28 adult New Zealand White rabbits, the influence of tension and size on muscle regeneration in tibialis anterior free muscle grafts (without vascular anastomoses) was examined 6 months after transplantation. Three laboratory models were studied: (1) whole dynamic (WD) graft (allowing ankle excursion and, therefore, variable dynamic physiologic tension), (2) whole static (WS) graft (constant, fixed length and, thus, only isometric tension), and (3) longitudinally sliced (reduced radius) dynamic (SD) model. Bilateral orthotopic grafts of the tibialis anterior muscle were performed in 24 rabbits (eight animals in each of the three different model groups). Controls consisted of normal tibialis anterior muscle from four age-matched rabbits. All tibialis anterior muscle grafts were examined histologically (fiber counts) and functionally (determined by in situ contractile properties under maximal stimulation conditions). The WD grafts demonstrated a significantly higher number of regenerated fibers per muscle cross section (4819 +/- 589) than the WS (2221 +/- 603) or SD (1919 +/- 732) grafts. The amount of tetanic tension in the WD grafts was 35 percent of the control and twice as much as that of the WS grafts (WD 1.0 +/- 0.2 kg versus WS 0.5 +/- 0.4 kg; p less than 0.05). The SD grafts produced approximately one-third as much maximum tetanic tension as the WD grafts (0.3 +/- 0.1 kg versus 1.0 +/- 0.2 kg), demonstrating that the amount of recovery was similar in these two dynamic models, since only the longitudinal middle third of the muscle was grafted in the SD model. Free muscle grafts under dynamic tension, which allows excursion, have shown a greater amount of muscle-fiber regeneration and restoration of function compared with a graft with fixed length. The positive effect of dynamic mechanical tension on small autogenous free muscle grafts (without vascular anastomoses) is clinically significant in the reconstruction of facial and hand neuromuscular deficits when blood vessels are not available for reanastomosis. Future studies using the tibialis anterior WD and SD transplant models will strengthen our understanding of the events of spontaneous revascularization and skeletal muscle regeneration.     

Morphological aspects of muscle fiber regeneration

Although striated muscle displays remarkable regenerative potential, the three-dimensional cytoarchitecture of the regenerated myofibers is different from that of myofibers formed during fetal development. It has been demonstrated with spaced, serial ultrathin sections that the regenerating myotubes that occur spontaneously (i.e., without secondary trauma) in dystrophic (dy2J) murine muscle and the regenerating fibers found in free whole-muscle transplants of normal, murine extensor digitorum longus muscles branch and recombine, forming a complex syncytium. Multiple motor end-plate regions are observed on the branched syncytia found in dystrophic muscle. Branched fibers persist in long-term grafts and are found with a frequency that indicates that they should be of physiological significance. Although the number of myofibers found in long-term grafts is approximately 68% of that found in control muscle, comparison of the diameter distributions of the regenerated muscle fibers with age-matched control fibers indicates that many of the regenerating fibers fail to achieve normal size. Type IIb fibers appear to be more growth inhibited than type IIa fibers. The size of the motoneuron pool to grafted muscles is smaller than that to control muscles.     

Triacylglycerol metabolism in rat skeletal muscle after exercise

The temporal relationships between triacylglycerol (TG) content and TG lipase activity in slow-twitch (STR) and fast-twitch red (FTR) muscles were determined in rats during recovery from a 2-h swim. Immediately after the exercise, plasma free fatty acid (FFA) was elevated and glycogen concentrations were decreased. TG content was decreased 40% in STR muscle and reduced 45% in FTR muscle. The TG concentration of STR muscle increased in a linear fashion throughout recovery so that control levels were reached within the first 24 h after exercise. TG lipase activity of STR muscle was elevated 36% above control immediately after the swim and continued to increase to 84% above control 24 h after the work. In STR muscle there was a net synthesis of TG, while lipase activity was elevated above that measured in muscle of control rats. TG content of FTR muscle remained 45% below control throughout the first 24 h of recovery, and TG lipase activity increased from 26% (P greater than 0.05) greater than control immediately after exercise to threefold above control 24 h after work. All parameters returned to control levels by 48 h of recovery. These data indicated that a net TG synthesis occurs in STR muscle when lipolytic activity is elevated. In FTR muscle, however, a gradual increase in TG lipase activity that occurs during the first 24 h of recovery accompanies a TG concentration well below the control level throughout this same time frame.(ABSTRACT TRUNCATED AT 250 WORDS)     

β-Hydroxy-β-methylbutyrate reduces myonuclear apoptosis during recovery from hind limb suspension-induced muscle fiber atrophy in aged rats

β-Hydroxy-β-methylbutyrate (HMB) is a leucine metabolite shown to reduce protein catabolism in disease states and promote skeletal muscle hypertrophy in response to loading exercise. In this study, we evaluated the efficacy of HMB to reduce muscle wasting and promote muscle recovery following disuse in aged animals. Fisher 344×Brown Norway rats, 34 mo of age, were randomly assigned to receive either Ca-HMB (340 mg/kg body wt) or the water vehicle by gavage (n = 32/group). The animals received either 14 days of hindlimb suspension (HS, n = 8/diet group) or 14 days of unloading followed by 14 days of reloading (R; n = 8/diet group). Nonsuspended control animals were compared with suspended animals after 14 days of HS (n = 8) or after R (n = 8). HMB treatment prevented the decline in maximal in vivo isometric force output after 2 wk of recovery from hindlimb unloading. The HMB-treated animals had significantly greater plantaris and soleus fiber cross-sectional area compared with the vehicle-treated animals. HMB decreased the amount of TUNEL-positive nuclei in reloaded plantaris muscles (5.1% vs. 1.6%, P < 0.05) and soleus muscles (3.9% vs. 1.8%, P < 0.05). Although HMB did not significantly alter Bcl-2 protein abundance compared with vehicle treatment, HMB decreased Bax protein abundance following R, by 40% and 14% (P < 0.05) in plantaris and soleus muscles, respectively. Cleaved caspase-3 was reduced by 12% and 9% (P < 0.05) in HMB-treated reloaded plantaris and soleus muscles, compared with vehicle-treated animals. HMB reduced cleaved caspase-9 by 14% and 30% (P < 0.05) in reloaded plantaris and soleus muscles, respectively, compared with vehicle-treated animals. Although, HMB was unable to prevent unloading-induced atrophy, it attenuated the decrease in fiber area in fast and slow muscles after HS and R. HMB's ability to protect against muscle loss may be due in part to putative inhibition of myonuclear apoptosis via regulation of mitochondrial-associated caspase signaling.     

Hindlimb unloading induces a collagen isoform shift in the soleus muscle of the rat

To determine whether hindlimb unloading (HU) alters the extracellular matrix of skeletal muscle, male Sprague-Dawley rats were subjected to 0 (n = 11), 1 (n = 11), 14 (n = 13), or 28 (n = 11) days of unloading. Remodeling of the soleus and plantaris muscles was examined biochemically for collagen abundance via measurement of hydroxyproline, and the percentage of cross-sectional area of collagen was determined histologically with picrosirius red staining. Total hydroxyproline content in the soleus and plantaris muscles was unaltered by HU at any time point. However, the relative proportions of type I collagen in the soleus muscle decreased relative to control (Con) with 14 and 28 days HU (Con 68 +/- 5%; 14 days HU 53 +/- 4%; 28 days HU 53 +/- 7%). Correspondingly, type III collagen increased in soleus muscle with 14 and 28 days HU (Con 32 +/- 5%; 14 days HU 47 +/- 4%; 28 days HU 48 +/- 7%). The proportion of type I muscle fibers in soleus muscle was diminished with HU (Con 96 +/- 2%; 14 days HU 86 +/- 1%; 28 days HU 83 +/- 1%), and the proportion of hybrid type I/IIB fibers increased (Con 0%; 14 days HU 8 +/- 2%; 28 days HU 14 +/- 2%). HU had no effect on the proportion of type I and III collagen or muscle fiber composition in plantaris muscle. The data demonstrate that HU induces a shift in the relative proportion of collagen isoform (type I to III) in the antigravity soleus muscle, which occurs concomitantly with a slow-to-fast myofiber transformation.     

Impaired Macrophage and Satellite Cell Infiltration Occurs in a Muscle-Specific Fashion Following Injury in Diabetic Skeletal Muscle

Background

Systemic elevations in PAI-1 suppress the fibrinolytic pathway leading to poor collagen remodelling and delayed regeneration of tibialis anterior (TA) muscles in type-1 diabetic Akita mice. However, how impaired collagen remodelling was specifically attenuating regeneration in Akita mice remained unknown. Furthermore, given intrinsic differences between muscle groups, it was unclear if the reparative responses between muscle groups were different.

Principal Findings

Here we reveal that diabetic Akita muscles display differential regenerative responses with the TA and gastrocnemius muscles exhibiting reduced regenerating myofiber area compared to wild-type mice, while soleus muscles displayed no difference between animal groups following injury. Collagen levels in TA and gastrocnemius, but not soleus, were significantly increased post-injury versus controls. At 5 days post-injury, when degenerating/necrotic regions were present in both animal groups, Akita TA and gastrocnemius muscles displayed reduced macrophage and satellite cell infiltration and poor myofiber formation. By 10 days post-injury, necrotic regions were absent in wild-type TA but persisted in Akita TA. In contrast, Akita soleus exhibited no impairment in any of these measures compared to wild-type soleus. In an effort to define how impaired collagen turnover was attenuating regeneration in Akita TA, a PAI-1 inhibitor (PAI-039) was orally administered to Akita mice following cardiotoxin injury. PAI-039 administration promoted macrophage and satellite cell infiltration into necrotic areas of the TA and gastrocnemius. Importantly, soleus muscles exhibit the highest inducible expression of MMP-9 following injury, providing a mechanism for normative collagen degradation and injury recovery in this muscle despite systemically elevated PAI-1.

Conclusions

Our findings suggest the mechanism underlying how impaired collagen remodelling in type-1 diabetes results in delayed regeneration is an impairment in macrophage infiltration and satellite cell recruitment to degenerating areas; a phenomena that occurs differentially between muscle groups.