Muscle Ischemia-reperfusion Injury Research Articles

UCHL1 Negatively Regulates Key Factors and Signals Involved in Skeletal Muscle Myogenesis

Ubiquitin C-terminal hydrolase L1 (UCHL1), a deubiquitinating enzyme that was originally discovered in neurons, is also expressed in skeletal muscle, but its functions remain to be fully understood. Skeletal muscle injury is prevalent in trauma and surgical procedures, and skeletal muscle ischemia-reperfusion (IR) injury is a common and significant public health problem. Myogenesis is a critical process involved in embryonic development, growth, and regeneration following skeletal muscle injury. In this study we report that UCHL1 negatively affects some key factors that are involved in myogenesis and mitophagy, both during muscle development as well as during the regeneration process following IR injury; we hypothesize that UCHL1 is a negative regulator of myogenesis. First, we observed that UCHL1 knockdown in C2C12 myoblasts resulted in increased differentiation and myotube formation. Furthermore, western blot showed that UCHL1 KD consistently upregulated myogenin and MyoD protein levels, key proteins involved in myogenesis, at multiple time points throughout differentiation of the myotubes. Consistent with this in vitro result, skeletal muscle specific knockout (smKO) of UCHL1 increased muscle fiber size in both 1- and 2-month-old mice (p=1.77x10−17, p=6.51x10−13). Additionally, 1- and 2- month-old UCHL1 smKO mice also exhibited increased protein levels of various myosin heavy chain isoforms. To induce IR injury, a tourniquet was placed on the hindlimb for 90 minutes and then removed, allowing for reperfusion for 3-12 days with the contralateral hindlimb serving as a control. Following skeletal muscle IR injury, myogenin and MyoD protein expression was upregulated in injured muscle from UCHL1 smKO mice. Functionally, UCHL1 smKO mice had increased muscle performance after injury compared to control mice when subjected to in situ contractile testing. Interestingly, several mitophagy markers including Parkin and DRP1 were upregulated in IR injured muscle from UCHL1 smKO mice compared to WT mice (p=0.0003, p=0.029). This data suggests that skeletal muscle UCHL1 may function as a negative regulator of myogenesis, both during growth, and repair following injury, possibly by decreasing mitochondrial activity. This work was funded by NIH grant R01HL147105. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Beneficial Effects of Insulin on Ischemia Reperfusion Injury in Human Skeletal Muscle.

PurposeExaggerated leucocyte activity is a crucial step in the pathophysiology of skeletal muscle ischemia-reperfusion injury (IRI). We tested the hypothesis that insulin, via its’ anti-leukocyte activity, attenuates skeletal muscle IRI in humans.MethodsMaterials and This randomized, blinded, placebo-controlled trial was conducted in patients with skeletal muscle ischemia who required revascularization. Treatment protocols were similar among them except for the insulin group, which received an infusion of insulin at 2.5 U/h. The degree of endothelial adhesiveness; leukocyte activity and pro-inflammatory status via P-selectin, tumor necrosis factor (TNF)-alpha, and myeloperoxidase (MPO) levels in the venous effluent; and clinical outcomes were measured.ResultsTwenty-four consenting patients were randomized to the insulin or control group. There were no significant differences between the two groups except for the median serum insulin level, which was higher in the insulin group (P<0.01). No serious intervention-related adverse events were observed. P-selectin (55.04-99.86 pg/mL; P<0.001), MPO (110.8-160.6 pg/mL; P<0.001), and TNF-alpha (12.16-36.01 pg/mL; P<0.001) levels demonstrated a significant increase post-reperfusion in the ‘control’ group, reaching a peak value at 2 hours post-reperfusion. The increase in all three markers from baseline was significantly diminished in the insulin group at the two-hour (P-selectin, P=0.001; MPO, P=0.001; TNF-alpha, P=0.005) and four-hour (P-selectin, P=0.003; MPO, P=0.002; TNF-alpha, P=0.01) intervals. The differences in clinical outcomes between the insulin and control groups were not statistically significant.ConclusionIn clinical practice, insulin has the potential to attenuate the severity of skeletal muscle IRI inhibiting P-selectin, MPO, and TNF-alpha levels.

Effect of pheniramine maleate on rat skeletal muscle ischemia-reperfusion injury.

Skeletal muscle ischemia-reperfusion injury (IRI) is a common clinical problem encountered after tourniquet ap-plication or replantation. This study investigated the effect of pheniramine maleate (Ph), which is frequently used in clinical practice to reduce IRI, and compared its efficacy in IRI with N-acetylcysteine (NAC), a molecule that has been shown to be effective in IRI. Twenty-eight male Sprague-Dawley rats were randomly divided into four groups (sham, ischemia-reperfusion [IR], IR+Ph, IR+NAC; n=7 rats per group). Ischemia was induced in the lower right extremities of rats for 3 h using a femoral artery clamp and an elastic tourniquet. Ph and NAC were administered intraperitoneally 15 min before ischemia was terminated. At 24 h after reperfusion, levels of thiobarbituric acid reactive substance (TBARS), catalase (CAT), myeloperoxidase (MPO), superoxide dismutase (SOD), polyadenosine diphosphate ribose polymerase (PARP), and neutrophil infiltration were evaluated. Inducible nitric oxide syn-thase (iNOS) density in muscle tissue was evaluated by immunohistochemical methods after 1 week. SOD, MPO, PARP, CAT, and TBARS levels in muscle tissue were significantly lower in the sham group compared with the other groups (p<0.001). All parameters except TBARS were lower in the NAC and Ph groups than in the IR group (p<0.001). Neu-trophil infiltration in the muscle tissue samples from the IR group was significantly increased compared with the NAC and Ph groups (p<0.05). iNOS staining was not observed in the sham and NAC groups. Ph is effective at reducing experimental rat skeletal muscle IRI.

Deficiency of myostatin protects skeletal muscle cells from ischemia reperfusion injury

Ischemia reperfusion (IR) injury plays a pivotal role in many diseases and leads to collateral damage during surgical interventions. While most studies focus on alleviating its severity in the context of brain, liver, kidney, and cardiac tissue, research as regards to skeletal muscle has not been conducted to the same extent. In the past, myostatin (MSTN), primarily known for supressing muscle growth, has been implicated in inflammatory circuits, and research provided promising results for cardiac IR injury mitigation by inhibiting MSTN cell surface receptor ACVR2B. This generated the question if interrupting MSTN signaling could temper IR injury in skeletal muscle. Examining human specimens from free myocutaneous flap transfer demonstrated increased MSTN signaling and tissue damage in terms of apoptotic activity, cell death, tissue edema, and lipid peroxidation. In subsequent in vivo MstnLn/Ln IR injury models, we identified potential mechanisms linking MSTN deficiency to protective effects, among others, inhibition of p38 MAPK signaling and SERCA2a modulation. Furthermore, transcriptional profiling revealed a putative involvement of NK cells. Collectively, this work establishes a protective role of MSTN deficiency in skeletal muscle IR injury.

Hydrogen sulfide‐loaded microbubbles combined with ultrasound mediate thrombolysis and simultaneously mitigate ischemia‐reperfusion injury in a rat hindlimb model

BackgroundThromboembolism and subsequent ischemia/reperfusion injury (IRI) remain major clinical challenges. ObjectivesTo investigate whether hydrogen sulfide (H2S)‐loaded microbubbles (hs‐Mbs) combined with ultrasound (US) radiation (hs‐Mbs+US) dissolve thrombi and simultaneously alleviate tissue IRI through local H2S release. Methodshs‐Mbs were manufactured and US‐triggered H2S release was recorded. White and red thromboembolisms were established ex vivo and in rats left iliac artery. All subjects randomly received control, US, Mbs+US, or hs‐Mbs+US treatment for 30 minutes. ResultsH2S was released from hs‐Mbs+US both ex vivo and in vivo. Compared with control and US, hs‐Mbs+US and Mbs+US showed comparable substantial decreases in thrombotic area, clot mass, and flow velocity increases for both ex vivo macrothrombi. In vivo, hs‐Mbs+US and Mbs+US caused similarly increased recanalization rates, blood flow velocities, and hindlimb perfusion for both thrombi compared with the other treatments, with no obvious influence on hemodynamics, respiration, and macrophage vitality. More importantly, hs‐Mbs+US substantially alleviated skeletal muscle IRI by reducing reactive oxygen species, cellular apoptosis, and proapoptotic Bax, caspase‐3, and caspase‐9 and increasing antiapoptotic Bcl‐2 compared with other treatments. In vitro, hypoxia/reoxygenation‐predisposed skeletal muscle cells and endothelial cells treated with normal saline solution exhibited similar trends, which were largely reversed by an H2S scavenger or an inhibitor of Akt phosphorylation. Conclusionhs‐Mbs+US effectively dissolved both white and red macrothrombi and simultaneously alleviated skeletal muscle IRI through the US‐triggered, organ‐specific release of H2S. This integrated therapeutic strategy holds promise for treating thromboembolic diseases and subsequent IRI.

Myogenic progenitor cell transplantation for muscle regeneration following hindlimb ischemia and reperfusion

BackgroundMuscle is severely affected by ischemia/reperfusion injury (IRI). Quiescent satellite cells differentiating into myogenic progenitor cells (MPC) possess a remarkable regenerative potential. We herein established a model of local application of MPC in murine hindlimb ischemia/reperfusion to study cell engraftment and differentiation required for muscle regeneration.MethodsA clamping model of murine (C57b/6 J) hindlimb ischemia was established to induce IRI in skeletal muscle. After 2 h (h) warm ischemic time (WIT) and reperfusion, reporter protein expressing MPC (TdTomato or Luci-GFP, 1 × 106 cells) obtained from isolated satellite cells were injected intramuscularly. Surface marker expression and differentiation potential of MPC were analyzed in vitro by flow cytometry and differentiation assay. In vivo bioluminescence imaging and histopathologic evaluation of biopsies were performed to quantify cell fate, engraftment and regeneration.Results2h WIT induced severe IRI on muscle, and muscle fiber regeneration as per histopathology within 14 days after injury. Bioluminescence in vivo imaging demonstrated reporter protein signals of MPC in 2h WIT animals and controls over the study period (75 days). Bioluminescence signals were detected at the injection site and increased over time. TdTomato expressing MPC and myofibers were visible in host tissue on postoperative days 2 and 14, respectively, suggesting that injected MPC differentiated into muscle fibers. Higher reporter protein signals were found after 2h WIT compared to controls without ischemia, indicative for enhanced growth and/or engraftment of MPC injected into IRI-affected muscle antagonizing muscle damage caused by IRI.ConclusionWIT-induced IRI in muscle requests increased numbers of injected MPC to engraft and persist, suggesting a possible rational for cell therapy to antagonize IRI. Further investigations are needed to evaluate the regenerative capacity and therapeutic advantage of MPC in the setting of ischemic limb injury.

Perfusion imaging with 320-slice spiral computed tomography and color-coded digital subtraction angiography for assessing acute skeletal muscle ischemia-reperfusion injury in a rabbit model

BackgroundIn recent years, skeletal muscle-related ischemia-reperfusion injury (IRI) has become more common. IRI can lead to severe limb injuries, multiple organ failure, and even death in some cases. However, there is still a lack of rapid and sensitive detection methods for IRI in skeletal muscle. This study aims to explore the value of computed tomography perfusion imaging (CTPI) and color-coded digital subtraction angiography (DSA) in assessing acute IRI of skeletal muscle in a rabbit model.MethodsFifty New Zealand white rabbits were randomly assigned to the ischemia-reperfusion (IR) group (n = 40) or sham group (n = 10). After 3 h of surgically-induced hindlimb ischemia, the IR group underwent reperfusion and CTPI and color-coded DSA were taken to assess the skeletal muscle at 0, 6, 12, or 24 h post-reperfusion. The data from CTPI and DSA in the right and left hindlimbs, blood flow (AF-R/L), blood volume (BV-R/L), contrast clearance rate (C-R/L) and the maximum contrast enhancement values (peak-R/L) were obtained. Serum superoxide dismutase (SOD), creatine kinase (CK), lactic dehydrogenase (LDH) and malondialdehyde (MDA) were measured. The statistical correlation between the above parameters (CTPI, color-coded DSA, and biochemical markers) was analyzed.ResultsThe mean value of AF-R/L, BV-R/L, C-R/L and peak-R/L decreased linearly from 1.07 ± 0.08 to 0.75 ± 0.11, 1.03 ± 0.06 to 0.85 ± 0.14, 0.93 ± 0.15 to 0.71 ± 0.18, and 1.07 ± 0.01 to 0.47 ± 0.04, respectively. The correlation coefficients between AF-R/L and SOD, CK, LDH and MDA were 0.57, − 0.44, − 0.60, and − 0.62, respectively (p < 0.001). The correlation coefficients between Peak-R/L and SOD, CK, LDH, MDA were 0.59, 0.68, 0.71 and 0.66, respectively (p < 0.001). The correlation coefficient between AF-R/L and Peak-R/L was 0.70 (p < 0.001).ConclusionBoth CTPI and color-coded DSA could dynamically assess skeletal muscle IRI in rabbits.

Mitochondrial transplantation ameliorates acute limb ischemia

ObjectiveAcute limb ischemia (ALI), the most challenging form of ischemia-reperfusion injury (IRI) in skeletal muscle tissue, leads to decreased skeletal muscle viability and limb function. Mitochondrial injury has been shown to play a key role in skeletal muscle IRI. In previous studies, we have demonstrated that mitochondrial transplantation (MT) is an efficacious therapeutic strategy to replace or to augment mitochondria damaged by IRI, allowing enhanced muscle viability and function in cardiac tissue. In this study, we investigated the efficacy of MT in a murine ALI model. MethodsC57BL/6J mice (male, 10-12 weeks) were used in a model of ALI. Ischemia was induced by applying a tourniquet on the left hindlimb. After 2 hours of ischemia, the tourniquet was released, and reperfusion of the hindlimb was re-established; either vehicle alone (n = 15) or vehicle containing mitochondria (n = 33) was injected directly into all the muscles of the hindlimb. Mitochondria were delivered at concentrations of 1 × 106 to 1 × 109 per gram wet weight to each muscle, and the animals were allowed to recover. Sham mice received no ischemia or injections but were anesthetized for 2 hours and allowed to recover. After 24 hours of recovery, limb function was assessed by DigiGait (Mouse Specifics Inc, Boston, Mass), and animals were euthanized; the gastrocnemius, soleus, and vastus medialis muscles were collected for analysis. ResultsAfter 24 hours of hindlimb reperfusion, infarct size (percentage of total mass) and apoptosis were significantly decreased (P < .001, each) in the gastrocnemius, soleus, and vastus medialis muscles in MT mice compared with vehicle mice for all mitochondrial concentrations (1 × 106 to 1 × 109 per gram wet weight). DigiGait analysis at 24 hours of reperfusion showed that percentage shared stance time was significantly increased (P < .001) and stance factor was significantly decreased (P = .001) in vehicle compared with MT and sham mice. No significant differences in percentage shared stance time (P = .160) or stance factor (P = .545) were observed between MT and sham mice. ConclusionsMT ameliorates skeletal muscle injury and enhances hindlimb function in the murine model of ALI.

Chronic intermittent hypobaric hypoxia attenuates skeletal muscle ischemia-reperfusion injury in mice

AimThe aim of this study was to investigate the protective effect of chronic intermittent hypobaric hypoxia (CIHH) against skeletal muscle ischemia-reperfusion (IR) injury and to determine the underlying mechanism. Main methodsC57BL/6 mice were randomly divided into 3 groups: skeletal muscle IR injury group (IR), CIHH pretreatment following IR group (IR + CIHH), and sham operation group (Sham). The skeletal muscle IR injury model was induced by the unilateral application of a tourniquet on a hind limb for 3 h and then releasing it for 24 h. CIHH pretreatment simulating a 5000-m altitude was applied 6 h per day for 28 days. The functional and morphological performance of IR-injured gastrocnemius muscle was evaluated using contraction force, H&E staining, and transmission electron microscopy. IR injury-induced CD68+ macrophage infiltration was assessed by immunofluorescence. TNFα levels in serum and muscle were measured by ELISA and western blotting, respectively. Apoptosis was examined by TUNEL staining and Cleaved Caspase-3 protein expression. Key findingsAcute IR injury resulted in reduced contraction tension, morphological destruction, macrophage infiltration, increased TNFα levels, and apoptosis in gastrocnemius muscle. CIHH pretreatment significantly ameliorated contraction function and morphological performance in IR-injured skeletal muscle. In addition, CIHH pretreatment resulted in marked decreases in CD68+ macrophage infiltration, TNFα levels, and apoptosis. SignificanceThese data demonstrated that CIHH has a protective effect against acute IR injury in skeletal muscle via inhibition of inflammation and apoptosis.

A histological Study Comparing the Possible Protective Mechanisms of Oxytocin and Vitamin E on ischemia reperfusion injury in Rat skeletal muscle

Background: Ischemia-reperfusion (IR) injury in skeletal muscle is a pathophysiology which affect quality of life.Aim of Work: To compare the possible protective mechanisms by which oxytocin and Vitamin E may exert on skeletal muscle IR injury in rat. Materials and Methods: 28 male rats were divided into: Control (Group A) of 10 rats and IR group (Group B) of 18 rats. Control group underwent the same surgical steps as IR group but without clamping the femoral artery followed by the injection of one of the following [saline (AI), oxytocin (AII) or vitamin E (AIII)]. In the IR group, lower limb ischemia was induced by clamping the femoral artery. After 4h of ischemia, the clamp was removed to allow reperfusion coinciding with the injection of one of the following [saline (BI), Oxytocin (0.5μg/kg) (BII) and Vitamin E (10mg/kg) (BIII)]. After 2 hours of reperfusion, Blood samples were drawn from the femoral vein to measure glutathione (GSH) and malondialdehyde (MDA). Gastrocnemius muscles were obtained for evaluation by light and electron microscopes. Cytochrome C, Myogenin immunohistochemistry and statistical analysis were applied.Results: A significant increase in GSH and significant decrease in MDA were reported in BII and BIII as compared to BI. Mean area % of cytochrome C in BII and BIII showed a significant decrease compared to BI. Mean number of myogenin immunopositive cells in BII was significantly higher compared to BI. Muscular damage was proved by light and electron microscopic findings in BI. While, BII and BIII encountered evident protection from muscular damage induced by IR injury. Conclusion: Oxytocin exerts weak protective antioxidant effect against IR injury. However, it activates stem cells to regenerate the damaged muscle. Vitamin E was better antioxidant which can react as early as 1 hour in the protection of the skeletal muscle against IR injury.

Influence of C(60) fullerene on the ischemia-reperfusion injury in the skeletal muscle of rat limb: mechanokinetic and biochemical analysis

Influence of C(60) fullerene on the ischemia-reperfusion injury in the skeletal muscle of rat limb: mechanokinetic and biochemical analysis

Research on the oxygen partial pressure with vacuum sealing drainage in rabbits

Objective To measure the oxygen partial pressure (PtO2) within a situation of vacuum sealing drainage (VSD) implement, and to discuss the potential mechanism of VSD alleviating skeletal muscle ischemia reperfusion (I/R) injury in rabbits. Methods Rabbits were randomly(random number) divided into three groups: sham (n=10), I/R (n=10) and I/R+VSD (n=10) groups. In the I/R group, left hind limb ischemia was induced by clamping the femoral artery and vein for 4 h, then released for 6 h, to allow reperfusion of the limb. VSD was only performed in the I/R+VSD group during the reperfusion period. All rabbits were administered a series test of PtO2 in different time points before and after reperfusion, and the left hind limb skeletal muscle and ear vein blood samples were immediately harvested for biochemical analyses, including hypoxia inducible factor 1 alpha (HIF-1α) mRNA and protein, and lactic acid (LA). One-way analysis of variance (ANOVA), Fisher’s least significant difference (LSD) test or Tamhane’s T2 test was used where appropriate to analyze the experimental results. Results The levels of PtO2 in the I/R and I/R+VSD groups were decreased after ischemia, but no significant difference was noted between the two groups (t=1.322, P=0.296). The PtO2 levels in the I/R+VSD group were less increased than those in the I/R group after reperfusion (t=2.015, P=0.046). The levels of HIF-1α mRNA and protein were increased after I/R injuries (F=10.120, P=0.002; F=36.480, P<0.01; F=6.960, P=0.015, F=4.470, P=0.035), and the levels in the I/R+VSD group were greater than those in the I/R group (t=1.799, P=0.048, t=5.911, P=0.019; t=1.878, P=0.046, t=2.609, P=0.030). The LA levels were significantly increased in the skeletal muscle and blood samples in the I/R and I/R+NPWT groups compared with those in the sham group at the final stage (F=9.540, P=0.002 and F=13.750, P<0.01), but the levels in the I/R+VSD group were less than those in the I/R group (t=2.263, P=0.040 and t=3.617, P=0.027). Conclusions Our results suggested that VSD technique decreased PtO2 in regional ischemic muscles and led to increases in HIF-1α mRNA and protein that subsequent accumulation of less LA in both of regional skeletal muscle and systemic blood circulation, which provided protection effect after I/R injury in rabbits. Key words: Trauma; Ischemia reperfusion injury; Vacuum sealing drainage; Oxygen partial pressure; Hypoxia inducible factor 1 alpha; Lactic acid

Color-coded Digital Subtraction Angiography for Assessing Acute Skeletal Muscle Ischemia-Reperfusion Injury in a Rabbit Model

This paper describes an ongoing investigation of imaging and characterization of ischemia-reperfusion (IR) and investigated the use of color-coded digital subtraction angiography (DSA) to assess reperfusion injury or potential injury. New Zealand white rabbits were subjected to right hindlimb ischemia (IR, n = 24) or sham operation (control, n = 6). After 3 hours, the IR rabbits underwent reperfusion and were assessed at 0, 6, 12, or 24 hours (n = 6 each). DSA of the bilateral vastus lateralis muscle of each animal was performed. The maximum contrast enhancement value of a consistent region of interest in the right and left hind limbs (peak enhancement-R/L) was determined. Associations between the relative ratio of the peak right limb to the peak left limb (peak-R/L) and the following blood indicators of IR injury were analyzed: lactic dehydrogenase (LDH), creatine kinase (CK), malondialdehyde (MDA), and superoxide dismutase (SOD). Serum LDH, CK, and MDA values in each IR group were significantly higher than those of the control group and were positively associated with the IR interval, whereas SOD was significantly lower and negatively associated. The mean peak-R/L decreased linearly with the IR interval from 1.07 ± 0.01 in the control group, and 0.93 ± 0.06, 0.79 ± 0.05, 0.65 ± 0.04, and 0.47 ± 0.04 at 0, 6, 12, and 24 hours in the IR groups. The coefficients of correlation between the peak-R/L and LDH, CK, MDA, SOD serum levels were -0.885, -0.908, -0.541, and 0.832, respectively. Color-coded DSA may be used for monitoring the dynamics of skeletal muscle IR injury.

Targeted Complement Inhibition Protects Vascularized Composite Allografts From Acute Graft Injury and Prolongs Graft Survival When Combined With Subtherapeutic Cyclosporine A Therapy.

Recipients of vascularized composite allografts require aggressive and lifelong immunosuppression, and because the surgery is usually performed in nonlife-threatening situations, the development of strategies to minimize immunosuppression is especially pertinent for this procedure. We investigated how complement affects acute graft injury, alloimmunity, and immunosuppressive therapy. Vascularized composite allografts were transplanted from Balb/C to C57BL/6 mice that were complement deficient (C3 or double C3a Receptor (R)/C5aR), or treated with a targeted complement inhibitor (CR2-Crry). Allografts were analyzed for acute inflammation and injury, subacute T cell response, and survival in the absence and presence of cyclosporine A (CsA) therapy. Allografts in C3-deficient or CR2-Crry-treated recipients were protected from skin and muscle ischemia-reperfusion injury (IRI). C3aR/C5aR-deficient recipients were more modestly protected. IgM and C3d colocalized within allografts from wild type and C3aR/C5aR-deficient recipients indicating IgM-mediated complement activation, and C3d deposition was almost absent in allografts from C3-deficient and CR2-Crry-treated recipients. Inflammatory cell infiltration and P-selectin expression was also significantly reduced in C3-deficient and CR2-Crry-treated recipients. Acute treatment with CR2-Crry or with 3 mg/kg per day CsA modestly, but significantly increased median allograft survival from 5.8 to 7.4 and 7.2 days, respectively. However, combined acute CR2-Crry treatment and CsA therapy increased mean graft survival to 17.2 days. Protection was associated with significantly reduced T cell infiltration of allografts and Tc1 cells in recipient spleens. Complement-mediated IRI augments graft allogenicity, and appropriate complement inhibition ameliorates IRI, decreases alloimmune priming and allows more immune-sparing CsA dosing.

The Protective Effects of Fasciotomy on Reperfusion Injury of Skeletal Muscle of Rabbits.

The authors aim to investigate protective effects of fasciotomy against ischemia reperfusion injury of skeletal muscle in rabbit and to compare the treatment effects of prereperfusion + fasciotomy and fasciotomy + postreperfusion against ischemia reperfusion injury of skeletal muscle. 24 healthy male Japanese white rabbits were randomly divided into 3 groups, and 4 hours' ischemia was established in these rabbits through surgery. Six hours' reperfusion was performed in group A; reperfusion + postfasciotomy was performed in group B; and prefasciotomy + reperfusion was performed in group C. Result showed that prefasciotomy and postfasciotomy could protect skeletal muscle against ischemia reperfusion injury, reduced MDA (malondialdehyde) expression, MPO (myeloperoxidase) expression, and apoptosis of muscle in the reperfused areas, increased Bcl-2 expression, and decreased Bax expression. The MDA and MPO levels in group B and group C were significantly lower than those in group A, and MDA and MPO levels in group C were significantly lower than those in group B. Prefasciotomy and postfasciotomy could protect against ischemia reperfusion injury in skeletal muscle. The protective effects of prefasciotomy against ischemia reperfusion injury are better than postfasciotomy.

Effect of different doses of pregabalin on skeletal muscle ischaemia-reperfusion injury in rats.

Analgesic, anti-inflammatory and anti-apoptotic effects of pregabalin have been shown previously. In this study, we investigated the protective effect of different doses of pregabalin on skeletal muscle IR injury in rats. 24 rats were randomly divided into 4 groups (Control, Ischaemia-Reperfusion (IR), IR-Pregabalin 50 mg, IR-Pregabalin 200 mg). Following IR, serum Ischemia Modified Albumin (IMA) and tissue Paraoxonase (PON) were studied and gastrocnemius muscle tissue was removed for histopathologic examination. Interstitial inflammation was higher in the IR group than in the control and Pregabalin 200 mg groups (p = 0.037, p = 0.037, respectively). Congestion was higher in the IR group than in the control, Pregabalin 50 and 200 mg groups (p = 0.001, p = 0.004, p = 0.004, respectively). PON was lower in the IR group than in the Control, Pregabalin 50 and 200 mg groups (p = 0.001, p = 0.007, p = 0.015, respectively). IMA was higher in the IR group than in the Control, Pregabalin 50 and 200 mg groups (p < 0.0001, all). We think that administration of pregabalin, more prominent at 200 mg, can reverse the injury that occurs in the skeletal muscle of IR-induced rats. Pregabalin can be safely used for analgesia in cases of IR (Tab. 2, Fig. 9, Ref. 41).

Administration of FTY720 during Tourniquet-Induced Limb Ischemia Reperfusion Injury Attenuates Systemic Inflammation.

Acute ischemia-reperfusion injury (IRI) of the extremities leads to local and systemic inflammatory changes which can hinder limb function and can be life threatening. This study examined whether the administration of the T-cell sequestration agent, FTY720, following hind limb tourniquet-induced skeletal muscle IRI in a rat model would attenuate systemic inflammation and multiple end organ injury. Sprague-Dawley rats were subjected to 1 hr of ischemia via application of a rubber band tourniquet. Animals were randomized to receive an intravenous bolus of either vehicle control or FTY720 15 min after band placement. Rats (n = 10/time point) were euthanized at 6, 24, and 72 hr post-IRI. Peripheral blood as well as lung, liver, kidney, and ischemic muscle tissue was analyzed and compared between groups. FTY720 treatment markedly decreased the number of peripheral blood T cells (p < 0.05) resulting in a decreased systemic inflammatory response and lower serum creatinine levels and had a modest but significant effect in decreasing the transcription of injury-associated target genes in multiple end organs. These findings suggest that early intervention with FTY720 may benefit the treatment of IRI of the limb. Further preclinical studies are necessary to characterize the short-term and long-term beneficial effects of FTY720 following tourniquet-induced IRI.

Colchicine protects rat skeletal muscle from ischemia/reperfusion injury by suppressing oxidative stress and inflammation.

Neutrophils play an important role in ischemia/reperfusion (IR) induced skeletal muscle injury. Microtubules are required for neutrophil activation in response to various stimuli. This study aimed to investigate the effects of colchicine, a microtubule-disrupting agent, on skeletal muscle IR injury in a rat hindlimb ischemia model. Twenty-one Sprague-Dawley rats were randomly allocated into three groups IR group, colchicine treated-IR (CO) group and sham operation (SM) group. Rats of both the IR and CO groups were subjected to 3 hr of ischemia by clamping the right femoral artery followed by 2 hr of reperfusion. Colchicine (1 mg/kg) was administrated intraperitoneally prior to hindlimb ischemia in the CO group. After 2 hr of reperfusion, we measured superoxide dismutase (SOD) and myeloperoxidase (MPO) activities, and malondialdehyde (MDA), tumor necrosis factor (TNF)-α and interleukin (IL)-1β levels in the muscle samples. Plasma creatinine kinase (CK) and lactate dehydrogenase (LDH) levels were measured. We also evaluated the histological damage score and wet/dry weight (W/D) ratio. The histological damage score, W/D ratio, MPO activity, MDA, TNF-α and IL-1β levels in muscle tissues were significantly increased, SOD activity was decreased, and plasma CK and LDH levels were remarkably elevated in both the IR and CO groups compared to the SM group (P<0.05). Colchicine treatment significantly reduced muscle damage and edema, oxidative stress and levels of the inflammatory parameters in the CO group compared to the IR group (P<0.05). Colchicine attenuates IR-induced skeletal muscle injury in rats.

Novel intracellular N-terminal truncated matrix metalloproteinase-2 isoform in skeletal muscle ischemia-reperfusion injury.

Ischemia-reperfusion injury (IRI) occurs when blood returns to tissues following a period of ischemia. Reintroduction of blood flow results in the production of free radicals and reactive oxygen species that damage cells. Skeletal muscle IRI is commonly seen in orthopedic trauma patients. Experimental studies in other organ systems have elucidated the importance of extracellular and intracellular matrix metalloproteinase-2 (MMP-2) isoforms in regulating tissue damage in the setting of oxidant stress resulting from IRI. Although the extracellular full-length isoform of MMP-2 (FL-MMP-2) has been previously studied in the setting of skeletal muscle IRI, studies investigating the role of the N-terminal truncated isoform (NTT-MMP-2) in this setting are lacking. In this study, we first demonstrated significant increases in FL- and NTT-MMP-2 gene expression in C2C12 myoblast cells responding to re-oxygenation following hypoxia in vitro. We then evaluated the expression of FL- and NTT-MMP-2 in modulating skeletal muscle IRI using a previously validated murine model. NTT-MMP-2, but not FL-MMP-2 expression was significantly increased in skeletal muscle following IRI. Moreover, the expression of toll-like receptors (TLRs) -2 and -4, IL-6, OAS-1A, and CXCL1 was also significantly up-regulated following IRI. Treatment with the potent anti-oxidant pyrrolidine dithiocarbamate (PDTC) significantly suppressed NTT-MMP-2, but not FL-MMP-2 expression and improved muscle viability following IRI. This data suggests that NTT-MMP-2, but not FL-MMP-2, is the major isoform of MMP-2 involved in skeletal muscle IRI.

Hydrogen-rich saline attenuates ischemia–reperfusion injury in skeletal muscle

BackgroundTo investigate the potential beneficial effect of hydrogen-rich saline (HRS) in ischemia–reperfusion (IR) injury of skeletal muscle. MethodsThree experimental groups were established in male Sprague–Dawley rats: (1) sham group, (2) IR with normal saline group, (3) and IR with HRS group. A rat model of skeletal muscle IR injury was induced by 3-h tourniquet occlusion on its left hind limb and 4-h reperfusion. Normal saline and HRS (1.0 mL/100 g) were administered intraperitoneally at 10 min before reperfusion, respectively. Muscle and serum samples were analyzed for detecting the levels of myeloperoxidase (MPO), superoxide dismutase (SOD), malondialdehyde (MDA), and hydroxyl radical (•OH). Muscle samples were assessed by wet/dry rate, hematoxylin and eosin histologic assessment, Bcl2, Bax, cytochrome C, LC3B, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling, and electron microscopy. ResultsThe wet/dry ratio increased significantly in the IR group (P < 0.01 compared with that in the sham group) and decreased significantly in IR with HRS groups (4.12 ± 0.14 versus 4.12 ± 0.14, P < 0.01 compared with that in the IR group). Muscle tissues and serum of the IR group had significantly increased levels of MPO, MDA, •OH content, and decreased SOD activities compared with the sham group (P < 0.01). The activity of SOD in the IR with HRS group was greatly elevated compared with that in the IR group (295.028 ± 9.288 versus 249.190 ± 5.450 in muscle tissues; 91.627 ± 2.604 versus 73.4045 ± 6.487 in serum; P < 0.01), whereas the levels of MPO, MDA, and •OH content were clearly reduced (MPO: 0.5649 ± 0.0724 versus 1.0984 ± 0.0824 in muscle tissues; 0.7257 ± 0.1232 versus 1.3147 ± 0.0531 in serum. MDA: 4.457 ± 0.650 versus 7.107 ± 0.597 in muscle tissues; 2.531 ± 0.434 versus 4.626 ± 0.237 in serum. •OH: 16.451 ± 0.806 versus 19.871 ± 0.594 in muscle tissues; 500.212 ± 7.387 versus 621.352 ± 7.591 in serum, P < 0.01). The integrated optical density of positive amethyst staining increased significantly in the IR group (P < 0.01 compared with that in the sham group) and decreased significantly in IR with HRS group (928.79 ± 234.537 versus 3005.972 ± 83.567, P < 0.01 compared with that in the IR group). Muscle tissues of the IR group had significantly increased levels of Bax, cytochrome C, LC3B content, and decreased Bcl2 activities compared with those in the sham group (P < 0.01). The activity of Bcl2 in the IR with HRS group was greatly elevated compared with that in the IR group (0.2635 ± 0.0704 versus 0.1242 ± 0.0662; P < 0.01), whereas the levels of Bax, cytochrome C, and LC3B content were clearly reduced (Bax: 0.3103 ± 0.0506 versus 0.5122 ± 0.0148; cytochrome C: 0.4194 ± 0.1116 versus 0.8127 ± 0.0166; LC3B: 0.5884 ± 0.0604 versus 1.3758 ± 0.0319; respectively, P < 0.01). ConclusionsHRS seems to be effective in attenuating IR injury in skeletal muscle via its antioxidant, anti-apoptosis, and anti-autophagy effect.