Rat Muscle Research Articles

Early Movement Restriction Affects FNDC5/Irisin and BDNF Levels in Rat Muscle and Brain.

Interaction with the environment appears necessary for the maturation of sensorimotor and cognitive functions in early life. In rats, a model of sensorimotor restriction (SMR) from postnatal day 1 (P1) to P28 has shown that low and atypical sensorimotor activities induced the perturbation of motor behavior due to muscle weakness and the functional disorganization of the primary somatosensory and motor cortices. In the present study, our objective was to understand how SMR affects the muscle-brain dialogue. We focused on irisin, a myokine secreted by skeletal muscles in response to exercise. FNDC5/irisin expression was determined in hindlimb muscles and brain structures by Western blotting, and irisin expression in blood and cerebrospinal fluid was determined using an ELISA assay at P8, P15, P21 and P28. Since irisin is known to regulate its expression, Brain-Derived Neurotrophic Factor (BDNF) levels were also measured in the same brain structures. We demonstrated that SMR increases FNDC5/irisin levels specifically in the soleus muscle (from P21) and also affects this protein expression in several brain structures (as early as P15). The BDNF level was increased in the hippocampus at P8. To conclude, SMR affects FNDC5/irisin levels in a postural muscle and in several brain regions and has limited effects on BDNF expression in the brain.

Role of GPR81 in regulating intramuscular triglyceride storage during aerobic exercise in rats.

Lactate, a metabolite of exercise, plays a crucial role in the body. In these studies, we aimed to investigate the role of G protein-coupled receptor 81 (GPR81), a specific receptor for lactate, in regulating lipid storage in the gastrocnemius muscle of rats. To achieve this, we measured the impact of sodium 3-hydroxybutyrate (3-OBA) concentration and time on the cAMP-PKA signaling pathway in the gastrocnemius muscles ofrats. Our investigation involved determining the effects of administering 3-OBA at a concentration of 3 mmol L-1 just 15 min before exercise. As expected, exercise led to a notable increase in intramuscular lactate concentration in rats. However, injecting 3-OBA prior to exercise yielded intriguing results. It not only further augmented the cAMP-PKA signaling pathway but also boosted the expression of lipolysis-related proteins such as hormone-sensitive lipase (HSL) and adipose triglyceride lipase (ATGL). Simultaneously, it decreased the expression of fat-synthesizing proteins, including acetyl CoA carboxylase (ACC) and fatty acid synthase (FAS), while increasing the protein expression of cytochrome c oxidase subunit Ⅳ(COX Ⅳ) and the activity of citrate synthetase (CS). Unfortunately, there was no significant change observed in intramuscular triglyceride (IMTG) content. In summary, our findings shed light on the role of lactate in partially regulating intramuscular triglycerides during exercise.

EFFECT OF AMMONIUM PYRROLIDINEDITHIOCARBAMATE ON THE DEVELOPMENT OF OXIDATIVE STRESS IN BICEPS FEMORIS MUSCLES OF RATS DURING MODELLED METABOLIC SYNDROME

Excessive consumption of high-calorie food, a sedentary lifestyle and increased psycho-emotional stress are risk factors for the development of metabolic syndrome. These factors are especially relevant for residents of countries with highly developed economies. Metabolic syndrome is accompanied not only by metabolic disorders, but also leads to the development of a systemic inflammatory response, which is associated with excessive production and circulation of cytokines in the blood. As a rule, the activation of the transcription factor NF-κB leads to an increase in the production of pro-inflammatory cytokines.
 The aim of this work is to determine the effect of the NF-κB transcription factor activation inhibitor on the activity of antioxidant enzymes, the production of superoxide anion radical, the content of oxidatively modified proteins, and the concentration of malondialdehyde in the biceps femoris muscle of rats who underwent experimental metabolic syndrome.
 The study included 24 sexually mature male Wistar rats weighing 200-260 g. The animals were divided into 4 groups of 6 animals each. The first group was the control group; the second group involved the rats exposed to metabolic syndrome simulated by adding a 20% fructose solution as the only source of drinking water to the standard vivarium diet for 60 days; the third group involved the animals received ammonium pyrrolidine dithiocarbamate in a dose of 76 mg/kg intraperitoneally 3 times a week for 60 days; the fourth group was exposed to the combined effect of the ammonium pyrrolidinedithiocarbamate administration during modeled metabolic syndrome. In this study we investigated a 10% homogenate of the biceps femoris muscle with the research focus on several parameters: superoxide anion radical production, superoxide dismutase and catalase activity, malondialdehyde concentration, and the presence of oxidatively modified proteins.
 Simulation of the metabolic syndrome led to the development of oxidative stress in the biceps muscle of rats that was accompanied by an increase in the production of the superoxide anion radical and a decrease in the activity of antioxidant enzymes. The administration of ammonium pyrrolidinedithiocarbamate during modeled metabolic syndrome caused a decrease in the basic production of superoxide anion radical, production of superoxide anion radical by the microsomal electron transport chain and production of superoxide anion radical by the mitochondrial electron transport chain by 30.28%, 26.21 %, and 27.00%, respectively, compared to the rats in metabolic syndrome group. Superoxide dismutase activity increased by 78.81% and catalase activity grew by 144.74% compared to the metabolic syndrome group. The concentration of free malondialdehyde under the administration of ammonium pyrrolidinedithiocarbamate during metabolic syndrome reduced by 51.80%, and the content of oxidatively modified proteins decreased by 31.74% compared to the metabolic syndrome group.
 The administration of ammonium pyrrolidinedithiocarbamate, an NF-κB transcription factor activation inhibitor, effectively prevents the development of oxidative stress in the biceps femoris muscle of rats with modeled metabolic syndrome.
 This study is a part of the initiative research project No. 0124U000092 "High- and low-intensity phenotypes of systemic inflammatory response: molecular mechanisms and new medical technologies for their prevention and correction".

Differential impact of unilateral stroke on the bihemispheric motor cortex representation of the jaw and tongue muscles in young and aged rats.

Dysphagia commonly occurs after stroke, yet the mechanisms of post-stroke corticobulbar plasticity are not well understood. While cortical activity associated with swallowing actions is bihemispheric, prior research has suggested that plasticity of the intact cortex may drive recovery of swallowing after unilateral stroke. Age may be an important factor as it is an independent predictor of dysphagia after stroke and neuroplasticity may be reduced with age. Based on previous clinical studies, we hypothesized that cranial muscle activating volumes may be expanded in the intact hemisphere and would contribute to swallowing function. We also hypothesized that older age would be associated with limited map expansion and reduced function. As such, our goal was to determine the impact of stroke and age on corticobulbar plasticity by examining the jaw and tongue muscle activating volumes within the bilateral sensorimotor cortices. Using the middle cerebral artery occlusion rat stroke model, intracortical microstimulation (ICMS) was used to map regions of sensorimotor cortex that activate tongue and jaw muscles in both hemispheres. Young adult (7 months) and aged (30 months) male F344 × BN rats underwent a stroke or sham-control surgery, followed by ICMS mapping 8 weeks later. Videofluoroscopy was used to assess oral-motor functions. Increased activating volume of the sensorimotor cortex within the intact hemisphere was found only for jaw muscles, whereas significant stroke-related differences in tongue activating cortical volume were limited to the infarcted hemisphere. These stroke-related differences were correlated with infarct size, such that larger infarcts were associated with increased jaw representation in the intact hemisphere and decreased tongue representation in the infarcted hemisphere. We found that both age and stroke were independently associated with swallowing differences, weight loss, and increased corticomotor thresholds. Laterality of tongue and jaw representations in the sham-control group revealed variability between individuals and between muscles within individuals. Our findings suggest the role of the intact and infarcted hemispheres in the recovery of oral motor function may differ between the tongue and jaw muscles, which may have important implications for rehabilitation, especially hemisphere-specific neuromodulatory approaches. This study addressed the natural course of recovery after stroke; future work should expand to focus on rehabilitation.

Circulating and Tissues Concentrations of Selected Trace Elements in Response to Moderate Physical Exercise: Protective Effects of Cucurbita pepo Seeds Oil Extraction

Background: Physical exercise and the subsequent recovery processes have been proposed to affect the essential hemostatic balance of trace elements, either by increasing degradation rates or by enhancing loss from the body. This might disturb the function of vital metabolic organs, such as muscles and liver, leading to unsatisfactory performance. It appears that taking advantage of mineral-rich herbal supplements could replenish the body's mineral sources. Objectives: The present study aimed to investigate the effects of Cucurbita pepo supplementation on exercise-induced dynamics of micronutrients in serum, liver, and skeletal muscle of Wistar rats. Methods: Forty male Wistar rats (aged 4 - 6 weeks old, weighing 130 - 150 g) were randomly divided into four groups of ten rats each: (1) Saline control group (SC), (2) training at 25 m/min group (ST), (3) pumpkin seed oil supplementation group (PC), and (4) pumpkin seed oil supplementation + Training at 25m/min (PT). Moderate-intensity aerobic exercise (25 m/min, eliciting approximately 65% VO2Peak) and consumption of pumpkin seed oil (4 ml/kg/day) were conducted for 6 weeks. The ICP-OES technique was used to measure plasma, liver, and skeletal muscle levels of zinc (Zn), iron (Fe), manganese (Mn), magnesium (Mg), and copper (Cu) approximately 48 to 72 hours after the final session of the training protocol. Data were analyzed using one-way ANOVA and post-hoc Bonferroni tests (P < 0.05). Results: No significant changes in serum Fe (P = 0.728) and Mn (P = 0.52) levels, as well as liver tissue levels of Zn (P = 0.633), Fe (P = 0.512), Mn (P = 0.194), Mg (P = 0.156), and Cu (P = 0.711), were observed among the intervention groups. However, all trace mineral element (TME) concentrations, except for Cu (P = 0.907) in the gastrocnemius tissue, significantly changed among the study groups (P < 0.05). Additionally, the results of the Bonferroni test showed that six weeks of moderate physical activity decreased the serum, liver, and gastrocnemius levels of TMEs, and subsequent supplementation with pumpkin seed oil played a replenishing role in preserving the resources. Conclusions: Pumpkin seed oil supplementation was demonstrated to be an effective ergogenic aid for the up-regulation and replenishment of exercise-induced TME excretion from the liver, gastrocnemius, and body fluids in healthy rats.

Laser-assisted synthesis of nano-hydroxyapatite and functionalization with bone active molecules for bone regeneration

The main goal of bone tissue engineering research is to replace the allogenic and autologous bone graft substitutes that can promote bone repair. Owing to excellent biocompatibility and osteoconductivity, hydroxyapatite is in extensive research and high demand for both medical and non-medical applications. Although various methods have been developed for the synthesis of hydroxyapatite, in the present study we have shown the use of nanosecond laser energy in the wet precipitation method of nano-hydroxyapatite (nHAP) synthesis without using ammonium solution or any other chemicals for pH maintenance. Here, the present study aimed to fabricate the nanohydroxyapatite using a nanosecond laser. The X-ray diffraction and Fourier transform infrared spectroscopy have confirmed the hydroxyapatite formation under laser irradiation in less time without aging. A transmission electron microscopy confirmed the nano size of synthesized nHAP, which is comparable to conventional nHAP. The length and width of the laser-assisted nHAP were found to be in the range of 50–200 nm and 15–20 nm, respectively, at various laser parameters. The crystallite size obtained by Debye Scherrer formulae was found to be in the range of ∼ 16–36 nm. In addition, laser-assisted nHAP based composite cryogel (nanohydroxyapatite/gelatin/collagen I) was synthesized and impregnated with bioactive molecules (bone morphogenic protein and zoledronic acid) that demonstrated significant osteogenic potential both in vitro in cell experiment and in vivo rat muscle pouch model (abdomen and tibia muscles). Dual-energy X-ray analysis, micro-CT, and histological analysis confirmed ectopic bone regeneration. Micro-CT based histomorphometry showed a higher amount (more than 10-fold) of mineralization for animal groups implanted with composite cryogels loaded with bioactive molecules compared to only composite cryogels groups. Our findings thus demonstrate a controlled and rapid synthetic method for the synthesis of nHAP with various physical, chemical, and biological properties exhibited as comparable to conventionally synthesized nHAP.

Novel Bioresorbable Drug-Eluting Mesh Scaffold for Therapy of Muscle Injury.

A novel bioresorbable drug-eluting polycaprolactone (PCL) mesh scaffold was developed, utilizing a solvent-cast additive manufacturing technique, to promote therapy of muscle injury. The degradation rate and mechanical properties strength of the PCL mesh were characterized after immersion in a buffer solution for different times. The in vitro release characteristics of vancomycin, ceftazidime, and lidocaine from the prepared mesh were evaluated using a high-performance liquid chromatography (HPLC) assay. In addition, the in vivo efficacy of PCL meshes for the repair of muscle injury was investigated on a rat model with histological examinations. It was found that the additively manufactured PCL meshes degraded by 13% after submission in buffered solution for four months. All PCL meshes with different pore sizes exhibited greater strength than rat muscle and survived through 10,000 cyclic loadings. Furthermore, the meshes could offer a sustained release of antibiotics and analgesics for more than 3 days in vitro. The results of this study suggest that drug-loaded PCL mesh exhibits superior ability to pure PCL mesh in terms of effectively promoting muscle repair in rat models. The histological assay also showed adequate biocompatibility of the resorbable meshes. The additively manufactured biodegradable drug-eluting meshes may be adopted in the future in humans for the therapy of muscle injuries.

Effects of age and diet consistency on the expression of myosin heavy-chain isoforms on jaw-closing and jaw-opening muscles in a rat model.

Skeletal craniofacial morphology can be influenced by changes in masticatory muscle function, which may also change the functional profile of the muscles. To investigate the effects of age and functional demands on the expression of Myosin Heavy-Chain (MyHC) isoforms in representative jaw-closing and jaw-opening muscles, namely the masseter and digastric muscles respectively. Eighty-four male Wistar rats were divided into four age groups, namely an immature (n = 12; 4-week-old), early adult (n = 24; 16-week-old), adult (n = 24; 26-week-old) and mature adult (n = 24; 38-week-old) group. The three adult groups were divided into two subgroups each based on diet consistency; a control group fed a standard (hard) diet, and an experimental group fed a soft diet. Rats were sacrificed, and masseter and digastric muscles dissected. Real-time quantitative polymerase chain reaction was used to compare the mRNA transcripts of the MyHC isoforms-Myh7 (MyHC-I), Myh2 (MyHC-IIa), Myh4 (MyHC-IIb) and Myh1 (MyHC-IIx)-of deep masseter and digastric muscles. In the masseter muscle, hypofunction increases Myh1 (26, 38 weeks; p < .0001) but decreases Myh4 (26 weeks; p = .046) and Myh2 (26 weeks; p < .0001) expression in adult rats. In the digastric muscle, hypofunction increases Myh1 expression in the mature adult rats (38 weeks; p < .0001), while Myh2 expression decreases in adult rats (26 weeks; p = .021) as does Myh4 (26 weeks; p = .001). Myh7 expression is increased in the digastric muscle of mature adult rats subjected to hypofunction (38 weeks; p = <.0001), while it is very weakly expressed in the masseter. In jaw-opening and jaw-closing muscles, differences in myosin expression between hard- and soft-diet-fed rats become evident in adulthood, suggesting that long-term alteration of jaw function is associated with changes in the expression of MyHC isoforms and potential fibre remodelling. This may give insight into the role of function on masticatory muscles and the resultant craniofacial morphology.

Celecoxib attenuates hindlimb unloading-induced muscle atrophy via suppressing inflammation, oxidative stress and ER stress by inhibiting STAT3.

Improving inflammation may serve as useful therapeutic interventions for the hindlimb unloading-induced disuse muscle atrophy. Celecoxib is a selective non-steroidal anti-inflammatory drug. We aimed to determine the role and mechanism of celecoxib in hindlimb unloading-induced disuse muscle atrophy. Celecoxib significantly attenuated the decrease in soleus muscle mass, hindlimb muscle function and the shift from slow- to fast-twitch muscle fibers caused by hindlimb unloading in rats. Importantly, celecoxib inhibited the increased expression of inflammatory factors, macrophage infiltration in damaged soleus muscle. Mechanistically, Celecoxib could significantly reduce oxidative stress and endoplasmic reticulum stress in soleus muscle of unloaded rats. Furthermore, celecoxib inhibited muscle proteolysis by reducing the levels of MAFbx, MuRF1, and autophagy related proteins maybe by inhibiting the activation of pro-inflammatory STAT3 pathway in vivo and in vitro. This study is the first to demonstrate that celecoxib can attenuate disuse muscle atrophy caused by hindlimb unloading via suppressing inflammation, oxidative stress and endoplasmic reticulum stress probably, improving target muscle function and reversing the shift of muscle fiber types by inhibiting STAT3 pathways-mediated inflammatory cascade. This study not only enriches the potential molecular regulatory mechanisms, but also provides new potential therapeutic targets for disuse muscle atrophy.

Simultaneous Effect of Eight Weeks High-fat Diet, Continuous Moderate-intensity Training, and High-intensity Intermittent Training on the Gene Expression Levels of PLIN2 and ATGL in the Twin Muscle of Male Wistar Rats

Background: A high-fat diet can cause obesity and endanger health. Therefore, the present study aimed to investigate the effect of eight weeks of high-fat diet, continuous moderate-intensity training, and high-intensity intermittent training on the gene expression levels of PLIN2 and ATGL in the twin muscles of male Wistar rats. Materials and Methods: Thirty-two male Wistar rats were divided into four groups (n=8): 1- control group with a standard diet (ND) 2- control group with a high-fat diet (HFD) 3- high-fat diet and continuous aerobic training with Moderate intensity (HFD+MICT) 4- high-fat diet and intermittent aerobic training (HFD+HIIT). For one training group, eight weeks of moderate-intensity continuous aerobic training (MICT), and for the other group, eight weeks of high-intensity intermittent aerobic training (HIIT) were performed on the treadmill. Then, 48 hours after the last training session, the twin muscle tissue was isolated to check the expression of PLIN2 and ATGL genes. Gene expression analysis was done by Real-time PCR technique. One-way analysis of variance (ANOVA) and Tukey's post-hoc test were used to analyze the collected data. Results: After eight weeks of training, the level of PLIN2 gene expression in the two training groups was significantly lower than in the HDF group, and the level of ATGL gene expression was significantly higher in both groups than in the HDF group (P&lt;0.05). Conclusion: It was shown that two exercises had a modulating effect on the expression of genes that are effective in the process of autophagy and the formation of fat cells.

Effects of hindlimb unloading on the mevalonate and mechanistic target of rapamycin complex 1 signaling pathways in a fast-twitch muscle in rats.

Fast-twitch muscles are less susceptible to disuse atrophy, activate the mechanistic target of rapamycin complex 1 (mTORC1) signaling pathway, and increase protein synthesis under prolonged muscle disuse conditions. However, the mechanism underlying prolonged muscle disuse-induced mTORC1 signaling activation remains unclear. The mevalonate pathway activates the mTORC1 signaling pathway via the prenylation and activation of Ras homolog enriched in brain (Rheb). Therefore, we investigated the effects of hindlimb unloading (HU) for 14 days on the mevalonate and mTORC1 signaling pathways in the plantaris muscle, a fast-twitch muscle, in adult male rats. Rats were divided into HU and control groups. The plantaris muscles of both groups were harvested after the treatment period, and the expression and phosphorylation levels of metabolic and intracellular signaling proteins were analyzed using Western blotting. We found that HU increased the expression of 3-hydroxy-3-methylglutaryl-coenzyme A reductase, the rate-limiting enzyme of the mevalonate pathway, and activated the mTORC1 signaling pathway without activating AKT, an upstream activator of mTORC1. Furthermore, HU increased prenylated Rheb. Collectively, these findings suggest that the activated mevalonate pathway may be involved in the activation of the Rheb/mTORC1 signaling pathway without AKT activation in fast-twitch muscles under prolonged disuse conditions.

Ultrasound-based assessment of the expression of inflammatory markers in the rectus femoris muscle of rats.

Ultrasonographic characteristics of skeletal muscles are related to their health status and functional capacity, but they still provide limited information on muscle composition during the inflammatory process. It has been demonstrated that an alteration in muscle composition or structure can have disparate effects on different ranges of ultrasonogram pixel intensities. Therefore, monitoring specific clusters or bands of pixel intensity values could help detect echotextural changes in skeletal muscles associated with neurogenic inflammation. Here we compare two methods of ultrasonographic image analysis, namely, the echointensity (EI) segmentation approach (EI banding method) and detection of selective pixel intensity ranges correlated with the expression of inflammatory regulators using an in-house developed computer algorithm (r-Algo). This study utilized an experimental model of neurogenic inflammation in segmentally linked myotomes (i.e., rectus femoris (RF) muscle) of rats subjected to lumbar facet injury. Our results show that there were no significant differences in RF echotextural variables for different EI bands (with 50- or 25-pixel intervals) between surgery and sham-operated rats, and no significant correlations among individual EI band pixel characteristics and protein expression of inflammatory regulators studied. However, mean numerical pixel values for the pixel intensity ranges identified with the proprietary r-Algo computer program correlated with protein expression of ERK1/2 and substance P (both 86-101-pixel ranges) and CaMKII (86-103-pixel range) in RF, and were greater (p < 0.05) in surgery rats compared with their sham-operated counterparts. Our findings indicate that computer-aided identification of specific pixel intensity ranges was critical for ultrasonographic detection of changes in the expression of inflammatory mediators in neurosegmentally-linked skeletal muscles of rats after facet injury.

Inhibitory effect of acupoint electrostimulation with different layers and intensities on muscular inflammatory pain and spinal WDR neuron activity.

To observe the analgesic effects of different levels and intensities of electrical stimulation on the local acupoints in the pain source area and their impact on wide dynamic range (WDR) neurons in the spinal dorsal horn, in order to provide a basis for selecting appropriate parameters for electroacupuncture (EA) stimulation. Wistar rats were used in 3 parts of the experiment. Complete Freund's adjuvant was used to establish a model of inflammation-induced pain in the gastrocnemius muscle. After modeling, 6 rats were randomly selected for multi-channel extracellular electrophysiological recording of the electrical activity of WDR neurons, to determine the threshold for activating the A-component (Ta) and the C-component (Tc), which were used as the intervention intensities for skin transcutaneous electrical acupoint stimulation (TEAS) or EA. Thirty-six rats were randomly divided into normal , model , TEAS-Ta , TEAS-Tc, EA-Ta , and EA-Tc groups, with 6 rats in each group. In the pain source area , Ta or Tc intensity of TEAS or EA intervention at"Chengshan"(BL57) was performed for 30 min each time, once a day, for 3 consecutive days. A small animal pressure pain measurement instrument was used to measure the mechanical pressure pain threshold of the gastrocnemius muscle in rats, and the Von Frey filament was used to measure the mechanical pain threshold of the footpad. Thirteen rats were randomly selected to observe the immediate responsiveness of WDR neurons to Ta/Tc intensity of EA or TEAS in BL57. The thresholds of TEAS to activate WDR neuron A-component or C-component were (2.43±0.57) mA and (7.00±1.34) mA, respectively, while the thresholds for EA to activate muscle WDR neuron A-component or C-component were (0.72±0.34) mA and (1.58±0.35) mA, respectively. After injection of CFA into the gastrocnemius muscle, compared with the normal group both the mechanical pressure pain threshold of the gastrocnemius muscle and the mechanical pain threshold of the footpad of rats in the model group were significantly decreased (P<0.001). After TEAS-Ta, TEAS-Tc or EA-Ta intervention in the BL57, both the mechanical pressure pain threshold of the gastrocnemius muscle and the mechanical pain threshold of the footpad were significantly higher than those in the model group (P<0.05, P<0.001). Compared with the normal group, the electrical threshold for evoking WDR neuron C-component discharge was significantly decreased (P<0.001) in the model group, while increased after TEAS-Ta, TEAS-Tc, or EA-Ta intervention (P<0.01) compared with the model group. The evoked discharge frequency of muscle WDR neurons decreased significantly after immediate intervention with TEAS-Ta, TEAS-Tc, or EA-Ta (P<0.01, P<0.05). EA-Tc had no significant improvement on the evoked electrical activity of WDR neurons or pain behavior. TEAS-Ta, TEAS-Tc, or EA-Ta can all alleviate the local and footpad mechanical pain in rats with muscle inflammation and inhibit the responsiveness of WDR neurons, indicating that different intensities are required for analgesic effects at different levels of acupoints in the pain source area.

Effect of robotic gait training on muscle and bone characteristics in spinal cord transected rats.

Osteoporosis and loss of muscle mass are secondary issues with spinal cord injury. Robotic gait training has provided evidence of increasing bone density and muscle mass, but its effect on bone strength is undetermined. The purpose of this study was to determine the effect of a 6-week robotic locomotion training program on skeletal muscle mass and bone characteristics. Twelve female Sprague-Dawley rats received a mid-thoracic spinal cord transection at 5 days old and at 3 weeks old were assigned to a Control or Trained Group. The Trained Group performed 5-min sessions on the Rat Stepper 5 days a week for 6 weeks with 90% of body weight supported. At the end of the 6 weeks, body mass was obtained and right femurs and four lower extremity muscles were harvested. Femur bone mineral density was measured with DXA and mechanical characteristics of the femur were determined via 3-point bending testing. Independent t-tests, effects sizes and percent differences were computed between the two groups (p < 0.05). The Trained Group had significantly larger normalized femur mass (p = 0.007) and normalized soleus muscle mass (p = 0.033) when compared to the Control Group. There was a medium or large effect size with the Trained Groups' femurs having larger mass, bone mineral density, rupture loads, cortical wall thickness, shaft cross sectional area, soleus mass, normalized gastrocnemius mass, and smaller shaft inner diameters compared to the Control Group. These changes may contribute to decreasing osteoporosis and fracture risk in those with spinal cord injuries.

Real-Time Tissue Classification Using a Novel Optical Needle Probe for Biopsy.

Core needle biopsy is a part of the histopathological process, which is required for cancerous tissue examination. The most common method to guide the needle inside of the body is ultrasound screening, which in greater part is also the only guidance method. Ultrasound screening requires user experience. Furthermore, patient involuntary movements such as breathing might introduce artifacts and blur the screen. Optically enhanced core needle biopsy probe could potentially aid interventional radiologists during this procedure, providing real-time information on tissue properties close to the needle tip, while it is advancing inside of the body. In this study, we used diffuse optical spectroscopy in a custom-made core needle probe for real-time tissue classification. Our aim was to provide initial characteristics of the smart needle probe in the differentiation of tissues and validate the basic purpose of the probe of informing about breaking into a desired organ. We collected optical spectra from rat blood, fat, heart, kidney, liver, lungs, and muscle tissues. Gathered data were analyzed for feature extraction and evaluation of two machine learning-based classifiers: support vector machine and k-nearest neighbors. Their performances on training data were compared using subject-independent k-fold cross-validation. The best classifier model was chosen and its feasibility for real-time automated tissue recognition and classification was then evaluated. The final model reached nearly 80% of correct real-time classification of rat organs when using the needle probe during real-time classification.

Up-regulated expression of two-pore domain K+ channels, KCNK1 and KCNK2, is involved in the proliferation and migration of pulmonary arterial smooth muscle cells in pulmonary arterial hypertension.

Pulmonary arterial hypertension (PAH) is a severe and rare disease in the cardiopulmonary system. Its pathogenesis involves vascular remodeling of the pulmonary artery, which results in progressive increases in pulmonary arterial pressure. Chronically increased pulmonary arterial pressure causes right ventricular hypertrophy and subsequent right heart failure. Pulmonary vascular remodeling is attributed to the excessive proliferation and migration of pulmonary arterial smooth muscle cells (PASMCs), which are induced by enhanced Ca2+ signaling following the up-/down-regulation of ion channel expression. In the present study, the functional expression of two-pore domain potassium KCNK channels was investigated in PASMCs from idiopathic PAH (IPAH) patients and experimental pulmonary hypertensive (PH) animals. In IPAH-PASMCs, the expression of KCNK1/TWIK1 and KCNK2/TREK1 channels was up-regulated, whereas that of KCNK3/TASK1 and KCNK6/TWIK2 channels was down-regulated. The similar up-regulated expression of KCNK1 and KCNK2 channels was observed in the pulmonary arterial smooth muscles of monocrotaline-induced PH rats, Sugen 5416/hypoxia-induced PH rats, and hypoxia-induced PH mice. The facilitated proliferation of IPAH-PASMCs was suppressed by the KCNK channel blockers, quinine and tetrapentylammonium. The migration of IPAH-PASMCs was also suppressed by these channel blockers. Furthermore, increases in the proliferation and migration were inhibited by the siRNA knockdown of KCNK1 or KCNK2 channels. The siRNA knockdown also caused membrane depolarization and subsequent decrease in cytosolic [Ca2+]. The phosphorylated level of c-Jun N-terminal kinase (JNK) was elevated in IPAH-PASMCs compared to normal-PASMCs. The increased phosphorylation was significantly reduced by the siRNA knockdown of KCNK1 or KCNK2 channels. Collectively, these findings indicate that the up-regulated expression of KCNK1 and KCNK2 channels facilitates the proliferation and migration of PASMCs via enhanced Ca2+ signaling and JNK signaling pathway, which is associated with vascular remodeling in PAH.

Effect of Supplementation with Omega-3 Polyunsaturated Fatty Acids on Metabolic Modulators in Skeletal Muscle of Rats with an Obesogenic High-Fat Diet.

Previous studies provided evidence of the benefits of omega-3 polyunsaturated fatty acids (ω-3 PUFA) on the cardiovascular system and inflammation. However, its possible effect on skeletal muscle is unknown. This study aimed to evaluate whether ω-3 PUFA reverses the dysregulation of metabolic modulators in the skeletal muscle of rats on a high-fat obesogenic diet. For this purpose, an animal model was developed using male Wistar rats with a high-fat diet (HFD) and subsequently supplemented with ω-3 PUFA. Insulin resistance was assessed, and gene and protein expression of metabolism modulators in skeletal muscle was also calculated using PCR-RT and Western blot. Our results confirmed that in HFD rats, zoometric parameters and insulin resistance were increased compared to SD rats. Furthermore, we demonstrate reduced gene and protein expression of peroxisome proliferator-activated receptors (PPARs) and insulin signaling molecules. After ω-3 PUFA supplementation, we observed that glucose (24.34%), triglycerides (35.78%), and HOMA-IR (40.10%) were reduced, and QUICKI (12.16%) increased compared to HFD rats. Furthermore, in skeletal muscle, we detected increased gene and protein expression of PPAR-α, PPAR-γ, insulin receptor (INSR), insulin receptor substrate 1 (ISR-1), phosphatidylinositol-3-kinase (PI3K), and glucose transporter 4 (GLUT-4). These findings suggest that ω-3 PUFAs decrease insulin resistance of obese skeletal muscle.

The temperature sensitivity of motor units in rat soleus

Temperature has a significant impact on the performance of the neuromuscular system and motor control processes. The smallest functional components of these systems are motor units (MUs), which may differ significantly between different muscles. The influence of temperature on the contractile properties of slow-twitch (S) MUs from soleus (SOL) muscles in rats was investigated under hypothermia (25 °C), normothermia (37 °C), and hyperthermia (41 °C). Hypothermia prolonged the twitch time parameters, decreased the rate of force development, increased the twitch-to-tetanus ratio, enhanced twitch force, and abolished post-tetanic depression. In contrast, hyperthermia did not alter twitch time parameters. Moreover, there was no effect on force despite the noted increase in post-tetanic depression and the twitch-to-tetanus ratio. Therefore, hypothermia induced more profound changes in S MUs compared with hyperthermia. The temperature effects in SOL MUs were compared to the effects previously reported for S MUs in the medial gastrocnemius (MG). The major differences between the S MUs of both muscles were the effects of temperature on twitch force, post-tetanic force modulation, twitch-to-tetanus ratio, and the slope of the force-frequency curve under hypothermia. Hyperthermia shortened twitch time parameters solely in the MG. In contrast, post-tetanic depression, twitch-to-tetanus ratio, and the slope of the force-frequency curve were influenced by hyperthermia only in SOL MUs. The different temperature effects of S MUs probably corresponded to differences in muscle architecture and their diverse functional tasks and enzyme activity. In summary, S MUs in SOL are more thermal-sensitive than their counterparts in MG.

Mechanical interaction of myosin and native thin filament in the disused rat soleus muscle

The disuse of skeletal limb muscles occurs in a variety of conditions, yet our comprehension of the molecular mechanisms involved in adaptation to disuse remains incomplete. We studied the mechanical characteristics of actin-myosin interaction using an in vitro motility assay and isoform composition of myosin heavy and light chains by dint of SDS-PAGE in soleus muscle of both control and hindlimb-unloaded rats. 14 days of hindlimb unloading led to the increased maximum sliding velocity of actin, reconstituted, and native thin filaments over rat soleus muscle myosin by 24 %, 19 %, and 20 %, respectively. The calcium sensitivity of the “pCa-velocity” relationship decreased. There was a 26 % increase in fast myosin heavy chain IIa (MHC IIa), a 22 % increase in fast myosin light chain 2 (MLC 2f), and a 13 % increase in fast MLC 1f content. The content of MLC 1s/v, typical for slow skeletal muscles and cardiac ventricles did not change. At the same time, MLC 1s, typical only for slow skeletal muscles, disappeared. The maximum velocity of soleus muscle native thin filaments was 24 % higher compared to control ones sliding over the same rabbit myosin. Therefore, both myosin and native thin filament kinetics could influence the mechanical characteristics of the soleus muscle. Additionally, the MLC 1s and MLC 1s/v ratio may contribute to the mechanical characteristics of slow skeletal muscle, along with MHC, MLC 2, and MLC 1 slow/fast isoforms ratio.

MiR-122/PPARβ axis is involved in hypoxic exercise and modulates fatty acid metabolism in skeletal muscle of obese rats

Hypoxic exercise is an effective intervention for obesity, because it promotes weight loss by regulating fatty acid (FA) metabolism. The regulation of peroxisome proliferator-activated receptor β (PPARβ) by miR-122 may be involved in this process, but the detailed mechanisms are unknown. In order to address this issue, we probed how miR-122 affected the expression of factors associated with FA metabolism in skeletal muscle of obese rats undergoing hypoxic training. By injecting adeno-associated virus 9 containing miR-122 overexpression vector or miR-122 inhibitor into skeletal muscles of rats with a 4-week hypoxic exercise regimen, the miR-122 expression level can be regulated. Body composition and blood lipid levels were analyzed, and PPARβ, carnitine palmitoyltransferase 1b (CPT1b), acetylCoA carboxylase 2 (ACC2), and FA synthase (FAS) mRNA and protein levels were evaluated using quantitative reverse transcription quantitative PCR(RT-qPCR) and Western blot analysis. We found that miR-122 overexpression increased low-density lipoprotein cholesterol (LDL-C) and triglyceride (TG) levels and decreased PPARβ, ACC2, and FAS expression. Conversely, miR-122 inhibition decreased TG level, increased high-density lipoprotein cholesterol (HDL-C) level, and upregulated PPARβ, ACC2, FAS, and CPT1b. These data indicated that the negative regulation of PPARβ by miR-122 promotes FA metabolism by altering the levels of the factors related to FA metabolism in skeletal muscle of obese rat under hypoxic training, thus providing molecular-level insight into the beneficial effects of this intervention.