Gene expression profiling of ventilation induced muscle wasting in rat plantaris
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ABSTRACT: RNASeq technology is applied to study the gene expression affected by ventilation or loading in plantaris. Non-loaded right plantaris group under short-term (6 hours to 4 days) or long-term (4.5 to 9 days) ventilation, as well as the corresponding loaded groups, are included besides the healthy control group.
Project description:RNASeq technology is applied to study the gene expression affected by ventilation in diaphragm and intercostal. Groups under short-term (6 hours to 4 days) or long-term (4.5 to 9 days) ventilation are included besides the healthy control group.
Project description:Acute quadriplegic myopathy (AQM) or critical illness myopathy (CIM) is frequently observed in intensive care unit (ICU) patients. In order to elucidate duration-dependent effects of the ICU intervention on molecular and functional networks that control the muscle wasting and weakness in AQM, gene expression profile was analyzed at time points varying from 6 hours to 14 days in a unique experimental rat model mimicking ICU conditions, i.e., post-synaptically paralyzed, mechanically ventilated and extensively monitored animals. A total of five sham operated controls and 23 experimental female Sprague-Dawley rats were included in this study. The experimental rats were anaesthetized, treated with the neuromuscular blocker (NMBA), α-cobrotoxin, mechanically ventilated and monitored for durations varying from 6h to 4 days (n=13), from 5 to 9 days (n=4), and from 9 to 14 days (n=6). Muscle biopsies were obtained from gastrocnemius muscle (proximal part) immediately after euthanasia and were quickly frozen in liquid propane cooled by liquid nitrogen, and stored at -80°C.RNA was extracted.
Project description:Critically ill intensive care unit (ICU) patients commonly develop severe muscle wasting and impaired muscle function, leading to delayed recovery, with subsequent increased morbidity and financial costs, and decrease quality of life of survivors. Acute Quadriplegic Myopathy (AQM) is one of the most common neuromuscular disorders associated with ICU-acquired muscle weakness. Although there are no available treatments for the ICU-acquired muscle weakness, it has been demonstrated that early mobilization can improve its prognosis and functional outcomes. This study aims at improving our understanding of the effects of passive mechanical loading on skeletal muscle structure and function by using a unique experimental rat ICU model allowing analyses of the temporal sequence of changes in mechanically ventilated and pharmacologically paralyzed animals at durations varying from 6 h to 14 days. Results show that passive mechanical loading alleviated the muscle wasting and the loss of force-generation associated with the ICU intervention, resulting in a doubling of the functional capacity of the loaded vs. unloaded muscles after a 2-week ICU intervention. We demonstrated that the improved maintenance of muscle structure and function is likely a consequence of a reduced oxidative stress, and a reduced loss of the molecular motor protein myosin. A complex temporal gene expression pattern, delineated by microarray analysis, was observed with loading-induced changes in transcript levels of sarcomeric proteins, muscle developmental processes, stress response, ECM/cell adhesion proteins and metabolism. Thus, the results from this study show that passive mechanical loading alleviates the severe negative consequences on muscle structure and function associated with mechanical silencing in ICU patients, strongly supporting early and intense physical therapy in immobilized ICU patients. This study aims to unravel the effects of passive mechanical loading on skeletal muscle structure and function in an experimental rat ICU model at duration varying between 6h and 14 days. A total of 23 experimental female Sprague-Dawley rats were included in this study. The experimental rats were anaesthetized, treated with the neuromuscular blocking agent (NMBA) M-NM-1-cobrotoxin, mechanically ventilated and monitored for durations varying from 6h to 4 days (n=13), from 5 to 8 days (n=4), and from 9 to 14 days (n=6). The left leg of the animal was activated for 6 hours at the shortest duration and 12 hours per day at durations 12 hours and longer throughout the experiment, using a mechanical lever arm that produced a continuous passive maximal ankle joint flexions-extensions at a speed of 13.3 cycles per minute. Muscle biopsies were obtained from gastrocnemius muscle (proximal part) immediately after euthanasia, were quickly frozen in liquid propane cooled by liquid nitrogen, and stored at -80M-BM-0C. RNA was extracted.
Project description:The study aims to characterize a new isolate that lacks mecA but is nevertheless resistant to methicillin in order to find a novel mechanism of methicillin resistance. Samples were hybridized on aminosilane coated slides with 70-mer oligos.
Project description:PSC is a chronic inflammatory condition of the biliary tree causing biliary strictures and fibrosis and predisposes to cholangiocarcinoma. Traditionally it is considered a primary disorder of the bile duct epithelia. Here we investigate whether injury to the arterial blood supply may contribute to PSC. Hepatic arteries were collected from 8 patients at the time of liver transplantation: 4 with PSC and 4 disease controls. Samples were used for RNA-sequencing, differential expression and pathways analyses.
Project description:Background: Skeletal muscle wasting and impaired muscle function in response to mechanical ventilation and immobilization in intensive care unit (ICU) patients are clinically challenging partly due to (i) the poorly understood intricate cellular and molecular networks; and (ii) the unavailability of an animal model mimicking this condition. By employing a unique porcine model mimicking the conditions in the ICU with long-term mechanical ventilation and immobilization, we have analyzed the expression profile of skeletal muscle biopsies taken at three time points during a five-day period. Results: We have analyzed the expression profile of skeletal muscle biopsies taken at three time points during a five-day period. Among the differentially regulated transcripts, extracellular matrix, energy metabolism, sarcomeric proteins and LIM protein mRNA levels were down-regulated while ubiquitin proteasome system, cathepsins, oxidative stress responsive genes and heat shock proteins (HSP) mRNAs were up-regulated Conclusions: We conclude that induction of HSPs may play an inherent temporary protective mechanism in skeletal muscle in the early stages of immobilization and mechanical ventilation. The proposed molecular events leading to our final hypothesis are illustrated in the manuscript. Keywords: Treatment, immobilization, muscle function. Sixteen female domestic piglets (Sus scrofa, average body weight 26.5 kg) were used in this study. All piglets were immobilized by anesthesia and mechanically ventilated (Servo 900C, Siemens-Elema, Solna, Sweden). The first m. biceps femoris biopsies (day 1) were obtained from all animals, after administering anesthetics. During the five day study period, four animals were sedated using isoflurane inhalation (Abbott Laboratories, Chicago, Il, USA, 0.8 – 1.3% end-tidal concentration) supplemented by intravenous bolus doses of morphine and ketamine as needed. Biopsies from the m. biceps femoris were obtained on two further separate occasions (days 3 and 5) in these animals. Core body temperature (blood) was maintained in the range of 38.5 – 40°C by a servocontrolled heating pad. The animals received intravenous crystalloid fluid (Ringer’s acetate) to maintain stable blood pressure and urinary output and a glucose infusion (Rehydrex, Fresenius Kabi, Stockholm, Sweden, 25 mg glucose /mL) in the range of 0.5 – 1.5 mg/kg/minute to decrease the effects of catabolism. Each animal received prophylactic streptomycin 750 mg/d and benzylpenicillin 600 mg/d (Streptocillin Vet, Boeringer-Ingelheim, Hellerup, Denmark). Arterial blood gas analysis as well as electrolytes and blood glucose levels were monitored regularly and kept in the normal range throughout the study period.
Project description:The liver regeneration process terminates when the normal liver-mass/body-weight ratio of 2.5 % has been re-established. We aimed to generate and analyse global expression profiles to study the genetic interactions in relation to liver regeneration, to illuminate the genetic interactions between genes regulating the cell cycle and apoptosis, and to clarify the role of TGF-M-CM-^_ signalling in the terminating phase of liver regeneration. Twelve pigs were randomised to either 60% liver resection (n=4), sham operation (n=4) or control animals (n=4). Liver biopsies were taken at time of resection, after three weeks and upon termination the sixth week. Global gene expression profiles in the biopsies were obtained using porcine oligonucleotide microarrays, representing approximately 20000 porcine genes. Microarray analysis revealed a clear dominance of genes regulating apoptosis towards the end of regeneration, compared to the sham and control groups. Caspase Recruitment Domain-Containing Protein 11 (CARD11) was upregulated six weeks after liver resection, suggesting the involvement of the caspase system at this time. Zinc Finger Protein (ZNF490) gene, with a potential negative effect on cell cycle progression and promotion of apoptosis, was only up regulated at three and six weeks after liver resection indicating a central role at this time. TGF-M-CM-^_ was not found to be significantly affected. CARD11 and ZNF490 appear to play a central role in the termination of liver regeneration in the present study. The lack of TGF-M-CM-^_ could indicate that signaling by TGF-M-CM-^_ is not required for termination of liver regeneration. time course, treatment comparison
Project description:Skeletal muscle possesses the ability to adapt its size in response to milieus, which is called plasticity. Overload like resistance training induces the increment of muscle mass called muscle hypertrophy. Muscle stem cells (also known as muscle stem cells) function to supply new nuclei for myofiber during the overload in muscle. Using compensatory hypertrophy in plantaris muscles, we isolated MuSCs from plantaris muscles 4 days after surgery. Control MuSCs were also prepared from sham plantaris muscles.