Diabetic cardiomyopathy (DCM) is a medical condition characterized by cardiac remodeling and dysfunction in individuals with diabetes mellitus. Sarcoplasmic reticulum (SR) and mitochondrial Ca²⁺ overload in cardiomyocytes have been recognized as biological hallmarks in DCM; however, the specific factors underlying these abnormalities remain largely unknown. In this study, we aimed to investigate the role of a cardiac-specific long noncoding RNA, D830005E20Rik (Trdn-as), in DCM. Our results revealed the remarkably upregulation of Trdn-as in the hearts of the DCM mice and cardiomyocytes treated with high glucose (HG). Knocking down Trdn-as in cardiac tissues significantly improved cardiac dysfunction and remodeling in the DCM mice. Conversely, Trdn-as overexpression resulted in cardiac damage resembling that observed in the DCM mice. At the cellular level, Trdn-as induced Ca²⁺ overload in the SR and mitochondria, leading to mitochondrial dysfunction. RNA-seq and bioinformatics analyses identified calsequestrin 2 (Casq2), a primary calcium-binding protein in the junctional SR, as a potential target of Trdn-as. Further investigations revealed that Trdn-as facilitated the recruitment of METTL14 to the Casq2 mRNA, thereby enhancing the m⁶A modification of Casq2. This modification increased the stability of Casq2 mRNA and subsequently led to increased protein expression. When Casq2 was knocked down, the promoting effects of Trdn-as on Ca²⁺ overload and mitochondrial damage were mitigated. These findings provide valuable insights into the pathogenesis of DCM and suggest Trdn-as as a potential therapeutic target for this condition.
Animals ; Diabetic Cardiomyopathies/pathology* ; RNA, Long Noncoding/genetics* ; Myocytes, Cardiac/metabolism* ; Mice ; Calsequestrin/genetics* ; Calcium/metabolism* ; Male ; Sarcoplasmic Reticulum/metabolism* ; Methyltransferases/metabolism* ; Mice, Inbred C57BL ; Mitochondria, Heart/metabolism* ; Disease Models, Animal ; Mitochondria/metabolism*

Atrial Ca²⁺ handling abnormalities, mainly involving the dysfunction of ryanodine receptor (RyR) and sarcoplasmic reticulum Ca²⁺-ATPase (SERCA), play a role in the pathogenesis of atrial fibrillation (AF). Previously, we found that the expression and function of transient receptor potential vanilloid subtype 4 (TRPV4) are upregulated in a sterile pericarditis (SP) rat model of AF, and oral administration of TRPV4 inhibitor GSK2193874 alleviates AF in this animal model. The aim of this study was to investigate whether oral administration of GSK2193874 could alleviate atrial Ca²⁺ handling abnormalities in SP rats. A SP rat model of AF was established by daubing sterile talcum powder on both atria of Sprague-Dawley (SD) rats after a pericardiotomy, to simulate the pathogenesis of postoperative atrial fibrillation (POAF). On the 3rd postoperative day, Ca²⁺ signals of atria were collected in isolated perfused hearts by optical mapping. Ca²⁺ transient duration (CaD), alternan, and the recovery properties of Ca²⁺ transient (CaT) were quantified and analyzed. GSK2193874 treatment reversed the abnormal prolongation of time to peak (determined mainly by RyR activity) and CaD (determined mainly by SERCA activity), as well as the regional heterogeneity of CaD in SP rats. Furthermore, GSK2193874 treatment relieved alternan in SP rats, and reduced its incidence of discordant alternan (DIS-ALT). More importantly, GSK2193874 treatment prevented the reduction of the S2/S1 CaT ratio (determined mainly by RyR refractoriness) in SP rats, and decreased its regional heterogeneity. Taken together, oral administration of TRPV4 inhibitor alleviates Ca²⁺ handling abnormalities in SP rats primarily by blocking the TRPV4-Ca²⁺-RyR pathway, and thus exerts therapeutic effect on POAF.
Administration, Oral ; Animals ; Atrial Fibrillation/etiology* ; Calcium/metabolism* ; Myocytes, Cardiac/metabolism* ; Pericarditis/pathology* ; Rats ; Rats, Sprague-Dawley ; Ryanodine Receptor Calcium Release Channel/pharmacology* ; Sarcoplasmic Reticulum/pathology* ; TRPV Cation Channels

OBJECTIVETo compare the effect of Shen-Fu Injection (SFI) and epinephrine on the expression of sarcoplasmic reticulum Ca(2+) ATPase 2a (SERCA2a) in a pig model with post-resuscitation myocardial dysfunction.

METHODSVentricular fibrillation (VF) was electrically induced in Wu-zhi-shan miniature pigs. After 8 min of untreated VF and 2 min of cardiopulmonary resuscitation (CPR), all animals were randomly administered a bolus injection of saline placebo (SA group, n=10), SFI (0.8 mg/kg, SFI group, n=10) or epinephrine (20 μg/kg, EPI group, n=10). After 4 min of CPR, a 100-J shock was delivered. If the defibrillation attempt failed to attain restoration of spontaneous circulation (ROSC), manual chest compressions were rapidly resumed for a further 2 min followed by a second defibrillation attempt. Hemodynamic variables were recorded, and plasma concentrations of catecholamines were measured. Adenylate cyclase (AC), cyclic adenosine monophosphate (cAMP) and the expressions of β1-adrenoceptor (AR) and SERCA 2a were determined.

RESULTSCardiac output, left ventricular dp/dtmax and negative dp/dtmax were significantly higher in the SFI group than in the SA and EPI groups at 4 and 6 h after ROSC. The expression of β1-AR and SERCA2a at 24 h after ROSC were significantly higher in the SFI group than in the SA and EPI groups (P<0.05 or P<0.01).

CONCLUSIONSThe administration of epinephrine during CPR decreased the expression of SERCA2a and aggravated postresuscitation myocardial function (P<0.01). SFI attenuated post-resuscitation myocardial dysfunction, and the mechanism might be related to the up-regulation of SERCA2a expression.

Adenylyl Cyclases ; metabolism ; Animals ; Blotting, Western ; Cardiac Output ; drug effects ; Cardiopulmonary Resuscitation ; Cyclic AMP ; metabolism ; Dopamine ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Enzyme-Linked Immunosorbent Assay ; Epinephrine ; blood ; Heart Ventricles ; drug effects ; metabolism ; physiopathology ; Hemodynamics ; drug effects ; Injections ; Male ; Myocardium ; enzymology ; pathology ; Norepinephrine ; blood ; Receptors, Adrenergic, beta-1 ; metabolism ; Sarcoplasmic Reticulum Calcium-Transporting ATPases ; metabolism ; Swine ; Swine, Miniature ; Up-Regulation ; drug effects

OBJECTIVETo observe the dynamic expression of calcium-sensing receptor(CaSR) in myocardium of diabetic rats.

METHODSThirty male Wistar rats were randomly divided into 3 groups including control, diabetic-4 week and diabetic-8 week groups(n = 10). The type 2 diabetes mellitus models were established by intraperitoneal injection of streptozotocin (STZ, 30 mg/kg) after high-fat and high-sugar diet for one month. The cardiac morphology was observed by electron microscope. Western blot analyzed the expression of CaSR, phospholamban (PLN), a calcium handling regulator, and Ca+-ATPase(SERCA) in cardiac tissues.

RESULTSCompared with control group, the expressions of CaSR and SERCA were decreased, while the expression of PLN was significantly increased in a time-dependent manner in diabetic groups. Meanwhile diabetic rats displayed abnormal cardiac structure.

CONCLUSIONThese results indicate that the CaSR expression of myocardium is reduced in the progression of DCM, and its potential mechanism may be related to the imnaired intracellular calcium homeostasis.

Animals ; Calcium-Binding Proteins ; metabolism ; Diabetes Mellitus, Experimental ; complications ; Diabetes Mellitus, Type 2 ; Diabetic Cardiomyopathies ; metabolism ; physiopathology ; Disease Progression ; Heart ; physiopathology ; Male ; Myocardium ; metabolism ; pathology ; Rats ; Rats, Wistar ; Receptors, Calcium-Sensing ; metabolism ; Sarcoplasmic Reticulum Calcium-Transporting ATPases ; metabolism ; Streptozocin

The aim of the present study was to investigate the abnormal calcium re-uptake function of myocardium sarcoplasmic reticulum (SR) in rabbits with heart failure, as well as potential mechanisms. Heart failure model was established in rabbits through aortic insufficiency and constriction of abdominal aorta. The SR Ca(2+) re-uptake function was measured with a calcium imaging device. The activity of myocardium SR calcium adenodine triphosphatase 2a (SERCA2a) was measured by inorganic phosphate. The protein expressions of SERCA2a, CaMKII, PKA, PP1α, phospholamban (PLB), PLB-Ser(16) and PLB-Thr(17) were evaluated by Western blot. The activities of PKA and CaMKII were detected by γ-(32)P substrate incorporation. The results showed that, compared with the sham operation group, the heart failure group exhibited reduced Ca(2+) re-uptake amount (P < 0.01) and the expression and activity of SERCA2a (P < 0.05 or P < 0.01), decreased expression of PLB and its phosphorylation status in sites of Ser(16) and Thr(17) (P < 0.05), increased expressions and activities of PKA and CaMKII (P < 0.05 or P < 0.01), and increased expression of PP1α (P < 0.05). These results suggest that the abnormal Ca(2+) re-uptake function in heart failure is related with reduced expression and activities of SERCA2a, as well as reduced expression of PLB and its phosphorylation status. Both PLB-Ser(16) and -Thr(17) may be involved in the regulation of myocardium SR calcium pump activity in heart failure.
Animals ; Calcium ; metabolism ; Calcium-Binding Proteins ; metabolism ; Heart Failure ; physiopathology ; Myocardium ; metabolism ; Phosphorylation ; Rabbits ; Sarcoplasmic Reticulum ; pathology

UNLABELLEDobjective: To investigate effects of buyang huanwu decoction (BYHWD) on the rats' myocardial hypertrophic model induced by abdominal aortic constriction, and to clarify the molecular regulatory mechanisms for sarcoplasmic reticulum calcium uptake.

METHODSHypertrophic myocardium rat model was induced by abdominal aorta constriction (AAC). Four weeks after modeling, rats were randomly divided into the sham-operation group (Group S), the AAC model group (Group M), the Enalapril group (Group E), and the BYHWD treatment group (Group BYHWD), respectively. The left ventricular systolic pressure (LVSP), left ventricular end-diastolic pressure (LVEDP), + dp/dtmax,-dp/dtmax, cardiac output (CO), heart mass index (HMI), and left ventricular mass index (LVMI) were observed in each group after 12-week medication. The serum contents of the atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) were detected using ELISA. The SERCA2a activity, the ex- pressions of SERCA2a, phospholamban (PLN), and PLN phosphorylation were observed finally.

RESULTSCompared with Group S, LVSP and LVEDP significantly increased,-dp/dtmax and CO obviously decreased, the myocardial tissue was obviously thickened, the serum contents of ANP and BNP increased, the activity and expression of SERCA2a decreased, the SERCA2a/PLN ratio and PLN phosphorylation degree decreased in Group M (all P <0.05). Compared with Group M, LVEDP obviously decreased, -dp/dtmax and CO obviously increased, the hypertrophy myocardial tissue was obviously lessened, the serum contents of ANP and BNP decreased, the activity of SERCA2a increased, the relative expression contents of SERCA2a, Ser16, and Thrl7 were elevated in Group BYHWD (all P <0.05). BYHWD had significant roles in elevating the SERCA2a/PLN ratio and PLN phosphorylation degree (P <0.05).

CONCLUSIONBYHWD could significantly improve hemodynamics of heart failure rats, elevate CO, lessen cardiac hypertrophy, and improve the capabilities for sarcoplasmic reticulum calcium uptake.

Animals ; Aortic Aneurysm, Abdominal ; metabolism ; pathology ; Calcium ; metabolism ; Constriction, Pathologic ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Hypertrophy, Left Ventricular ; metabolism ; pathology ; Male ; Myocardium ; pathology ; Rats ; Rats, Wistar ; Sarcoplasmic Reticulum ; metabolism

OBJECTIVETo investigate the molecular mechanisms of diaphragm injury in rats with liver cirrhosis.

METHODSThirty adult male Sprague-Dawley rats were randomized into control group (n=10) and carbon tetrachloride-induced liver cirrhosis group (LC group, n=20). In the 9th week, the rat body weight and diaphragm to body weight ratio were measured, and the parameters of diaphragm contractility including peak twitch tension (Pt), maximum tetanic tension (Po), time to peak contraction (CT), half relaxation time (1/2RT), and force-frequency curve were assessed using a Medlab-U/4C biological signal collecting system. The activities of superoxide dismutase (SOD), succinic dehydrogenase (SDH) and myeloperoxidase (MPO) and malondiadehyde (MDA) content in the diaphragm were detected. The mRNA expression levels of sarcoplasmic reticulum calcium ATPase (SERCA) and cytoskeletal proteins (titin and nebulin) in the diaphragm were detected by RT-PCR, and the diaphragm ultrastructure was examined with electron microscopy.

RESULTSCompared with those in the control group, body weight, diaphragm to body weight ratio, Pt, Po, and tetanic force under the stimulus frequency of 10, 20, 40, 60, 100 Hz were all significantly decreased (P<0.01), while CT and 1/2RT were significantly prolonged in LC group (P<0.01). SOD and SDH activities were significantly lowered (P<0.01) while the contents of MDA and MPO activity were significantly increased in LC group (P<0.01) with significantly decreased SERCA, titin and nebulin mRNA expressions in the diaphragm (P<0.01). Electron microscopy of the diaphragm in LC group revealed myofibrillar degeneration, absence of the Z line, and mitochondria swelling and edema.

CONCLUSIONLiver cirrhosis increases free radicals and aggravates inflammatory response and lipid peroxidation in the diaphragm, thus leading to mitochondrial damages and decreased expressions of cytoskeletal proteins and SERCA to cause diaphragmatic dysfunction.

Animals ; Body Weight ; Carbon Tetrachloride ; Connectin ; metabolism ; Diaphragm ; metabolism ; Lipid Peroxidation ; Liver ; enzymology ; pathology ; Liver Cirrhosis ; metabolism ; Male ; Muscle Contraction ; Muscle Proteins ; metabolism ; Oxidation-Reduction ; Rats ; Rats, Sprague-Dawley ; Sarcoplasmic Reticulum Calcium-Transporting ATPases ; metabolism

Muscle unloading due to long-term exposure of weightlessness or simulated weightlessness causes atrophy, loss of functional capacity, impaired locomotor coordination, and decreased resistance to fatigue in the antigravity muscles of the lower limbs. Besides reducing astronauts' mobility in space and on returning to a gravity environment, the molecular mechanisms for the adaptation of skeletal muscle to unloading also play an important medical role in conditions such as disuse and paralysis. The tail-suspended rat model was used to simulate the effects of weightlessness on skeletal muscles and to induce muscle unloading in the rat hindlimb. Our series studies have shown that the maximum of twitch tension and the twitch duration decreased significantly in the atrophic soleus muscles, the maximal tension of high-frequency tetanic contraction was significantly reduced in 2-week unloaded soleus muscles, however, the fatigability of high-frequency tetanic contraction increased after one week of unloading. The maximal isometric tension of intermittent tetanic contraction at optimal stimulating frequency did not alter in 1- and 2-week unloaded soleus, but significantly decreased in 4-week unloaded soleus. The 1-week unloaded soleus, but not extensor digitorum longus (EDL), was more susceptible to fatigue during intermittent tetanic contraction than the synchronous controls. The changes in K+ channel characteristics may increase the fatigability during high-frequency tetanic contraction in atrophic soleus muscles. High fatigability of intermittent tetanic contraction may be involved in enhanced activity of sarcoplasmic reticulum Ca(2+)-ATPase (SERCA) and switching from slow to fast isoform of myosin heavy chain, tropomyosin, troponin I and T subunit in atrophic soleus muscles. Unloaded soleus muscle also showed a decreased protein level of neuronal nitric oxide synthase (nNOS), and the reduction in nNOS-derived NO increased frequency of calcium sparks and elevated intracellular resting Ca2+ concentration ([Ca2+]i) in unloaded soleus muscles. High [Ca2+]i activated calpain-1 which induced a higher degradation of desmin. Desmin degradation may loose connections between adjacent myofibrils and further misaligned Z-disc during repeated tetanic contractions. Passive stretch in unloaded muscle could preserve the stability of sarcoplasmic reticulum Ca2+ release channels by means of keeping nNOS activity, and decrease the enhanced protein level and activity of calpain to control levels in unloaded soleus muscles. Therefore, passive stretch restored normal appearance of Z-disc and resisted in part atrophy of unloaded soleus muscles. The above results indicate that enhanced fatigability of high-frequency tetanic contraction is associated to the alteration in K+ channel characteristics, and elevated SERCA activity and slow to fast transition of myosin heavy chain (MHC) isoforms increases fatigability of intermittent tetanic contraction in atrophic soleus muscle. The sarcomeric damage induced by tetanic contraction can be retarded by stretch in atrophic soleus muscles.
Animals ; Calcium Signaling ; Calpain ; metabolism ; Desmin ; metabolism ; Muscle Contraction ; Muscle Fatigue ; Muscle, Skeletal ; physiopathology ; Muscular Atrophy ; physiopathology ; Myosin Heavy Chains ; metabolism ; Rats ; Sarcoplasmic Reticulum ; pathology ; Sarcoplasmic Reticulum Calcium-Transporting ATPases ; metabolism ; Weightlessness Simulation

The troponin I subunit (TnI) was used as a molecular marker to explore the relationship between the resting intracellular Ca(2+) concentration and myofibril degradation in muscle fibers. The isolated soleus muscle strips of rats were treated by caffeine and H2O2. Caffeine is an opener to increase the calcium release channel open probability of sarcoplasmic reticulum (SR) in contraction phase. H2O2 induces a calcium leak of SR calcium release channel in relaxation phase. The expression and degradation of TnI were detected by Western blot. The resting tension of tetanic contraction and expression of TnI were not changed, but the developed tension was lowered in isolated soleus muscle strips during 40 min of calcium-free Krebs perfusion. Low concentrations of caffeine (1 and 5 mmol/L) perfusion induced a transient increase in resting tension during fatigue period, but did not alter the extent of fatigue, recovery rate after fatigue and expression of TnI in muscle strips. High concentration of caffeine (10 mmol/L) perfusion induced a progressive increase in resting tension, a higher rate of fatigue and a decrease in recovery rate after fatigue in muscle strips. There was a detectable degradation of TnI in soleus after 10 mmol/L caffeine treatment. H2O2 perfusion facilitated a progressive increase in resting tension in a dose-dependent manner, but did not influence the fatigue rate of tetanic contraction. The recovery rate after fatigue showed a quick resumption before decline during H2O2 perfusion. Degradation of TnI occurred in 5 and 10 mmol/L H2O2-treated soleus muscles. Since resting tension is dependent on intracellular Ca(2+) concentration, the above-mentioned results suggest that SR Ca(2+) leakage in relaxation phase may induce a degradation of TnI in skeletal muscle fibers.
Animals ; Caffeine ; pharmacology ; Calcium ; metabolism ; Calcium Channels ; metabolism ; Hydrogen Peroxide ; pharmacology ; In Vitro Techniques ; Muscle Fibers, Skeletal ; metabolism ; Rats ; Sarcoplasmic Reticulum ; pathology ; Troponin I ; metabolism

Sarcoplasmic reticulum is a principal subcellular organelle which regulates calcium homeostasis, protein synthesis, and apoptosis of cardiomyocytes. Endoplasmic reticulum (ER) stress is defined as the perturbation of ER function which is caused by the alterations in the ER environment, such as the perturbation of Ca(2+) homeostasis, elevated protein synthesis, the deprivation of glucose, altered glycosylation, and the accumulation of misfolded proteins. Moderate ER stress is able to restore cellular homeostasis, i.e., to exert a compensatory effect on cardiomyocytes. However, intense or persistent ER stress may cause ER stress-induced apoptosis, which shifts the hypertrophied myocardium to failure, and affects the pathogenesis and development of myocardial hypertrophy. The article reviewed the role of ER stress response in the pathogenesis and development of myocardial hypertrophy.
Animals ; Apoptosis ; Endoplasmic Reticulum Stress ; Homeostasis ; Hypertrophy ; pathology ; Myocardium ; pathology ; Myocytes, Cardiac ; pathology ; Protein Biosynthesis ; Sarcoplasmic Reticulum ; pathology