MiR-34a targets MTUS1 to regulate cardiac fibrosis in rats

To observe the effects of moxibustion at "Feishu" (BL13) and "Xinshu" (BL15) on the circular RNA of exon 2-5 of the Pan3 gene (circPAN3), forkhead box O3 (FOXO3), and Bcl-2/adenovirus E1B19kDa-interacting protein 3 (BNIP3) in rats with chronic heart failure (CHF), and explore the potential mechanisms of moxibustion in alleviating myocardial fibrosis. Ten rats of 60 male SPF-grade SD rats were randomly assigned into a normal group. The remaining rats underwent left anterior descending coronary artery (LAD) ligation to establish the CHF model. Forty successfully modeled rats were randomly divided into a model group, a moxibustion group, a rapamycin (RAPA) group, and a moxibustion+RAPA group, with 10 rats in each group. The moxibustion group received mild moxibustion at bilateral "Feishu" (BL13) and "Xinshu" (BL15), 30 min per session. The RAPA group received intraperitoneal injection of the autophagy activator RAPA (1 mg/kg). The moxibustion+RAPA group first received RAPA injection, followed by mild moxibustion at bilateral "Feishu" (BL13) and "Xinshu" (BL15). All interventions were administered once daily for 4 consecutive weeks. After the intervention, cardiac ultrasound was used to measure ejection fraction (EF) and left ventricular fractional shortening (FS). Serum placental growth factor (PLGF) level was determined by ELISA. Myocardial tissue morphology and collagen volume were assessed using hematoxylin-eosin (HE) staining and Masson's trichrome staining. The expression levels of circPAN3, FOXO3, and BNIP3 mRNA in myocardial tissue were detected by real-time PCR, while FOXO3 and BNIP3 protein expression levels were analyzed by Western blot. Compared with the normal group, the model group exhibited myocardial cell disorder, severe fibrosis, and increased collagen volume (P<0.01), along with significantly decreased EF, FS, and circPAN3 mRNA expression in myocardial tissue (P<0.01), and the serum PLGF level, as well as FOXO3 and BNIP3 mRNA and protein expression in myocardial tissue were increased (P<0.01). Compared with the model group, the moxibustion group showed reduced myocardial fibrosis, decreased collagen volume (P<0.01), increased EF, FS, and circPAN3 mRNA expression in myocardial tissue (P<0.01), and decreased serum PLGF level as well as FOXO3 and BNIP3 mRNA and protein expression in myocardial tissue (P<0.01). Compared with the model group, the RAPA group showed further deterioration in these parameters (P<0.01). Compared with the RAPA group, the moxibustion+RAPA group exhibited alleviation of myocardial fibrosis, reduced collagen volume (P<0.01), increased EF, FS, and circPAN3 mRNA expression in myocardial tissue (P<0.01), and decreased serum PLGF level as well as FOXO3 and BNIP3 mRNA and protein expression in myocardial tissue (P<0.01). Moxibustion could alleviate myocardial fibrosis in CHF rats, possibly through upregulation of myocardial circPAN3 expression, downregulation of FOXO3 and BNIP3 expression, and inhibition of excessive myocardial autophagy.

Bamboo leaf extract ameliorates cardiac fibrosis possibly via alleviating inflammation, oxidative stress and apoptosis

This study aims to explore the mechanism of Anshen Dingxin Granules in improving isoproterenol(ISO)-induced ventricular premature beats(VPB) in rats. Sixty rats were randomly divided into six groups: a control group, a model group, a low-dose Anshen Dingxin Granules group, a middle-dose Anshen Dingxin Granules group, a high-dose Anshen Dingxin Granules group, and a propranolol group. The control and model groups were given physiological saline. Two hours after gavage, ISO was injected to induce VPB in all groups except the control group. The injection continued for 7 days. Afterward, the VPB occurrence in each group was monitored by electrocardiogram. Histopathological changes in myocardial tissues were observed by HE and Masson staining. Ultrastructural changes in myocardial tissues were examined by transmission electron microscopy(TEM). The serum superoxide dismutase(SOD), malondialdehyde(MDA), and glutathione(GSH) levels were measured using biochemical methods. The hypoxia-inducible factor-1α(HIF-1α) protein expression in myocardial tissues was analyzed by immunohistochemistry. The HIF-1α, nuclear factor E2-related factor 2(Nrf2), heme oxygenase 1(HO-1), and glutathione peroxidase 4(GPX4) protein expressions in myocardial tissues were detected by Western blot. The Nrf2, HO-1, and GPX4 mRNA expressions in myocardial tissues were measured by PCR. The results show that, compared with the control group, the model group exhibits frequent VPBs and significantly increased arrhythmia scores(P&lt;0.01), significantly increased heart weight index(HWI)(P&lt;0.01), significant pathological damage in myocardial tissues, elevated levels of the oxidative stress marker MDA(P&lt;0.01), and decreased SOD and GSH levels(P&lt;0.01), increased HIF-1α protein expression(P&lt;0.01), decreased Nrf2, HO-1, and GPX4 protein expressions(P&lt;0.01), as well as decreased Nrf2, HO-1, and GPX4 mRNA expressions(P&lt;0.01). Compared with the model group, the propranolol group and the high-dose Anshen Dingxin Granules group exhibit delayed onset and reduced frequency of VPBs(P&lt;0.01). Besides, in the two groups of the above comparison, a significant decrease occurs as follows: arrhythmia scores(P&lt;0.05), HWI(P&lt;0.05 or P&lt;0.01), myocardial tissue pathological damage, oxidative stress indicator MDA level(P&lt;0.05 or P&lt;0.01), HIF-1α protein expression(P&lt;0.01) while significant increase is found in the items below: SOD and GSH levels(P&lt;0.01), Nrf2, HO-1, and GPX4 protein expressions(P&lt;0.05 or P&lt;0.01), Nrf2, HO-1, and GPX4 mRNA expressions(P&lt;0.05 or P&lt;0.01). In conclusion, Anshen Dingxin Granules can improve ISO-induced VPB in rats. This effect may stem from HIF-1 signaling pathway regulation, which inhibits oxidative stress and in turn reduces myocardial cell damage.

ABSTRACTCardiac fibrosis, especially in the infarct border zone, leads to decreased cardiac compliance, impaired systolic and diastolic function, resulting in heart failure. M6A methylation plays a role in fibrosis development. However, its underlying mechanism remains poorly understood. This study explores the role and molecular mechanisms of m6A methylation in regulating cardiac fibrosis after myocardial infarction (MI). A mouse myocardial fibrosis model post‐MI was established by ligating the left coronary artery. Corresponding gene knockdown was achieved in vitro or in vivo using short hairpin RNA or fibroblast‐specific AAV9 virus. Echocardiography assessed cardiac function in mice, while Masson staining determined the degree of collagen deposition post‐MI. The meRIP‐Seq kit detected mRNA methylation levels in myocardial tissue and hypoxia‐treated cardiac fibroblasts. Expression of RNA methylation‐related enzymes, fibrosis‐related proteins, and SMOC2 expression in the myocardial tissue or cardiac fibroblasts were detected using western blotting. Actinomycin D assessed SMOC2 mRNA stability. Results demonstrated increased levels of m6A methylation and METTL3 expression in myocardial fibrosis tissue post‐MI and in hypoxia‐treated cardiac fibroblasts. In vivo METTL3 downregulation reduced the fibrotic area and improved cardiac function, while METTL3 downregulation in vitro can alleviate cardiac fibroblast proliferation and differentiation after hypoxia. Mechanistically, METTL3 promoted SMOC2 mRNA stability by increasing its m6A methylation level, thereby regulating cardiac fibroblast proliferation and differentiation. Together, our work uncovers a critical link between METTL3 and SMOC2, providing insight into the functional importance of the mRNA m6A methylation and its modulators in cardiac fibrosis post MI.

This study aimed to investigate the mechanism and target sites of Shenfu Injection in the intervention of chronic heart fai-lure based on the PI3K/Akt/mTOR autophagy signaling pathway. The chronic heart failure model was induced in rats by subcutaneous injection of isoproterenol. The model rats were randomly divided into model group, Shenfu Injection group, and 3-methyladenine autophagy inhibitor(3-MA) group. A normal group was also set up. After 15 days of administration, cardiac function indexes of the rats were detected by echocardiography. The serum N-terminal pro-B-type natriuretic peptide(NT-proBNP) levels were measured using the ELISA. HE and Masson staining was performed to observe the morphological changes in myocardial tissues, and electron microscopy was used to observe the autophagosomes in myocardial tissues. Western blot was conducted to measure the changes in autophagy-related proteins(LC3 Ⅱ/Ⅰ and p62), PI3K, Akt, mTOR, and phosphorylation levels. The results showed that compared with normal group, model group in rats led to reduced cardiac function, significant activation of cardiac autophagy, increased fibrotic lesions in myocardial tissues, structural disorder of the myocardium, increased autophagosomes, and cytoplasmic vacuolization. Compared with model group, Shenfu Injection group in rats led to cardiac function significantly improved, myocardial fibrosis decreased, and the number of autophagosomes and cytoplasmic vacuolization decreased. The phosphorylation levels of PI3K, Akt, and mTOR were significantly increased(P&lt;0.01). In the 3-MA group, autophagy was inhibited through the activation of the PI3K/Akt/mTOR signaling pathway, resulting in improved cardiac function, reduced myocardial fibrosis, and no significant cytoplasmic vacuolization. The findings suggest that Shenfu Injection can activate the PI3K/Akt/mTOR signaling pathway and inhibit autophagy, thereby improving cardiac function.

To investigate the effects of Luhong Yixin Granules on myocardial fibrosis in rats with heart failure and its possible mechanism, a total of 60 male Wistar rats were randomly divided into the control group, model group, and low-, medium-and high-dose Luhong Yixin Granules groups, with 12 rats in each group. Except for those in the control group, rats in the other groups were induced by intraperitoneal injection of doxorubicin(DOX) into a rat model. After the Luhong Yixin Granules were dissolved in the same amount of normal saline, they were given by gavage at low, medium and high doses(2.8, 5.6, 11.2 g·kg~(-1)·d~(-1)), and the control group and the model group were given the same amount of normal saline by gavage for 40 days. After the end of dosing, echocardiography was used to measure left ventricular ejection fraction(LVEF) and left ventricular fractional shortening(LVFS). Rat body weight(BW) and heart weight(HW) were calculated as HW/BW. Enzyme-linked immunosorbent assay was used to measure the levels of interleukin-6(IL-6), interleukin-17(IL-17), tumor necrosis factor-α(TNF-α), transforming growth factor-β1(TGF-β1), growth stimulation expressed gene 2 protein(ST2), N-terminal pro-B-type natriuretic peptide(NT-proBNP), galectin-3(Gal-3) and creatine kinase isoenzyme(CK-MB) in serum. Hematoxylin-eosin(HE) staining and Masson staining were used to observe the pathological morphology of myocardial tissue. Western blot and quantitative real-time polymerase chain reaction were used to detect the protein and mRNA expression levels of IL-6, IL-17, TNF-α, TGF-β1, Smad3, Smad7, α-smooth muscle actin(α-SMA), and collagen Ⅰ(COL-Ⅰ), respectively. RESULTS:: showed that compared with those in the control group, LVEF, LVFS, and HW/BW in the model group were decreased(P&lt;0.05), and the levels of IL-6, IL-17, TNF-α, TGF-β1, ST2, NT-proBNP, Gal-3, and CK-MB were increased(P&lt;0.05). HE staining showed inflammatory changes in myocardial tissue; Masson staining showed decreases in the cross-sectional area and ventricular cavity area of the heart, and myocardial fibrosis of varying degrees(P&lt;0.05). The protein and mRNA expression of IL-6, IL-17, TNF-α, TGF-β1, Smad3, α-SMA, and COL-Ⅰ were increased(P&lt;0.05), and the protein and mRNA expression of Smad7 protein was decreased(P&lt;0.01). Compared with those in the model group, LVEF, LVFS and HW/BW of the low-, medium-and high-dose Luhong Yixin Granules groups were increased(P&lt;0.05), and the levels of IL-6, IL-17, TNF-α, TGF-β1, ST2, NT-proBNP, Gal-3 and CK-MB were decreased(P&lt;0.05). HE staining showed gradually reduced inflammatory changes of myocardial tissue, and Masson staining showed increased cross-sectional area and ventricular cavity area of the heart and decreased area of myocardial fibrosis(P&lt;0.05). The protein and mRNA expression levels of IL-6, IL-17, TNF-α, TGF-β1, Smad3, α-SMA, and COL-Ⅰ were decreased(P&lt;0.05), while the protein and mRNA expression levels of Smad7 were increased(P&lt;0.05). Luhong Yixin Granules may be of great value in the treatment of heart failure by regulating the TGF-β1/Smads signaling pathway, inhibiting the expression of inflammation-related proteins, reducing the deposition of extracellular matrix, and alleviating myocardial fibrosis.

Chronic kidney disease (CKD) is a major contributor to the development of heart failure with preserved ejection fraction (HFpEF), whereas the underlying mechanism of cardiorenal HFpEF is still elusive. The aim of this study was to investigate the role of cardiac fibrosis in a rat model of cardiorenal HFpEF and explore whether treatment with Telmisartan, an inhibitor of renin-angiotensin-aldosterone system (RAAS), can ameliorate cardiac fibrosis and preserve diastolic function in cardiorenal HFpEF. Male rats were subjected to 5/6 subtotal nephrectomy (SNX) or sham operation (Sham), and rats were allowed four weeks to recover and form a stable condition of CKD. Telmisartan or vehicle was then administered p.o. (8 mg/kg/d) for 12 weeks. Blood pressure, brain natriuretic peptide (BNP), echocardiography, and cardiac magnetic resonance imaging were acquired to evaluate cardiac structural and functional alterations. Histopathological staining, real-time polymerase chain reaction (PCR) and western blot were performed to evaluate cardiac remodeling. SNX rats showed an HFpEF phenotype with increased BNP, decreased early to late diastolic transmitral flow velocity (E/A) ratio, increased left ventricular (LV) hypertrophy and preserved ejection fraction (EF). Pathology revealed increased cardiac fibrosis in cardiorenal HFpEF rats compared with the Sham group, while chronic treatment with Telmisartan significantly decreased cardiac fibrosis, accompanied by reduced markers of fibrosis (collagen I and collagen III) and profibrotic cytokines (α-smooth muscle actin, transforming growth factor-β1, and connective tissue growth factor). In addition, myocardial inflammation was decreased after Telmisartan treatment, which was in a linear correlation with cardiac fibrosis. Telmisartan also reversed LV hypertrophy and E/A ratio, indicating that Telmisartan can improve LV remodeling and diastolic function in cardiorenal HFpEF. In conclusion, cardiac fibrosis is central to the pathology of cardiorenal HFpEF, and RAAS modulation with Telmisartan is capable of alleviating cardiac fibrosis and preserving diastolic dysfunction in this rat model.

This study observed the effect of Chaihu Jia Longgu Muli Decoction on myocardial fibrosis in the rat model of myocardial infarction(MI) and explored the effect of this decoction on ventricular remodeling induced by myocardial fibrosis after MI based on transforming growth factor-β1(TGF-β1)/Smad proteins(Smads) pathway, aiming to reveal the therapeutic mechanism of this decoction. MI model rats were established by coronary artery ligation and assigned into sham, model, and low-dose, medium-dose, and high-dose(2.09, 4.19, and 8.37 g·kg~(-1), respectively) Chaihu Jia Longgu Muli Decoction groups, with 10 rats in each group. Chaihu Jia Longgu Muli Decoction groups were administrated with the decoction at different doses by gavage, and the sham and model groups received an equal volume of distilled water once daily for 4 weeks. The cardiac function of rats after MI was assessed via echocardiography. Myocardial fiber deposition and collagen volume fraction(CVF) were observed by Sirius red staining and Masson staining. Ultrastructure changes of myocardial cells were observed by transmission electron microscopy. Creatine kinase isoenzymes(CK-MB), cardiac troponin T(cTnT), and soluble growth stimulation expressed gene 2(sST2) levels were measured by ELISA. Western blot was employed to assess the protein levels of collagen Ⅰ(Col Ⅰ) and α-smooth muscle actin(α-SMA) associated with myocardial fibrosis, and TGF-β1, Smad3, and Smad7 associated with the TGF-β1/Smads pathway. Compared with the sham group, the model group had significantly decreased body weight, compromised cardiac function, abnormal ventricular structure, collagen fiber deposition, increased fibrosis area and CVF, seriously damaged ultrastructure of myocardium, a part of mitochondria being swollen or even ruptured, disarrangement and lysis of myofibril, and broken myotome and myofilament with emergence of space. Furthermore, the model group showed risen levels of cTnT, sST2 and CK-MB, obvious myocardial fibrosis, up-regulated protein levels of pro-fibrotic cytokines TGF-β1 and Smad3, down-regulated protein level of the inhibitory cytokine Smad7, and increased expression of myocardial fibrosis-related proteins Col Ⅰ and α-SMA. Compared with the model group, Chaihu Jia Longgu Muli Decoction groups showed increased body weight, improved cardiac function and ventricular structure, reduced collagen fiber deposition, fibrosis area, and CVF, intact ultrastructure of myocardium, normal mitochondrial structure, and well-arranged myofibril and myofilament. In addition, Chaihu Jia Longgu Muli Decoction groups presented declined cTnT, sST2, and CK-MB levels, down-regulated protein levels of TGF-β1 and Smad3, up-regulated protein level of Smad7, and reduced expression of Col Ⅰ and α-SMA. Chaihu Jia Longgu Muli Decoction may reduce excessive deposition of extracellular matrix by regulating the expression of proteins in the TGF-β1/Smads pathway, thereby alleviating myocardial fibrosis and ameliorating ventricular remodeling.

Objective To observe the intervention effect of vitamin C (Vit C) and adenosine triphosphate (ATP) on myocardial fibrosis in rats. Methods Forty male SD rats were selected, body weight were 125-140 g, and they were divided into 8 groups according to body weight using a random number table method. Four rats for control group, 3 rats for model group, 6 rats for Vit C early group, 6 rats for ATP early group, 6 rats for Vit C + ATP early group, 5 rats for Vit C late group, 5 rats for ATP late group, and 5 rats for Vit C + ATP late group. Rats in model group and these intervention groups were induced with doxorubicin (2 mg/kg each week) for 6 weeks, and control group was given the same amount of normal saline. All early groups were intragastrically administered with Vit C (200 mg·kg-1·d-1), ATP (45 mg·kg-1·d-1) and Vit C + ATP (200 mg·kg-1·d-1 + 45 mg·kg-1·d-1) in the fourth week; these late groups were intragastrically administered with the same dose in the sixth week; each group was continuously administered for 21 days. Three days after the last intervention, cardiac ultrasonography was performed in all surviving rats, and left ventricular end diastolic diameter(LVEDD), left ventricular end systolic diameter (LVESD), left ventricular ejection fraction (LVEF), and left ventricular fractional shortening (LVFS) were recorded. The rats were sacrificed and the hearts were taken. HE staining and Masson staining were used to observe the pathological changes of myocardial tissue and the collagen volume fraction (CVF) was calculate. Serum cardiac troponin I (CTn-I) and type I procollagen amino terminal peptide (PINP) levels were determined by enzyme-linked immunosorbent assay (ELISA). Results Compared with control group [(3.65 ± 0.25) mm, (80.63 ± 3.03)%, (43.57 ± 2.54)%], LVESD [(5.07 ± 0.58), (4.06 ± 0.68), (4.71 ± 0.43), (4.87 ± 0.44), (4.79 ± 0.59), (5.07 ± 0.62), (4.97 ± 0.29) mm] of model group and each intervention groups were increased, LVEF [(62.17 ± 4.92)%, (71.28 ± 3.54)%, (65.03 ± 3.35)%, (59.81 ± 2.45)%, (60.42 ± 9.22)%, (60.15 ± 3.06)%, (60.65 ± 2.05)%], and LVFS [(30.05 ± 2.95)%, (36.44 ± 2.90)%, (31.63 ± 2.15)%, (26.95 ± 1.05)%, (28.35 ± 6.84)%, (27.79 ± 2.41)%, (28.38 ± 1.42)%] were decreased (P 0.05). Conclusions Vit C can reduce myocardial fibrosis and improve cardiac function in the early stage. The effect of ATP alone to improve fibrosis is not obvious. Key words: Vitamin C; ATP; Myocardial fibrosis

To observe the effect of electroacupuncture(EA)pretreatment on transient receptor potential vanilloid 1(TRPV1)/calcitonin gene-related peptide(CGRP)signal and nuclear factor-κB p65 (NF-κB p65) protein expression in myocardial tissue of acute myocardial ischemic injury (AMI) rats, and to investigate the possible mechanism of electroacupuncture pretreatment against AMI. A total of 60 adult male SD rats were randomly divided into blank control, sham operation, model and EA pretreatment groups, 15 rats in each group. The acute myocardial ischemia model was established by ligating the left anterior descending (LAD)branch of the coronary artery in the model group and EA pretreatment group, while threading but no ligating at left anterior descending branch of the coronary artery was applied in the sham operation group. In the EA pretreatment group, bilateral "Neiguan"(PC6) acupoints were selected, with intensity of 2 mA and frequency of 2 Hz/100 Hz, for 20 min, once daily for 7 days before modeling. Electrocardiogram (ECG) was recorded by physiological signal acquisition system, and the ST segment potential offset values of standard Ⅱ lead were analyzed before surgery，30 min and 24 h after operation. The TTC staining was used to observe the percentage of myocardial infarction area. The HE staining was used to observe the pathological changes of myocardial tissue and the degree of inflammatory cell infiltration. And Western blot was used to detect TRPV1/CGRP signal and NF-κB p65 protein expression levels in myocardial tissue. Compared with the sham operation group, the ECG-J point potential in the model group was significantly increased at 30 min and decreased at 24 h after operation (P<0.05), myocardial infarction area increased significantly (P<0.05), the myocardial fibers were obviously disordered, inflammatory cell infiltration was obvious, and the expressions of TRPV1，CGRP and NF-κB p65 proteins were all increased (P<0.05). Compared with the model group, the EA pretreatment group was decreased in the ECG-J point potential at 30 min after operation(P<0.05), significantly reduced in myocardial infarction area (P<0.05), improved in the morphology of myocardial fibers, reduced ininflammatory cell infiltration, and increased in the protein expressions of TRPV1 and CGRP in myocardium (P<0.05), significantly decreased in the protein expression of NF-κB p65 (P<0.05). EA pretreatment may enhance TRPV1/CGRP signaling, down-regulate NF-κB p65 protein expression, reduce myocardial inflammatory response status, improve AMI injury, and reduce myocardial infarction area.

Quercetin exhibits numerous pharmacological effects, including the capacity for cardioprotection. This study aimed to investigate whether quercetin or its glycoside derivative rutin has any protective action against isoproterenol (ISO) induced cardiac fibrosis, and investigate the structure-activity relationship. Male Wistar rats were injected subcutaneously with ISO (15 mg·(kg body mass)(-1)) to induce experimental cardiac fibrosis. The cardioprotective effect of co-treatment with quercetin (25 or 50 mg·kg(-1)) or rutin (25 or 50 mg·kg(-1)) was investigated in ISO-induced cardiac fibrosis in rats. The administration of quercetin and rutin signifcantly decreased the cardiac weight index and myocardial enzyme activity, increased the activity of superoxide dismutase in the serum, and inhibited the ISO-induced increase in angiotensin II and aldosterone in the plasma. Furthermore, overexpression of transforming growth factor β1 (TGF-β1), connective tissue growth factor (CTGF), and excessive deposition of extracellular matrix (ECM) in isoproterenol-treated myocardial tissues were normalized by quercetin and rutin. Our results suggest that both quercetin and rutin exhibited cardioprotective effects in cardiac fibrosis induced by ISO in the rat heart. Moreover, the effects of rutin are weaker than quercetin at the same dose. The mechanism of these effects may be related to antioxidative stress, inhibition of the renin-angiotensin-aldosterone system, decrease in the expression of TGF-β1 and CTGF, and the subsequent reduction in the deposition of the ECM.

Objective To study the expression of transforming growth factor-β1 (TGF-β1) in myocardial tissue and the concentration of serum B-type natriuretic peptide (BNP) in myocardial remodeling of SD rats induced by isoproterenol (ISO) and the effects of carvedilol intervention. Methods According to random number table, 30 male SD rats were divided into 3 groups: (1)after 9 g/L saline was injected with 5 mL/(kg·d) for 10 days, 9 g/L saline was used at the dose of 10 mL/(kg·d) by way of gavage for 4 weeks in the control group (n=10); (2) after ISO was injected with 5 mg/(kg·d) for 10 days, 9 g/L saline was used at the dose of 10 mL/(kg·d) by way of gavage for 4 weeks in the model group (n=10); (3)after ISO was injected with 5 mg/(kg·d) for 10 days, carvedilol was used at the dose of 10 mg/(kg·d) by way of gavage for 4 weeks in the treatment group (n=10). Then, all the rats were killed and the cardiac weight index (CWI) was measured.The pathological changes of myocardial tissue were observed through HE staining and Masson staining.The mRNA expression of TGF-β1 was detected by adopting reverse transcription-polymerase chain reaction.The protein of TGF-β1 was detected by means of immunohistochemistry and Western blot.The concentration of serum BNP was tested by adopting enzyme-linked immuno sorbent assay. Results (1)Pathological changes by light microscope: no significant pathological changes were observed in myocardial tissue of the control group.The denaturization, hypertrophy, edema, necrosis of myocardial cells and collagen fibers increased in myocardial tissue of the model group were more serious than those of the treatment group.(2) CWI: CWI of the model group[(3.31±0.07) mg/g] was significantly higher than that of the treatment group[(3.03±0.04) mg/g], and CWI of the treatment group was significantly higher than that of the control group[(2.98±0.10) mg/g]. There was significant difference among 3 groups (F=54.383, P=0.000). There was also significant difference between any 2 groups (all P<0.01). (3) The level of TGF-β1 of myocardial tissue: the level of TGF-β1 of the model group was significantly higher than that of the treatment group, and the level of TGF-β1 of the treatment group was significantly higher than that of the control group by using three detection methods.There was significant difference among 3 groups (F=13.62, P=0.000). There was also significant difference between any 2 groups (all P<0.01). (4) The concentration of serum BNP: the concentration of serum BNP of the model group[(61.13±2.00) ng/L] was significantly higher than that of the treatment group[(57.08±1.52) ng/L], and the concentration of serum BNP of the treatment group was significantly higher than that of the control group[(51.56±1.80) ng/L]. There was significant difference among 3 groups (F=72.81, P=0.000). There also was significant difference between any 2 groups (all P<0.01). Conclusions The expression of TGF-β1 in myocardial tissue was up-regulated and the concentration of serum BNP was increased in myocardial remodeling of SD rats induced by ISO.The carvedilol intervention can down-regulate the expression of TGF-β1 and decrease the concentration of BNP, then inhibit myocardial remodeling, and improve cardiac function. Key words: Transforming growth factor-β1; B-type natriuretic peptide; Myocardial remodeling; Carvedilol

To investigate the effect of isoflurane on myocardial ischemia-reperfusion injury through the p38 mitogen-activated protein kinase (MAPK) signaling pathway. A total of 36 specific-pathogen-free (SPF) Sprague-Dawley rats were randomly divided into sham group (n=12), model group (n=12) and isoflurane group (n=12). In model group and isoflurane group, the myocardial ischemia-reperfusion injury model was established via the ligation of left anterior descending coronary artery (ischemia for 30 min and reperfusion for 3 h). In sham group, the left anterior descending coronary artery was not ligated, but the chest was opened and threaded using the same method. After ischemia, the rats in isoflurane group were inhaled with isoflurane. The cardiac function of rats in each group was detected before ischemia (T0) and once every 2 h after reperfusion (T1-T4) for a total of 5 times, and the cardiac function indexes included ejection fraction (EF), fractional shortening (FS), left ventricular systolic pressure (LVSP) and left ventricular end-diastolic pressure (LVEDP). After the rats were executed, the myocardial infarction tissues were taken for hematoxylin-eosin (HE) staining and 2,3,5-triphenyltetrazolium chloride (TTC) staining to observe the morphological changes in tissues and the degrees of myocardial ischemia and infarction. The malondialdehyde (MDA) content and superoxide dismutase (SOD) activity in myocardial cells in the infarction site in each group were detected using the MDA and SOD kits. Moreover, the expression levels of related proteins in the p38 MAPK signaling pathway in myocardial cells in the infarction site were detected via Western blotting. In model group, the cardiac function was significantly damaged (p<0.01), there was significant pathological damage in the myocardium, the area of myocardial infarction was significantly increased (p<0.01), the MDA content was significantly increased (p<0.01), the SOD activity declined obviously (p<0.01), and the expression levels of p-p38 and p-tau protein were significantly increased (p<0.01) compared with those in control group. After intervention with isoflurane, the cardiac function of rats was significantly improved (p<0.01), the pathological damage in myocardial tissues was alleviated, the area of myocardial infarction was reduced (p<0.01), the MDA content declined (p<0.01), the SOD activity was increased (p<0.01), and the expression levels of p-p38 and p-tau protein were decreased (p<0.01). Isoflurane can, through inhibiting the p38 MAPK signaling pathway, effectively protect the cardiac function of rats from myocardial ischemia-reperfusion injury, reduce the area of myocardial infarction, alleviate the pathological damage in myocardial cells and reduce the oxidative stress response.

This study investigated the mechanisms and targets of Shenfu Injection in the intervention in chronic heart failure(CHF) through the NOD-like receptor thermal protein domain associated protein 3(NLRP3)/caspase-1 signaling pathway. A CHF model was induced in rats by subcutaneous injection of isoproterenol. Model rats were randomly divided into a model group, a Shenfu Injection group, and a MCC950(NLRP3 inhibitor) group, and a blank group was also set up as a control. After 15 days of treatment, echocardiography was performed to measure cardiac function parameters [left ventricular ejection fraction(LVEF) and left ventricular fractional shortening(LVFS)]. Enzyme-linked immunosorbent assay(ELISA) was used to measure serum levels of N-terminal pro-brain natriuretic peptide(NT-proBNP), interleukin(IL)-1β, and IL-18. Hematoxylin-eosin(HE) and Masson staining were used to observe morphological changes in myocardial tissues, and Western blot was used to measure the expression levels of NLRP3/caspase-1 pathway-related proteins [NLRP3, caspase-1, apoptosis-associated speck-like protein containing a CARD(ASC), gasdermin D(GSDMD), IL-1β, and IL-18]. The study found that isoproterenol-induced CHF in rats resulted in decreased cardiac function, worsened myocardial fibrosis, increased expression levels of NLRP3, ASC, caspase-1, GSDMD-N, IL-1β, and IL-18 in myocardial tissues, elevated serum inflammatory factors, and induced myocardial cell pyroptosis. Following Shenfu Injection intervention, the Shenfu Injection group showed significantly improved LVEF and LVFS, a significant decrease in NT-proBNP, a marked downregulation of NLRP3, ASC, caspase-1, GSDMD-N, IL-1β, and IL-18 protein expression levels, reduced serum inflammatory factors IL-1β and IL-18 expression in CHF rats, and a decrease in the rate of TUNEL-positive cells. Shenfu Injection can significantly improve cardiac function in CHF, inhibit myocardial fibrosis, and alleviate the progression of myocardial cell pyroptosis through the inhibition of the NLRP3/caspase-1 pathway.

This study aims to investigate the activation of the PPARγ/NF-κB pathway and its influence on macrophage M2 polarization induced by acupuncture at the Neiguan acupoint. Additionally, it seeks to explore the potential mechanisms by which electroacupuncture treatment may facilitate the reduction of inflammation in rats subjected to myocardial ischemia-reperfusion injury (MIRI). SD rats were randomly assigned to four groups: sham operation, model, electroacupuncture, and inhibitor, with 12 rats in each group. The left anterior descending branch of the coronary artery was ligated to establish the MIRI rat model. Electroacupuncture intervention at the bilateral Neiguan acupoints commenced 24 h post-chest closure, lasting for 30 min once daily over three consecutive days. Myocardial infarction was assessed through electrocardiography, while cardiac function was evaluated via echocardiography 24 h after modeling. The morphology and structure of myocardial tissues were examined using HE staining. Myocardial tissue levels of interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), interleukin-10 (IL-10) and transforming growth factor-β (TGF-β) were measured by ELISA in each group of rats. The expression of the M1 macrophage marker inducible nitric oxide synthase (iNOS) and the M2 macrophage marker arginase-1 (Arg-1) in myocardial tissues was analyzed using immunohistochemistry (IHC). Detection of macrophage polarization status in myocardial tissue by flow cytometry. Additionally, peroxisome proliferator-activated receptor gamma (PPARγ) and nuclear transcription factor-κB (NF-κB) expression in myocardial tissues was assessed using Western blotting (WB). Compared with the sham-operated group, rats in the model group exhibited a significant decrease in ejection fraction (EF) (P < 0.01), along with notable myocardial fiber damage characterized by inflammatory cell infiltration. Additionally, there was an elevated expression of IL-6, TGF-β, iNOS, CD-86 and phosphorylated p65 (p-p65) in myocardial tissue (P < 0.01, P < 0.01, P < 0.05, P < 0.05 and P < 0.01, respectively). In contrast, rats in the electroacupuncture group demonstrated an increase in EF (P < 0.01) compared to the model group. Myocardial fiber damage was significantly ameliorated, inflammatory cell infiltration was reduced. Furthermore, the expression of IL-10, TGF-β, Arg-1, CD-163 and PPARγ in cardiac muscle tissue was increased (P < 0.01, P < 0.01, P < 0.05, P < 0.05 and P < 0.01, respectively). Conversely, when compared to the electroacupuncture group, the EF of rats in the inhibitor group was significantly reduced (P < 0.05), with pronounced myocardial fiber damage and accompanying inflammatory cell infiltration. Additionally, IL-6, TGF-β, iNOS CD-86 and p-p65 expression in myocardial tissue was increased (P < 0.01, P < 0.01, P < 0.05, P < 0.05 and P < 0.01, respectively). Electroacupuncture may ameliorate myocardial MIRI by activating the PPARγ/NF-κB pathway, promoting polarization of macrophages towards the M2 type, and reducing inflammatory damage in myocardial tissues.