Analysis of international competitiveness of the China's licorice industry from the perspective of global trade.

ETHNOPHARMACOLOGICAL RELEVANCE: Licorice is a multi-purpose plant raw material, which is widely used in the pharmaceutical industry and food industry, cosmetic industry, etc. It has a wide application in various countries and regions around the world. AIM OF STUDY: This paper studied the trade situation of licorice-related products among major countries and regions in the world, providing a practical reference for the sustainable development of the global licorice industry. MATERIALS AND METHODS: The licorice trade data of licorice-related products came from the United Nations Commodity Trade Database and China Customs data. We analyzed the world's major trading countries by using international market share (IMS), trade competitiveness index (TC), average export price (AEP) and average import price (AIP), and analyzed global trade flows with chord diameter. RESULTS: Uzbekistan, Kazakhstan and Iran mainly export licorice raw materials and low value-added products. China is both a producer and a consumer of licorice raw materials and licorice products. The processing trade of the licorice industry in China has advantages, and the structure of import and export trade has been continuously improved. The United States, France, Germany and other developed countries are still important consumers who rely on the intellectual property rights and brand advantages of licorice products, which have stronger global trade radiation capacity. CONCLUSIONS: China's trade structure has been optimized and its industrial competitiveness has been enhanced. China's experience can be used for reference by other countries, especially those with rich licorice resources among the SCO members.

Fasting improves tolerance to acute hypoxia in rats.

Acute high-altitude illness seriously threatens the health and lives of people who rapidly ascend to high altitudes, but there is currently no particularly effective method for the prevention or treatment of acute high-altitude illness. In the present study, we found that fasting preconditioning effectively improved the survival rate of rats exposed to a simulated altitude of 7620 m for 24 h, and a novel animal model of rapid adaptation to acute hypoxia was established. Compared with control treatment, fasting preconditioning activated AMPK, induced autophagy, decreased ROS levels, and inhibited NF-κB signaling in the cardiac tissues of rats. Our results suggested that fasting effectively improved the acute hypoxia tolerance of rats, which was gradually enhanced with prolongation of fasting. In addition, the acute hypoxia tolerance of young rats was significantly higher than that of adult rats. These experimental results lay the foundation for achieving rapid adaptation to acute hypoxia in humans.

Elevated ROS depress mitochondrial oxygen utilization efficiency in cardiomyocytes during acute hypoxia.

Mitochondria are important sites for the production of ATP and the generation of ROS in cells. However, whether acute hypoxia increases ROS generation in cells or affects ATP production remains unclear, and therefore, monitoring the changes in ATP and ROS in living cells in real time is important. In this study, cardiomyocytes were transfected with RoGFP for ROS detection and MitGO-Ateam2 for ATP detection, whereby ROS and ATP production in cardiomyocytes were respectively monitored in real time. Furthermore, the oxygen consumption rate (OCR) of cardiomyocytes was measured. Similar results were produced for adult and neonatal rat cardiomyocytes. Hypoxia (1% O2) reduced the basal OCR, ATP-linked OCR, and maximal OCR in cardiomyocytes compared with these OCR levels in the cardiomyocytes in the normoxic group (21% O2). However, ATP-linked OCR, normalized to maximal OCR, was increased during hypoxia, indicating that the electron leakage of complex III exacerbated the increase of ATP-linked oxygen consumption during hypoxia and vice versa. Combined with the result that cardiomyocytes expressing MitGO-Ateam2 showed a significant decrease in ATP production during hypoxia compared with that of normoxic group, acute hypoxia might depress the mitochondrial oxygen utilization efficiency of the cardiomyocytes. Moreover, cardiomyocytes expressing Cyto-RoGFP or IMS-RoGFP showed an increase in ROS generation in the cytosol and the mitochondrial intermembrane space (IMS) during hypoxia. All of these results indicate that acute hypoxia generated more ROS in complex III and increased mitochondrial oxygen consumption, leading to less ATP production. In conclusion, acute hypoxia depresses the mitochondrial oxygen utilization efficiency by decreasing ATP production and increasing oxygen consumption as a result of the enhanced ROS generation at mitochondrial complex III.

Mitochondrial electron transport chain, ROS generation and uncoupling (Review).

The mammalian mitochondrial electron transport chain (ETC) includes complexes I­IV, as well as the electron transporters ubiquinone and cytochrome c. There are two electron transport pathways in the ETC: Complex I/III/IV, with NADH as the substrate and complex II/III/IV, with succinic acid as the substrate. The electron flow is coupled with the generation of a proton gradient across the inner membrane and the energy accumulated in the proton gradient is used by complex V (ATP synthase) to produce ATP. The first part of this review briefly introduces the structure and function of complexes I­IV and ATP synthase, including the specific electron transfer process in each complex. Some electrons are directly transferred to O2 to generate reactive oxygen species (ROS) in the ETC. The second part of this review discusses the sites of ROS generation in each ETC complex, including sites IF and IQ in complex I, site IIF in complex II and site IIIQo in complex III, and the physiological and pathological regulation of ROS. As signaling molecules, ROS play an important role in cell proliferation, hypoxia adaptation and cell fate determination, but excessive ROS can cause irreversible cell damage and even cell death. The occurrence and development of a number of diseases are closely related to ROS overproduction. Finally, proton leak and uncoupling proteins (UCPS) are discussed. Proton leak consists of basal proton leak and induced proton leak. Induced proton leak is precisely regulated and induced by UCPs. A total of five UCPs (UCP1­5) have been identified in mammalian cells. UCP1 mainly plays a role in the maintenance of body temperature in a cold environment through non­shivering thermogenesis. The core role of UCP2­5 is to reduce oxidative stress under certain conditions, therefore exerting cytoprotective effects. All diseases involving oxidative stress are associated with UCPs.

Comparison of hypoxic effects induced by chemical and physical hypoxia on cardiomyocytes.

The degree and duration of chemical hypoxia induced by sodium dithionite (Na2S2O4) have not been reported. It is not yet clear how much reduction in the O2 concentration (physical hypoxia) can lead to hypoxia in cultured cardiomyocytes. In this study, oxygen microelectrodes were used to measure changes in the O2 concentration in media containing different concentrations of Na2S2O4. Then, hypoxic effects of 0.8, 1.0, and 2.0 mM Na2S2O4 or 1%, 3%, and 5% O2 in cultured cardiomyocytes from neonatal rats were observed and compared. The results showed that the O2 concentration failed to remain constant by Na2S2O4 treatment during the 180-minute observation period. Only the 2.0 mM Na2S2O4 group significantly increased the expression of hypoxia-inducible factor 1α (HIF-1α) and hypoxic responses. Notably, 3% O2 only significantly increased the expression of HIF-1α in cardiomyocytes, while 1% O2 not only increased the expression of HIF-1α but also increased the apoptotic rate in cardiomyocytes. These results suggest that Na2S2O4 is not suitable for establishing a hypoxic model in cultured neonatal rat cardiomyocytes, and neonatal rat cardiomyocytes cultured at or below 1% O2 induced significant hypoxic effects, which can be used as a starting O2 concentration for establishing a hypoxic cell model.

Identification of differential gene expression profile from peripheral blood cells of military pilots with hypertension by RNA sequencing analysis.

BACKGROUND: Elevated blood pressure is an important risk factor for cardiovascular disease and is also an important factor in global mortality. Military pilots are at high risk of cardiovascular disease because they undergo persistent noise, high mental tension, high altitude hypoxia, high acceleration and high calorie diet. Hypertension is the leading cause of cardiovascular disease in military pilots. In this study, we want to identify key genes from peripheral blood cells of military pilots with hypertension. Identification of these genes may help diagnose and control hypertension and extend flight career for military pilots. METHODS: We use RNA sequencing technology, bioinformatics analysis and Western blotting to identify key genes from peripheral blood cells of military pilots with hypertension. RESULTS: Our study detected 121 up-regulated genes and 623 down-regulated genes in the peripheral blood mononuclear cells (PBMCs) from hypertensive military pilots. We have also identified 8 important genes (NME4, PNPLA7, GGT5, PTGS2, IGF1R, NT5C2, ENTPD1 and PTEN), a number of gene ontology categories and biological pathways that may be associated with military pilot hypertension. CONCLUSIONS: Our study may provide effective means for the prevention, diagnosis and treatment of hypertension for military pilot and extend their flight career.

Simulated microgravity hampers Notch signaling in the fight against myocardial ischemia­reperfusion injury.

The gravitational field is an important determinant of cardiovascular function. Exposure to microgravity during spaceflight may lead to a series of maladaptive alterations in the cardiovascular system. The authors have previously demonstrated that microgravity can increase the susceptibility to myocardial ischemia­reperfusion (IR) injury under simulated microgravity. Although Notch1 signaling protects against myocardial IR injury, whether Notch1 protects against myocardial IR injury under simulated weightlessness remains unknown. The present study is designed to investigate the role of the Notch1 receptor in myocardial IR injury under simulated weightlessness. The differences in Notch signaling expression and myocardial infarct size following myocardial IR were compared between normal rats and tail­suspended rats that were kept in 30˚ head­down tilt and hindlimb unloading position. The data revealed low expression levels of Notch1 receptor and its endogenous ligand Jagged1 in normal adult rat hearts. However, significantly higher expression of Notch1 was observed in the border zone compared with the infarcted area and the remote zone following myocardial IR. Notch1 expression was notably reduced in the infarcted hearts of tail­suspended rats compared with the control group. Conversely, the myocardial infarct size was significantly increased in tail­suspended rats compared with the control rats. In conclusion, these data suggested that the proper function of Notch signaling may be hampered under simulated microgravity.

Glycosylated CD147 reduces myocardial collagen cross-linking in cardiac hypertrophy.

The mechanism of transition from chronic pressure overload-induced cardiac hypertrophy to heart failure is still unclear. Angiotensin II (Ang II) may be an important factor that mediates the transition in the end-stage of cardiac hypertrophy. In the present study, Goldblatt two-kidney one-clip (2K1C) rat model was used to simulate Ang II-induced hypertension. The elevated Ang II not only induced the concentric hypertrophy of left ventricle and cardiac fibrosis, but also increased the expression and glycosylation of CD147 in 2K1C rats. The left ventricular structure and function detected by echocardiogram showed a sign of the transition from cardiac hypertrophy to heart failure in 16 weeks of 2K1C rats. Ang II can activate N-acetylglucosamine transferase V (GnT-V), a key enzyme for CD147 glycosylation. Retinoic acid, an agonist of GnT-V, further increased glycosylated CD147, and activated matrix metalloproteinase-2/-9 (MMP-2 and MMP-9) in the hypertrophied left ventricle of 2K1C rat. Meanwhile, collagen cross-linking in the hypertrophied left ventricle significantly reduced in 2K1C rats. On the contrary, tunicamycin, an inhibitor of N-glycan biosynthesis, inhibited glycosylation of CD147 and activity of MMP-2 and MMP-9, and then maintained a stable of collagen cross-linking in the 2K1C rat hearts. The above results suggested that Ang II increased glycosylated CD147 which activated MMP-2 and MMP-9. Collagens were degraded by the activated MMPs and then reduced collagen cross-linking. Finally, the hypertrophied left ventricle was progressively dilated in chronic pressure overload due to losing the limitation of collagen cross-linking. Therefore, the compensated hypertrophy of left ventricle gradually transited to congestive heart failure.

Simulated microgravity increases myocardial susceptibility to ischemia-reperfusion injury via a deficiency of AMP-activated protein kinase.

Gravitation is an important factor in maintaining cardiac contractility. Our study investigated whether simulated microgravity increases myocardial susceptibility to ischemia-reperfusion (IR) injury. Using the Langendorff-perfused heart model with 300 beats/min pacing, 4-week tail suspension (SUS) and control (CON) male Sprague-Dawley rats (n = 10 rats/group) were subjected to 60 min of left anterior descending coronary artery (LAD) occlusion followed by 120 min of reperfusion. Left ventricular end-systolic pressure (LVESP), left ventricular end-diastolic pressure (LVEDP), creatine kinase (CK) and lactate dehydrogenase (LDH) activity, and infarct size were assessed. Data demonstrated that there were significantly increased LVEDP, CK, LDH, and infarct size in SUS compared with CON (P < 0.05), accompanied by decreased LVESP (P < 0.05). Furthermore, TUNEL-positive cardiomyocytes were higher in SUS than that in CON (P < 0.01), and AMP-activated protein kinase (AMPK) phosphorylation and Bcl-2/Bax in SUS were less compared with CON (P < 0.05). Similarly, isolated hearts pre-treated with A-769662 exhibited better recovery of cardiac function, increased AMPK phosphorylation, and reduced necrosis and apoptosis. Furthermore, AMPKα protein showed a significant suppression in 4-week hindlimb unweighting rats. These results suggest that AMPK deficiency increases myocardial susceptibility to IR injury in rats subjected to simulated microgravity.

Nitric Oxide Protects L-Type Calcium Channel of Cardiomyocyte during Long-Term Isoproterenol Stimulation in Tail-Suspended Rats.

The aim of this study was to investigate the effects of nitric oxide (NO) and reactive oxygen species (ROS) on L-type calcium channel (LTCC) gating properties of cardiomyocytes during long-term isoproterenol (ISO) stimulation. Expression and activity of nNOS as well as S-nitrosylation of LTCC α1C subunit significantly decreased in the myocardium of SUS rats. Long-term ISO stimulation increased ROS in cardiomyocytes of SUS rats. ISO-enhanced calcium current (I Ca,L) in the SUS group was less than that in the CON group. The maximal I Ca,L decreased to about 80% or 60% of initial value at the 50th minute of ISO treatment in CON or SUS group, respectively. Specific inhibitor NAAN of nNOS reduced maximal I Ca,L to 50% of initial value in the CON group; in contrast, NO donor SNAP maintained maximal I Ca,L in SUS group to similar extent of CON group after 50 min of ISO treatment. Long-term ISO stimulation also changed steady-state activation (P < 0.01), inactivation (P < 0.01), and recovery (P < 0.05) characteristics of LTCC in SUS group. In conclusion, NO-induced S-nitrosylation of LTCC α1C subunit may competitively prevent oxidation from ROS at the same sites. Furthermore, LTCC can be protected by NO during long-term ISO stimulation.

[Effects of tamoxifen on CD147 glycosylation and MMPs in the diabetic rat myocardium].

OBJECTIVE: Over the last few decades, diabetic cardiomyopathy has been identified as a significant contributor in cardiac morbidity. However, the mechanisms of diabetic cardiomyopathy have not been clarified. METHODS: In the present study, a diabetic rat model was induced by the intraperitoneal injection of streptozotocin. The myocardial CD147 expression and extent of glycosylation, as well as thematrixmetalloproteinases(MMPs) expression and activity, were observed in the diabetic and synchronous rats. RESULTS: The results showed that CD147 located on sarcolemma of cardiomyocytes. The myocardial CD147 expression and glycosylation were significantly increased in the diabetic rats as compared with the control. Expression of MMP-2 protein, MMP-2 and MMP-9 activity were also increased in left ventricular myocardium in the diabetic rats. Tamoxifen only inhibited the enhanced expression of myocardial CD147 in the diabetic rats, but not in synchronous control rats. Tamoxifen inhibited glycosylation of myocardial CD147 in both diabetic and control rats. The inhibition of tamoxifen on CD147 glycosylation was stronger than on the expression in the myocardium. The extent of myocardial CD147glycosylation was positively related toMMP-2 and MMP-9 activity. Tamoxifen induced an inhibition of myocardial MMP-2 and MMP-9 activity in the control and diabetic rats. CONCLUSION: These results indicate that myocardial CD147 expression, especially the extent of glycosylation, regulates MMP-2 and MMP-9 activity, then accelerates cardiac pathological remodeling inducing diabetic cardiomyopathy. Tamoxifen inhibits myocardial CD147 glycosylation and further depress the activity of MMPs. Therefore, tamoxifen may protect the diabetic rats against diabetic myocardium.

Thymosin alpha 1 suppresses proliferation and induces apoptosis in breast cancer cells through PTEN-mediated inhibition of PI3K/Akt/mTOR signaling pathway.

Thymosin alpha 1 (Tα1), an immunoactive peptide, has been shown to inhibit cell proliferation and induce apoptosis in human leukemia, non-small cell lung cancer, melanoma, and other human cancers. However, the response and molecular mechanism of breast cancer cells exposed to Tα1 remain unclear. PTEN, a tumor suppressor gene, is frequently mutated in a variety of human cancers. In the present study, we aimed to investigate the biological roles of PTEN in the growth inhibition of human breast cancer cells exposed to Tα1. Using wild-type and mutant PTEN-expressing cells, we found a strong correlation between PTEN status and Tα1-mediated growth inhibition of breast cancer cells. The growth inhibition effect was more pronounced in breast cancer cells in which Tα1 enhanced PTEN expression, whereas endogenous PTEN knockdown reversed the growth inhibition effect of Tα1 in breast cancer cells. Further investigation revealed that PTEN up-regulation, which was induced by Tα1, can inhibit the activation of the PI3K/Akt/mTOR signaling pathway, leading to the growth inhibition of breast cancer cells. The addition of the synergy between Tα1 and the inhibition of PI3K/Akt/mTOR activation could strongly block cell viability in PTEN down-regulated breast cancer cells. PTEN-overexpressing cells not only up-regulated Bax and cleaved caspase-3/9 and PARP expression but also down-regulated Bcl-2 compared to the treatment with Tα1 alone. Together these findings suggest that PTEN mediates Tα1-induced apoptosis through the mitochondrial death cascade and inhibition of the PI3K/Akt/mTOR signaling pathway in breast cancer cells.

Nuclear Translocation of Calpain-2 Mediates Apoptosis of Hypertrophied Cardiomyocytes in Transverse Aortic Constriction Rat.

Apoptosis of cardiomyocytes plays an important role in the transition from cardiac hypertrophy to heart failure. Hypertrophied cardiomyocytes show enhanced susceptibility to apoptosis. Therefore, the aim of this study was to determine the susceptibility to apoptosis and its mechanism in hypertrophied cardiomyocytes using a rat model of transverse abdominal aortic constriction (TAC). Sixteen weeks of TAC showed compensatory and pathological hypertrophy in the left ventricle. TUNEL-positive nuclei were significantly increased in TAC with angiotensin II (Ang II) treatment. Calpain inhibitor, PD150606, effectively inhibited Ang II-induced apoptosis of hypertrophied cardiomyocytes. Ang II increased nuclear translocation of intracellular Ca(2+) activated calpain-2 in hypertrophied cardiomyocytes. Ang II enhanced the interaction between activated calpain-2 and Ca(2+)/calmodulin-dependent protein kinase II δB (CaMKIIδB), and promoted the degradation of CaMKIIδB by calpain-2 in the nuclei of hypertrophied cardiomyocytes. Consequently, the depressed CaMKIIδB downregulated the expression of antiapoptotic Bcl-2 leading to mitochondrial depolarization and release of cytochrome c led to apoptosis of hypertrophied cardiomyocytes. In conclusion, hypertrophied cardiomyocytes show increased susceptibility to apoptosis during Ang II stimulation via nuclear calpain-2 and CaMKIIδB pathway.

ROS-Induced Nuclear Translocation of Calpain-2 Facilitates Cardiomyocyte Apoptosis in Tail-Suspended Rats.

Isoproterenol (ISO) induced nuclear translocation of calpain-2 which further increased susceptibility of cardiomyocyte apoptosis in tail-suspended rats. The underlying mechanisms remain elusive. In the present study, the results showed that ISO (10 nM) significantly elevated NADPH oxidases (NOXs) activity and NOXs-derived ROS productions which induced nuclear translocation of calpain-2 in cardiomyocytes of tail-suspended rats. In contrast, the inhibition of NADPH oxidase or cleavage of ROS not only reduced ROS productions, but also resisted nuclear translocation of calpain-2 and decreased ISO-induced apoptosis of cardiomyocyte in tail-suspended rats. ISO also increased the constitutive binding between calpain-2 and Ca(2+)/calmodulin-dependent protein kinase II δB (CaMK II δB) in nuclei, concomitant with the promotion of CaMK II δB degradation and subsequent down-regulation of Bcl-2 mRNA expression and the ratio of Bcl-2 to Bax protein in tail-suspended rat cardiomyocytes. These effects of ISO on cardiomyocytes were abolished by a calpain inhibitor PD150606. Inhibition of calpain significantly reduced ISO-induced loss of the mitochondrial membrane potential, cytochrome c release into the cytoplasm, as well as the activation of caspase-3 and caspase-9 in mitochondrial apoptotic pathway. In summary, the above results suggest that ISO increased NOXs-derived ROS which activated nuclear translocation of calpain-2, subsequently nuclear calpain-2 degraded CaMK II δB which reduced the ratio of Bcl-2 to Bax, and finally the mitochondria apoptosis pathway was triggered in tail-suspended rat cardiomyocytes. Therefore, calpain-2 may represent a potentially therapeutic target for prevention of oxidative stress-associated cardiomyocyte apoptosis.

Impaired translocation of GLUT4 results in insulin resistance of atrophic soleus muscle.

Whether or not the atrophic skeletal muscle induces insulin resistance and its mechanisms are not resolved now. The antigravity soleus muscle showed a progressive atrophy in 1-week, 2-week, and 4-week tail-suspended rats. Hyperinsulinemic-euglycemic clamp showed that the steady-state glucose infusion rate was lower in 4-week tail-suspended rats than that in the control rats. The glucose uptake rates under insulin- or contraction-stimulation were significantly decreased in 4-week unloaded soleus muscle. The key protein expressions of IRS-1, PI3K, and Akt on the insulin-dependent pathway and of AMPK, ERK, and p38 on the insulin-independent pathway were unchanged in unloaded soleus muscle. The unchanged phosphorylation of Akt and p38 suggested that the activity of two signal pathways was not altered in unloaded soleus muscle. The AS160 and GLUT4 expression on the common downstream pathway also was not changed in unloaded soleus muscle. But the GLUT4 translocation to sarcolemma was inhibited during insulin stimulation in unloaded soleus muscle. The above results suggest that hindlimb unloading in tail-suspended rat induces atrophy in antigravity soleus muscle. The impaired GLUT4 translocation to sarcolemma under insulin stimulation may mediate insulin resistance in unloaded soleus muscle and further affect the insulin sensitivity of whole body in tail-suspended rats.

[Dynamic assembly of intercalated disc during postnatal development in the rat myocardium].

The intercalated disc (ICD) complex of cardiomyocyte consists of fascia adherens, desmosomes and gap junctions which are mainly constructed by their transmembrane proteins: N-cadherin (N-cad), desmoglein-2 (DSG2) and connexin 43 (Cx43), respectively. The aim of this study was to observe the dynamic changes in colocalization of N-cad, DSG2 and Cx43 with each other in the rat left ventricular myocardium at 1, 7, 14, 28 and 90 day(s) after birth (P1, P7, P14, P28 and P90) using immunofluorescent staining. The results showed that, N-cad, DSG2 and Cx43 located all around the plasma membrane at the P1. These proteins accumulated to the long ends of cardiomyocytes, indicating preliminary formation of the ICD at the P7. The localization of three proteins at the ICD increased progressively, but their lateral localization showed an inverse trend from the P14 to P90. However, Cx43 still kept a certain amount of lateral localization in cardiomyocytes even at the P90 as compared with N-cad and DSG2. Quantitative colocalization of proteins was analyzed by the stereological method. Total percentage of colocalization of N-cad with DSG2 was 33.5% at the P1, and increased to 38.6% at the P7, 9.4% in ICD and 29.2% in lateral side. The total percentage of colocalization of N-cad with DSG2 increased to 65.7% at the P90, ICD colocalization increasing to 60.5% and lateral colocalization decreasing to 5.2%. Total percentage of colocalization of N-cad with Cx43 increased from 10.3% at the P1 to 37.1% at the P90, and only ICD colocalization increased, but lateral colocalization kept about 5%. The colocalization pattern of DSG2 with Cx43 was similar to that of N-cad with Cx43. Total percentage of colocalization of N-cad with DSG2 was higher than those of N-cad or DSG2 with Cx43. The above results suggest that the formation of mechanical junctions at the ICD of cardiomyocyte is prior to that of electrochemistry junctions during postnatal development. In other words, cardiomyocyte growth needs a stable mechanical environment at first.

Multisite phosphorylation of Bcl-2 via protein kinase Cδ facilitates apoptosis of hypertrophic cardiomyocytes.

Activated protein kinase Cδ (PKCδ) associated with cardiac hypertrophy moves from the cytoplasm to the mitochondria and subsequently triggers the apoptotic signalling pathway. The underlying mechanisms remain unknown. The aim of the present study was to investigate whether mitochondrial translocation of PKCδ phosphorylates multiple sites of Bcl-2, resulting in an imbalance between Bcl-2 and Bak or Bax, thus enhancing the susceptibility of hypertrophic cardiomyocytes to angiotensin II (AngII)-induced apoptosis. Chronic pressure overload was induced by transverse aortic constriction (TAC) in rats. The apoptotic rate increased in hypertrophied cardiomyocytes. In AngII-treated hearts (10 nmol/L, 60 min), there was an increase in the number of TERMINAL deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end-labelling (TUNEL)-positive cells; PKCδ inhibition with 500 nmol/L δV1-1 for 60 min prevented the AngII-induced increase in apoptosis. In the hypertrophied myocardium, PKCδ expression increased, whereas that of Bcl-2 decreased compared with the synchronous control. Treatment of hearts with 10 nmol/L AngII for 60 min activated PKCδ and induced translocation of PKCδ to the mitochondria, where activated PKCδ facilitated the phosphorylation of Bcl-2 at serine-87 and serine-70 sites. The multisite phosphorylated Bcl-2 was released from the mitochondria, and exhibited reduced affinity for Bak and Bax. The imbalance between Bcl-2 and Bak/Bax induced the release of mitochondrial cytochrome c and then activated the caspase 3 apoptotic pathway during AngII stimulation (10 nmol/L, 60 min) of hypertrophied cardiomyocytes. Inhibition of PKCδ reduced these effects of AngII. The results suggest that PKCδ can counteract the anti-apoptotic effect of Bcl-2 and may promote cardiomyocyte apoptosis through multisite phosphorylation of Bcl-2 in hypertrophied cardiomyocytes.

Porcine reproductive and respiratory syndrome virus nonstructural protein 4 antagonizes beta interferon expression by targeting the NF-κB essential modulator.

UNLABELLED: Porcine reproductive and respiratory syndrome virus (PRRSV) is a highly infectious pathogen that causes severe diseases in pigs and great economic losses to the swine industry worldwide. Type I interferons (IFNs) play a crucial role in antiviral immunity. In the present study, we demonstrated that infection with the highly pathogenic PRRSV strain JXwn06 antagonized type I IFN expression induced by poly(I·C) in both porcine alveolar macrophages (PAMs) and blood monocyte-derived macrophages (BMo). Subsequently, we showed that the inhibition of poly(I·C)-induced IFN-ß production by PRRSV was dependent on the blocking of NF-κB signaling pathways. By screening PRRSV nonstructural and structural proteins, we demonstrated that nonstructural protein 4 (nsp4), a viral 3C-like serine protease, significantly suppressed IFN-ß expression. Moreover, we verified that nsp4 inhibited NF-κB activation induced by signaling molecules, including RIG-I, VISA, TRIF, and IKKß. nsp4 was shown to target the NF-κB essential modulator (NEMO) at the E349-S350 site to mediate its cleavage. Importantly, nsp4 mutants with defective protease activity abolished its ability to cleave NEMO and inhibit IFN-ß production. These findings might have implications for our understanding of PRRSV pathogenesis and its mechanisms for evading the host immune response. IMPORTANCE: Porcine reproductive and respiratory syndrome virus (PRRSV) is a major agent of respiratory diseases in pigs. Like many other viruses, PRRSV has evolved a variety of strategies to evade host antiviral innate immunity for survival and propagation. In this study, we show that PRRSV nsp4 is a novel antagonist of the NF-κB signaling pathway, which is responsible for regulating the expression of type I interferons and other crucial cytokines. We then investigated the underlying mechanism used by nsp4 to suppress NF-κB-mediated IFN-ß production. We found that nsp4 interfered with the NF-κB signaling pathway through the cleavage of NEMO (a key regulator of NF-κB signaling) at the E349-S350 site, leading to the downregulation of IFN-ß production induced by poly(I·C). The data presented here may help us to better understand PRRSV pathogenesis.

[In vitro heart models simulating in vivo cardiac ischemia-reperfusion injury].

The aim of this study was to compare in vivo and several in vitro cardiac ischemia-reperfusion (I-R) myocardial injury models, and choose a superior in vitro cardiac I-R model. Sprague-Dawley (SD) rats were randomly grouped into in vivo, Langendorff, Langendorff + pacing, and working heart groups. Left anterior descending (LAD) coronary artery was ligated for 60 min and then reperfused for 120 min in in vivo and in vitro rat hearts. Cardiac function and myocardial infarct size were measured by using pressure transducer and TTC/Evans blue double staining, respectively. The results showed that heart rate was greater in in vivo model than those in the three in vitro models. Coronary flows were dropped after LAD ligation and could recover at early phase of releasing LAD ligation in I-R models of the isolated working heart, Langendorff and Langendorff with 300 beats/min of electrical stimulation. Left ventricular end-systolic pressure (LVESP) decreased during ischemia, and partially restored during reperfusion in the three in vitro models. Left ventricular end-diastolic pressure (LVEDP) increased during ischemia in the three in vitro models. LVEDP was significantly higher in the isolated working heart than those in Langendorff models during ischemia, whereafter decreased slowly during reperfusion. LVEDP elevated further in the initiation of reperfusion period and then decreased, but did not recover to normal levels during reperfusion in Langendorff and Langendorff + pacing groups. Left ventricular myocardial infarct size was (60.4 ± 5.4)% in in vivo I-R model, which was significantly higher than that in Langendorff model and the isolated working heart. Notably, there was no significant difference in myocardial infarct size between in vivo model and Langendorff model with electrical stimulation. These results suggest that Langendorff I-R model with 300 beats/min of electrical stimulation can simulate the in vivo I-R myocardial injury.

[Depressed cardiac output at higher pacing rate in isolated working heart of rat].

OBJECTIVE: To observe the regulation of heart rate to cardiac pump function in the phase of negative force-frequency relationship and their possible mechanisms. METHODS: The left ventricular pressure, aortic pressure, and cardiac output were measured in isolated working heart of rat from 240 to 300 beats/min of pacing rate. RESULTS: Cardiac output of isolated working heart was decreased by a proximally 20% (P < 0.01) with the increase in the pacing rate from 240 to 300 beats/min. Left ventricular end-systolic pressure (LVESP) was declined by 4.8% (P < 0.05), but left ventricular end-diastolic pressure (LVEDP) was elevated by 139% (P < 0.01) with an increase in the pacing rate. Left atrium was enlarged at 300 beats/min of pacing rate. The time from peak to 75% relaxation in left ventricular pressure was shortened with the increased pacing rate. Pressure at aortic valve close was raised (P < 0.01) and ejection duration was shortened with the increased pacing rate (P < 0.01). CONCLUSION: Those above results suggest that there are different mechanisms between the depressed cardiac output at higher heart rate and negative force-frequency relationship. The frequency-dependent acceleration of relaxation facilitates the decline of left ventricular pressure, and then may elevate the pressure of aortic valve close in the condition that the shape of aortic pressure curve stays the same. Therefore, the ejection duration is shortened at higher pacing rate. The shortened ejection duration may induce a decrease in stroke volume of the left ventricle. The increment of heart rate is not enough to compensate the decreased stroke volume. Finally, cardiac output shows a decrease at higher heart rate.