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For connected graphs $G$ and $H$, the $H$-structure connectivity $\kappa (G; H)$ (resp. $H$-substructure connectivity $\kappa ^{s}(G; H)$) of $G$ is the minimum cardinality of a set of subgraphs $\mathcal{F}$ of $G$ such that each is isomorphic to $H$ (resp. to a connected subgraph of $H$) so that $G-\mathcal{F}$ is disconnected or singleton. In this paper, we consider $P_t$-structure connectivity and $P_t$-substructure connectivity of augmented $k$-ary $n$-cubes $AQ_{n,k}$ for $n\geq 2$, $k\geq 3$ and $1\leq t\leq 4n-2$. We obtain that $\kappa (AQ_{n,k}; P_t)=\kappa ^s(AQ_{n,k}; P_t)=\frac{4n-2}{t}+1$ for $t\mid 4n-2$, $t\nmid 2n-1$, $t>6$, $n\geq 3$ and $k\geq 4$; $\kappa (AQ_{n,k}; P_t)=\kappa ^s(AQ_{n,k}; P_t)=\lceil \frac{4n-2}{t}\rceil $, in other cases.

Cardiac microvascular dysfunction is associated with cardiac hypertrophy and can eventually lead to heart failure. Dysregulation of long non-coding RNAs (lncRNAs) has recently been recognized as one of the key mechanisms involved in cardiac hypertrophy. However, the potential roles and underlying mechanisms of lncRNAs in cardiac microvascular dysfunction have not been explicitly delineated. Our results confirmed that cardiac microvascular dysfunction was related to cardiac hypertrophy and ferroptosis of cardiac microvascular endothelial cells (CMECs) occurred during cardiac hypertrophy. Using a combination of in vivo and in vitro studies, we identified a lncRNA AABR07017145.1, named as lncRNA AAB for short, and revealed that lncRNA AAB was upregulated in the hearts of cardiac hypertrophy rats as well as in the Ang II-induced CMECs. Importantly, we found that lncRNA AAB sponged and sequestered miR-30b-5p to induce the imbalance of MMP9/TIMP1, which enhanced the activation of transferrin receptor 1 (TFR-1) and then eventually led to the ferroptosis of CMECs. Moreover, we have developed a delivery system based on neutrophil membrane (NM)-camouflaged mesoporous silica nanocomplex (MSN) for inhibition of cardiac hypertrophy, indicating the potential role of silenced lncRNA AAB (si-AAB) and overexpressed miR-30b-5p as the novel therapy for cardiac hypertrophy., Graphical abstract Neutrophil membrane-camouflaged core-shell structure for siRNA of lncRNA AAB inhibits CMEC ferroptosis in cardiac hypertrophy rat NM + si-AAB + MSN targets injured CMECs. Upon the uptake by CMECs, NM + si-AAB + MSN is decomposed and releases siRNA into cytoplasm. si-AAB increases miR-30b-5p, and then miR-30b-5p regulates the MMP9/TIMP1 balance, which leads to suppression of the activation of TFR-1 to reduce iron toxicity, and ultimately inhibits CMEC ferroptosis., lncRNA AAB sponged and sequestered miR-30b-5p to induce the imbalance of MMP9/TIMP1, which enhanced the activation of TFR-1 and then eventually led to the ferroptosis of CMECs in cardiac hypertrophy. We have developed a nanocomplex that can be targeted to delivery si-AAB or miR-30b-5p to injured CMECs, which inhibits CMEC ferroptosis.

Circular RNAs (circRNAs) play critical regulatory roles in cancer biological processes. Nevertheless, the contributions and underlying mechanisms of circRNAs to pancreatic ductal adenocarcinoma (PDAC) remain largely unexplored. Dysregulated circRNAs between cancerous tissues and matched adjacent normal tissues were identified by circRNA microarray in PDAC. The biological effect of hsa_circ_007367 both in vitro and in vivo was demonstrated by gain- and loss-of-function experiments. Further, dual-luciferase reporter and RNA pull-down assays were performed to confirm the interaction among hsa_circ_007367, miR-6820-3p, and Yes-associated protein 1 (YAP1). The expression of hsa_circ_007367 and YAP1 were detected by in situ hybridization (ISH) and immunohistochemistry (IHC) using tissue microarray (TMA) in 128 PDAC samples. We first identified that a novel circRNA, hsa_circ_0007367, was markedly upregulated in PDAC tissues and cells. Functionally, in vivo and in vitro data indicated that hsa_circ_0007367 promotes the proliferation and metastasis of PDAC. Mechanistically, we confirmed that hsa_circ_0007367 could facilitate the expression of YAP1, a well-known oncogene, by sponging miR-6820-3p, which function as a tumor suppresser in PDAC cells. The results of ISH and IHC demonstrated that hsa_circ_0007367 and YAP1 were upregulated in PDAC tissues. Furthermore, clinical data showed that higher hsa_circ_0007367 expression was correlated with advanced histological grade and lymph node metastasis in PDAC patients. In conclusion, our findings reveal that hsa_circ_0007367 acts as an oncogene via modulating miR-6820-3p/YAP1 axis to promote the progression of PDAC, and suggest that hsa_circ_0007367 may serve as a potential therapeutic target for treatment of PDAC.

The RNA-binding protein QKI belongs to the hnRNP K-homology domain protein family, a well-known regulator of pre-mRNA alternative splicing and is associated with several neurodevelopmental disorders. Qki is found highly expressed in developing and adult hearts. By employing the human embryonic stem cell (hESC) to cardiomyocyte differentiation system and generating QKI-deficient hESCs (hESCs-QKIdel) using CRISPR/Cas9 gene editing technology, we analyze the physiological role of QKI in cardiomyocyte differentiation, maturation, and contractile function. hESCs-QKIdel largely maintain normal pluripotency and normal differentiation potential for the generation of early cardiogenic progenitors, but they fail to transition into functional cardiomyocytes. In this work, by using a series of transcriptomic, cell and biochemical analyses, and the Qki-deficient mouse model, we demonstrate that QKI is indispensable to cardiac sarcomerogenesis and cardiac function through its regulation of alternative splicing in genes involved in Z-disc formation and contractile physiology, suggesting that QKI is associated with the pathogenesis of certain forms of cardiomyopathies., RNA binding protein Quaking (QKI) is known for its broad function in pre-mRNA splicing and modification and its association with several neurodevelopmental disorders. Here the authors reveal that QKI-mediated regulation of RNA splicing is indispensable to cardiac development and contractile physiology.

The objective of this essay is to forge a more explicit link between the “visual turn” and the “practice turn” in entrepreneurship research. Specifically, we explore three key aspects of mobilizing visual methods for studying entrepreneurship-as-practice (EaP), i.e., data sources, collection strategies, and analytical perspectives, highlighting the important theoretical and empirical promises that visual methods hold for said research. This essay bears implications for researchers and educators working at the intersection of entrepreneurship research, the practice theory, and visual methods.

Cardiac hypertrophy, a response of the heart to increased workload, is a major risk factor for heart failure. Myostatin (MSTN) is an inhibitor of myogenesis, regulating the number and size of skeletal myocytes. In recent years, cardiomyocyte autophagy also has been considered to be involved in controlling the hypertrophic response. However, less is known about the detailed mechanism of MSTN on cardiac hypertrophy via regulation of cardiomyocyte autophagy. In this study, we found that the deletion of MSTN potentiated abdominal aorta coarctation (AAC) and angiotensin II (Ang II)-induced pathological cardiac hypertrophy and cardiac autophagy; however, AAC and Ang II-induced cardiac hypertrophic phenotype and cardiac autophagy were dramatically diminished by MSTN in vivo and in vitro. Mechanistically, the anti-hypertrophic and anti-autophagic effects mediated by MSTN in response to pathological stimuli were associated with the direct inactivation of activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) and activation of the peroxisome proliferator-activated receptor gamma (PPARγ)/nuclear factor κB (NF-κB) signaling pathway. Additionally, miR-128 aggravated the progression of cardiac hypertrophy through suppressing its target PPARγ. Furthermore, MSTN downregulated miR-128 expression induced by AAC and Ang II. Taken together, MSTN significantly blunts pathological cardiac hypertrophy and dysfunction, at least in part, by inhibiting excessive cardiac autophagy via blocking AMPK/mTOR and miR-128/PPARγ/NF-κB signaling pathways.

This research investigates the extent to which organizational change initiatives may lead to divergent patterns of sensemaking among organizational members. Drawing on the symbolic convergence theory, we performed an in-depth fantasy theme analysis of organization members’ rhetoric around an organizational change at a private university. Our analysis uncovers six fantasy themes and two corresponding fantasy types, which lead to no rhetorical vision. The lack of cognitive convergence between change initiators and change recipients suggests the inherent incompatibility between managerial and employee fantasies around organizational change, barring the exceptions of dual-responsibility change recipients (e.g., faculty members who also assume administrative responsibilities), who tend to adopt the change initiator rhetoric. Overall, this study informs our extant knowledge of change sensemaking with novel theoretical and methodological insights and bears implications for organizational change researchers and practitioners alike.

The connectivity is an important parameter to evaluate the robustness of a network. As a generalization, structure connectivity and substructure connectivity of graphs were proposed. For connected graphs $G$ and $H$, the $H$-structure connectivity $\kappa(G; H)$ (resp. $H$-substructure connectivity $\kappa^{s}(G; H)$) of $G$ is the minimum cardinality of a set of subgraphs $F$ of $G$ that each is isomorphic to $H$ (resp. to a connected subgraph of $H$) so that $G-F$ is disconnected or the singleton. As popular variants of hypercubes, the $n$-dimensional folded hypercubes $FQ_{n}$ and augmented cubes $AQ_{n}$ are attractive interconnected network prototypes for multiple processor systems. In this paper, we obtain that $\kappa(FQ_{n};K_{1,m})=\kappa^{s}(FQ_{n};K_{1,m})=\lceil\frac{n+1}{2}\rceil$ for $2\leqslant m\leqslant n-1$, $n\geqslant 7$, and $\kappa(AQ_{n};K_{1,m})=\kappa^{s}(AQ_{n};K_{1,m})=\lceil\frac{n-1}{2}\rceil$ for $4\leqslant m\leqslant \frac{3n-15}{4}$.

Cardiac hypertrophy is a common pathological change frequently accompanied by chronic hypertension and myocardial infarction. Nevertheless, the pathophysiological mechanisms of cardiac hypertrophy have never been elucidated. Recent studies indicated that miR‐103 expression was significantly decreased in heart failure patients. However, less is known about the role of miR‐103 in cardiac hypertrophy. The present study was designed to investigate the relationship between miR‐103 and the mechanism of pressure overload‐induced cardiac hypertrophy. TRPV3 protein, cardiac hypertrophy marker proteins (BNP and β‐MHC) and autophagy associated proteins (Beclin‐1 and LC3‐II) were up‐regulated, as well as, miR‐103 expression and autophagy associated proteins (p62) were down‐regulated in cardiac hypertrophy models in vivo and in vitro respectively. Further results indicated that silencing TRPV3 or forcing overexpression of miR‐103 could dramatically inhibit cell surface area, relative fluorescence intensity of Ca2+ signal and the expressions of BNP, β‐MHC, Beclin‐1 and LC3‐II, but promote p62 expression. Moreover, TRPV3 protein was decreased in neonatal rat ventricular myocyte transfected with miR‐103, but increased by AMO‐103. Co‐transfection of the miR‐103 with the luciferase reporter vector into HEK293 cells caused a sharp decrease in luciferase activity compared with transfection of the luciferase vector alone. The miR‐103‐induced depression of luciferase activity was rescued by an AMO‐103. These findings suggested that TRPV3 was a direct target of miR‐103. In conclusion, miR‐103 could attenuate cardiomyocyte hypertrophy partly by reducing cardiac autophagy activity through the targeted inhibition of TRPV3 signalling in the pressure‐overloaded rat hearts.

Cardiac hypertrophy is a compensatory response to mechanical stimuli and neurohormonal factors, ultimately progresses to heart failure. The proteins of some transient receptor potential (TRP) channels, Ca2+‐permeable nonselective cation channel, are highly expressed in cardiomyocytes, and associated with the occurrence of cardiac hypertrophy. Transient receptor potential vanilloid 3 (TRPV3) is a member of TRP, however, the functional role of TRPV3 in cardiac hypertrophy remains unclear. TRPV3 was elevated in pathological cardiac hypertrophy, but not in swimming exercise‐induced physiological cardiac hypertrophy in rats. TRPV3 expression was also increased in Ang II–induced cardiomyocyte hypertrophy in vitro, which was remarkably increased by carvacrol (a nonselective TRPV channel agonist), and reduced by ruthenium red (a nonselective TRPV channel antagonist). Interestingly, we found that activated TRPV3 in Ang II–induced cardiomyocyte hypertrophy was accompanied with increasing intracellular calcium concentration, promoting calcineurin, and phosphorylated CaMKII protein expression, and enhancing NFATc3 nuclear translocation. However, blocking or knockdown of TRPV3 could inhibit the expressions of calcineurin, phosphorylated CaMKII and NFATc3 protein by Western blot. In conclusion, the activation of TRPV3 aggravated pathological cardiac hypertrophy through calcineurin/NFATc3 signalling pathway and correlated with the protein expression levels of calcineurin, phosphorylated CaMKII and NFATc3, revealing that TRPV3 might be a potential therapeutic target for cardiac hypertrophy.

Cardiac fibrosis is a common pathologic change along with pressure overload. Recent studies indicated that transient receptor potential (TRP) channels played multiple roles in heart. However, the functional role of transient receptor potential vanilloid-3 (TRPV3) in cardiac fibrosis remained unclear. The present study was designed to investigate the relationship between TRPV3 activation and pressure overload-induced cardiac fibrosis. Pressure overload rats were successfully established by abdominal aortic constriction (AAC), and cardiac fibrosis was simulated by 100 nM angiotensin II (Ang II) in neonatal cardiac fibroblasts. Echocardiographic parameters, cardiac fibroblast proliferation, cell cycle, intracellular calcium concentration ([Ca2+] i ), and the protein expressions of collagen I, collagen III, transforming growth factor beta 1 (TGF-β1), cyclin E, and cyclin-dependent kinase 2 (CDK2) were measured. Echocardiographic and histological measurements suggested that the activation of TRPV3 exacerbated the cardiac dysfunction and increased interstitial fibrosis in pressure overload rats. Further results showed that TRPV3 activation upregulated the expressions of collagen I, collagen III, TGF-β1, cyclin E, and CDK2 in vivo and in vitro. At the same time, blocking TGF-β1 pathway could partially reverse the effect of TRPV3 activation. These results suggested that TRPV3 activation exacerbated cardiac fibrosis by promoting cardiac fibroblast proliferation through TGF-β1/CDK2/cyclin E pathway in the pressure-overloaded rat hearts.

As a generalization of vertex connectivity, for connected graphs $G$ and $T$, the $T$-structure connectivity $\kappa (G; T)$ (resp. $T$-substructure connectivity $\kappa ^{s}(G; T)$) of $G$ is the minimum cardinality of a set of subgraphs $F$ of $G$ that each is isomorphic to $T$ (resp. to a connected subgraph of $T$) so that $G-F$ is disconnected. For $n$-dimensional hypercube $Q_{n}$, Lin et al. showed $\kappa (Q_{n};K_{1,1})=\kappa ^{s}(Q_{n};K_{1,1})=n-1$ and $\kappa (Q_{n};K_{1,r})=\kappa ^{s}(Q_{n};K_{1,r})=\lceil \frac{n}{2}\rceil $ for $2\leq r\leq 3$ and $n\geq 3$ (Lin, C.-K., Zhang, L.-L., Fan, J.-X. and Wang, D.-J. (2016) Structure connectivity and substructure connectivity of hypercubes. Theor. Comput. Sci., 634, 97–107). Sabir et al. obtained that $\kappa (Q_{n};K_{1,4})=\kappa ^{s}(Q_{n};K_{1,4})= \lceil \frac{n}{2}\rceil $ for $n\geq 6$ and for $n$-dimensional folded hypercube $FQ_{n}$, $\kappa (FQ_{n};K_{1,1})=\kappa ^{s}(FQ_{n};K_{1,1})=n$, $\kappa (FQ_{n};K_{1,r})=\kappa ^{s}(FQ_{n};K_{1,r})= \lceil \frac{n+1}{2}\rceil $ with $2\leq r\leq 3$ and $n\geq 7$ (Sabir, E. and Meng, J.(2018) Structure fault tolerance of hypercubes and folded hypercubes. Theor. Comput. Sci., 711, 44–55). They proposed an open problem of determining $K_{1,r}$-structure connectivity of $Q_n$ and $FQ_n$ for general $r$. In this paper, we obtain that for each integer $r\geq 2$, $\kappa (Q_{n};K_{1,r})$ $=\kappa ^{s}(Q_{n};K_{1,r})$ $=\lceil \frac{n}{2}\rceil $ and $\kappa (FQ_{n};K_{1,r})=\kappa ^{s}(FQ_{n};K_{1,r})= \lceil \frac{n+1}{2}\rceil $ for all integers $n$ larger than $r$ in quare scale. For $4\leq r\leq 6$, we separately confirm the above result holds for $Q_n$ in the remaining cases.

Cardiac microvascular endothelial cells (CMECs), derived from coronary circulation microvessel, are the main barrier for the exchange of energy and nutrients between myocardium and blood. However, microvascular I/R injury is a severely neglected topic, and few strategies can reverse this pathology. In this study, we investigated the mechanism of shear stress in microvascular I/R injury, and try to elucidate the downstream signaling pathways that inhibit CMECs apoptosis to reduce I/R injury. Our results demonstrated that shear stress inhibited the apoptosis protein, increased PECAM-1 expression and eNOS phosphorylation in hypoxia reoxygenated (H/R) CMECs. The mechanism of shear stress was related to up-regulated expression of YAP, the increased number of YAP entering the nucleus by dephosphorylation, the reduced number of TUNEL positive cells, increased miR-206 and inhibited protein level of PDCD4 in CMECs. However, siRNA-mediated knockdown of YAP abolished the protective effects of shear stress on CMECs apoptosis, similar results obtained from administration with AMO-miR-206, and also prevented PDCD4 (target gene of miR-206) increasing when treatment with both AMO-miR-206 and mimics-miR-206. In vivo, restoring the blood fluid with nitroglycerin (NTG) to mimic in vitro shear stress levels, which subsequently improved cardiac function, reduced infarcted area, lowered microvascular perfusion defects. Functional investigations clearly illustrated that increased the protein expression of PECAM-1 and eNOS phosphorylation, activated YAP, strengthened miR-206 expression, and suppressed PDCD4 expression. In summary, this study confirmed that shear stress reversed CMECs apoptosis, relieved microvascular I/R injury, the mechanism of which involving through YAP/miR-206/PDCD4 signaling pathway to finally suppress myocardial I/R injury.

ω-Hydroxy oleic acid is an important intermediate for the synthesis of certain polyesters and polyamides. In this study, a functional CYP153A/putidaredoxin (Pdx)/putidaredoxin reductase (Pdr) hybrid system was engineered for improved ω-hydroxylation activity towards oleic acid. By the combination of site-directed saturation mutagenesis (SDSM) and iterative saturation mutagenesis (ISM), a best mutant (Variant II) was obtained with mutations at two sites (S120 and P165) at the Pdx interaction interface with CYP153A, and one site (S453) in the substrate binding pocket. The in vitro-reconstituted activity of Variant II with purified Pdx and Pdr was 2.7-fold that of the template, while the whole cell transformation activity was 2.0-fold that of the template. A 96-well format-based screening scheme for CYP153A was also developed, which should be useful for engineering of other P450s with low activity. Kinetic analyses indicated that the activity improvement for CYP153A variants largely resulted from enhanced electron transfer. This further demonstrates the importance of the electron transfer between P450s and the non-native redox partners for the overall performance of hybrid P450 systems.

Objective Cardiac microvascular damage is significantly associated with the development of cardiac hypertrophy (CH). Researchers found that allicin could inhibit CH, but the relationship between cardiac microvessel and the inhibition of allicin on CH has not been reported. We aimed to investigate the effect of allicin on the function of cardiac microvascular endothelial cells (CMECs) in CH rat. Materials and Methods The hemodynamic parameters were measured by BL-420F biological function experimental system and the indicators of the ventricular structure and function were measured by echocardiographic system. MTT assay was performed to assess the cell viability. Nitrite detection was performed to detect nitric oxide content. The morphology and molecular characteristics were detected by electron micrographs, immunofluorescence, quantitative real-time polymerase chain reaction (qRT-PCR), western blot. Wound healing experiment, analysis of tube formation and shear adaptation were performed to assess CMECs migration ability, angiogenesis and shear-responsiveness respectively. Result Our findings have identified that microvascular density was decreased by observing the expression of platelet endothelial cell adhesion molecule-1 (PECAM-1) in CH rats. Interestingly, allicin improved the distribution and expression of PECAM-1. Meanwhile, allicin enhanced the migration and angiogenesis ability of CMECs, activated PECAM-1-PI3K-AKT-eNOS signaling pathway, however, the role of allicin was disappear after PECAM-1 was silenced. Allicin decreased the expression of caspase-3 and receptor interacting protein 3 (RIP3), inhibited necroptosis, and increased the levels of Angiopoietin-2 (Ang-2) and platelet-derived growth factor receptor-β (PDGFR-β). Under 10 dyn/cm2 condition, allicin advanced the modification ability of CMECs's shear-adaptation by activating PECAM-1. Conclusion Allicin provided cardioprotection for CH rats by improving the function of CMECs through increasing the expression of PECAM-1.

Competing endogenous RNA (ceRNA) have received wide attention because they are a novel way to regulate genes through sharing microRNAs (miRNAs) that are crucial for complex processes in many diseases. However, no systematic analysis of ceRNA mechanism in cardiovascular disease (CVD) is known. To gain insights into the global properties of ceRNAs in multi-CVDs, we constructed the global view of mRNA-related ceRNA cross-talk in eight major CVDs from ~2,800 samples. We found common features that could be used to uncover similarities among different CVDs and highlighted a common core ceRNA network across CVDs. Comparative analysis of hub ceRNAs in each network revealed three types of hubs, which might play key roles in diverse biological processes. Importantly, by combining CVD-related pathway genes with ceRNA-ceRNA interactions, common modules that might exert functions in specific mechanisms were identified. In addition, our study investigated a potential mechanistic linkage between pathway cross-talk and ceRNA cross-talk. In summary, this study uncovered and systematically characterized global properties of mRNA-related ceRNA cross-talks across CVDs, which may provide a new layer for exploring biological mechanisms and shed new light on cardiology.

Objective Cerebral ischemia is the most common type of neuronal injury and is characterized by a reduction in the function and number of hippocampal neurons. Carvacrol has a significant neuroprotective effect in cerebral ischemia. However, the mechanisms by which carvacrol affects cerebral ischemia, especially with respect to the regulation of neuronal damage by iron levels, have never been systematically studied. This study aimed to reveal the mechanisms by which carvacrol protects against hippocampal neuron impairment after ischemic stroke in gerbils. Materials and methods The Morris water maze test was performed to evaluate learning and memory impairments. Iron ion content and oxidative stress index were detected by the kit. MTT assay was performed to assess the cell viability. The morphology and molecular characteristics were detected by electron micrographs and western blot. Results In the present study, we demonstrated the neuroprotective effects of carvacrol in vivo and in vitro. The Morris water maze test showed that the learning and memory abilities of the gerbils treated with carvacrol were significantly improved. Lipid peroxide injury was evaluated by measuring the levels of lipid peroxide biomarkers; the results indicated that carvacrol decreased the level of lipid peroxide in ischemic gerbil brain tissue. Histopathological examinations and western blotting were performed to evaluate injury in neurons, and carvacrol reduced cell death. Moreover, ferroptosis in the hippocampus was evaluated by measuring the levels of proteins involved in this iron-dependent form of regulated cell death. These results indicated that carvacrol reduced cell death and that carvacrol inhibited ferroptosis by increasing the expression of glutathione peroxidase 4(GPx4). This study showed that carvacrol may be a valuable drug for treating cerebral ischemia. Conclusion Carvacrol provides protection for hippocampal neurons against I/R in gerbils by inhibiting ferroptosis through increasing the expression of GPx4.

Background Cardiac hypertrophy is an adaptive response of the myocardium to pressure or volume overload. Recent evidences indicate that allicin can prevent cardiac hypertrophy. However, it is not clear whether allicin alleviates cardiac hypertrophy by inhibiting autophagy. Purpose We aimed to investigate the effects of allicin on pressure overload-induced cardiac hypertrophy, and further to clarify the related mechanism. Study design/methods Cardiac hypertrophy was successfully established by abdominal aortic constriction (AAC) in rats, and cardiomyocytes hypertrophy was simulated by angiotensin II (Ang II) in vitro. Hemodynamic parameters were monitored by organism function experiment system in vivo. The changes of cell surface area were observed using HE and immunofluorescence staining in vivoand in vitro, respectively. The expressions of cardiac hypertrophy relative protein (BNP and β-MHC), autophagy marker protein (LC3-II and Beclin-1), Akt, PI3K and ERK were detected by western blot. Results Allicin could improve cardiac function, and reduce cardiomyocytes size, and decrease BNP and β-MHC protein expressions. Further results showed that allicin could lower LC3-II and Beclin-1 protein expressions both in vivo and in vitro experiments. And pharmacological inhibitor of mTOR, rapamycin could antagonize the effects of allicin on Ang II-induced cardiac hypertrophy and autophagy. Simultaneously, allicin could promote the expressions of p-Akt, p-PI3K and p-ERK protein. Conclusion These findings reveal a novel mechanism of allicin attenuating cardiac hypertrophy which allicin could inhibit excessive autophagy via activating PI3K/Akt/mTOR and MAPK/ERK/mTOR signaling pathways.

Hybrid P450 systems in which P450 monooxygenases are reconstituted with non-native or surrogate redox partners have become important for the engineering of this class of versatile enzymes. P450sca-2 from Streptomyces carbophilus stereoselectively hydroxylates mevastatin to yield pravastatin, a cholesterol-lowering drug. While S. carbophilus has been successfully applied in the industrial biotransformation process for pravastatin, the molecular study and engineering of P450sca-2 has been very limited. We have previously established a functional P450sca-2/Pdx/Pdr hybrid system. In this study, on the basis of a more active P450sca-2 mutant (R8-5C), five sites located in the substrate binding pocket, substrate access entrance, and presumed Pdx interaction interface were rationally chosen, and systematically subjected to site-directed saturation mutagenesis (SDSM), and three rounds of iterative saturation mutagenesis (ISM). A best mutant (Variant III) was obtained, which showed a whole cell biotransformation activity (377.5 mg/L) and an overall apparent kcat (6.37 min−1) that was 7.1- and 10.0-fold that of the starting template R8-5C, respectively. Kinetic characterization revealed that most of the improvements seen for the SDSM and ISM mutants came from enhanced overall electron transfer, with the two sites at the interface between P450sca-2 and Pdx (T119 and N363) being most critical. Our study underscores the important role of electron transfer in a hybrid P450 system, and also demonstrates the utility of ISM in optimizing the redox partner interface. This should facilitate engineering of this and other important hybrid P450 systems. Biotechnol. Bioeng. 2013;110: 2815–2825. © 2013 Wiley Periodicals, Inc.

Cardiac fibrosis is a common pathologic change along with pressure overload. Recent studies indicated that transient receptor potential (TRP) channels played multiple roles in heart. However, the functional role of transient receptor potential vanilloid-3 (TRPV3) in cardiac fibrosis remained unclear. The present study was designed to investigate the relationship between TRPV3 activation and pressure overload-induced cardiac fibrosis. Pressure overload rats were successfully established by abdominal aortic constriction (AAC), and cardiac fibrosis was simulated by 100 nM angiotensin II (Ang II) in neonatal cardiac fibroblasts. Echocardiographic parameters, cardiac fibroblast proliferation, cell cycle, intracellular calcium concentration ([Ca

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Hybrid P450 systems in which P450 monooxygenases are reconstituted with non-native or surrogate redox partners have become important for the engineering of this class of versatile enzymes. P450sca-2 from Streptomyces carbophilus stereoselectively hydroxylates mevastatin to yield pravastatin, a cholesterol-lowering drug. While S. carbophilus has been successfully applied in the industrial biotransformation process for pravastatin, the molecular study and engineering of P450sca-2 has been very limited. We have previously established a functional P450sca-2/Pdx/Pdr hybrid system. In this study, on the basis of a more active P450sca-2 mutant (R8-5C), five sites located in the substrate binding pocket, substrate access entrance, and presumed Pdx interaction interface were rationally chosen, and systematically subjected to site-directed saturation mutagenesis (SDSM), and three rounds of iterative saturation mutagenesis (ISM). A best mutant (Variant III) was obtained, which showed a whole cell biotransformation activity (377.5 mg/L) and an overall apparent k(cat) (6.37 min⁻¹) that was 7.1- and 10.0-fold that of the starting template R8-5C, respectively. Kinetic characterization revealed that most of the improvements seen for the SDSM and ISM mutants came from enhanced overall electron transfer, with the two sites at the interface between P450sca-2 and Pdx (T119 and N363) being most critical. Our study underscores the important role of electron transfer in a hybrid P450 system, and also demonstrates the utility of ISM in optimizing the redox partner interface. This should facilitate engineering of this and other important hybrid P450 systems.

P450sca-2 is an industrially important enzyme that stereoselectively converts mevastatin into pravastatin. However, little information or engineering efforts have been reported for this enzyme or its redox partner. In this study, we successfully reconstituted the P450sca-2 activity in Escherichia coli by co-expression with putidaredoxin reductase (Pdr) and putidaredoxin (Pdx) from the Pseudomonas putida cytochrome P450cam system. With an HPLC-based screening assay, random mutagenesis was applied to yield a mutant (R8-5C) with a pravastatin yield of the whole-cell biotransformation 4.1-fold that of the wild type. P450sca-2 wild-type and R8-5C were characterized in terms of mevastatin binding and hydroxylation, electron transfer, and circular dichroism spectroscopy. R8-5C showed an active P450 expression level that was 3.8-fold that of the wild type, with relatively smaller changes in the apparent k(cat)/K(M) with respect to the substrate mevastatin (1.3-fold) or Pdx (1.5-fold) compared with the wild type. Thus, the increase in the pravastatin yield of the whole-cell biotransformation primarily came from the improved active P450 expression, which has resulted largely from better heme incorporation, although none of the six mutations of R8-5C are located near the heme active site. These results will facilitate further engineering of this P450sca-2 system and provide useful clues for improving other hybrid P450 systems.