3-methoxy aroylhydrazones – free radicals scavenging,anticancer and cytoprotective potency

Objective: This study aimed to determine the capability of newly designed 3-methoxy derivatives of salicylaldehyde benzoylhydrazone to influence the oxidative stress processes and to test their in vitro cytotoxicity.

Methods: We have used chemiluminescent and spectrophotometric model systems containing different types of reactive oxygen species (OH^●, OCl^─ and O₂^─●). The hydrazones effect on the viability of Hep-G2, HEK-293 and SH-SY5Y cell lines was determined via MTT assay.

Results: The comparative analysis of the C₅₀ values of the chemiluminescent investigation demonstrated moderate activity against the hydroxyl radicals (C₅₀?>?50?μmol/L) and remarkable reactivity in the systems containing a superoxide radical and a hypochlorous anion (C₅₀?● generation and consequent 2′-deoxyribose oxidative damage excluded the possibility of quenching effect and proved the direct interaction of the studied compounds with that generated in the system reactive oxygen species (ROS). The encapsulation of the studied derivatives into chitosan-alginate particles led to the protection and stabilization of their antioxidant activity as revealed by a one-month study using the ABTS ^●+ method. The cytotoxic study revealed less pronounced effects against the non-malignant cell line (HEK-293) compared to Hep-G2 and SH-SY-5Y cells.

Discussion: The incorporation of a hydroxyl group in the hydrazide part of a parent molecule which relates to better antioxidant effect in most of the studied systems is associated with higher IC₅₀ values in all cytotoxicity experiments and relates to the cytoprotective effect against N-methyl-D-aspartate-induced excitotoxicity in SH-SY5Y human neuronal cells.     

Binding properties of Ru(II) polypyridyl complexes with poly(U)·poly(A)*poly(U) triplex: the ancillary ligand effect on third-strand stabilization

The binding properties of [RuL₂(mip)]²⁺ {where L is 1,10-phenanthroline (phen) or 4,7-dimethyl-1,10-phenanthrollne (4,7-dmp) and mip is 2′-(3″,4″-methylenedioxyphenyl)imidazo[4′,5′-f][1,10]phenanthroline} with regard to the triplex RNA poly(U)·poly(A)*poly(U) were investigated using various biophysical techniques and quantum chemistry calculations. In comparison with [Ru(4,7-dmp)₂(mip)]²⁺, remarkably higher binding affinity of [Ru(phen)₂(mip)]²⁺ for the triplex RNA poly(U)·poly(A)*poly(U) was achieved by changing the ancillary ligands. The stabilization of the Hoogsteen-base-paired third strand was improved by about 10.9 °C by [Ru(phen)₂(mip)]²⁺ against 6.6 °C by [Ru(4,7-dmp)₂(mip)]²⁺. To the best of our knowledge, [Ru(phen)₂(mip)]²⁺ is the first metal complex able to raise the third-strand stabilization of poly(U)·poly(A)*poly(U) from 37.5 to 48.4 °C. The results reveal that the ancillary ligands have an important effect on third-strand stabilization of the triplex RNA poly(U)·poly(A)*poly(U) when metal complexes contain the same intercalative ligands.     

More pronounced salt dependence and higher reactivity for platination of the hairpin r(CGCGUUGUUCGCG) compared with d(CGCGTTGTTCGCG)

The DNA interference pathways exhibited by cisplatin and related anticancer active metal complexes have been extensively studied. Much less is known to what extent RNA interaction pathways may operate in parallel, and perhaps contribute to both antineoplastic activity and toxicity. The present study was designed with the aim of comparing the reactivity of two model systems comprising RNA and DNA hairpins, r(CGCGUUGUUCGCG) and d(CGCGTTGTTCGCG), towards a series of platinum(II) complexes. Three platinum complexes were used as metallation reagents; cis-[PtCl(NH₃)₂(OH₂)]⁺ (1), cis-[PtCl(NH₃)(c-C₆H₁₁NH₂)(OH₂)]⁺ (2), and trans-[PtCl(NH₃)(quinoline)(OH₂)]⁺ (3). The reaction kinetics were studied at pH 6.0, 25 °C, and 1.0 mM ≤ I ≤ 500 mM. For both types of nucleic acid targets, compound 3 was found to react about 1 order of magnitude more rapidly than compounds 1 and 2. Further, all platinum compounds exhibited a more pronounced salt dependence for the interaction with r(CGCGUUGUUCGCG). Chemical and enzymatic cleavage studies revealed similar interaction patterns with r(CGCGUUGUUCGCG) after long exposure times to 1 and 2. A substantial decrease of cleavage intensity was found at residues G4 and G7, indicative of bifunctional adduct formation. Circular dichroism studies showed that platinum adduct formation leads to a structural change of the ribonucleic acid. Thermal denaturation studies revealed platination to cause a decrease of the RNA melting temperatures by 5–10 °C. Our observations therefore suggest that RNA is a kinetically competitive target to DNA. Furthermore, platination causes destabilization of RNA structural elements, which may lead to deleterious intracellular effects on biologically relevant RNA targets.Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Evidence for ditopic coordination of phosphate diesters to [Mg(15-crown-5)]<Superscript>2+</Superscript>. Implications for magnesium biocoordination chemistry

The interaction of a series of phosphate diesters and triesters (1=diphenyl phosphate, 2=dimethyl phosphate, 3=bis(2-ethylhexyl) phosphate, 4=trimethyl phosphate, 5=methyldiphenyl phosphate, 6=triphenyl phosphate) with [Mg(15-crown-5)]²⁺ (15-crown-5=1,4,7,10,13-pentaoxocyclopentadecane) was studied as a simplified model for the interaction of aqueous Mg²⁺ ion with phosphate-containing biomolecules such as RNA. Using electrospray mass spectrometry, we confirm the formation of 1:1 adducts in the gas phase. Proton and ³¹P NMR titration data were used to construct binding isotherms, and a 1:1 binding equilibrium was fit to the isotherms at room temperature to estimate the binding affinities. The binding affinity data are consistent with ditopic coordination of neutral dialkyl phosphate ligands to the [Mg(15-crown-5)]²⁺ unit. This involves inner-sphere coordination to the Mg²⁺ via an oxygen atom, which is complemented by a weak hydrogen-bonding interaction with the crown ether ligand. Ditopic interaction is consistent with low-temperature NMR spectra showing four different configurations for 1 coordinated to [Mg(15-crown-5)]²⁺, which are interpreted in terms of hindered rotation around the Mg–O_phos bond. Thermochemical analysis of the binding affinity data suggests that the second-shell interaction contributes only about 1 kcal/mol to the binding free energy, so additional factors, such as steric constraints, must be operative to give a preferred phosphate orientation in this system. However, the experimental data do suggest that second-shell interactions contribute as much as 40% of the total binding energy, consistent with the pronounced ability of aqueous Mg²⁺ to form salt-bridges linking secondary and tertiary elements of RNA structure.Abbreviations OTf trifluoromethanesulfonate - ESI-MS electrospray mass spectrometry     

利用转录组测序筛选低切应力作用下人脐静脉内皮细胞的差异表达基因

目的:通过转录组测序分析获得不同切应力作用下人脐静脉内皮细胞的基因的表达谱,为进一步探索切应力影响内皮细胞形态和功能的机制提供依据。方法:以人脐静脉内皮细胞为材料,通过Streamer系统建立6通道可调控切应力的流体动力学细胞模型,以层流切应力(15 dynes/cm~2)为对照,以低切应力(0.1 dynes/cm~2)为实验组,分别加载细胞18 h。提取总RNA逆转录合成cDNA,建立文库,以二代测序平台Illumina HiSeq中进行扩增和测序。结果:序列比对结果显示,有19986个基因比对上,新转录本分析显示各组新转录本数约占总转录本数的50%。基因表达差异分析显示,较对照组,低切应力组表达上调基因983个,表达下调基因701个。GO分析显示,有18499个基因得到了归类注释,绝大多数基因富集到生物学过程。KEGG分析显示,富集Top20的信号通路与细胞周期、DNA复制和细胞分裂、细胞应激和凋亡等生物学过程相关。结论:低切应力作用不仅仅激活内皮细胞中细胞的增殖相关基因,同时也涉及到DNA损伤修复和凋亡相关基因。     

smc5基因敲除斑马鱼肝脏转录组学分析

摘要 目的：探讨smc5基因敲除对斑马鱼肝脏基因表达谱的影响,进一步明确smc5突变对斑马鱼代谢的影响。方法：用CRISPR/Cas9技术构建smc5基因敲除斑马鱼模型,取3个月的smc5^-/-和野生型斑马鱼肝脏进行转录组测序,创建基因表达谱文库,观察smc5基因敲除后斑马鱼肝脏基因表达谱的变化,将筛选出的差异表达基因进行功能富集,并运用荧光定量PCR对KEGG通路中显著的差异表达基因进行验证。结果：成功构建出7号外显子上2碱基缺失造成移码突变的smc5基因敲除斑马鱼模型。RNA-seq发现smc5^-/-斑马鱼的肝脏基因表达谱变化显著,包含p53的多个通路激活,如细胞周期和凋亡。糖酵解、脂肪酸降解与代谢、丙酮酸代谢等相关通路显著下调。荧光定量PCR结果与RNA-seq结果一致。结论：smc5基因敲除下调斑马鱼肝脏糖脂代谢。本研究结果为进一步研究SMC5基因在糖脂代谢调控中的潜在机制奠定基础。     

The Osmolyte TMAO Stabilizes Native RNA Tertiary Structures in the Absence of Mg: Evidence for a Large Barrier to Folding from Phosphate Dehydration

The stabilization of RNA tertiary structures by ions is well known, but the neutral osmolyte trimethylamine oxide (TMAO) can also effectively stabilize RNA tertiary structure. To begin to understand the physical basis for the effects of TMAO on RNA, we have quantitated the TMAO-induced stabilization of five RNAs with known structures. So-called m values, the increment in unfolding free energy per molal of osmolyte at constant KCl activity, are ∼ 0 for a hairpin secondary structure and between 0.70 and 1.85 kcal mol^− 1m^− 1 for four RNA tertiary structures (30-86 nt). Further analysis of two RNAs by small-angle X-ray scattering and hydroxyl radical probing shows that TMAO reduces the radius of gyration of the unfolded ensemble to the same endpoint as seen in titration with Mg²⁺ and that the structures stabilized by TMAO and Mg²⁺ are indistinguishable. Remarkably, TMAO induces the native conformation of a Mg²⁺ ion chelation site formed in part by a buried phosphate, even though Mg²⁺ is absent. TMAO interacts weakly, if at all, with KCl, ruling out the possibility that TMAO stabilizes RNA indirectly by increasing salt activity. TMAO is, however, strongly excluded from the vicinity of dimethylphosphate (unfavorable interaction free energy, + 211 cal mol^− 1m^− 1 for the potassium salt), an ion that mimics the RNA backbone phosphate. We suggest that formation of RNA tertiary structure is accompanied by substantial phosphate dehydration (loss of 66-173 water molecules in the RNA structures studied) and that TMAO works principally by reducing the energetic penalty associated with this dehydration. The strong parallels we find between the effects of TMAO and Mg²⁺ suggest that RNA sequence is more important than specific ion interactions in specifying the native structure.     

Cross flow filtration of RNA extracts by hollow fiber membrane

The feasibility of using ultrafiltration to concentrate RNA was investigated. Ultrafiltration flux and solute retentivity were affected by membrane cut-off and inlet pressure. The RNA can be concentrated by selecting a hydraulically permeable membrane (PM 30) and then operating the system at a sufficiently high pressure (2.07 · 10⁵ Nm⁻²) and a high recirculation rate (150 cm⁻¹). The use of this procedure was limited to a high-flux regime. However, a very high retentivity of macrosolute was achieved. The effect of transmembrane pressure on the apparent mass transfer coefficient in the “gel polarization model” was discussed and related to ultrafiltration flux.     

Mg2+ Impacts the Twister Ribozyme through Push-Pull Stabilization of Nonsequential Phosphate Pairs

RNA molecules perform a variety of biological functions for which the correct three-dimensional structure is essential, including as ribozymes where they catalyze chemical reactions. Metal ions, especially Mg²⁺, neutralize these negatively charged nucleic acids and specifically stabilize RNA tertiary structures as well as impact the folding landscape of RNAs as they assume their tertiary structures. Specific binding sites of Mg²⁺ in folded conformations of RNA have been studied extensively; however, the full range of interactions of the ion with compact intermediates and unfolded states of RNA is challenging to investigate, and the atomic details of the mechanism by which the ion facilitates tertiary structure formation is not fully known. Here, umbrella sampling combined with oscillating chemical potential Grand Canonical Monte Carlo/molecular dynamics simulations are used to capture the energetics and atomic-level details of Mg²⁺-RNA interactions that occur along an unfolding pathway of the Twister ribozyme. The free energy profiles reveal stabilization of partially unfolded states by Mg²⁺, as observed in unfolding experiments, with this stabilization being due to increased sampling of simultaneous interactions of Mg²⁺ with two or more nonsequential phosphate groups. Notably, these results indicate a push-pull mechanism in which the Mg²⁺-RNA interactions actually lead to destabilization of specific nonsequential phosphate-phosphate interactions (i.e., pushed apart), whereas other interactions are stabilized (i.e., pulled together), a balance that stabilizes unfolded states and facilitates the folding of Twister, including the formation of hydrogen bonds associated with the tertiary structure. This study establishes a better understanding of how Mg²⁺-ion interactions contribute to RNA structural properties and stability.     

Zn2+ Promoted Hydrolysis of RNA

Abstract

The Zn²⁺ promoted hydrolysis of phosphodiester bonds of RNA has been studied by using a series of model compounds from dinucleoside monophosphates to oligo- and polynucleotides. The results will be discussed with respect to complex formation between the metal ion catalysts and substrates.     

Sequence specific hydrogen bond of DNA with denaturants affects its stability: Spectroscopic and simulation studies

BackgroundSeveral different small molecules have been used to target the DNA helix in order to treat the diseases caused by its mutation. Guanidinium(Gdm⁺) and urea based drugs have been used for the diseases related to central nervous system, also as the anti-inflammatory and chemotherapeutic agent. However, the role of Gdm⁺ and urea in the stabilization/destabilization of DNA is not well understood.MethodsSpectroscopic techniques along with molecular dynamics (MD) simulation have been performed on different sequences of DNA in the presence of guanidinium chloride (GdmCl) and urea to decode the binding of denaturants with DNA and the role of hydrogen bond with the different regions of DNA in its stability/destability.Results and conclusionOur study reveals that, Gdm⁺ of GdmCl and urea both intrudes into the groove region of DNA along with the interaction with its phosphate backbone. However, interaction of Gdm⁺ and urea with the nucleobases in the groove region is different. Gdm⁺ forms the intra-strand hydrogen bond with the central region of the both sequences of DNA whereas inter-strand hydrogen bond along with water assisted hydrogen bond takes place in the case of urea. The intra-strand hydrogen bond formation capability of Gdm⁺ with the nucleobases in the minor groove of DNA decreases its groove width which probably causes the stabilization of B-DNA in GdmCl. In contrast, the propensity of the formation of inter-strand hydrogen bond of urea with the nucleobases in the groove region of DNA without affecting the groove width destabilizes B-DNA as compared to GdmCl. This study depicts that the opposite effect of GdmCl and urea on the stability is a general property of B-DNA. However, the extent of stabilization/destabilization of DNA in Gdm⁺ and urea depend on its sequence probably due to the difference in the intra/inter-strand hydrogen bonding with different bases present in both the sequences of DNA.General significanceThe information obtained from this study will be useful for the designing of Gdm⁺ based drug molecule which can target the DNA more specifically and selectively.     

Perturbation of Hydrogen Bonds in the Adenine…Thymine Base Pair by Na+, Mg2+, Ca2+ and NH4 + Cations

Abstract

Perturbation of the hydrogen bonds in the adenine…thymine base pair by Na⁺, Mg²⁺, Ca²⁺ and NH₄ ⁺ cations has been investigated by means of ab initio SCF calculations with the STO-3G basis set. The geometry of adenine…thymine, as well as those of the perturbed pairs were optimized. Approach of any cation to thymine at 06 leads to destabilization of the adenine…thy mine pair; divalent cations (Mg²⁺, Ca²⁺) have a profound effect on the structure of the base pair. The approach of a cation to other available sites (thymine: O2, adenine N1 and N3) leads, on the other hand, to stabilization of the base pair. If a water molecule is placed between the cation and the base pair, the structure and stability of the base pair are changed only negligibly.     

N-[2-Naphthyl]-glycine hydrazide,a potent inhibitor of DNA-dependent RNA polymerase ofMycobacterium tuberculosis H37RV

N-[2-Naphthyl]-glycine hydrazide has been shown for the first time as a potent inhibitor of the DNA-dependent RNA polymerase (EC 2.7.7.6) ofMycobacterium tuberculosis H₃₇R_v. At a concentration of 10^-9 M, the compound shows maximum inhibition of the enzyme, the inhibition being less at higher concentrations. It is suggested that the novel type of inhibition pattern may be due to hydrophobic interactions occurring between the molecules of the compound at higher concentrations. The finding that there is a shift in the λ_max of the compound could also account for this phenomenon. The effect of this compound was also tested on DNA-dependent RNA polymerases from an eukaryotic fungus,Microsporum canis. At a concentration of 10⁻⁹ M it inhibits RNA polymerase II (32%) but not RNA polymerasesI andIII     

A Remarkable Stabilization of Complexes Formed by 2,6-Diaminopurine Oligonucleotide N3′→P5′ Phosphoramidates

Abstract

2′-Deoxyribo- and ribo-oligonucleotide N3′→P5′phosphoramidates containing 2,6-diaminopurine nucleosides were synthesized. Thermal denaturation experiments demonstrated a significant stabilization of the complexes formed by these compounds with DNA and RNA complementary strands, relative to adenosine-containing phosphoramidate counterparts. The increase in melting temperature of the complexes reached up to 6.9 °C per substitution. The observed stabilization was attributed to the apparent synergistic effects of N-type sugar puckering of the oligonucleotide N3′→5′ phosphoramidate backbone, and the ability of 2,6-diaminopurine bases to form three hydrogen bonds.     

Specificity of Phage Display Selected Peptides for Modified Anticodon Stem and Loop Domains of tRNA

Protein recognition of RNA has been studied using Peptide Phage Display Libraries, but in the absence of RNA modifications. Peptides from two libraries, selected for binding the modified anticodon stem and loop (ASL) of human tRNA^Lys3 having 2-thiouridine (s²U₃₄) and pseudouridine (Ψ₃₉), bound the modified human ASL^Lys3(s²U₃₄;Ψ₃₉) preferentially and had significant homology with RNA binding proteins. Selected peptides were narrowed to a manageable number using a less sensitive, but inexpensive assay before conducting intensive characterization. The affinity and specificity of the best binding peptide (with an N-terminal fluorescein) were characterized by fluorescence spectrophotometry. The peptide exhibited the highest binding affinity for ASL^Lys3(s²U₃₄;Ψ₃₉), followed by the hypermodified ASL^Lys3 (mcm⁵s²U₃₄;ms²t⁶A₃₇) and the unmodified ASL^Lys3, but bound poorly to singly modified ASL^Lys3 constructs (Ψ₃₉, ms²t⁶A_37, s²U₃₄), ASL^Lys1,2 (t⁶A₃₇) and Escherichia coli ASL^Glu (s²U₃₄). Thus, RNA modifications are potentially important recognition elements for proteins and can be targets for selective recognition by peptides.     

Zn2+ -Promoted Hydrolysis of 3′,5′-Dinucleoside Monophosphates and Polyribonucleotides. The Effect of Nearest Neighbours on the Cleavage of Phosphodiester Bonds

Abstract

Pseudo first-order rate constants for the Zn²⁺ -promoted cleavage of 15 different dinucleoside monophosphates, 4 different ribo homopogmers and RNA III from baker's yeast have been determined. Furthermore, the distribution of various nucleosides at the 3′-and 5′-terminus of the oligomeric hdrolysls products of RNA has been quantified. On these bases, the effect of nearest neighbours on the metal-ion-promoted hydrolysis of the internucleosidic phosphodiester bonds of RNA is discussed.     

Synthesis,characterization and nucleic acid binding studies of mononuclear copper(II) complexes derived from azo containing O,O donor ligands

Abstract

Azo linked salicyldehyde and a new 2-hydroxy acetophenone based ligands (HL¹ and HL²) with their copper(II) complexes [Cu(L¹)₂] (1) and [Cu(L²)₂] (2) were synthesized and characterized by spectroscopic methods such as ¹H, 13C NMR, UV–Vis spectroscopy and elemental analyses. Calculation based on Density Functional Theory (DFT), have been performed to obtain optimized structures. Binding studies of these copper (II) complexes with calf thymus DNA (ct-DNA) and torula yeast RNA (t-RNA) were analyzed by absorption spectra, emission spectra and Viscosity studies and Molecular Docking techniques. The absorption spectral study indicated that the copper(II) complexes of 1 and 2 had intrinsic binding constants with DNA or RNA in the range of 7.6?±?0.2?×?10³?M^?1 or 6.5?±?0.3?×?10³M^?1 and 5.7?±?0.4?×?10⁴ M^?1 or 1.8?±?0.5?×?10³ M^?1 respectively. The synthesized compounds and nucleic acids were simulated by molecular docking to explore more details mode of interaction of the complexes and their orientations in the active site of the receptor.     

NMR Studies of a Lead Ribozyme and Its Non-Cleavable Analogue

Abstract

The structure of a lead ribozyme, which consists of two RNA strands, at neutral pH has been studied by NMR. Nearly all resonances of imino protons, base protons (H2, H5, H6 and H8) and sugar protons (H1′ and H2′) were assigned sequentially. Interesting structural features which deviate from the standard structure were found for the residues at an active site which consists of an internal loop. No indication of stable G:A base pairs was found in the loop. The effect of addition of Pb²⁺ was studied by the use of a non-cleavable analogue in which the cytidine at a cleavage site is replaced by 2′-O-methylcytidine. It was suggested that Pb²⁺ binds close to the cleavage site and that the structural change induced by Pb²⁺ is moderate and localized.

     

Assembly of Functional Ribonucleoprotein Complexes by AU-rich Element RNA-binding Protein 1 (AUF1) Requires Base-dependent and -independent RNA Contacts

AU-rich element RNA-binding protein 1 (AUF1) regulates the stability and/or translational efficiency of diverse mRNA targets, including many encoding products controlling the cell cycle, apoptosis, and inflammation by associating with AU-rich elements residing in their 3′-untranslated regions. Previous biochemical studies showed that optimal AUF1 binding requires 33–34 nucleotides with a strong preference for U-rich RNA despite observations that few AUF1-associated cellular mRNAs contain such extended U-rich domains. Using the smallest AUF1 isoform (p37^AUF1) as a model, we employed fluorescence anisotropy-based approaches to define thermodynamic parameters describing AUF1 ribonucleoprotein (RNP) complex formation across a panel of RNA substrates. These data demonstrated that 15 nucleotides of AU-rich sequence were sufficient to nucleate high affinity p37^AUF1 RNP complexes within a larger RNA context. In particular, p37^AUF1 binding to short AU-rich RNA targets was significantly stabilized by interactions with a 3′-purine residue and largely base-independent but non-ionic contacts 5′ of the AU-rich site. RNP stabilization by the upstream RNA domain was associated with an enhanced negative change in heat capacity consistent with conformational changes in protein and/or RNA components, and fluorescence resonance energy transfer-based assays demonstrated that these contacts were required for p37^AUF1 to remodel local RNA structure. Finally, reporter mRNAs containing minimal high affinity p37^AUF1 target sequences associated with AUF1 and were destabilized in a p37^AUF1-dependent manner in cells. These findings provide a mechanistic explanation for the diverse population of AUF1 target mRNAs but also suggest how AUF1 binding could regulate protein and/or microRNA binding events at adjacent sites.     

Xin-Ji-Er-Kang protects myocardial and renal injury in hypertensive heart failure in mice

BackgroundXin-Ji-Er-Kang (XJEK) as a herbal formula of traditional Chinese medicine (TCM) has shown the protective effects on myocardial function as well as renal function in mouse models of myocardial infarction.Hypothesis/PurposeWe investigated the effects of XJEK on cardiovascular- and renal-function in a heart failure mouse model induced by high salt (HS) and the associated mechanisms.Study designFor the purpose of assessing the effects of XJEK on a hypertensive heart failure model, mice were fed with 8% high salt diet. XJEK was administered by oral gavage for 8 weeks. Cardiovascular function parameters, renal function associated biomarkers and XJEK's impact on renin-angiotensin–aldosterone system (RAAS) activation were assessed. To determine the underlying mechanism, the calpain1/junctophilin-2 (JP2)/sarcoplasmic reticulum Ca²⁺ ATPase (SERCA2a) pathway was further studied in AC16 cells after angiotensin II-challenge or after calpastatin small interfering RNA (siRNA) transfection.ResultsMice on HS-diet exhibited hypertensive heart failure along with progressive kidney injury. Similar to fosinopril, XJEK ameliorated hypertension, cardiovascular-and renal- dysfunction in mice of HS-diet group. XJEK inhibited HS-induced activation of RAAS and reversed the abnormal expression pattern of calpain1and JP2 protein in heart tissues. XJEK significantly improved cell viability of angiotensin II-challenged AC16 cells. Moreover, XJEK's impact on calpain1/JP2 pathway was partly diminished in AC16 cells transfected with calpastatin siRNA.ConclusionXJEK was found to exert cardiovascular- and renal protection in HS-diet induced heart failure mouse model. XJEK inhibited HS-diet induced RAAS activation by inhibiting the activity and expression of calpain1 and protected the junctional membrane complex (JMC) in cardiomyocytes.