Injection Of Matrix Metalloproteinase-9 Leads To Ventricular Remodeling
HindawiDisease MarkersVolume 2022, Article ID 1659771, 12 pageshttps://doi.org/10.1155/2022/1659771Research ArticleInjection of Matrix Metalloproteinase-9 Leads toVentricular RemodelingEnzheng Zhu ,1 Congcong Yuan,1 Simiao Hu,1 Yiling Liao,1 Bowei Li,1 Yuliang Zhou,1and Wanxing Zhou 1,21Department of Cardiology, The First Affiliated Hospital of Guangdong Pharmaceutical University, Guangdong Province,510080, China2Metabolic Disease Research Center of Integrated Chinese and Western Medicine, Guangdong Province, 510080, ChinaCorrespondence should be addressed to Wanxing Zhou; zhouwanx2015@sina.comReceived 4 July 2022; Accepted 6 September 2022; Published 20 September 2022Academic Editor: Azizah UgusmanCopyright 2022 Enzheng Zhu et al. This is an open access article distributed under the Creative Commons Attribution License,which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.Objective. Previous studies have found that some ventricular remodeling is accompanied by increased matrix metalloproteinase-9(MMP-9) in vivo, and MMP-9 inhibitors can reduce ventricular remodeling. However, there is still no direct evidence that MMP9 causes ventricular remodeling. In this study, MMP-9 was injected into rats to observe whether MMP-9 caused ventricularremodeling, thereby providing direct evidence of MMP-9-induced ventricular remodeling. Methods. Forty-eight eight-week-oldmale Wistar rats were randomly divided, by weight, into control, low-, medium-, and high-dose MMP-9 groups and wereadministered normal saline or recombinant rat MMP-9 0.7, 1.4, or 2.1 ng/g, respectively, via intraperitoneal injection, twice perweek. On the 28th day, six rats were randomly selected from each group (Stage I). The remaining rats continued receivinginjections until the 56th day (Stage II). Echocardiography was performed to observe cardiac structure and function, and the leftventricular mass index (LVWI) was calculated. Myocardial pathological changes and the collagen volume fraction (CVF) wereobserved by HE and VG staining in myocardial tissue. MMP-9 levels in serum were tested using ELISA. Myocardial MMP-9levels were measured using Western blots, and the myocardial expression levels of MMP-9 mRNA were assessed using RTPCR. Results. During Stage I, serum MMP-9 and myocardial MMP-9 mRNA levels are increased; hypertrophiccardiomyocytes, disorderly arrangement of fibers, and endochylema dissolution are observed in the medium- and high-dosegroups. The left ventricular weight index (LVWI) and myocardial MMP-9 increased, and the collagen volume fraction (CVF)reduced in the high-dose group. In Stage II, the left ventricular end-diastolic volume (LVEDV) and diameter (LVIDd) arehigher, and CVF decreased in the medium- and high-dose groups. Myocardial pathological lesions intensified. Serum MMP-9in the model groups and myocardial MMP-9 in the medium- and high-dose groups are increased. Conclusions. Injection ofMMP-9 can lead to ventricular remodeling.1. IntroductionVentricular remodeling is a series of improper ventriclestructural changes generated when the body deals with various cardiovascular damage types, including hypertension,cardiomyopathy, and myocardial infarction (MI). Ventricular remodeling includes myocardial parenchymal remodeling and myocardial interstitial remodeling [1]. Therefore,characterizing the mechanism of ventricular remodeling isa critical step toward overcoming cardiovascular diseases.The mechanism underlying ventricular remodeling involvesthe renin-angiotensin-aldosterone system (RAAS) andexcessive sympathetic nerve activation. Outstanding clinicaleffects have been achieved with the targeted use ofangiotensin-converting enzyme inhibitors (ACEI), angiotensin II receptor blockers (ARBs), and β-blockers. However,the inflexion point of ventricular remodeling control hasnot yet been identified, and the morbidity and mortalityassociated with cardiac failure continue to increase [2]. Theglobal number of patients with cardiac failure increased to
223 million in 2020 [3], indicating a lack of understanding ofthe mechanisms underlying ventricular remodeling. Thus,further understanding of the mechanisms underlying ventricular remodeling is urgently required.Matrix metalloproteinase-9 (MMP-9) is a member of thematrix metalloproteinase (MMP) family. In recent years,research evidence has indicated that MMP-9 is involved inthe development of the fetal heart, angiogenesis, woundhealing, and other physiological processes but is also associated with ventricular remodeling in various cardiovasculardiseases [4]. For example, MMP-9 gene knockout or inhibition can significantly relieve ventricular remodeling in MI.MMP-9 inhibition can also mitigate the development ofmyocarditis and cardiomyopathy. Different ventricularremodeling processes in hypertension are accompanied byvariations in myocardial MMP-9 levels [4, 5]. However,according to the “evidence chain” requirements for determining pathogenic factors, a specific factor is considered apathogenic factor if (1) there is a significant increase in theincidence of the disease among individuals directly exposedto the pathogenic factor, and (2) inhibition of the factorcan reduce or cure the disease. Therefore, this study injectedMMP-9 into healthy rats to observe whether MMP-9 causedventricular remodeling, aiming to provide direct evidence ofMMP-9-induced ventricular remodeling. These findingsprovide a basis for further exploration of the mechanismunderlying ventricular remodeling and the search for newdrug therapeutic targets.2. Materials and Methods2.1. Experimental Animals and Grouping. In total, 48 Specific Pathogen-Free (SPF) level, healthy eight-week-old maleWistar rats (160-200 g) from the experimental animal centreof Southern Medical University were used in this study (certificate of quality number SCXX (Guangdong) 2011-0015).All animals were fed a conventional diet and housed in theSPF-level animal house of the First Affiliated Hospital ofGuangdong Pharmaceutical University, with access to sufficient food and water. The rats were housed under a 12 hlight/dark cycle, and the housing bedding was changed twiceweekly. The experiment began after one week of adaptation.The rats were randomly divided into four groups accordingto body weight stratification: control, low dose, mediumdose, and high dose (n 6 in each group). The control groupwas given normal saline (2 ml), and the model groups weretreated with 2 ml recombinant rat MMP-9 (rMMP-; purchased from the R&D systems Company, USA) at differentdoses through intraperitoneal injection twice per week:low-dose group 0.7 ng/body weight (g), medium-dose group1.4 ng/body weight (g), and high-dose group 2.1 ng/bodyweight (g). By the end of the fourth week, six rats in eachgroup were randomly selected into the Stage I group(n 24), and modeling injections ceased for this group ofrats. The remaining six rats in each group continued toreceive the modeling injections until the end of the eighthweek and were denoted the Stage II group (n 24). The testparameters for each group were the same. This study wasapproved by the Animal Ethics Committee of the First Affil-Disease Markersiated Hospital of Guangdong Pharmaceutical University(No. GYFY20161121010).2.2. Blood Pressure Test. All rats underwent blood pressuremeasurements one day before the commencement of theexperiments and at the completion (i.e., Stage I group onday 28 and Stage II group on day 56) under sober conditions. Blood pressure measurements were taken using aninvasive caudal artery sphygmomanometer (ZH-HX-Z)(purchased from China Anhui Zhenghua Bio-instrumentCompany). Blood pressure measurements were performeduniformly after 10:00 am. During each measurement, thetest table and sphygmomanometer were stored in the samestatic and windless environment, and the indoor temperature was maintained at 37 C. All rats were kept static for5 min before blood pressure measurements.2.3. Evaluation of Ventricular Remodeling. Rats were anaesthetized using 2% isoflurane at the end of either Stage I orII. The left ventricle structure and functions were examinedusing cardiac ultrasound (Philips HP5500), according to apreviously described method [6, 7]. Cardiac ultrasound ofall animals was performed by the same physician who wasblind to the experimental grouping. Rats were then anaesthetized with sodium pentobarbital (50 mg/kg, intraperitoneally(i.p.)). The left ventricle was separated and weighed. Basedon the weight, the LVWI was calculated according to the following formula: LVWI left ventricle weight ðLVWÞ ðmgÞ/body weight ðBWÞ ðgÞ. The left ventricular rat myocytes werecut into three parts (upper, middle, and lower) along the horizontal plane. The middle part was fixed in 4% paraformaldehyde and then underwent dehydration, the addition oftransparency reagent, conventional paraffin embedding, andslicing (4 μm). Tissue slices were submitted to HE staining.Myocardial cells and myocardial matrix fiber morphologyand arrangement were examined under an HE staining microscope. Changes to myocardium collagen fibers were observedusing VG staining (Baso Diagnosis Company, Zhuhai). Fivevisual fields without small coronary arteries were examinedin each slice. The Image-Pro Plus color pathological graphicanalysis system measured the collagen area. The collagen volume fraction (CVF) of the left ventricle was calculated as follows: CVF collagen area of the left ventricle/measured areaof view. Five CVFs were scored for each slice, and the meanwas calculated.3. Test of Serum MMP-9, Myocardial MMP-9,and Myocardial MMP-9 mRNA3.1. ELISA. MMP-9 levels in serum were determined usingELISA. After the rats were sacrificed, the abdomen was cut,and the aorta abdominalis was separated. Blood was collected in a non-anticoagulant blood collection tube and keptstatic for 30 min. The blood samples were then centrifugedfor 10 min at 4 C and 1500 rpm, and the serum was collectedand placed immediately on ice. All samples were kept in arefrigerator at -80 C for later use, preventing repeated freezing and thawing. Serum MMP-9 was detected using a rat
Disease MarkersMMP-9ELISA kit (R&D Company, USA), according to themanufacturer’s instructions.3.2. MMP-9 Contents of Myocardium Detected UsingWestern Blot. Myocardial MMP-9 was determined usingWestern blots: 60 mg of myocardial tissue was collected andground using liquid nitrogen to extract proteins for thebicinchoninic acid (BCA) protein assay kit (Beyotime Biotechnology, Haimen, China) quantification. Thirty micrograms ofprotein samples was electrophoresed and separated using 10%sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE). The proteins were later transferred to a polyvinylidene fluoride (PVDF) membrane. After soaking in milkfor 45 min, the anti-rat MMP-9 antibody was added anddiluted at a ratio of 1 : 1000 (Abcam Company, UK). Themembrane was then incubated on a shaker at 4 C overnight.Following this, the membrane was cleaned with PBST for30 min. The secondary antibody with horseradish peroxidaselabelling was then added, diluted at a ratio of 1 : 5000 (AbcamCompany, UK), and the membrane was incubated for onehour. The membrane was then cleaned again with PBST for30 min. Finally, the reaction was strengthened with a chemiluminescence reagent. Samples were compressed and exposedusing X-ray, and the images were analyzed.3.3. Myocardial MMP-9 mRNA Detected by RT-PCR. Trizolwas used to separate myocardial tissue and process the samples. Next, 1 μl RNA was collected, and an ultraviolet spectrophotometer measured its optical density (OD) at260 nm. The RNA concentration ðμg/mlÞ OD260 40 100. The RT reaction of cDNA was completed with AMVreverse transcriptase. The RT-PCR system was run as follows: 2 μl RNA, 4 μl RNase free ddH2O, and 1 μl Oligo dT,72 C, 3 min. Later, the cDNA was cooled on ice, 5 μl dNTP,5 μl RT buffer, and 1 μl AMV reverse transcriptase wereadded and reacted at 42 C for one hour. The RT productswere kept at -20 C for later use. The RT cDNA was usedas the template, and primers were used for PCR amplification. The target gene primers were designed according tothe literature and were synthesized by Sangon. The primerswere as follows: for MMP-9 5′-CCTGGAGACCTGAGAACCAATC-3′, for MMP-9 Rev5′-CCACCCGAGTGTAACCATAGC-3′, for β-actin 5′-AAGGTCGCTCGGATGGTTAT-3′, and β-actin Rev5′-GGGAGTCCTCGTGGTAGTGATA-3′. The amplification program was 94 C 2 min; 94 C30 s, 55 C 30 s, and 72 C 50 s for 30 cycles, followed by72 C 3 min. The PCR products were tested using 2% agarosegel electrophoresis (AGE).3.4. Statistical Analysis. SPSS 21.0 data analysis software(IBM, Armonk, NY, USA) was used to analyze the data statistically. Normality and homogeneity of variance tests werefirst carried out. Each group’s normally distributed measurement data were represented by the mean standarddeviation ( x SD); nonnormally distributed data were represented by the median. Intergroup comparisons were performed using a one-way analysis of variance (ANOVA)under the assumption of homogeneity of variance. TheLSD test was used for pairwise comparisons. Nonparametric3tests were used for data that violated the assumption ofhomogeneity of variance. The Pearson test was used for correlation analysis. Values of p 0:05 were taken to indicate astatistically significant difference.4. Results4.1. No Difference in Baseline Values; Experimental SurvivalRate 100%. The survival rate of the rats in the model andcontrol groups was 100%. There were no significant differences among the groups in terms of mean body weight andbaseline blood pressure level (p 0:05) (Table 1).4.2. Injection of MMP-9 Can Cause Changes toVentricular Remodeling4.2.1. Injection of MMP-9 Can Increase LVWI as a Functionof Time and Dosage. The LVWI of the high-dose group issignificantly higher than that of the control and low-dosegroups in Stage I and Stage II rats (p 0:05). Compared tothe control group, the medium-dose group and low-dosegroup exhibit an upward trend in LVWI in Stage I and StageII rats (p 0:05) (shown in Figure 1).4.2.2. Injection of MMP-9 Can Enlarge the Left Ventricle.There are no significant differences in the cardiac ultrasonictesting indices between the various model groups in Stage I(p 0:05). In Stage II, the left ventricular end-diastolic volume (LVEDV) and left ventricular end-diastolic diameter(LVIDd) of the high-dose group are far higher than thoseof the control and low-dose groups (p 0:05). Comparedto the control group, the medium-dose group exhibited significant increases in the LVEDV and LVIDd (p 0:05). Theposterior diastolic wall thickness (LVPWd) of the mediumdose group shows an increasing trend compared to that ofthe control group (p 0:05). No significant differences areobserved in the other indices (shown in Figure 2).4.2.3. HE Staining Shows Myocardial Damage and StructuralDisorder. In Stage I rats, compared to the control group, themedium-dose and high-dose groups present with evidentcardiomyocyte hypertrophy and disordered arrangement,accompanied by breakage of myocardial fibers, endochylemadissolution, and necrosis. There are no significant changes inthe myocardial cells of the low-dose group (shown inFigures 3(a)–3(d)).In the Stage II rats, compared to the control group, allmodel groups presented with breakage, damage, and myolysis of myocardial fibers, and myocardial cells had disappeared. Moreover, these lesions intensified with increasingMMP-9 levels (shown in Figures 3(e)–3(h)).4.2.4. VG Staining Shows a Reduction in Myocardial MatrixCollagen Volume Fraction (CVF). The myocardial matrix CVFof the high-dose group decreases significantly compared tothat of the control and the low-dose groups, for both Stage Iand II rats (p 0:05). In Stage II rats, the myocardial matrixCVF of the medium-dose group is far lower than that of thecontrol group (p 0:05). The low-dose group was not
4Disease MarkersTable 1: Body weights and blood pressure measurements.Stage I (n 24)Stage II (n 24)Initial weight (g) Initial SBP (mmHg) Initial DBP (mmHg) Initial weight (g) Initial SBP (mmHg) Initial DBP (mmHg)Control207 16:7121 7:881 4:6205 19:9124 9:480 6:0Low dose206 16:8123 5:881 5:0207 14:5122 8:481 4:5Mediumdose210 14:8120 6:479 6:5205 18:4121 7:379 5:2High dose209 14:3118 8:478 6:2209 17:3119 9:780 4:9Notes: data are presented as the mean SD. SBP: systolic pressure; DBP: diastolic pressure.3⁎#LVWI (mg/g)⁎#214w4w con4wlo trom w- led doiusm e4w -dohigh se-dose8wc8 o8w w lo ntrom w- led doiuse8w m-dohigh se-dose0Stage IStage IIFigure 1: Changes in the left ventricular mass index in Stage I andStage II rats. Notes: results are expressed as the average magnitudeof each value within a group of animals SD (n 6/group). represents p 0:05 compared to the control group. # represents p 0:05 compared to the low-dose group.significantly different from the control group in both Stages Iand II rats (p 0:05) (shown in Figures 4(a)–4(i)).4.2.5. Injection of MMP-9 Can Increase Blood Pressure. ForStage I rats, compared with the control group, the lowdose group exhibited no significant increase in blood pressure (p 0:05). In contrast, the medium- and high-dosegroups show significant increases in systolic blood pressure(SBP) (p 0:05). The high-dose group also has significantlyhigher SBP (p 0:05) than the low-dose group. In the StageII rats, the increases in SBP are similar to those in Stage Irats. SBP is significantly increased in the medium- andhigh-dose groups (p 0:05), with a more significant increasein the high-dose group. Diastolic blood pressure (DBP) issignificantly increased in all three model groups (p 0:05)(shown in Figure 5).4.2.6. Injection of MMP-9 Can Increase Myocardial MMP-9,Serum MMP-9, and Myocardial MMP-9 mRNA. The highdose group displayed significant increases in myocardialMMP-9 and serum MMP-9 compared to the control andlow-dose groups, in both Stage I and II rats (p 0:05). InStage I rats, the myocardial MMP-9 mRNA of the highdose group is significantly higher than that of the controlgroup (p 0:05) but not significantly higher than the otherdose groups. There are no significant differences in myocardial MMP-9 mRNA among the groups in the Stage II rats(p 0:05).Compared to the control group, the medium-dose groupshows far higher serum MMP-9 and myocardial MMP-9mRNA in Stage I rats (p 0:05). In Stage II rats, the serumMMP-9 level is further increased (p 0:05), as was the myocardial MMP-9 level (p 0:05), but there are no significantdifferences in myocardial MMP-9 mRNA among the different groups (p 0:05).In both stages, the low-dose group presents an increasingtrend in myocardial MMP-9 levels, serum MMP-9 levels,and myocardial MMP-9 mRNA compared with the controlgroup (p 0:05). The serum MMP-9 level of the low-dosegroup is significantly increased compared to that of the control group in Stage II rats (p 0:05) (shown in Figures 6(a)–6(d)).4.2.7. Myocardial MMP-9 Levels Positively Correlated withLVWI, LVIDd, and LVEDV and Negatively Correlated withLVEF. Correlation analysis showed that the myocardialMMP-9 level is significantly positively correlated with LVWI,LVIDd, and LVEDV, with correlation coefficients of 0.57,0.62, and 0.56, respectively (p 0:01). Myocardial MMP-9level is negatively correlated with LVEF, with a correlationcoefficient of -0.35 (p 0:05) (shown in Figures 7(a)–7(d)).4.2.8. Serum MMP-9 Positively Correlated with LVWI andLVIDd and Negatively Correlated with LVEF. Correlationanalysis shows that serum MMP-9 level is significantly positively correlated with LVWI and LVIDd, with correlationcoefficients of 0.44 and 0.33 (p 0:05), respectively. SerumMMP-9 level is negatively correlated with LVEF, with a coefficient of -0.37 (p 0:05) (shown in Figures 7(e)–7(g)).5. DiscussionVentricular remodeling includes myocardial parenchymalremodeling and myocardial interstitial remodeling. Histologically, the former is characterized by cardiomyocyte hypertrophy, myocardial apoptosis, and fibroblast proliferation.The latter is characterized by extracellular matrix degradation, collagen fibrinolysis, and sedimentation. In general, itmanifests as increased LVWI, myocardial hypertrophy,decreased E/A peak values of the mitral diastolic blood flowspectrum, and decreased LVEF [8]. In this study, after
0.210.14HR (bpm)Stage IStage IStage I602000.20.00.350.070.00Stage II2.5Stage I2.02.01.50.51.00.50.00.0Stage IIStage I57380.8Stage II2.5Stage I0.62.01.51.0Stage IIStage IStage IStage IIIVSs (mm)6.53.05.22.421.20.600.00.0Stage IIStage IStage II950.4Stage I760.3190Stage IIStage I0.50.20.00.0Stage I8w8 c8w w l ontm ow role d -doi8w u m - s ehi d o sgh e-dose4wco4w 4w ntrm low ole d -diu os4w m - ehi d o sgh e-dose4wc4w 4w ontm low role d -diu os4w m - ehi d o sgh e-dose8wc8o8w w nm low trole d -diu os8w m - ehi d o sgh e-doseLVIDs (mm)2.64wc4w 4w ontm low roled -diu os4w m- ehi dosgh e-dose8wc8w 8w ontm low roled -diu os8w m-d ehi osegh-dose1.03.94w4 c4w w l ontm ow- roled doius4w m- ehi dosgh e-dose8wc8w 8w l ontm ow roled -diu os8w m- ehi dosgh e-doseStage IILVPWs (mm)2.5CO (1/min)8w8w conlo trowe d -d liu ose8w m hi d o sgh e-dose⁎LVEDV (ml)Stage Im44wc4w 4w ontm low roled -diu os4w m-d ehi osegh-dose8wc8w 8w l ontm ow roled -diu os8w m- ehi dosgh e-dose1.5LVPWd (mm)Stage I8w4wc4w 4w ontm low role d -diu os4w m - d ehi o s egh-doseLVIDd (mm)64wc4w 4w ontm low roled -diu os4w m-d ehi osgh e-dose8w8w con8wl tm ow- roled diu ose8w mhi -dogh se-dose40LVEF (%)4wc4w 4w ontm low roled -diu os4w m-d ehi osegh-dose8w8w co8wl ntm ow- roleddiu os8w m- ehi dosgh e-doseIVSd (mm)⁎#4wc4w 4w ontm low roled -diu os4w m-d ehi osgh e-dose8wc88w w l ontm ow roled -diu os8w m- ehi dosgh e-dose0.4E/A4w4w 4w conm low troled -diu o4w m- sehi dosgh e-dose8wco8w 8w ntrm low oled -doi8w um sehi -dogh se-doseLVFS (%)84wc4w 4w ontm low roled -diu os4w m- ehi dosgh e-dose8wco8w8wntm low roled -diu os8w m- ehi dosgh e-dose4w4 c4w w l ontm ow roled -diu os4w m-d ehi osgh e-dose8wco88w w nm low troled -diu os8w m-d ehi osegh-doseSV (ml )104wc4w 4w ontm low roled -diu os4w m- ehi dosgh e-dose8wc8w 8w ontm low roled -diu os8w m-d ehi osegh-dose4wc4w 4w ontm low roled -diu os4w m-d ehi osgh e-dose8w8w con8wl tm ow roled -diu os8w m- ehi dosgh e-doseIVSDV (ml)Disease Markers51.81.23.3Stage II2.21.10.0Stage II0.20.10.01.0Stage II0.8⁎ ⁎#0.60.4Stage II0.28450360270180900Stage IIFigure 2: Changes in general cardiac structure and functions of Stage I and Stage II rats. Notes: LVIDd: left ventricular end-diastolicdiameter; LVIDs: left ventricular end-systolic diameter; IVSd: interventricular septum diastolic diameter; IVSs: interventricular septumsystole diameter; LVPWd: diastole left ventricular posterior wall diameter; LVPWs: systole left ventricular wall diameter; LVFS: leftventricular shortening fraction; LVEF: left ventricular ejection fraction; CO: cardiac output; SV stroke volume; E/A: bicuspid valve bloodpeak E/peak A; LVEDV: left ventricular end-diastolic volume; LVSDV: left ventricular end-systolic volume; HR: heart rate. Results areexpressed as the average magnitude of each value within a group of animals SD (n 6/group). represents p 0:05 compared to thecontrol group. # represents p 0:05 compared to the low-dose group.
6Disease Markers100 𝜇m100 𝜇m100 𝜇m4W control4W low-dose(a)(b)100 𝜇m4W medium-dose(c)100 𝜇m100 𝜇m4W high-dose(d)100 𝜇m100 𝜇m8W control8W low-dose8W medium-dose8W high-dose(e)(f)(g)(h)Figure 3: Light microscope changes in the myocardial structure of each group in Stage I and Stage II rats (HE staining, normal lightmicroscope, 100). In Stage I rats (a–d), compared with the control group, the medium- and high-dose groups showed pathologicalchanges in myocardial ventricular remodeling, such as cell hypertrophy, disarrangement, cytoplasmic lysis, and necrosis. In contrast, thelow-dose group had no significant change. The model observed pathological changes in ventricular remodeling in the Stage II rats (e–h)compared with the control group.intraperitoneal injection of recombinant MMP-9 in healthyrats, myocardial fiber fracture, cytoplasmic lysis, and structural disorder were observed histologically. Myocardialinterstitial CVF is significantly decreased. In general,increases in the ventricular mass index (LVWI), left ventricular end-diastolic volume (LVEDV), and left ventricularend-diastolic diameter (LVIDd) are observed. These findingssuggest that ventricular remodeling occurred in rats afterMMP-9 injection.Further, MMP-9 and MMP-9 mRNA levels in serumand myocardial tissue are correlated with changes in theventricular remodeling indices. The results demonstratedthat the medium-dose and high-dose groups exhibited significant increases in serum MMP-9 in the fourth week. Incontrast, myocardial MMP-9 is increased significantly onlyin the high-dose group. In the eighth week, serum MMP-9in the medium-dose and high-dose groups is furtherincreased. Moreover, myocardial MMP-9 in the two groupsalso increased significantly. These results indicate that serumMMP-9 increased earlier than myocardial MMP-9, consistent with the assertion that intraperitoneal-injected exogenous drugs are absorbed by the blood and then enter themyocardial tissue. This finding is similar to the previouslyreported order of elevated plasma concentrations in anin vivo model constructed by intraperitoneal injection ofexogenous drugs [9, 10]. Thus, these findings indicate thatthe in vivo MMP-9 elevation observed here is primarilyderived from exogenous sources. Secondly, in the currentstudy, the degree of ventricular remodeling (e.g., increasesin LVWI, LVEDV, LVIDd, and the degree of myocardial celldestruction and decreases in myocardial interstitial CVF)increased in a dose- and time-dependent manner withMMP-9 injections. Regarding the correlations between theventricular remodeling indices and the myocardial tissue andserum MMP-9 levels, it is noted that myocardial MMP-9was significantly higher in the high-dose group in the fourthweek. However, the changes to the ventricular remodelingindices, such as LVWI, LVEDV, LVIDd, myocardial cells,and interstitium, were not sufficiently noticeable at this time.The relevant indicators of ventricular remodeling did notexhibit statistically significant changes until the eighth week.The lag in myocardial remodeling behind the elevation inMMP-9 is consistent with the temporal characteristics ofMMP-9 as the cause of ventricular remodeling.Moreover, myocardial and serum MMP-9 levels correlate significantly and positively with LVWI, LVIDd, andLVEDV. LVEF is significantly negatively correlated withmyocardial and serum MMP-9. The myocardial MMP-9level correlates better with the ventricular remodelingparameters than the serum MMP-9 levels. This further supports the assertion that myocardial remodeling is related tothe local myocardial MMP-9 elevation. However, the correlation between LVEF and MMP-9 is lower than the correlations for the above cardiac structural indicators, an indexconsidered more closely related to structural changes inearly ventricular remodeling than functional decline. Inaddition, the results of this study indicate that MMP-9 injections resulted in MMP-9 mRNA elevation, suggesting theinduction of endogenous MMP-9 expression. However, thisphenomenon was only observed in the first-stage group and
Disease Markers750 𝜇m50 𝜇m(a)4W medium-dose4W medium-dose(c)(d)(b)50 𝜇m50 𝜇m50 𝜇m8W low-dose(e)(f)8W medium-dose(g)50 𝜇m8W high-dose(h)54⁎#3⁎⁎#2104w4w con4w lo trom w- led doiu se4w m-dhi osegh-dose8w8 co8w w lo ntrom w- led doiuse8w m-dhi osegh-doseCollagen volume fraction (%)8W control50 𝜇m50 𝜇m4W low-dose4W controlStage IStage II(i)Figure 4: The morphology and distribution of myocardial collagen in Stage I (a–d) and Stage II (e–h) rats (VG staining, normal lightmicroscope, 200); (i) the CVF of the different groups in Stage I and Stage II. Notes: collagenous fibers are red, and muscle fibers areyellow. Results are expressed as the average magnitude of each value within a group of animals SD (n 6/group). represents p 0:05compared to the control group. # represents p 0:05 compared to the low-dose group.not in the second-stage group, suggesting that the inductionof endogenous MMP-9 expression is transient and does notlast long. This outcome also suggests that the furtherincrease in myocardial MMP-9 in the second-stage groupwas mainly due to continuous exogenous input. Finally, thisstudy’s finding of ventricular remodeling after MMP-9 injection is consistent with previously reported ventricularremodeling with elevated MMP-9 characteristics and theaction of MMP-9 itself [4].As a proteolytic enzyme, MMP-9 can act on several substrates in the extracellular matrix, including cleaved collagen,thereby decreasing collagen content [4]. MMP-9 can alsoinduce cardiomyocyte hypertrophy, apoptosis, death, andmyocardial myosin heavy chain rupture, damaging the myocardial structure and contractile ability [11–13]. After injec-tion of MMP-9, the high MMP-9 ventricular remodeling(LVR-hMMP-9) model rats exhibited myocardial cell damage,reductions in CVF, disordered arrangement, and expansion ofcardiac chambers, consistent with the characteristics of MMP9 ventricular remodeling. There was also a significant increasein blood pressure after MMP-9 injection, and hypertensioncan also lead to ventricular remodeling.The main manifestations of hypertension-induced ventricular remodeling are increased collagen fiber content inthe interstitium, ventricular wall hypertrophy, and reducedleft ventricular diameter [14]. However, in the currentmodel, CVF decreased and left ventricular diameterincreased. Therefore, although blood pressure is significantlyincreased in the model group, elevated blood pressure doesnot seem to be the primary cause of ventricular remodeling.
8Disease Markers160⁎⁎⁎#8040⁎⁎⁎75DBP (mmHg)120SBP (mmHg)100⁎#502504w4w con4wl trm ow- ole d doius4w m - ehi dos
ventricular remodeling, aiming to provide direct evidence of MMP-9-induced ventricular remodeling. These findings provide a basis for further exploration of the mechanism underlying ventricular remodeling and the search for new drug therapeutic targets. 2. Materials and Methods 2.1. Experimental Animals and Grouping. In total, 48 Spe-
CONTENTS CONTENTS Notation and Nomenclature A Matrix A ij Matrix indexed for some purpose A i Matrix indexed for some purpose Aij Matrix indexed for some purpose An Matrix indexed for some purpose or The n.th power of a square matrix A 1 The inverse matrix of the matrix A A The pseudo inverse matrix of the matrix A (see Sec. 3.6) A1 2 The square root of a matrix (if unique), not elementwise
A Matrix A ij Matrix indexed for some purpose A i Matrix indexed for some purpose Aij Matrix indexed for some purpose An Matrix indexed for some purpose or The n.th power of a square matrix A 1 The inverse matrix of the matrix A A The pseudo inverse matrix of the matrix A (see Sec. 3.6) A1/2 The square root of a matrix (if unique), not .
CONTENTS CONTENTS Notation and Nomenclature A Matrix Aij Matrix indexed for some purpose Ai Matrix indexed for some purpose Aij Matrix indexed for some purpose An Matrix indexed for some purpose or The n.th power of a square matrix A 1 The inverse matrix of the matrix A A The pseudo inverse matrix of the matrix A (see Sec. 3.6) A1/2 The square root of a matrix (if unique), not elementwise
CONTENTS CONTENTS Notation and Nomenclature A Matrix A ij Matrix indexed for some purpose A i Matrix indexed for some purpose Aij Matrix indexed for some purpose An Matrix indexed for some purpose or The n.th power of a square matrix A 1 The inverse matrix of the matrix A A The pseudo inverse matrix of the matrix A (see Sec. 3.6) A1 2 The sq
injection) Code injection attacks: also known as "code poisoning attacks" examples: Cookie poisoning attacks HTML injection attacks File injection attacks Server pages injection attacks (e.g. ASP, PHP) Script injection (e.g. cross-site scripting) attacks Shell injection attacks SQL injection attacks XML poisoning attacks
Further Maths Matrix Summary 1 Further Maths Matrix Summary A matrix is a rectangular array of numbers arranged in rows and columns. The numbers in a matrix are called the elements of the matrix. The order of a matrix is the number of rows and columns in the matrix. Example 1 [is a ] 3 by 2 or matrix as it has 3 rows and 2 columns. Matrices are .
cyanocobalamin injection usp. baclofen injection lidocaine hci injection (1%, 2%) piperacillin and tazobactam for injection dimenhydrinate injection usp with preservative. dimenhydrinate injection usp diazepam injection usp. naloxone hydrochloride
Diesel fuel-injection systems: An overview Fields of application 4 Technical requirements 4 Injection-pump designs 6 Mechanically-controlled (governed) axial-piston distributor fuel-injection pumps VE Fuel-injection systems 8 Fuel-injection techniques 9 Fuel supply and delivery 12 Mechanical engine-speed control (governing) 22 Injection timing 29
