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Since the COVID-19 outbreak, raccoon dogs have been suggested as a potential intermediary in transmitting SARS-CoV-2 to humans. To understand their role in the COVID-19 pandemic and the species barrier for SARS-CoV-2 transmission to humans, we analyzed how their ACE2 protein interacts with SARS-CoV-2 spike protein. Biochemical data showed that raccoon dog ACE2 is an effective receptor for SARS-CoV-2 spike protein, though not as effective as human ACE2. Structural comparisons highlighted differences in the virus-binding residues of raccoon dog ACE2 compared to human ACE2 (L24Q, Y34H, E38D, T82M, R353K), explaining their varied effectiveness as receptors for SARS-CoV-2. These variations contribute to the species barrier that exists between raccoon dogs and humans regarding SARS-CoV-2 transmission. Identifying these barriers can help assess the susceptibility of other mammals to SARS-CoV-2. Our research underscores the potential of raccoon dogs as SARS-CoV-2 carriers and identifies molecular barriers that affect the virus's ability to jump between species. Author summary: Raccoon dogs are thought to have been a crucial link in the chain that led to the spread of the COVID-19 virus to humans. The key to a coronavirus moving from one species to another is its spike protein's ability to bind to receptors on the cells of the new host. In our research, we investigated the interaction between the spike protein of the COVID-19 virus and the ACE2 receptor in raccoon dogs, using both structural and biochemical techniques. We found that the ACE2 receptor in raccoon dogs does indeed work as an effective receptor for the virus, but there are critical differences when compared to the human ACE2 receptor, which may act as barriers to the virus making the jump between raccoon dogs and humans. To our knowledge, this study is the first in-depth examination of these cross-species transmission barriers for the COVID-19 virus, shedding light on how other mammals might be impacted by the virus. Our findings indicate that raccoon dogs could potentially carry and pass the COVID-19 virus on to humans, providing foundational knowledge for how the virus can navigate between different species. [ABSTRACT FROM AUTHOR]

Transmembrane proteins have exhibited a significant correlation with glioblastoma multiforme (GBM). The current study elucidates the roles of transmembrane protein 150A (TMEM150A) in GBM. Data on patients with GBM were collected from The Cancer Genome Atlas and Xena databases. The objective was to identify the expression levels of TMEM150A in patients with GBM, and evaluate its diagnostic and prognostic values, accomplished using the receiver operating characteristic and survival analyses. On a cellular level, Cell Counting Kit-8, Wound healing, and Transwell experiments were performed to gauge the impact of TMEM150A on cell growth and migration. The study further investigated the correlation between TMEM150A expression and immune status, along with ribonucleic acid (RNA) modifications in GBM. The findings demonstrated TMEM150A overexpression in the cancerous tissues of patients with GBM, with an area under the curve value of 0.95. TMEM150A overexpression was significantly correlated with poor prognostic indicators. TMEM150A overexpression and isocitrate dehydrogenase (IDH) mutation status were predictive of poor survival time among patients with GBM. In vitro experiments indicated that suppressing TMEM150A expression could inhibit GBM cell proliferation, migration, and invasion. Moreover, TMEM150A overexpression was associated with stromal, immune, and estimate scores, immune cells (such as the T helper (Th) 17 cells, Th2 cells, and regulatory T cells), cell markers, and RNA modifications. Therefore, TMEM150A overexpression might serve as a promising biomarker for predicting poor prognosis in patients with GBM. Inhibiting TMEM150A expression holds the potential for improving the survival time of patients with GBM. [ABSTRACT FROM AUTHOR]

Background: Expression levels of transmembrane protein 41A (TMEM41A) are related to the progression of malignant tumors. However, the association between TMEM41A expression and endometrial carcinoma (EC) remains unclear. This study aims to identify the roles of TMEM41A expression in the prognosis of patients with EC and its correlation with EC progression. Methods: The TMEM41A expression and its correlation with the survival of patients with EC were assessed. Cox regression analysis was used to identify the prognostic factors, while nomograms were used to examine the association between the prognostic factors and the survival of patients with EC. Finally, the link between TMEM41A level and immune microenvironment and RNA modifications was investigated in EC. Results: TMEM41A was overexpressed in EC. TMEM41A overexpression could diagnose the EC and evaluate the poor prognosis of patients. Overexpression of TMEM41A was associated with clinical stage, age, weight, histological subtype, tumor grade, and survival status of patients with EC. Clinical stage, age, tumor grade, radiotherapy, and TMEM41A overexpression were factors of poor prognosis in patients with EC. The nomograms revealed the correlation between the TMEM41A level and survival time of patients with EC at 1, 3, and 5 years. Furthermore, TMEM41A overexpression was significantly correlated with the level of the stromal score, immune score, estimate score, NK CD56 bright cells, iDC, NK cells, eosinophils, pDC, T cells, TReg, cytotoxic cells, mast cells, Th17 cells, neutrophils, aDC, NK CD56 dim cells, TFH, Th2 cells, CD8 T cells, macrophages, immune cell markers, and RNA modifications. Conclusions: TMEM41A is overexpressed in EC tissues and is associated with the prognosis, immune microenvironment, and RNA modification. Our preliminary studies indicate that overexpression of TMEM41A can potentially serve as a biomarker for EC treatment. [ABSTRACT FROM AUTHOR]

An ancient enzyme family responsible for the catabolism of the prebiotic chemical cyanuric acid (1,3,5-triazine-2,4,6-triol) was recently discovered and is undergoing proliferation in the modern world due to industrial synthesis and dissemination of 1,3,5-triazine compounds. Cyanuric acid has a highly stabilized ring system such that bacteria require a unique enzyme with a novel fold and subtle active site construction to open the ring. Each cyanuric acid hydrolase monomer consists of three isostructural domains that coordinate and activate the three-fold symmetric substrate cyanuric acid for ring opening. We have now solved a series of X-ray structures of an engineered, thermostable cyanuric acid ring-opening enzyme at 1.51 ~ 2.25 Å resolution, including various complexes with the substrate, a tight-binding inhibitor, or an analog of the reaction intermediate. These structures reveal asymmetric interactions between the enzyme and bound ligands, a metal ion binding coupled to conformational changes and substrate binding important for enzyme stability, and distinct roles of the isostructural domains of the enzyme. The multiple conformations of the enzyme observed across a series of structures and corroborating biochemical data suggest importance of the structural dynamics in facilitating the substrate entry and the ring-opening reaction, catalyzed by a conserved Ser-Lys dyad. [ABSTRACT FROM AUTHOR]

Purpose: To determine the value of dual-source computed tomography (DSCT) in depicting the morphological characteristics and diagnosing the associated malformations for patients with transposition of the great arteries (TGA) before surgery. Materials and methods: Twenty-five patients with TGA who underwent DSCT and transthoracic echocardiography (TTE) examination were retrospectively reviewed. The morphological types of TGA, the spatial relationship between the pulmonary artery and the aorta, as well as coronary artery-associated abnormalities were assessed by DSCT. In contrast to TTE, the diagnostic accuracy of associated malformations on DSCT were analyzed and calculated with reference to surgical or digital subtraction angiography (DSA) findings. Effective doses (EDs) were also calculated. Results: Among the 25 patients, 12 (48%) had ventricular septal defects and left ventricular outflow tract stenosis. Sixteen patients (16/25, 64%) had great arteries with an oblique spatial relationship on DSCT. In addition, we found seven patients (7/25, 28%) with coronary artery malformation, including five with an abnormal coronary origin and two with signs of a myocardial bridge. According to DSA or surgical findings, DSCT was superior to TTE in demonstrating extracardiac anomalies (sensitivity, anomalies of great vessels: 100% vs. 93.33%, other anomalies: 100% vs. 46.15%). The mean estimated ED for those aged <10 years was <2 mSv (1.59 ± 0.95 mSv). Conclusions: DSCT can achieve an overall assessment of patients with TGA, including any associated malformations as well as the identification of the spatial relationship of the great arteries. DSCT can therefore be considered as an alternative imaging modality for surgical decision making. [ABSTRACT FROM AUTHOR]

Photodynamic antimicrobial chemotherapy (PACT), as a novel and effective therapeutic modality to eradicate drug resistant bacteria without provoking multidrug resistance, has attracted increasing attention. This study examined the antimicrobial efficacy of the novel cationic amino acid-porphyrin conjugate 4I with four lysine groups against two different clinical isolated strains (drug sensitive and multidrug resistant) of the Acinetobacter baumannii species and its toxicity on murine dermal fibroblasts in vitro, as well as the therapeutic effect of PACT on acute, potentially lethal multidrug resistant strain excisional wound infections in vivo. The PACT protocol exposed 4I to illumination, exhibiting high antimicrobial efficacy on two different strains due to a high yield of reactive oxygen species (ROS) and non-selectivity to microorganisms. The photoinactivation effects of 4I against two different strains were dose-dependent. At 3.9 μM and 7.8 μM, PACT induced 6 log units of inactivation of sensitive and multidrug resistant strains. In contrast, 4I alone and illumination alone treatments had no visibly antimicrobial effect. Moreover, cytotoxicity tests revealed the great safety of the photosensitizer 4I in mice. In the in vivo study, we found 4I-mediated PACT was not only able to kill bacteria but also accelerated wound recovery. Compared with non-treated mice, over 2.89 log reduction of multidrug resistant Acinetobacter baumannii strain was reached in PACT treat mice at 24 h post-treatment. These results imply that 4I-mediated PACT therapy is an effective and safe alternative to conventional antibiotic therapy and has clinical potential for superficial drug-resistant bacterial infections. [ABSTRACT FROM AUTHOR]

Purpose: To evaluate the feasibility and diagnostic accuracy of retrospective electrocardiographically (ECG)-gated dual-source computed tomography (DSCT) for the assessment of double outlet right ventricle (DORV) and associated multiple malformations in pediatric patients. Materials and Methods: Forty-seven patients <10 years of age with DORV underwent retrospective ECG-gated DSCT. The location of the ventricular septal defect (VSD), alignment of the two great arteries, and associated malformations were assessed. The feasibility of retrospective ECG-gated DSCT in pediatric patients was assessed, the image quality of DSCT and the agreement of the diagnosis of associated malformations between DSCT and transthoracic echocardiography (TTE) were evaluated, the diagnostic accuracies of DSCT and TTE were referred to surgical results, and the effective doses were calculated. Results: Apart from DORV, 109 associated malformations were confirmed postoperatively. There was excellent agreement (κ = 0.90) for the diagnosis of associated malformations between DSCT and TTE. However, DSCT was superior to TTE in demonstrating paracardiac anomalies (sensitivity, coronary artery anomalies: 100% vs. 80.00%, anomalies of great vessels: 100% vs. 88.57%, separate thoracic and abdominal anomalies: 100% vs. 76.92%, respectively). Combined with TTE, DSCT can achieve excellent diagnostic performance in intracardiac anomalies (sensitivity, 91.30% vs. 100%). The mean image quality score was 3.70 ± 0.46 (κ = 0.76). The estimated mean effective dose was < 1 mSv (0.88 ± 0.34 mSv). Conclusions: Retrospective ECG-gated DSCT is a better diagnostic tool than TTE for pediatric patients with complex congenital heart disease such as DORV. Combined with TTE, it may reduce or even obviate the use of invasive cardiac catheterization, and thus expose the patients to a much lower radiation dose. [ABSTRACT FROM AUTHOR]

Background and Aims: There is no prognostic model that is reliable and practical for patients who have received curative liver resection (CLR) for hepatocellular carcinoma (HCC). This study aimed to establish and validate a Surgery-Specific Cancer of the Liver Italian Program (SSCLIP) scoring system for those patients. Methods: 668 eligible patients who underwent CLR for HCC from five separate tertiary hospitals were selected. The SSCLIP was constructed from a training cohort by adding independent predictors that were identified by Cox proportional hazards regression analyses to the original Cancer of the Liver Italian Program (CLIP). The prognostic performance of the SSCLIP at 12 and 36-months was compared with data from existing models. The patient survival distributions at different risk levels of the SSCLIP were also assessed. Results: Four independent predictors were added to construct the SSCLIP, including age (HR = 1.075, 95%CI: 1.019–1.135, P = 0.009), albumin (HR = 0.804, 95%CI: 0.681–0.950, P = 0.011), prothrombin time activity (HR = 0.856, 95%CI: 0.751–0.975, P = 0.020) and microvascular invasion (HR = 19.852, 95%CI: 2.203–178.917, P = 0.008). In both training and validation cohorts, 12-month and 36-month prognostic performance of the SSCLIP were significantly better than those of the original CLIP, model of end-stage liver disease-based CLIP, Okuda and Child-Turcotte-Pugh score (all P < 0.05). The stratification of risk levels of the SSCLIP showed an enhanced ability to differentiate patients with different outcomes. Conclusions: A novel SSCLIP to predict survival of HCC patients who received CLR based on objective parameters may provide a refined, useful prognosis algorithm. [ABSTRACT FROM AUTHOR]

Cyanuric acid hydrolase (CAH) catalyzes the hydrolytic ring-opening of cyanuric acid (2,4,6-trihydroxy-1,3,5-triazine), an intermediate in s-triazine bacterial degradation and a by-product from disinfection with trichloroisocyanuric acid. In the present study, an X-ray crystal structure of the CAH-barbituric acid inhibitor complex from Azorhizobium caulinodans ORS 571 has been determined at 2.7 Å resolution. The CAH protein fold consists of three structurally homologous domains forming a β-barrel-like structure with external α-helices that result in a three-fold symmetry, a dominant feature of the structure and active site that mirrors the three-fold symmetrical shape of the substrate cyanuric acid. The active site structure of CAH is similar to that of the recently determined AtzD with three pairs of active site Ser-Lys dyads. In order to determine the role of each Ser-Lys dyad in catalysis, a mutational study using a highly sensitive, enzyme-coupled assay was conducted. The 109-fold loss of activity by the S226A mutant was at least ten times lower than that of the S79A and S333A mutants. In addition, bioinformatics analysis revealed the Ser226/Lys156 dyad as the only absolutely conserved dyad in the CAH/barbiturase family. These data suggest that Lys156 activates the Ser226 nucleophile which can then attack the substrate carbonyl. Our combination of structural, mutational, and bioinformatics analyses differentiates this study and provides experimental data for mechanistic insights into this unique protein family. [ABSTRACT FROM AUTHOR]

Background & Aims: Hepatitis B surface antigen (HBsAg) seroclearance and seroconversion are regarded as favorable outcomes of chronic hepatitis B (CHB). This study aimed to develop artificial neural networks (ANNs) that could accurately predict HBsAg seroclearance or seroconversion on the basis of available serum variables. Methods: Data from 203 untreated, HBeAg-negative CHB patients with spontaneous HBsAg seroclearance (63 with HBsAg seroconversion), and 203 age- and sex-matched HBeAg-negative controls were analyzed. ANNs and logistic regression models (LRMs) were built and tested according to HBsAg seroclearance and seroconversion. Predictive accuracy was assessed with area under the receiver operating characteristic curve (AUROC). Results: Serum quantitative HBsAg (qHBsAg) and HBV DNA levels, qHBsAg and HBV DNA reduction were related to HBsAg seroclearance (P<0.001) and were used for ANN/LRM-HBsAg seroclearance building, whereas, qHBsAg reduction was not associated with ANN-HBsAg seroconversion (P = 0.197) and LRM-HBsAg seroconversion was solely based on qHBsAg (P = 0.01). For HBsAg seroclearance, AUROCs of ANN were 0.96, 0.93 and 0.95 for the training, testing and genotype B subgroups respectively. They were significantly higher than those of LRM, qHBsAg and HBV DNA (all P<0.05). Although the performance of ANN-HBsAg seroconversion (AUROC 0.757) was inferior to that for HBsAg seroclearance, it tended to be better than those of LRM, qHBsAg and HBV DNA. Conclusions: ANN identifies spontaneous HBsAg seroclearance in HBeAg-negative CHB patients with better accuracy, on the basis of easily available serum data. More useful predictors for HBsAg seroconversion are still needed to be explored in the future. [ABSTRACT FROM AUTHOR]

Cardiac ischemia and reperfusion (I/R) injury occurs because the acute increase in oxidative/inflammatory stress during reperfusion culminates in the death of cardiomyocytes. Currently, there is no drug utilized clinically that attenuates I/R injury in patients. Previous studies have demonstrated degranulation of mast cell contents into the interstitium after I/R. Using a dog model of I/R, we tested the role of chymase, a mast cell protease, in cardiomyocyte injury using a specific oral chymase inhibitor (CI). 15 adult mongrel dogs had left anterior descending artery occlusion for 60 min and reperfusion for 100 minutes. 9 dogs received vehicle and 6 were pretreated with a specific CI. In vivo cardiac microdialysis demonstrated a 3-fold increase in interstitial fluid chymase activity in I/R region that was significantly decreased by CI. CI pretreatment significantly attenuated loss of laminin, focal adhesion complex disruption, and release of troponin I into the circulation. Microarray analysis identified an I/R induced 17-fold increase in nuclear receptor subfamily 4A1 (NR4A1) and significantly decreased by CI. NR4A1 normally resides in the nucleus but can induce cell death on migration to the cytoplasm. I/R caused significant increase in NR4A1 protein expression and cytoplasmic translocation, and mitochondrial degradation, which were decreased by CI. Immunohistochemistry also revealed a high concentration of chymase within cardiomyocytes after I/R. In vitro, chymase added to culture HL-1 cardiomyocytes entered the cytoplasm and nucleus in a dynamin-dependent fashion, and promoted cytoplasmic translocation of NR4A1 protein. shRNA knockdown of NR4A1 on pre-treatment of HL-1 cells with CI significantly decreased chymase-induced cell death and mitochondrial damage. These results suggest that the beneficial effects of an orally active CI during I/R are mediated in the cardiac interstitium as well as within the cardiomyocyte due to a heretofore-unrecognized chymase entry into cardiomyocytes. [ABSTRACT FROM AUTHOR]

Integration of the retrovirus linear DNA genome into the host chromosome is an essential step in the viral replication cycle, and is catalyzed by the viral integrase (IN). Evidence suggests that IN functions as a dimer that cleaves a dinucleotide from the 3′ DNA blunt ends while a dimer of dimers (tetramer) promotes concerted integration of the two processed ends into opposite strands of a target DNA. However, it remains unclear why a dimer rather than a monomer of IN is required for the insertion of each recessed DNA end. To help address this question, we have analyzed crystal structures of the Rous sarcoma virus (RSV) IN mutants complete with all three structural domains as well as its two-domain fragment in a new crystal form at an improved resolution. Combined with earlier structural studies, our results suggest that the RSV IN dimer consists of highly flexible N-terminal domains and a rigid entity formed by the catalytic and C-terminal domains stabilized by the well-conserved catalytic domain dimerization interaction. Biochemical and mutational analyses confirm earlier observations that the catalytic and the C-terminal domains of an RSV IN dimer efficiently integrates one viral DNA end into target DNA. We also show that the asymmetric dimeric interaction between the two C-terminal domains is important for viral DNA binding and subsequent catalysis, including concerted integration. We propose that the asymmetric C-terminal domain dimer serves as a viral DNA binding surface for RSV IN. [ABSTRACT FROM AUTHOR]

We have shown that Dicer processes 7SL RNA into different fragments ranging from ∼20 to more than 200 nucleotides. Here we addressed the molecular functions of these 7SL RNA fragments and found that some of them functioned as dominant-negative regulators of the full-length 7SL RNA, interfering with signal recognition particle (SRP) complex formation. Transfection of these 7SL RNA fragments inhibited the expression of cell surface glycoproteins, the targeting of a reporter protein to the endoplasmic reticulum, and the secretion of secreted alkaline phosphatase. These results suggest that some Dicer-processed 7SL RNA fragments interfered with SRP-mediated protein targeting. Moreover, we showed that Dicer knockdown enhanced SRP-mediated protein targeting and that transfection of a mixture of the 7SL RNA fragments partially restored this effect. Our data indicate that Dicer can fine-tune the efficiency of SRP-mediated protein targeting via processing a proportion of 7SL RNA into fragments of different lengths. [ABSTRACT FROM AUTHOR]

Cyanuric acid hydrolase (CAH) catalyzes the hydrolytic ring-opening of cyanuric acid (2,4,6-trihydroxy-1,3,5-triazine), an intermediate in s-triazine bacterial degradation and a by-product from disinfection with trichloroisocyanuric acid. In the present study, an X-ray crystal structure of the CAH-barbituric acid inhibitor complex from Azorhizobium caulinodans ORS 571 has been determined at 2.7 Å resolution. The CAH protein fold consists of three structurally homologous domains forming a β-barrel-like structure with external α-helices that result in a three-fold symmetry, a dominant feature of the structure and active site that mirrors the three-fold symmetrical shape of the substrate cyanuric acid. The active site structure of CAH is similar to that of the recently determined AtzD with three pairs of active site Ser-Lys dyads. In order to determine the role of each Ser-Lys dyad in catalysis, a mutational study using a highly sensitive, enzyme-coupled assay was conducted. The 10⁹-fold loss of activity by the S226A mutant was at least ten times lower than that of the S79A and S333A mutants. In addition, bioinformatics analysis revealed the Ser226/Lys156 dyad as the only absolutely conserved dyad in the CAH/barbiturase family. These data suggest that Lys156 activates the Ser226 nucleophile which can then attack the substrate carbonyl. Our combination of structural, mutational, and bioinformatics analyses differentiates this study and provides experimental data for mechanistic insights into this unique protein family.

Hairpin telomeres of bacterial linear chromosomes are generated by a DNA cutting-rejoining enzyme protelomerase. Protelomerase resolves a concatenated dimer of chromosomes as the last step of chromosome replication, converting a palindromic DNA sequence at the junctions between chromosomes into covalently closed hairpins. The mechanism by which protelomerase transforms a duplex DNA substrate into the hairpin telomeres remains largely unknown. We report here a series of crystal structures of the protelomerase TelA bound to DNA that represent distinct stages along the reaction pathway. The structures suggest that TelA converts a linear duplex substrate into hairpin turns via a transient strand-refolding intermediate that involves DNA-base flipping and wobble base-pairs. The extremely compact di-nucleotide hairpin structure of the product is fully stabilized by TelA prior to strand ligation, which drives the reaction to completion. The enzyme-catalyzed, multistep strand refolding is a novel mechanism in DNA rearrangement reactions., Competing Interests: The authors have declared that no competing interests exist.

Background and Objectives: Serum alanine aminotransferase (ALT) activity is the most common tool for the assessment of liver diseases. However, it is not clear whether the current normal ALT range really discriminate patients with or without liver diseases. The present study was to establish a new normal range of ALT and examine its ability to identify patients with hepatitis B or nonalcoholic fatty liver disease (NAFLD) in Chinese Han population., Methods: 53037 adults were included in this study from January 1st 2008 to August 31st 2010. The 95th percentile of ALT in population with relative low risk factors for liver diseases was set as the new upper limits of normal ALT in gender-specific manner., Results: The 95(th) percentile levels at low risk factors for liver diseases were achieved at 35 U/L for men and 23 U/L for women. The concordance statistics for detection were 0.873 (95%CI: 0.865-0.881) for HBV and 0.932 (95%CI: 0.927-0.937) for NAFLD in men while 0.857 (95%CI: 0.850-0.864) for HBV and 0.909 (95%CI: 0.903-0.915) for NAFLD in women. The median sensitivity of the current used ALT upper limit (40 U/L) was 6.6% for HBV and 29.7% for NAFLD and median specificity was 98.7% for men and 99.4% for women. Using our new-derived thresholds, the sensitivities ranged from 35.3% to 61.1% and the specificities were 94.8% for men and 94.6% for women., Conclusions: Our results suggest that upper limits of ALT 35 U/L for men and 23 U/L for women in Chinese Han population. Re-consideration of normal limits of ALT should be recommended., Trial Registration: ChiCTR.org ChiCTR-OCS-11001173.

Background: The clinical problem of a "pure volume overload" as in isolated mitral or aortic regurgitation currently has no documented medical therapy that attenuates collagen loss and the resultant left ventricular (LV) dilatation and failure. Here, we identify a potential mechanism related to upregulation of the kallikrein-kinin system in the volume overload of aortocaval fistula (ACF) in the rat., Methodology/principal Findings: LV interstitial fluid (ISF) collection, hemodynamics, and echocardiography were performed in age-matched shams and 4 and 15 wk ACF rats. ACF rats had LV dilatation and a 2-fold increase in LV end-diastolic pressure, along with increases in LV ISF bradykinin, myocardial kallikrein and bradykinin type-2 receptor (BK(2)R) mRNA expression. Mast cell numbers were increased and interstitial collagen was decreased at 4 and 15 wk ACF, despite increases in LV ACE and chymase activities. Treatment with the kallikrein inhibitor aprotinin preserved interstitial collagen, prevented the increase in mast cells, and improved LV systolic function at 4 wk ACF. To establish a cause and effect between ISF bradykinin and mast cell-mediated collagen loss, direct LV interstitial bradykinin infusion in vivo for 24 hrs produced a 2-fold increase in mast cell numbers and a 30% decrease in interstitial collagen, which were prevented by BK(2)R antagonist. To further connect myocardial stretch with cellular kallikrein-kinin system upregulation, 24 hrs cyclic stretch of adult cardiomyocytes and fibroblasts produced increased kallikrein, BK(2)R mRNA expressions, bradykinin protein and gelatinase activity, which were all decreased by the kallikrein inhibitor-aprotinin., Conclusions/significance: A pure volume overload is associated with upregulation of the kallikrein-kinin system and ISF bradykinin, which mediates mast cell infiltration, extracellular matrix loss, and LV dysfunction-all of which are improved by kallikrein inhibition. The current investigation provides important new insights into future potential medical therapies for the volume overload of aortic and mitral regurgitation.