Tal Teitz1, Jennifer J. Stanke1,2, Sara Federico2,3, Cori L. Bradley2, Rachel Brennan2, Jiakun Zhang2, Melissa D. Johnson4, Jan Sedlacik5, Madoka Inoue1, Ziwei M. Zhang4, Sharon Frase6, Jerold E. Rehg7, Claudia M. Hillenbrand5, David Finkelstein8, Christopher Calabrese4, Michael A. Dyer2,9,10, Jill M. Lahti1,11
1 Department of Tumor Cell Biology, St. Jude Children\\\'s Research Hospital, Memphis, Tennessee, United States of America,
2 Department of Developmental Neurobiology, St. Jude Children\\\'s Research Hospital, Memphis, Tennessee, United States of America,
3 Department of Hematology/Oncology, St. Jude Children\\\'s Research Hospital, Memphis, Tennessee, United States of America,
4 Animal Imaging Center, St. Jude Children\\\'s Research Hospital, Memphis, Tennessee, United States of America,
5 Department of Radiological Sciences, St. Jude Children\\\'s Research Hospital, Memphis, Tennessee, United States of America,
6 Cell and Tissue Imaging, St. Jude Children\\\'s Research Hospital, Memphis, Tennessee, United States of America,
7 Department of Pathology, St. Jude Children\\\'s Research Hospital, Memphis, Tennessee, United States of America,
8 Information Sciences, St. Jude Children\\\'s Research Hospital, Memphis, Tennessee, United States of America,
9 Department of Ophthalmology, University of Tennessee Health Science Center, Memphis, Tennessee, United States of America,
10 Howard Hughes Medical Institute, Chevy Chase, Maryland, United States of America,
11 Department of Molecular Sciences, University of Tennessee Health Science Center, Memphis, Tennessee, United States of America
PLoS One. 2011 Apr 29;6(4):e19133
Preclinical models of pediatric cancers are essential for testing new chemotherapeutic combinations for clinical trials. The most widely used genetic model for preclinical testing of neuroblastoma is the TH-MYCN mouse. This neuroblastoma-prone mouse recapitulates many of the features of human neuroblastoma. Limitations of this model include the low frequency of bone marrow metastasis, the lack of information on whether the gene expression patterns in this system parallels human neuroblastomas, the relatively slow rate of tumor formation and variability in tumor penetrance on different genetic backgrounds. As an alternative, preclinical studies are frequently performed using human cell lines xenografted into immunocompromised mice, either as flank implant or orthtotopically. Drawbacks of this system include the use of cell lines that have been in culture for years, the inappropriate microenvironment of the flank or difficult, time consuming surgery for orthotopic transplants and the absence of an intact immune system.
Here we characterize and optimize both systems to increase their utility for preclinical studies. We show that TH-MYCN mice develop tumors in the paraspinal ganglia, but not in the adrenal, with cellular and gene expression patterns similar to human NB. In addition, we present a new ultrasound guided, minimally invasive orthotopic xenograft method. This injection technique is rapid, provides accurate targeting of the injected cells and leads to efficient engraftment. We also demonstrate that tumors can be detected, monitored and quantified prior to visualization using ultrasound, MRI and bioluminescence. Finally we develop and test a \\\\\\\"standard of care\\\\\\\" chemotherapy regimen. This protocol, which is based on current treatments for neuroblastoma, provides a baseline for comparison of new therapeutic agents.
The studies suggest that use of both the TH-NMYC model of neuroblastoma and the orthotopic xenograft model provide the optimal combination for testing new chemotherapies for this devastating childhood cancer.
Amanda G. Linkous, Eugenia M. Yazlovitskaya, Dennis E. Hallahan
Neuro-Oncology Branch, National Institutes of Health, Bethesda, MD (AGL); Division of Nephrology, Department of Medicine, and Vanderbilt-Ingram Cancer Center, Vanderbilt University, Nashville, TN (EMY); Alvin Siteman Cancer Center and Department of Radiation Oncology, Washington University in St Louis, St Louis, MO (DEH)
Journal of the National Cancer Institute. 2010 Aug 20. [Epub ahead of print]
Lung cancer and glioblastoma multiforme are highly angiogenic and, despite advances in treatment, remain resistant to therapy. Cytosolic phospholipase A2 (cPLA2) activation contributes to treatment resistance through transduction of prosurvival signals. We investigated cPLA2 as a novel molecular target for antiangiogenesis therapy.
Glioblastoma (GL261) and Lewis lung carcinoma (LLC) heterotopic tumor models were used to study the effects of cPLA2 expression on tumor growth and vascularity in C57/BL6 mice wild type for (cPLA2α+/+) or deficient in (cPLA2α–/–) cPLA2α, the predominant isoform in endothelium (n = 6–7 mice per group). The effect of inhibiting cPLA2 activity on GL261 and LLC tumor growth was studied in mice treated with the chemical cPLA2 inhibitor 4-[2-[5-chloro-1-(diphenylmethyl)-2-methyl-1H-indol-3-yl]-ethoxy]benzoic acid (CDIBA). Endothelial cell proliferation and function were evaluated by Ki-67 immunofluorescence and migration assays in primary cultures of murine pulmonary microvascular endothelial cells (MPMEC) isolated from cPLA2α+/+ and cPLA2α–/– mice. Proliferation, invasive migration, and tubule formation were assayed in mouse vascular endothelial 3B-11 cells treated with CDIBA. Effects of lysophosphatidylcholine, arachidonic acid, and lysophosphatidic acid (lipid mediators of tumorigenesis and angiogenesis) on proliferation and migration were examined in 3B-11 cells and cPLA2α–/– MPMEC. All statistical tests were two-sided.
GL261 tumor progression proceeded normally in cPLA2α+/+ mice, whereas no GL261 tumors formed in cPLA2α–/– mice. In the LLC tumor model, spontaneous tumor regression was observed in 50% of cPLA2α–/– mice. Immunohistochemical examination of the remaining tumors from cPLA2α–/– mice revealed attenuated vascularity (P .001) compared with tumors from cPLA2α+/+ mice. Inhibition of cPLA2 activity by CDIBA resulted in a delay in tumor growth (eg, LLC model: average number of days to reach tumor volume of 700 mm3, CDIBA vs vehicle: 16.8 vs 11.8, difference = 5, 95% confidence interval = 3.6 to 6.4, P = .04) and a decrease in tumor size (eg, GL261 model: mean volume on day 21, CDIBA vs vehicle: 40.1 vs 247.4 mm3, difference = 207.3 mm3, 95% confidence interval = 20.9 to 293.7 mm3, P = .021). cPLA2 deficiency statistically significantly reduced MPMEC proliferation and invasive migration (P = .002 and P = .004, respectively). Compared with untreated cells, cPLA2α–/– MPMEC treated with lysophosphatidylcholine and lysophosphatidic acid displayed increased cell proliferation (P = .011) and invasive migration (P < .001).
In these mouse models of brain and lung cancer, cPLA2 and lysophospholipids have key regulatory roles in tumor angiogenesis. cPLA2 inhibition may be a novel effective antiangiogenic therapy.
James P.B. O'Connor1,2, Richard A.D. Carano3, Andrew R. Clamp2, Jed Ross3, Calvin C.K. Ho3, Alan Jackson1, Geoff J.M. Parker1, Chris J. Rose1, Franklin V. Peale4, Michel Friesenhahn5, Claire L. Mitchell1,2, Yvonne Watson1, Caleb Roberts1, Lynn Hope2, Sue Cheung1, Hani Bou Reslan3, Mary Ann T. Go6, Glenn J. Pacheco6, Xiumin Wu7, Tim C. Cao3, Sarajane Ross6, Giovanni A. Buonaccorsi1, Karen Davies1, Jurjees Hasan2, Paula Thornton2, Olivia del Puerto8, Napoleone Ferrara7, Nicholas van Bruggen3 and Gordon C. Jayson2
1 Imaging Science and Biomedical Engineering, School of Cancer and Imaging Sciences, University of Manchester and
2 Cancer Research UK and University of Manchester Department of Medical Oncology, Christie Hospital, Manchester, United Kingdom
3 Biomedical Imaging Group, Department of Tumor Biology and Angiogenesis,
6 Translational Oncology and
7 Ferrara Laboratory, Genentech, Inc., South San Francisco, California; and
8 Roche Products Limited, Hertfordshire, United Kingdom
Clinical Cancer Research. 2009 Nov 2; 15(21):667482
Little is known concerning the onset, duration, and magnitude of direct therapeutic effects of antivascular endothelial growth factor (VEGF) therapies. Such knowledge would help guide the rational development of targeted therapeutics from bench to bedside and optimize use of imaging technologies that quantify tumor function in early-phase clinical trials.
Preclinical studies were done using ex vivo microcomputed tomography and in vivo ultrasound imaging to characterize tumor vasculature in a human HM-7 colorectal xenograft model treated with the anti-VEGF antibody G6-31. Clinical evaluation was by quantitative magnetic resonance imaging in 10 patients with metastatic colorectal cancer treated with bevacizumab.
Microcomputed tomography experiments showed reduction in perfused vessels within 24 to 48 h of G6-31 drug administration (P ≤ 0.005). Ultrasound imaging confirmed reduced tumor blood volume within the same time frame (P = 0.048). Consistent with the preclinical results, reductions in enhancing fraction and fractional plasma volume were detected in patient colorectal cancer metastases within 48 h after a single dose of bevacizumab that persisted throughout one cycle of therapy. These effects were followed by resolution of edema (P = 0.0023) and tumor shrinkage in 9 of 26 tumors at day 12.
These data suggest that VEGF-specific inhibition induces rapid structural and functional effects with downstream significant antitumor activity within one cycle of therapy. This finding has important implications for the design of early-phase clinical trials that incorporate physiologic imaging. The study shows how animal data help interpret clinical imaging data, an important step toward the validation of image biomarkers of tumor structure and function.
angiogenesis, biomarker, imaging, VEGF
Mary E. Loveless, MS, Xia Li, PhD, Jessica Huamani, MS, Andrej Lyshchik, MD, PhD, Benoit Dawant, PhD, Dennis Hallahan, MD, John C. Gore, PhD, Thomas E. Yankeelov, PhD
Institute of Imaging Science (M.E.L., X.L., B.D., J.C.G., T.E.Y.) and Departments of Biomedical Engineering (M.E.L., D.H., J.C.G., T.E.Y.), Electrical Engineering and Computer Science (X.L., B.D.), Radiation Oncology (J.H., D.H.), Radiology and Radiological Sciences (A.L., J.C.G., T.E.Y.), Physics and Astronomy (J.C.G., T.E.Y.), and Cancer Biology (T.E.Y.), Vanderbilt University, Nashville, Tennessee USA
Journal of ultrasound in medicine. 2008 Dec; 27(12):1699-709
The purpose of this study was to develop a method for assessing tumor vascularity in a preclinical model of breast cancer using contrast-enhanced ultrasonography.
Eight mice were injected with 67NR breast cancer cells on their hind limbs and imaged with ultrasonography 8 days later. Mice were injected with an ultrasound contrast agent (UCA), and a sequence of images of the resultant backscattered echoes was recorded before and after high-power \"destruction\" pulses for each of multiple parallel planes. From these, data maps of the maximum contrast enhancement (within each time course) were constructed for each pixel, which enabled reconstruction of high-resolution coregistered sections into a 3-dimensional (3D) volume reflecting tumor vascularity. Additional studies were performed to determine the duration and repeatability of image enhancement, and images were correlated with conventional 3D power Doppler measurements.
The lifetime of the UCA in vivo was found to be 4.3 +/- 1.09 minutes (mean +/- SD). The 3D contrast-enhanced ultrasonographic technique produced images that correlated well with power Doppler images in specific regions but also depicted additional regions of flow surrounding the power Doppler signal. The mean correlation coefficient between voxel measurements of the central slice for each animal was 0.64 +/- 0.07 (P < .01). In addition, sequential studies in each animal were reproducible.
A method producing high-resolution volumetric assessments of tumor vascularity in a preclinical model of breast cancer is shown that correlates with other ultrasonographic measures of blood flow, which may provide greater sensitivity to the microvasculature.
Yan Ding, Elissa A. Boguslawski, Bree D. Berghuis, John J. Young, Zhongfa Zhang, Kim Hardy, Kyle Furge, Eric Kort, Arthur E. Frankel, Rick V. Hay, James H. Resau and Nicholas S. Duesbery
Laboratories of Cancer and Developmental Cell Biology, Analytical, Cellular and Molecular Microscopy, Cancer Genetics, Noninvasive Imaging and Radiation Biology, and Computational Biology, Van Andel Research Institute, Grand Rapids, Michigan and 6 Cancer Research Institute of Scott & White Memorial Hospital, Temple, Texas
Molecular Cancer Therapeutics. 2008 March 1;7: 648-658.
We hypothesized that signaling through multiple mitogen-activated protein kinase (MAPK) kinase (MKK) pathways is essential for the growth and vascularization of soft-tissue sarcomas, which are malignant tumors derived from mesenchymal tissues. We tested this using HT-1080, NCI, and Shac fibrosarcoma-derived cell lines and anthrax lethal toxin (LeTx), a bacterial toxin that inactivates MKKs. Western blots confirmed that LeTx treatment reduced the levels of phosphorylated extracellular signal-regulated kinase and p38 MAPK in vitro. Although short treatments with LeTx only modestly affected cell proliferation, sustained treatment markedly reduced cell numbers. LeTx also substantially inhibited the extracellular release of angioproliferative factors including vascular endothelial growth factor, interleukin-8, and basic fibroblast growth factor. Similar results were obtained with cell lines derived from malignant fibrous histiocytomas, leiomyosarcomas, and liposarcomas. In vivo, LeTx decreased MAPK activity and blocked fibrosarcoma growth. Growth inhibition correlated with decreased cellular proliferation and extensive necrosis, and it was accompanied by a decrease in tumor mean vessel density as well as a reduction in serum expression of angioproliferative cytokines. Vital imaging using high-resolution ultrasound enhanced with contrast microbubbles revealed that the effects of LeTx on tumor perfusion were remarkably rapid (<24 h) and resulted in a marked reduction of perfusion within the tumor but not in nontumor tissues. These results are consistent with our initial hypothesis and lead us to propose that MKK inhibition by LeTx is a broadly effective strategy for targeting neovascularization in fibrosarcomas and other similar proliferative lesions.
Jiri Neuzil, Marco Tomasetti, Yan Zhao, Lan-Feng Dong, Marc Birringer, Xiu-Fang Wang, Pauline Low, Kun Wu, Brian A. Salvatore, and Steven J. Ralph
Apoptosis Research Group, School of Medical Science, Griffith University, Southport, Queensland, Australia (J.N., L.F.D., X.F.W.); Molecular Therapy Group, Institute of Molecular Genetics, Czech Academy of Sciences, Prague, Czech Republic (J.N.); Department of Nutrition and Food, Harbin Medical University, Harbin, Heilongjiang Province, China (Y.Z., K.W.); Department of Molecular Pathology and Innovative Therapies, Polytechnic University of Marche, Ancona, Italy (M.T.); Institute of Human Nutrition, Friedrich Schiller University, Jena, Germany (M.B.); Genomic Research Centre, School of Medical Science, Griffith University, Southport, Queensland, Australia (P.L., S.J.R.); and Department of Chemistry and Physics, Louisiana State University, Shreveport, Louisiana (B.A.S.)
Molecular Pharmacology. 2007 May;71(5):1185-99. Epub 2007 Jan 12
The search for a selective and efficient anticancer agent for treating all neoplastic disease has yet to deliver a universally suitable compound(s). The majority of established anticancer drugs either are nonselective or lose their efficacy because of the constant mutational changes of malignant cells. Until recently, a largely neglected target for potential anticancer agents was the mitochondrion, showing a considerable promise for future clinical applications. Vitamin E (VE) analogs, epitomized by alpha-tocopheryl succinate, belong to the group of \"mitocans\" (mitochondrially targeted anticancer drugs). They are selective for malignant cells, cause destabilization of their mitochondria, and suppress cancer in preclinical models. This review focuses on our current understanding of VE analogs in the context of their proapoptotic/anticancer efficacy and suggests that their effect on mitochondria may be amplified by modulation of alternative pathways operating in parallel. We show here that the analogs of VE that cause apoptosis (which translates into their anticancer efficacy) generally do not possess antioxidant (redox) activity and are prototypical of the mitocan group of anticancer compounds. Therefore, by analogy to Oscar Wilde\'s play The Importance of Being Earnest, we use the motto in the title \"the importance of being redox-silent\" to emphasize an essentially novel paradigm for cancer therapy, in which redox-silence is a prerequisite property for most of the anticancer activities described in this communication.
Pezold JC, Zinn K, Talbert MA, Desmond R, Rosenthal EL.
Department of Surgery, Division of Otolaryngology - Head and Neck Surgery, University of Alabama at Birmingham, Birmingham, Ala., USA.
ORL J Otorhinolaryngol Relat Spec.2006 Apr; 68(3):159-63. Epub 2006 Feb 6
Background: The murine orthotopic oral cavity tumor model allows evaluation of tumor growth and invasion. Currently, serial measurements of tissue growth are difficult to obtain since invasive procedures or animal sacrifice is necessary to evaluate tumor size. High-resolution ultrasound (Vevo 660, VisualSonics) was evaluated as a noninvasive method to monitor tumor size in vivo.
Methods: Sixteen immunodeficient mice, age 9 weeks, were injected transcervically with a human squamous cell carcinoma cell line into the tongue, and tumor volume was assessed by high-frequency ultrasound at 11 days. The animals were subsequently sacrificed and the tumors processed for histology. Tumor size was then calculated by caliper measurement in two dimensions.
Results: Tumor dimensions obtained using ultrasound were found to significantly correlate with the histologic measurements (Spearman coefficient 0.90, p < 0.0001). Tumor dimensions were on average larger using ultrasound versus histologic measurements, although this was not significantly different than zero (95% confidence interval -13.96 to 62.37 mm(2)).
Conclusions: High-resolution ultrasound accurately measures tumor volume in the murine orthotopic oral cavity tumor model without sacrifice.
Soban Umar, MD, PhD, Jong-Hwan Lee, MD, PhD, Enno de Lange, PhD, Andrea Iorga, BSc, Rod Partow-Navid, BSc, Aneesh Bapat, BSc, Arnoud van der Laarse, PhD, Rajeev Saggar, MD, Rajan Saggar, MD, Dirk L. Ypey, PhD, Hrayr S. Karagueuzian, PhD, FACC and Mansoureh Eghbali, PhD
From the Departments of Anesthesiology (S.U., A.I., R.P.-N., M.E.), Medicine (J.-H.L., E.d.L., A.B., Rajeev Saggar, Rajan Saggar, H.S.K.), the Division of Pulmonary and Critical Care Medicine (Rajeev Saggar, Rajan Saggar), and Cardiovascular Research Laboratories (S.U., J.-H.L., E.d.L., A.I., R.P.-N., A.B., H.S.K., M.E.), David Geffen School of Medicine at UCLA, Los Angeles, CA; the Department of Anesthesiology and Pain Medicine, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea (J.-H.L.); and the Department of Cardiology, Leiden University Medical Center, Leiden, The Netherlands (A.v.d.L., D.L.Y.)
Circulation: Arrhythmia and Electrophysiology 2012 Feb 1;5(1):181-90. Epub 2011 Dec 22
Right ventricular failure (RVF) in pulmonary hypertension (PH) is associated with increased incidence of sudden death by a poorly explored mechanism. We test the hypothesis that PH promotes spontaneous ventricular fibrillation (VF) during a critical post-PH onset period characterized by a sudden increase in mortality.
METHODS AND RESULTS:
Rats received either a single subcutaneous dose of monocrotaline (MCT, 60 mg/kg) to induce PH-associated RVF (PH, n=24) or saline (control, n=17). Activation pattern of the RV-epicardial surface was mapped using voltage-sensitive dye in isolated Langendorff-perfused hearts along with single glass-microelectrode and ECG-recordings. MCT-injected rats developed severe PH by day 21 and progressed to RVF by approximately day 30. Rats manifested increased mortality, and ≈30% rats died suddenly and precipitously during 23–32 days after MCT. This fatal period was associated with the initiation of spontaneous VF by a focal mechanism in the RV, which was subsequently maintained by both focal and incomplete reentrant wave fronts. Microelectrode recordings from the RV-epicardium at the onset of focal activity showed early afterdepolarization–mediated triggered activity that led to VF. The onset of the RV cellular triggered beats preceded left ventricular depolarizations by 23±8 ms. The RV but not the left ventricular cardiomyocytes isolated during this fatal period manifested significant action potential duration prolongation, dispersion, and an increased susceptibility to depolarization-induced repetitive activity. No spontaneous VF was observed in any of the control hearts. RVF was associated with significantly reduced RV ejection fraction (P<0.001), RV hypertrophy (P<0.001), and RV fibrosis (P<0.01). The hemodynamic function of the LV and its structure were preserved.
PH-induced RVF is associated with a distinct phase of increased mortality characterized by spontaneous VF arising from the RV by an early afterdepolarization–mediated triggered activity.
pulmonary hypertension, right ventricular failure, early afterdepolarization, optical mapping, ventricular fibrillation
Eleonora Mezzaroma1,2,3, Stefano Toldo1,2, Daniela Farkas2, Ignacio M. Seropian1,2,3, Benjamin W. Van Tassell2,3, Fadi N. Salloum1, Harsha R. Kannan1,2, Angela C. Menna1,2, Norbert F. Voelkel1,2, and Antonio Abbate1,2
1 VCU Pauley Heart Center,
2 VCU Victoria Johnson Center, and
3 School of Pharmacy, Virginia Commonwealth University, Richmond, VA 23298
Proceedings of the National Academy of Sciences of the United States of America (PNAS). 2011 Dec 6;108(49):19725-30. Epub 2011 Nov 21
Acute myocardial infarction (AMI) initiates an intense inflammatory response that promotes cardiac dysfunction, cell death, and ventricular remodeling. The molecular events underlying this inflammatory response, however, are incompletely understood. In experimental models of sterile inflammation, ATP released from dying cells triggers, through activation of the purinergic P2X7 receptor, the formation of the inflammasome, a multiprotein complex necessary for caspase-1 activation and amplification of the inflammatory response. Here we describe the presence of the inflammasome in the heart in an experimental mouse model of AMI as evidenced by increased caspase-1 activity and cytoplasmic aggregates of the three components of the inflammasome-apoptosis speck-like protein containing a caspase-recruitment domain (ASC), cryopyrin, and caspase-1, localized to the granulation tissue and cardiomyocytes bordering the infarct. Cultured adult murine cardiomyocytes also showed the inducible formation of the inflammasome associated with increased cell death. P2X7 and cryopyrin inhibition (using silencing RNA or a pharmacologic inhibitor) prevented the formation of the inflammasome and limited infarct size and cardiac enlargement after AMI. The formation of the inflammasome in the mouse heart during AMI causes additional loss of functional myocardium, leading to heart failure. Modulation of the inflammasome may therefore represent a unique therapeutic strategy to limit cell death and prevent heart failure after AMI.
interleukin-1, NALP3, NLRP3, pyroptosis
Naohiro Yamaguchi1, Asima Chakraborty1, Daniel A. Pasek1, Jeffery D. Molkentin2, and Gerhard Meissner1
1 Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill, North Carolina; and
2 Division of Molecular Cardiovascular Biology, Cincinnati Children\'s Hospital Medical Center, University of Cincinnati, Cincinnati, Ohio
American Journal of Physiology. Heart and Circulatory Physiology. 2011 Jun;300(6):H2187-95. Epub 2011 Mar 18
Mice with three amino acid mutations in the calmodulin binding domain of type-2 ryanodine receptor ion channel (Ryr2(ADA/ADA) mice) have impaired intracellular Ca(2+) handling and cardiac hypertrophy with death at an early age. In this report, the role of signaling molecules implicated in cardiac hypertrophy of Ryr2(ADA/ADA) mice was investigated. Calcineurin A-ß (CNA-ß) and nuclear factor of activated T cell (NFAT) signaling were monitored in mice carrying either luciferase transgene driven by NFAT-dependent promoter or knockout of CNA-ß. NFAT transcriptional activity in Ryr2(ADA/ADA) hearts was not markedly upregulated at embryonic day 16.5 compared with wild-type but significantly increased at postnatal days 1 and 10. Ablation of CNA-ß extended the life span of Ryr2(ADA/ADA) mice and enhanced cardiac function without improving sarcoplasmic reticulum Ca(2+) handling or suppressing the expression of genes implicated in cardiac hypertrophy. Embryonic day 16.5 Ryr2(ADA/ADA) mice had normal heart weights with no major changes in Akt1 and class II histone deacetylase phosphorylation and myocyte enhancer factor-2 activity. In contrast, phosphorylation levels of Erk1/2, p90 ribosomal S6 kinases (p90RSKs), and GSK-3ß were increased in hearts of embryonic day 16.5 homozygous mutant mice. The results indicate that an impaired calmodulin regulation of RyR2 was neither associated with an altered CNA-ß/NFAT, class II histone deacetylase (HDAC)/MEF2, nor Akt signaling in embryonic day 16.5 hearts; rather increased Erk1/2 and p90RSK phosphorylation levels likely leading to reduced GSK-3ß activity were found to precede development of cardiac hypertrophy in mice expressing dysfunctional ryanodine receptor ion channel.
KEYWORDS: calmodulin, calcineurin/nuclear factor of activated T cell, class II histone deacetylases/myocyte enhancer factor 2, extracellular signal-regulated kinase, glycogen synthetase kinase 3