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High grade gliomas are malignant brain tumors that arise in the central nervous system, in patients of all ages. Currently, the standard of care, entailing surgery and chemo radiation, exhibits a survival rate of 14-17 months. Thus, there is an urgent need to develop new therapeutic strategies for these malignant brain tumors. Currently, immunotherapies represent an appealing approach to treat malignant gliomas, as the pre-clinical data has been encouraging. However, the translation of the discoveries from the bench to the bedside has not been as successful as with other types of cancer, and no long-lasting clinical benefits have been observed for glioma patients treated with immune-mediated therapies so far. This review aims to discuss our current knowledge about gliomas, their molecular particularities and the impact on the tumor immune microenvironment. Also, we discuss several murine models used to study these therapies pre-clinically and how the model selection can impact the outcomes of the approaches to be tested. Finally, we present different immunotherapy strategies being employed in clinical trials for glioma and the newest developments intended to harness the immune system against these incurable brain tumors.

Gliomas are one of the most lethal types of cancers accounting for ∼80% of all central nervous system (CNS) primary malignancies. Among gliomas, glioblastomas (GBM) are the most aggressive, characterized by a median patient survival of fewer than 15 months. Recent molecular characterization studies uncovered the genetic signatures and methylation status of gliomas and correlate these with clinical prognosis. The most relevant molecular characteristics for the new glioma classification are IDH mutation, chromosome 1p/19q deletion, histone mutations, and other genetic parameters such as ATRX loss, TP53, and TERT mutations, as well as DNA methylation levels. Similar to other solid tumors, glioma progression is impacted by the complex interactions between the tumor cells and immune cells within the tumor microenvironment. The immune system’s response to cancer can impact the glioma’s survival, proliferation, and invasiveness. Salient characteristics of gliomas include enhanced vascularization, stimulation of a hypoxic tumor microenvironment, increased oxidative stress, and an immune suppressive milieu. These processes promote the neuro-inflammatory tumor microenvironment which can lead to the loss of blood-brain barrier (BBB) integrity. The consequences of a compromised BBB are deleteriously exposing the brain to potentially harmful concentrations of substances from the peripheral circulation, adversely affecting neuronal signaling, and abnormal immune cell infiltration; all of which can lead to disruption of brain homeostasis. In this review, we first describe the unique features of inflammation in CNS tumors. We then discuss the mechanisms of tumor-initiating neuro-inflammatory microenvironment and its impact on tumor invasion and progression. Finally, we also discuss potential pharmacological interventions that can be used to target neuro-inflammation in gliomas.

Glioblastoma (GBM) is the most common and aggressive primary brain tumor in the adult population and it carries a dismal prognosis. Inefficient drug delivery across the blood brain barrier (BBB), an immunosuppressive tumor microenvironment (TME) and development of drug resistance are key barriers to successful glioma treatment. Since gliomas occur through sequential acquisition of genetic alterations, gene therapy, which enables to modification of the genetic make-up of target cells, appears to be a promising approach to overcome the obstacles encountered by current therapeutic strategies. Gene therapy is a rapidly evolving field with the ultimate goal of achieving specific delivery of therapeutic molecules using either viral or non-viral delivery vehicles. Gene therapy can also be used to enhance immune responses to tumor antigens, reprogram the TME aiming at blocking glioma-mediated immunosuppression and normalize angiogenesis. Nano-particles-mediated gene therapy is currently being developed to overcome the BBB for glioma treatment. Another approach to enhance the anti-glioma efficacy is the implementation of viro-immunotherapy using oncolytic viruses, which are immunogenic. Oncolytic viruses kill tumor cells due to cancer cell-specific viral replication, and can also initiate an anti-tumor immunity. However, concerns still remain related to off target effects, and therapeutic and transduction efficiency. In this review, we describe the rationale and strategies as well as advantages and disadvantages of current gene therapy approaches against gliomas in clinical and preclinical studies. This includes different delivery systems comprising of viral, and non-viral delivery platforms along with suicide/prodrug, oncolytic, cytokine, and tumor suppressor-mediated gene therapy approaches. In addition, advances in glioma treatment through BBB-disruptive gene therapy and anti-EGFRvIII/VEGFR gene therapy are also discussed. Finally, we discuss the results of gene therapy-mediated human clinical trials for gliomas. In summary, we highlight the progress, prospects and remaining challenges of gene therapies aiming at broadening our understanding and highlighting the therapeutic arsenal for GBM.

Background Gliomas are the most common primary brain tumors. High-Grade Gliomas have a median survival (MS) of 18 months, while Low-Grade Gliomas (LGGs) have an MS of approximately 7.3 years. Seventy-six percent of patients with LGG express mutated isocitrate dehydrogenase (mIDH) enzyme. Survival of these patients ranges from 1 to 15 years, and tumor mutational burden ranges from 0.28 to 3.85 somatic mutations/megabase per tumor. We tested the hypothesis that the tumor mutational burden would predict the survival of patients with tumors bearing mIDH. Methods We analyzed the effect of tumor mutational burden on patients’ survival using clinical and genomic data of 1199 glioma patients from The Cancer Genome Atlas and validated our results using the Glioma Longitudinal AnalySiS consortium. Results High tumor mutational burden negatively correlates with the survival of patients with LGG harboring mIDH (P = .005). This effect was significant for both Oligodendroglioma (LGG-mIDH-O; MS = 2379 vs 4459 days in high vs low, respectively; P = .005) and Astrocytoma (LGG-mIDH-A; MS = 2286 vs 4412 days in high vs low respectively; P = .005). There was no differential representation of frequently mutated genes (eg, TP53, ATRX, CIC, and FUBP) in either group. Gene set enrichment analysis revealed an enrichment in Gene Ontologies related to cell cycle, DNA-damage response in high versus low tumor mutational burden. Finally, we identified 6 gene sets that predict survival for LGG-mIDH-A and LGG-mIDH-O. Conclusions we demonstrate that tumor mutational burden is a powerful, robust, and clinically relevant prognostic factor of MS in mIDH patients.

Gliomas are infiltrating brain tumors associated with high morbidity and mortality. Current standard of care includes radiation, chemotherapy and surgical resection. Today, survival rates for malignant glioma patients remain dismal and unchanged for decades. The glioma microenvironment is highly immunosuppressive and consequently this has motivated the development of immunotherapies for counteracting this condition, enabling the immune cells within the tumor microenvironment to react against this tumor.Areas covered: The authors discuss immunotherapeutic strategies for glioma in phase-I/II clinical trials and illuminate their mechanisms of action, limitations and key challenges. They also examine promising approaches under preclinical development.Expert opinion: In the last decade there has been an expansion in immune-mediated anti-cancer therapies. In the glioma field, sophisticated strategies have been successfully implemented in preclinical models. Unfortunately, clinical trials have not yet yielded consistent results for glioma patients. This could be attributed to our limited understanding of the complex immune cell infiltration and its interaction with the tumor cells, the selected time for treatment, the combination with other therapies and the route of administration of the agent. Applying these modalities to treat malignant glioma is challenging, but many new alternatives are emerging to by-pass these hurdles. Fil: Garcia Fabiani, Maria Belen. University of Michigan; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Ventosa, Maria. University of Michigan; Estados Unidos Fil: Comba, Andrea. University of Michigan; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Candolfi, Marianela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; Argentina Fil: Nicola Candia, Alejandro Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; Argentina Fil: Alghamri, Mahmoud S.. University of Michigan; Estados Unidos Fil: Kadiyala, Padma. University of Michigan; Estados Unidos Fil: Carney, Stephen. University of Michigan; Estados Unidos Fil: Faisal, Syed M.. University of Michigan; Estados Unidos Fil: Schwendeman, Anna. University of Michigan; Estados Unidos Fil: Moon, James J.. University of Michigan; Estados Unidos Fil: Scheetz, Lindsay. University of Michigan; Estados Unidos Fil: Lahann, Joerg. University of Michigan; Estados Unidos Fil: Mauser, Ava. University of Michigan; Estados Unidos Fil: Lowenstein, Pedro R.. University of Michigan; Estados Unidos Fil: Castro, Maria Gabriela. University of Michigan; Estados Unidos

Mutant isocitrate-dehydrogenase-1 (IDH1-R132H; mIDH1) is a hallmark of adult gliomas. Lower grade mIDH1 gliomas are classified into two molecular subgroups: (i) 1p/19q co-deletion/TERT-promoter mutations or (ii) inactivating mutations in α-thalassemia/mental retardation syndrome X-linked (ATRX) andTP53.This work, relates to the gliomas’ subtype harboring mIDH1,TP53andATRXinactivation. IDH1-R132H is a gain-of-function mutation that converts α-ketoglutarate into 2-hydroxyglutarate (D-2HG). The role of D-2HG within the tumor microenvironment of mIDH1/mATRX/mTP53 gliomas remains unexplored. Inhibition of 2HG, when used as monotherapy or in combination with radiation and temozolomide (IR/TMZ), led to increased median survival (MS) of mIDH1 glioma bearing mice. Also, 2HG inhibition elicited anti-mIDH1 glioma immunological memory. In response to 2HG inhibition, PD-L1 expression levels on mIDH1-glioma cells increased to similar levels as observed in wild-type-IDH1 gliomas. Thus, we combined 2HG inhibition/IR/TMZ with anti-PDL1 immune checkpoint-blockade and observed complete tumor regression in 60% of mIDH1 glioma bearing mice. This combination strategy reduced T-cell exhaustion and favored the generation of memory CD8+T-cells. Our findings demonstrate that metabolic reprogramming elicits anti-mIDH1 glioma immunity, leading to increased MS and immunological memory. Our preclinical data supports the testing of IDH-R132H inhibitors in combination with IR/TMZ and anti-PDL1 as targeted therapy for mIDH1/mATRX/mTP53 glioma patients.Brief SummaryInhibition of 2-Hydroxyglutrate in mutant-IDH1 glioma in the genetic context of ATRX and TP53 inactivation elicits metabolic-reprograming and anti-glioma immunity.

Mutant isocitrate dehydrogenase 1 (IDH1-R132H; mIDH1) is a hallmark of adult gliomas. Lower grade mIDH1 gliomas are classified into 2 molecular subgroups: 1p/19q codeletion/TERT-promoter mutations or inactivating mutations in α-thalassemia/mental retardation syndrome X-linked (ATRX) and TP53. This work focuses on glioma subtypes harboring mIDH1, TP53, and ATRX inactivation. IDH1-R132H is a gain-of-function mutation that converts α-ketoglutarate into 2-hydroxyglutarate (D-2HG). The role of D-2HG within the tumor microenvironment of mIDH1/mATRX/mTP53 gliomas remains unexplored. Inhibition of D-2HG, when used as monotherapy or in combination with radiation and temozolomide (IR/TMZ), led to increased median survival (MS) of mIDH1 glioma–bearing mice. Also, D-2HG inhibition elicited anti–mIDH1 glioma immunological memory. In response to D-2HG inhibition, PD-L1 expression levels on mIDH1-glioma cells increased to similar levels as observed in WT-IDH gliomas. Thus, we combined D-2HG inhibition/IR/TMZ with anti–PDL1 immune checkpoint blockade and observed complete tumor regression in 60% of mIDH1 glioma–bearing mice. This combination strategy reduced T cell exhaustion and favored the generation of memory CD8(+) T cells. Our findings demonstrate that metabolic reprogramming elicits anti–mIDH1 glioma immunity, leading to increased MS and immunological memory. Our preclinical data support the testing of IDH-R132H inhibitors in combination with IR/TMZ and anti-PDL1 as targeted therapy for mIDH1/mATRX/mTP53 glioma patients.

Gliomas are the most common primary brain tumors. High Grade Gliomas have a median survival of 18 months, while Low Grade Gliomas (LGG) have a median survival of ∼7.3 years. Seventy-six percent of patients with LGG express mutated isocitrate dehydrogenase (mIDH1) enzyme (IDH1R132H). Survival of these patients ranges from 1-15 years, and tumor mutational burden ranges from 8 to 447 total somatic mutations per tumor. We tested the hypothesis that the tumor mutational burden would predict survival of patients with tumors bearing mIDH1R132H. We analyzed the effect of tumor mutational burden on patients’ survival using clinical and genomic data of 1199 glioma patients from The Cancer Genome Atlas and validated our results using the Chinese Glioma Genome Atlas. High tumor mutational burden negatively correlates with survival of patients with LGG harboring IDH1R132H (pmIDH1 patients. In the TCGA data, median survival of the high mutational burden group was 76 months, while in the low mutational burden group it was 136 months; pTP53, ATRX, CIC, FUBP) in either group. Gene set enrichment analysis revealed an enrichment in Gene Ontologies related to Cell cycle, DNA damage response in high vs low tumor mutational burden. Finally, we identified a 19 gene signature that predicts survival for patients from both databases. In summary, we demonstrate that tumor mutational burden is a powerful, robust, and clinically relevant prognostic factor of median survival in mIDH1 patients.

Introduction: The field of neuro-oncology has experienced significant advances in recent years. More is known now about the molecular and genetic characteristics of glioma than ever before. This knowledge leads to the understanding of glioma biology and pathogenesis, guiding the development of targeted therapeutics and clinical trials. The goal of this review is to describe the state of basic, translational, and clinical research as it pertains to biological and synthetic pharmacotherapy for gliomas. Areas covered: Challenges remain in designing accurate preclinical models and identifying patients that are likely to respond to a particular targeted therapy. Preclinical models for therapeutic assessment are critical to identify the most promising treatment approaches. Expert opinion: Despite promising new therapeutics, there have been no significant breakthroughs in glioma treatment and patient outcomes. Thus, there is an urgent need to better understand the mechanisms of treatment resistance and to design effective clinical trials. Fil: Altshuler, David B.. University of Michigan; Estados Unidos Fil: Kadiyala, Padma. University of Michigan; Estados Unidos Fil: Nuñez, Felipe J.. University of Michigan; Estados Unidos Fil: Nuñez, Fernando M.. University of Michigan; Estados Unidos Fil: Carney, Stephen. University of Michigan; Estados Unidos Fil: Alghamri, Mahmoud S.. University of Michigan; Estados Unidos Fil: Garcia Fabiani, Maria Belen. University of Michigan; Estados Unidos Fil: Asad, Antonela Sofía. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; Argentina Fil: Nicola Candia, Alejandro Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; Argentina Fil: Candolfi, Marianela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; Argentina Fil: Lahann, Joerg. University of Michigan; Estados Unidos Fil: Moon, James J.. University of Michigan; Estados Unidos Fil: Schwendeman, Anna. University of Michigan; Estados Unidos Fil: Lowenstein, Pedro R.. University of Michigan; Estados Unidos Fil: Castro, Maria G.. University of Michigan; Estados Unidos

Glioma is one of the most aggressive tumors. The median survival of the most aggressive form (Glioblastoma (GBM)) is approximately 14–20 months. The current standard of care includes tumor resection, chemotherapy and radiation, nevertheless, the incidence of recurrence remains high and there is a critical need for developing new therapeutic strategies. T-cell mediated immunotherapy that triggers an anti-tumor T cell-mediated memory response is a promising approach since it will not only attack the primary tumor but also prevent recurrence. Multiple immunotherapeutic strategies against glioma are currently being tested in clinical trials. We have developed an immune-mediated gene therapy (Thymidine kinase plus Fms-like tyrosine kinase 3 ligand: TK/Flt3L) which induces a robust anti-tumor T cell response leading to tumor regression, long-term survival and immunological memory in glioblastoma models. Efficacy of the anti-glioma T cell therapy is determined by anti-tumor specific effector T cells (Han et al., 2014). Therefore, assessing effector T cell activation status and function are critical readouts for assessing the effectiveness of the therapy. Here, we detail methodologies to evaluate tumor specific T-cell responses using a genetically engineered Sleeping Beauty transposase-mediated glioma model. We first describe the glioma model and the generation of neurospheres (NS) that express the surrogate antigen cOVA. Then, we describe functional assays to determine anti-tumor T-cell response.

Despite advances in uncovering the molecular mechanisms that mediate glioma progression and the implementation of novel therapeutic modalities, patients’ prognosis remains dismal. This is due to both systemic and local tumor induced immune suppression. We are particularly interested in the role played by infiltrating immunosuppressive myeloid derived suppressor cells (MDSCs) in the glioma tumor microenvironment (TME). This immunosuppressive TME also interferes with the effectiveness of immunotherapies against glioma. Development of multipronged treatment approaches is imperative when aiming to generate a robust anti-glioma immune response. Evaluating the inhibitory potential of MDSCs within the TME is an important aspect for developing effective treatments for glioma. Herein, we discuss methodology to assess the inhibitory effects of MDSCs isolated from the TME using a mouse glioma model.

Gliomas are the most common primary brain tumors; patients exhibit a poor prognosis. Mutations in isocitrate dehydrogenase (mIDH) are present in most patients with lower grade glioma (LGG), and are correlated with better prognosis and survival. We postulated that mIDH1 induces epigenetic reprogramming leading to alteration in immune cells’ function. To examine the role of mIDH1 in the tumor immune microenvironment (TME), we generated LGG glioma models using the sleeping beauty system (Koschmann et al., 2016, Nunez et al., 2019). We show that mIDH1 gliomas exhibit increased levels of CD11b+ Gr1+ myeloid derived suppressor cells in the tumor, BM, circulation and spleen of mice. We found that mIDH1 modifies the cytokines’ repertoire in the glioma microenvironment altering the phenotype and function of the tumor infiltrating CD45+/CD11b+/Gr-1+ myeloid cells, rendering them non-immunosuppressive. Production of these cells results from activation of the granulocytic differentiation program in the BM. This novel mechanism is mediated by tumor-derived granulocyte-colony stimulating factor (G-CSF) which elicits expansion and differentiation of hematopoietic stem cells, skewing hematopoiesis towards the immature myeloid lineage. Moreover, mIDH1 glioma derived G-CSF causes mobilization of hematopoietic stem cells (HSCs) and myeloid progenitors (MPs) from BM to spleen. Blocking G-CSF in mIDH1 bearing mice significantly restored HSCs, and MPs frequencies in the spleen to levels encountered in wtIDH1 glioma. Interestingly, blocking G-CSF restored the inhibitory function of the granulocytic CD11b+ Gr-1+ in mIDH1, and shortened the median survival (MS) of mIDH1 bearing mice to the same MS encountered in wtIDH1 glioma. Our results provide insights into novel epigenetic alterations triggered by mIDH1 which regulate myeloid cells’ heterogeneity and immunosuppression; a feature that can be harnessed to develop novel immunotherapeutic strategies.

Pediatric high grade gliomas (pHGGs) have a median survival (MS) of 9–15 months and are the most common malignant brain tumors in children. No significant improvement in the MS of these patients has been registered in decades. Thus, a representative in vivo model to study these tumors is critical. Recurrent mutations in genes encoding histones H3.3 and H3.1 have been described, which are distinctive of pHGGs. To stablish a mouse model for pHGG harboring the mutation H3.3-G34R, which co-occurs with inactivating mutations in ATRX and TP53, we used the Sleeping Beauty transposase system (H3.3-G34R group). The MS of mice harboring H3.3-G34R mutation was significantly longer than the MS of the WT-H3.3 control group (p< 0.0001). We performed RNA-Sequencing of these tumors which yielded 233 differentially expressed genes between these two groups (p< 0.05). Gene set enrichment analysis of the upregulated genes in H3.3-G34R group showed significant GOs related to immune response activation and response to IFNγ. Thus, we next studied the tumor microenvironment (TME) to determine the distinctive immune cell populations, at the phenotypic and functional level. We observed that the H3.3-G34R TME exhibited less frequency of Gr-1+/CD11b+ cells (myeloid-derived suppressor cells: MDSC). MDSC are immature myeloid cells that impair T cell functions and the adaptive immune response. Moreover, the H3.3-G34R TME had increased numbers of total macrophages (p< 0.05), a higher proportion of M1 anti-tumor macrophages, and higher levels of CD8+ T cells (p< 0.01). Also, the H3.3-G34R renders glioma cells more sensitive to IFNγ regulation as it induced a higher pSTAT1/STAT1 ratio in H3.3-G34R cells. These results shed light on the interaction between the immune system and the H3.3-G34R glioma cells and how this could be harnessed to develop immune mediated therapies specific for pHGG harboring the H3.3-G34R mutation.

Molecular characterization studies have led to uncover the genetic signatures and methylation status of gliomas and correlate these with clinical prognosis. These studies have illustrated that mutation in isocitrate dehydrogenase (mIDH1), and far less common mIDH2, enzymes present in most patients with low grade and secondary glioma. Moreover, glioma patients with mIDH1 show better prognosis and survival. We hypothesize that mIDH1 impacts tumor immunity by altering the phenotype and function of tumor infiltrating immune cells. To examine the role of mIDH1, we generated glioma models containing the following genetic lesions; NRASG12V, shp53, and shATRx with or without mIDH1. The mouse models investigated are: mIDH1 tumors (NRASG12V+ shp53+ shATRX+ mIDH1) or wtIDH1 tumors (NRASG12V+ shp53+ shATRX+ wtIDH1). Mice bearing mIDH1 tumors exhibited longer median survival (MS) compared to the tumors harboring wtIDH1 (MS=163 vs. MS=70 days post-injection). Transcriptome analyses showed a positive enrichment in immune-stimulatory related gene families in mIDH1 tumors vs. wtIDH1 tumors. Flow cytometric characterization of immune cells showed higher expansion of CD11b+Gr-1+ myeloid cells in the tumor microenvironment (TME) and spleen of mIDH1 compared to wtIDH1 tumors (%MDSC/CD45+: 47.0% vs. 18.06%; P

Aminopeptidase-A (APA) is a less well-studied enzyme of the renin-angiotensin system. We propose that it is involved in cardiac angiotensin (ANG) metabolism and its pathologies. ANG-(1–7) can ameliorate remodeling after myocardial injury. The aims of this study are to 1) develop mass spectrometric (MS) approaches for the assessment of ANG processing by APA within the myocardium; and 2) investigate the role of APA in cardiac ANG-(1–7) metabolism after myocardial infarction (MI) using sensitive MS techniques. MI was induced in C57Bl/6 male mice by ligating the left anterior descending (LAD) artery. Frozen mouse heart sections (in situ assay) or myocardial homogenates (in vitro assay) were incubated with the endogenous APA substrate, ANG II. Results showed concentration- and time-dependent cardiac formation of ANG III from ANG II, which was inhibited by the specific APA inhibitor, 4-amino-4-phosphonobutyric acid. Myocardial APA activity was significantly increased 24 h after LAD ligation (0.82 ± 0.02 vs. 0.32 ± 0.02 ρmol·min−1·μg−1, MI vs. sham, P < 0.01). Both MS enzyme assays identified the presence of a new peptide, ANG-(2–7), m/z 784, which accumulated in the MI (146.45 ± 6.4 vs. 72.96 ± 7.0%, MI vs. sham, P < 0.05). Use of recombinant APA enzyme revealed that APA is responsible for ANG-(2–7) formation from ANG-(1–7). APA exhibited similar substrate affinity for ANG-(1–7) compared with ANG II { Km(ANG II) = 14.67 ± 1.6 vs. Km[ANG-(1–7)] = 6.07 ± 1.12 μmol/l, P < 0.05}. Results demonstrate a novel role of APA in ANG-(1–7) metabolism and suggest that the upregulation of APA, which occurs after MI, may deprive the heart of cardioprotective ANG-(1–7). Thus APA may serve as a potentially novel therapeutic target for management of tissue remodeling after MI.

Angiotensin-converting enzyme 2 (ACE2) is present in the heart and thought to exert protective functions. We conducted studies in ACE2 deficient mice to determine whether enzyme loss would exacerbate the cardiac and vascular pathological responses to chronic subcutaneous (sc) angiotensin II (Ang II) infusion. Eight-week-old male ACE2 knockout (KO) and wild type (WT) mice were infused with Ang II (1000 ng/kg per min, 4 weeks) using mini-osmotic pumps. Blood pressure (radiotelemetry), cardiac function (echocardiography, echo), cardiac/aortic structure (histology, collagen, and oxidative stress), and vascular inflammation were examined. Before Ang II infusion, ACE2 KO mice showed unaltered cardiac function and blood pressure. After 4 weeks of Ang II infusion, the mean arterial pressure (MAP) increased from 96 ± 2 to 136 ± 17 mm Hg (∼40%) in WT and from 104 ± 5 to 141 ± 13 mm Hg (∼ 35%) in ACE2 KO. While there were no differences in MAP between groups, the ACE2 KO responded differently to the hypertensive stimulus. Echo analysis revealed severe myocardial dysfunction in Ang II-infused ACE2 KO (Ang ACE2 KO). Ejection fraction was lower (39% versus 50%) as was fractional shortening (27% versus 38%) in ACE2 KO versus WT, respectively. Cardiac dysfunction was associated with hypertrophic cardiomyopathy shown by increased left-ventricular wall thickness, average cardiomyocyte cross-sectional area, and heart weight/body weight ratio. Collagen staining in the myocardium and aorta revealed increased collagen in Ang ACE2 KO, suggestive of remodeling. Results also showed enhanced oxidative stress in the myocardium and aorta of Ang ACE2 KO. There was a 3-fold elevation in macrophage inflammatory protein 1α (MIP 1α) in the aorta of ACE2 KO. Studies in the ACE2 KO model reveal the importance of ACE2 in the maladaptive cardiac and aortic responses to Ang II stimulation, seen as enhanced remodeling using physiological, structural, and biochemical markers. Results document a cardio- and vascular-protective role of ACE2 under pathological conditions.

Angiotensin Type 1 (AT1) receptors are involved in cardiovascular pathology. Lozartan, the AT1 receptor blocker, in combination with exercise has been shown to be effective in improving cardiac performance. The goal of the study was to investigate the role of AT1a receptors on cardiac function, and exercise tolerance in response to aerobic exercise using AT1a receptor knockout (AT1aKO) mice. An exercise wheel system was used for the exercise paradigm. Male (C57BL/6) wild type (WT) and AT1aKO mice were randomly assigned to four groups: WT control (n=6), WT exercise (WTEX, n=8), AT1a KO (KO) control (n=5), and AT1a KO exercise (KOEX, n=8). Mice were forced to run at a velocity of 8 m/min for 1 hour, 3d/wk, for 7 wks. Echocardiography was conducted at baseline and 7 wks. Lactate was measured during several exercise sessions. Electrolytes and cardiac histology were assessed post-sacrifice. Results showed a significant increase in ejection fraction (EF%) in KOEX (72.5±1.5%) vs. WTEX and KO control (63.4±1.2% and 63.2±2.2%, respectively). Mitral valve assessment revealed a marked decrease in E-wave velocity in WTEX compared to WT control at baseline (74±1.9 vs.90±3.5 cm/s, p p p 3 - ] in KOEX vs. WTEX (21.7±0.6 vs. 13.7±1.8 mmol/l). Cl - was lower in KOEX compared to WTEX (102.5±2.5 vs. 113.5±2.7mmol/l, p