Papers by David Goukassian
Pathophysiology and pharmacological management of asthma from a nature-nurture perspective
Primary Health Care, 2013
Journal of Investigative Dermatology, 1999
The tumor suppressor protein p53 participates in DNA repair and cell cycle regulation in response... more The tumor suppressor protein p53 participates in DNA repair and cell cycle regulation in response to injuries like ultraviolet (UV) irradiation. We have previously reported that the thymidine dinucleotide (pTpT), a common target for DNA photoproduct formation by UV light, mimics many effects of UV irradiation in cultured skin-derived cells, at least in part through the activation of p53. In this report we compare the effects of solar-simulated irradiation and pTpT on p53 and p53-regulated proteins involved in cellular growth arrest and DNA repair in cultured human dermal fibroblasts. We find that, like UV irradiation, pTpT M ore than half of all human cancers, including invasive cutaneous squamous cell carcinomas (Brash et al, 1991) and basal cells carcinomas, contain mutations of the p53 gene (Ziegler et al, 1993; Hollstein et al, 1994). The p53 nuclear phosphoprotein has been shown generally to participate in DNA repair and specifically to mediate cellular responses to UV irradiation, a major environmental genotoxin and cutaneous carcinogen. p53 transcriptionally upregulates at least six genes involved directly in DNA repair, increasing the time available for repair prior to DNA replication and cell division, and/or eliminating severely damaged cells from the tissue: p21, MDM2, GADD45, Bax, cyclin G, and IGF-BP3 (Ko and Prives, 1996). In addition, the genes encoding transforming growth factor-α (TGF-α) (
The FASEB Journal, Mar 26, 2002
Age-related decline in DNA repair capacity (DRC) is associated with decreased constitutive levels... more Age-related decline in DNA repair capacity (DRC) is associated with decreased constitutive levels of p53 and other nucleotide excision repair proteins. To determine whether pretreatment of cells with small DNA oligonucleotides compensates for decreased DRC in the elderly, fibroblasts from donors of different ages were pretreated with thymidine dinucleotide (pTT), a 5´ phosphorylated 9 base oligonucleotide (p9mer) or diluent alone for 48 h, then UV-irradiated with solar-simulated light. Western blot analysis revealed age-associated decreases of 40%-80% between newborn and old adult donor cells in the constitutive protein levels of p53, p21, XPA, RPA, ERCC1, and PCNA. Treatment with pTT or p9mer up-regulated these proteins by 200%-650% at 24, 48, and 72 h. Moreover, pretreatment with oligonucleotides significantly increased the removal rate of photoproducts as determined by reacting DNA with thymine dimer-specific antibodies: 40±5% vs. 20±9% and 15±11% remained after 24 h in diluent, pTT and p9mer treated cells, respectively. Oligonucleotide-treated adult cells removed thymine dimers at least as rapidly as diluent treated newborn cells, demonstrating that pTT and p9mer completely corrected the age-associated decrease in DRC. Our studies suggest that topical oligonucleotide treatment may enhance DRC in older adults and thus reduce the carcinogenic risk from solar UV irradiation in this age group.
Journal of Investigative Dermatology, Mar 1, 2000
Oral Session 07: Non-Cancer Risk Radiation-associated cardiovascular risks for future deep-space missions
Background: During the future Moon and Mars missions, astronauts will be exposed to space radiati... more Background: During the future Moon and Mars missions, astronauts will be exposed to space radiation (IR) for extended time. The majority of space flight-associated risks identified for the cardiovascular (CV) system to date were determined shortly after low Earth orbit (LEO) short- and long-duration space flights that include: serious cardiac dysrhythmias, compromised orthostatic CV response and manifestation of previously asymptomatic CV disease. Further ground-based experiments using a surrogate model of microgravity supported the space flight data for significant cardiac remodeling due to prolonged exposure to microgravity. These symptoms were determined to be a consequence of adaptation to microgravity that could be ameliorated by a post-mission exercise program, and were not identified as risk factors that were causatively related to space IR. Long-term degenerative effects of cosmic IR during and after space flights on CV system are unknown. It was suggested that due to GCR, each cell in an astronaut’s body will be traversed by 1 H every 3 days, helium ( 2 He) nuclei every few weeks and high charge and energy (HZE) nuclei (e.g. 28 Si, 56 Fe) every few months. Despite the fact that only 1% of GCR is composed of ions heavier than helium, ~41% of the IR dose-equivalent is predicted to be HZE particles with 13% being from 56 Fe particles, only. During an exploration-class space mission to Mars, astronauts will not have access to comprehensive healthcare services for a period of at least 2–3 years. Since the majority of experienced astronauts are middle-aged (average age is 46, and the range is 33–58 years), they are at risk for developing serious CV events which could be lifethreatening for the astronaut and mission-threatening for NASA. Therefore, it is important to evaluate the effects and potential CV risks caused by space IR. We hypothesized that: (i) low-dose space IR-induced biological responses may be long-lasting and are IR type-dependent; (ii) IR may increase CV risks in the aging heart (IR + AGING model) and affect the heart recovery after an adverse CV event, such as acute myocardial infarct (IR + AGING + AMI model). Methods: Eight- to 9-month-old C57BL/6N male mice were IR once with proton ( 1 H) 50 cGy, 1 GeV/n or iron ( 56 Fe) 15 cGy, 1 GeV/n. We evaluated IR-induced biological tissue responses—underlying molecular mechanisms, calcium handling, signal transduction, gene expression and cardiac fibrosis. Cardiac function was assessed by echocardiography (ECHO) and hemodynamic measurements (HEMO) as detailed in Fig. 1. AMI was induced by ligation of left anterior descending coronary artery 1 and 3 months post-IR as detailed in Fig. 2. Results: In the IR + AGING model, cardiac function was not different among the control and 1 H-IR group, whereas left ventricular end-diastolic pressure (LVEDP) was significantly increased in 56 Fe mice 1 and 3 months post-IR. There was a small but statistically significant (P 200% increases, P< 0.02) and 400% decreases in p-p38 MAPK (P < 0.05), suggesting activation of compensatory mechanisms in [Ca 2+ ]i handling in these hearts. By 3 months, compared with control, 1 H- and 56 Fe-IR hearts had 200–500% (P< 0.02) decreases in SERCA2a and more than 200% decreases in p-Creb-1 (P< 0.02), suggesting reduced capacity in intracellular
Abstract 184: TNF-TNFR1/p55 or TNFR2/p75 Receptor-Ligand Interactions Inhibit Early and Increase Delayed Radio-Biological Bystander Responses in BM-Derived EPCs
Circulation Research, Aug 3, 2012
TNF-α (TNF) binds two receptors TNFR1-p55 and TNFR2-p75. Ionizing radiation (IR) increases tissue... more TNF-α (TNF) binds two receptors TNFR1-p55 and TNFR2-p75. Ionizing radiation (IR) increases tissue TNF levels. TNF signaling regulates numerous cyto- and chemokines, known to mediate radiation-induced non-targeted effects (NTE), a phenomenon where cells that are not directly “hit” by IR exhibit IR effects as a result of signals received from nearby or distant IR cells. Little is known about the role of p55 or p75 in regulating NTEs in bone marrow (BM)-derived EPCs. We hypothesized that inhibition of TNF signaling either via p55, or p75 may alter TNF-mediated inflammatory responses, inducing NTEs. Medium transfer experiments (MTE) were performed in WT, p55 knockout(KO) and p75KO BM-derived EPC ex-vivo, where one set of EPCs was irradiated with 1Gy of γ-IR, then IR-conditioned medium (CM) was collected from these dishes at 1, 5, 24h and 3, 5d post-IR. Filtered (0.22µm) IR-CM was transferred to naïve non-IR same genotype EPCs and 24h post-incubation naïve EPCs were processed for p-y-H2AX staining for presence and decay of double strand breaks (DSB). CM from IR EPCs were processed for ELISA profiling (16 genes) and IR EPCs were processed for transcriptional profiling. In WT EPC the peak of detectable mean p-y-H2AX foci/cell were at 24h, whereas in p55 and p75KOs p-y-H2AX were the lowest at 24h (9±0.8 vs 4.8±0.6 and 3.7±0.5, p&amp;amp;lt;0.01 and 0.001, WT vs p55 and p75KOs). This finding indicates that altered TNF signaling inhibits early NTEs (hours) in EPCs. Compared to WT, delayed (5 days) NTEs were amplified in naïve p55 and p75KO EPCs (3.8±0.4 vs 8.5±1 and 5.9±0.8, p&amp;amp;lt;0.02 and 0.01, WT vs p55 and p75KOs), suggesting significant role for TNF signaling in mediating delayed NTEs. ELISA profiling of 16 proteins in CM over 5 days post-IR showed 200-1600% increases (p&amp;amp;lt;0.002, p55KO vs WT) in cumulative levels of TNF, IFNr, IL6, EGF, MIP-1α, GM-SCF, Rantes, and 200-1000%, increases (p&amp;amp;lt;0.05, p75KO vs WT) in IL1α, IL1β, G -CSF, MCP-1, SCF. Transcriptional profiling of γ-IR EPC revealed 139 genes (&amp;amp;gt;2 fold) up or down regulated, clustered into 9 groups. Other aspects of gene array data are being considered. We conclude that TNF ligand-receptor axis regulates NTEs in naïve EPCs and suggest that restoring TNF signaling balance could be used to prevent delayed NTEs in distant from primary IR target tissues.
Frontiers in Physiology, Nov 19, 2023
Lifetime evaluation of left ventricular structure and function in male ApoE null mice after gamma... more Lifetime evaluation of left ventricular structure and function in male ApoE null mice after gamma and space-type radiation exposure.
Abstract 19530: Cardiac Fibroblasts Derived Exosomes Enhanced Endothelial to Mesenchymal Transition and Associated Fibrotic Signaling
Circulation, Nov 11, 2016
Background: Endothelial to mesenchymal transition (EndMT), play critical role in pressure overloa... more Background: Endothelial to mesenchymal transition (EndMT), play critical role in pressure overload-induced pathological fibrosis. Recent studies suggested the contribution of fibroblasts-derived ex...
PLOS ONE, Oct 22, 2014
Previous epidemiologic data demonstrate that cardiovascular (CV) morbidity and mortality may occu... more Previous epidemiologic data demonstrate that cardiovascular (CV) morbidity and mortality may occur decades after ionizing radiation exposure. With increased use of proton and carbon ion radiotherapy and concerns about space radiation exposures to astronauts on future long-duration exploration-type missions, the long-term effects and risks of low-dose charged particle irradiation on the CV system must be better appreciated. Here we report on the long-term effects of wholebody proton (1 H; 0.5 Gy, 1 GeV) and iron ion (56 Fe; 0.15 Gy, 1GeV/nucleon) irradiation with and without an acute myocardial ischemia (AMI) event in mice. We show that cardiac function of proton-irradiated mice initially improves at 1 month but declines by 10 months post-irradiation. In AMI-induced mice, prior proton irradiation improved cardiac function restoration and enhanced cardiac remodeling. This was associated with increased pro-survival gene expression in cardiac tissues. In contrast, cardiac function was significantly declined in 56 Fe ion-irradiated mice at 1 and 3 months but recovered at 10 months. In addition, 56 Fe ion-irradiation led to poorer cardiac function and more adverse remodeling in AMI-induced mice, and was associated with decreased angiogenesis and pro-survival factors in cardiac tissues at any time point examined up to 10 months. This is the first study reporting CV effects following low dose proton and iron ion irradiation during normal aging and post-AMI. Understanding the biological effects of charged particle radiation qualities on the CV system is necessary both for the mitigation of space exploration CV risks and for understanding of long-term CV effects following charged particle radiotherapy.
Abstract 305: Tumor Necrosis Factor-α p75 Receptor, Satellite-Cell Survival, Activation and Muscle Regeneration
Circulation, Oct 16, 2007
Aging is a risk factor for ischemic diseases. TNF-α, a pro-inflammatory cytokine, is expressed in... more Aging is a risk factor for ischemic diseases. TNF-α, a pro-inflammatory cytokine, is expressed in ischemic tissues and is known to modulate angiogenesis. Little is known about the role of TNF-α receptors (TNFR1/p55 and TNFR2/p75) in angiogenic signaling and muscle regeneration. We studied neovascularization in the hind limb ischemia (HLI) model in young and old TNFR2/p75 knockout (p75KO) and wild type (WT) age-matched controls. Between days 7–10 post-HL surgery 100% of old p75KOs experienced auto-amputation of the operated limbs, whereas none of the age-matched WT mice exhibited HL necrosis. Poor blood flow recovery in p75KOs was associated with decreased capillary density and significant reduction in the expression of VEGF mRNA transcripts in ischemic tissue. Compared to presurgery, on days 1–10 post-HL surgery there was 6–10-fold increase in the number of satellite-cells (embrionic NCAM staining) in WT mice, whereas in p75KOs after day 1 through day 10 satellite cells were not detecable. Indeed, p75KO tissue showed increased and prolonged (via day 10) inflammation - neutrophil (MPO-1) and macrophage (F/480) infiltration. Transplantation of WT/GFP (+) BM mononuclear cells into γ-irradiated p75KOs one month prior to HL surgery prevented limb loss, suggesting that ischemia-induced neovascularization and mobilization of BM-derived cells is mediated, at least in part, via TNFR2/p75 expressed in BM-derived cells. In the same BM transplantation model we evaluated the rate of proliferation (Ki67 + cells) of resident GFP (−) vs BM-derived GFP (+) cells. We found that in both WT and p75KO ischemic tissue Ki67 (+) cells almost exclusively were GFP (+), indicating that only BM-derived cells proliferate in the ischemic tissue. Interestingly, Ki67/GFP (+) cells started to appear in WT tissue by day 3 through day 21, whereas in p75KO tissue first proliferative activity was detected on day 28, suggesting extremely delayed recovery and regenaration in p75KO tissue. Our study suggests that, signaling through p75 receptor is required for collateral vessel development in ischemia-induced neovascularization as well as plays a critical role in muscle regeneration and suggest a potential gene target, which could be used to improve the repair of ischemic tissue in adults.
Abstract 18514: Epigenetic Regulation of Chromatin Remodeler CHD5 in Stem Cells During Cardiac Differentiation and Stem Cell Mediated Myocardial Repair and Function After Myocardial Infarction
Circulation, Nov 14, 2017
Poor differentiation ability might be the barrier for the stem cell based therapy in ischemic myo... more Poor differentiation ability might be the barrier for the stem cell based therapy in ischemic myocardium. Epigenetic regulation play a critical role in this cell fate commitment, however, the mecha...
Abstract 18967: Inhibition of Circulating Exosome Improved Ischemic Hindlimb Repair in Type Diabetic db/db Mice
Circulation, Nov 14, 2017
Background: Recent studies have demonstrated that exosomes from diabetic animals/cells have detri... more Background: Recent studies have demonstrated that exosomes from diabetic animals/cells have detrimental effects on the post-injury vascular repair. Here, we tested the hypothesis that systemic exosome inhibition in vivo improves blood flow recovery in ischemic hindlimbs of diabetic db/db following surgical ischemia. Methods and Results: Exosomes were isolated from plasma of non-diabetic db/+ and diabetic db/db mice by standard ultracentrifugation method. Unilateral hindlimb ischemia surgery was conducted by ligation of left femoral artery in 12-week old, male db/+ and db/db mice. Exosome inhibitor GW-4869 (GW, 2 μg/g body weight) was administrated by intraperitoneal injection every other day for 4 weeks starting from one week before the hindlimb ischemia (HLI) surgery. HLI mice injected with vehicle served as controls. Mice were divided into four groups: 1) db/+ + vehicle; 2) db/db+ vehicle; 3) db/+ GW; 4) db/db + GW. We found that systemic administration of GW decreased necrosis and loss of toe/toenail, ...
JCI insight, Aug 8, 2019
Podoplanin, a small mucine-type transmembrane glycoprotein, has been recently shown to be express... more Podoplanin, a small mucine-type transmembrane glycoprotein, has been recently shown to be expressed by lymphangiogenic, fibrogenic and mesenchymal progenitor cells in the acutely and chronically infarcted myocardium. Podoplanin binds to CLEC-2, a C-type lectin-like receptor 2 highly expressed by CD11b high cells following inflammatory stimuli. Why podoplanin expression appears only after organ injury is currently unknown. Here, we characterize the role of podoplanin in different stages of myocardial repair after infarction and propose a podoplaninmediated mechanism in the resolution of post-MI inflammatory response and cardiac repair. Neutralization of podoplanin led to significant improvements in the left ventricular functions and scar composition in animals treated with podoplanin neutralizing antibody. The inhibition of the interaction between podoplanin and CLEC-2 expressing immune cells in the heart enhances the cardiac performance, regeneration and angiogenesis post MI. Our data indicates that modulating the interaction between podoplanin positive cells with the immune cells after myocardial infarction positively affects immune cell recruitment and may represent a novel therapeutic target to augment post-MI cardiac repair, regeneration and function.
Abstract 12916: Exosomal RNAS as Potential Biomarkers of Cardiovascular Risk in NASA Astronauts
Circulation, Nov 19, 2019
Abstract 16570: Interleukin-10 Deficiency Impairs Reparative Properties of Bone Marrow-Derived Endothelial Progenitor Cell Exosomes
Circulation, Nov 14, 2017
Circulation Research, Jul 25, 2003
Tumor necrosis factor-␣ (TNF-␣) is expressed locally in the vessel wall after angioplasty and ind... more Tumor necrosis factor-␣ (TNF-␣) is expressed locally in the vessel wall after angioplasty and induces growth arrest and apoptosis in endothelial cells (ECs), thereby delaying reendothelialization. Prior studies have shown that direct antagonism of TNF-␣, using a systemically administered soluble receptor, can enhance endothelial recovery and reduce neointimal thickening. These studies have also shown that downregulation of the transcription factor E2F1 was a key mechanism of TNF's effect on ECs. We now show that Ad-E2F1 overexpression at sites of balloon injury accelerates functional endothelial recovery, consistent with the prior in vitro findings. Moreover these studies also reveal divergent effects of TNF-␣ and overexpression of E2F1 on ECs versus VSMCs. TNF-␣ exposure of VSMCs had no affect on proliferation or apoptosis, in contrast to the effect seen in ECs. In Ad-E2F1-transduced VSMCs, however, TNF-␣-induced marked apoptosis in contrast to the survival effect seen in ECs. Finally, these studies suggest that differential activation of NF-B may play a key role in mediating these opposing effects. Nuclear translocation and transcriptional activity of NF-B was markedly attenuated in Ad-E2F1-transduced VSMCs, whereas it remained active in similarly treated ECs when the cells were exposed to TNF-␣. These studies reveal that overexpression of Ad-E2F1 primes VSMCs to TNF-␣-induced apoptosis. Furthermore, E2F1 potentiates VSMC death by blocking antiapoptotic signaling pathway through inhibition of NF-B activation. The divergent responses of VSMCs and ECs to E2F1 overexpression provide unique therapeutic possibilities: simultaneously targeting the cell cycle of two different cell types, within same tissue microenvironment resulting in opposite and biologically complimentary effects. (Circ Res. 2003;93:162-169.
Abstract 353: Bone Marrow Fibroblast Progenitor Cell-derived Exosomes Activate Resident Fibroblast and Augment Pressure Overload Induced Cardiac Fibrosis in IL10KO Mice
Circulation Research, Jul 21, 2017
Background: Activated fibroblasts (myoFBs) play critical role in cardiac fibrosis, however, their... more Background: Activated fibroblasts (myoFBs) play critical role in cardiac fibrosis, however, their origin in diseased heart remains uncertain. Previous studies suggest the contribution of bone marrow fibroblasts progenitor cells (FPC) in pressure overload (PO)-induced cardiac fibrosis and inflammation acts as catalyst in this process. Recently others and we have shown that paracrine mediators packaged in exosomes play important role in cardiac pathophysiology. Thus, we hypothesized that exosome-derived from IL10KO-FPC augments PO-induced resident cardiac fibroblast activation and therefore, aggravate cardiac fibrosis. Methods and Results: Cardiac fibrosis was induced in Wild-type (WT) and IL10-knockout (IL10KO) mice by transverse aortic constriction (TAC). TAC-induced left ventricular (LV) dysfunction and fibrosis were further exaggerated in IL10KO mice. PO-enhanced FPC (Prominin1 + cells) mobilization and homing in IL10KO mice compared to WT mice. To establish the IL10KO-FPC paracrine signaling, exosomes were isolated from WT and IL10KO BM-FPC culture media and characterized for proteins/miRNA. IL10 KO FPC-exosomes showed altered packaging of signature fibrotic miR and proteins. To explore whether FPC-exosomes modulate resident fibroblast activation, adult cardiac fibroblasts were treated with WT and IL10KO FPC-derived exosomes. IL10KO-FPC-derived exosomes exaggerate TGFβ 2 -induced activation of adult fibroblasts. These data suggest that fibrotic remodeling factors (miRs and/or proteins) packaged in IL10KO-FPC exosomes are sufficient to enhance the resident cardiac fibroblast activation and mediate cardiac fibrotic remodeling IL10 treatment significantly inhibits TGFβ 2 -induced FPC to myoFBs transition. Conclusion: Taken together, our findings suggest that paracrine factors secreted by BM-FPC augment resident cardiac fibroblast activation and fibrosis in pressure overloaded myocardium and IL10 negatively regulates this process. Ongoing investigations using molecular approaches will provide a better understanding on the mechanistic and therapeutic aspects of IL10 on PO-induced cardiac fibrosis and heart failure.
American Journal of Pathology, Nov 1, 2009
UV-irradiated skin and UV-induced tumors overexpress the inducible isoform of cyclooxygenase-2 (C... more UV-irradiated skin and UV-induced tumors overexpress the inducible isoform of cyclooxygenase-2 (Cox-2), and Cox-2 inhibition reduces photocarcinogenesis. To evaluate photoprotective effects of Polypodium leucotomos extract (PL), hairless Xpc ؉/؊ mice were fed for 10 days with PL (300 mg/kg) or vehicle then UV-irradiated, once. By 24 hours, UVinduced Cox-2 levels were increased in vehicle-fed and PL-fed mice, whereas by 48 and 72 hours, Cox-2 levels were four-to fivefold lower in PL-fed mice (P < 0.05). p53 expression/activity was increased in PL-fed versus vehicle-fed then UV-irradiated mice. UV-induced inflammation was decreased in PL-fed mice, as shown by ϳ60% decrease (P < 0.001) in neutrophil infiltration at 24 hours, and macrophages by ϳ50% (<0.02) at 24 and 48 hours. By 72 hours, 54 ؎ 5% cyclobutane pyrimidine dimers remained in vehiclefed versus 31 ؎ 5% in PL-fed skin (P < 0.003). The number of 8-hydroxy-2-deoxyguanosine-positive cells were decreased before UV irradiation by ϳ36% (P < 0.01), suggesting that PL reduces constitutive oxidative DNA damage. By 6 and 24 hours, the number of 8-hydroxy-2-deoxyguanosine-positive cells were ϳ59% (P < 0.01) and ϳ79% (P < 0.03) lower in PL-fed versus vehicle-fed mice. Finally, UV-induced mutations in PL-fed-mice were decreased by ϳ25% when assessed 2 weeks after the single UV exposure. These data demonstrate that PL extract supplementation affords the following photoprotective effects: p53 activation and reduction of acute inflammation via Cox-2 enzyme inhibition, increased cyclobutane pyrimidine dimer removal, and reduction of oxidative DNA damage.
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Papers by David Goukassian