title: Angiotensin-converting enzyme 2 (ACE2) activator diminazene aceturate ameliorates endotoxin-induced uveitis in mice.
abstract: OBJECTIVE
  Uveitis is a common cause of vision loss. The renin angiotensin system (RAS), which plays a vital role in cardiovascular system, is a potent mediator of inflammation and has been implicated in the pathogenesis of uveitis. A newly identified axis of RAS, ACE2/Ang-(1-7)/Mas, has emerged as a novel target because it counteracts the deleterious effect of angiotensin II. The purpose of this study was to investigate the effect of endogenous ACE2 activation in preventing endotoxin-induced uveitis (EIU) in mice.
  
  
  METHODS
  ACE2 activator diminazene aceturate (DIZE) was administered both systemically and locally. For systemic administration, female BALB/c mice received intraperitoneal injection of DIZE (60 mg/kg body weight [BW]) for 2 days prior to lipopolysaccharide (LPS) intravitreal injection (125 ng) to induce uveitis. For local study, DIZE was given at 0.5, 0.1, and 0 mg/mL as eyedrops six times per day for 2 days before LPS injection. The anterior segment of the mice was examined at 12, 24, 48, and 72 hours after LPS injection, and clinical scores were determined at the same time. Morphology and infiltrating inflammatory cells were evaluated after 24 hours. The mRNA levels of inflammatory cytokines were analyzed by real-time RT-PCR. ACE2 activity was determined using a self-quenching fluorescent substrate.
  
  
  RESULTS
  At 24 hours, the clinical score of mice treated with DIZE systemically was significantly lower (mean, ∼1.75) than the saline vehicle group (mean, ∼4) (P < 0.001). Histological examination showed 63.4% reduction of infiltrating inflammatory cells in the anterior segment and 57.4% reduction in the posterior segment of DIZE-treated eyes. The number of CD45(+) inflammatory cells in the vitreous of the DIZE-treated group was decreased (43.3%) compared to the vehicle group (P < 0.01). The mRNA levels of inflammatory cytokines were significantly reduced in the DIZE-treated group (P < 0.01, P < 0.001). The number of infiltrating inflammatory cells was also significantly reduced in eyes that received topical administration of DIZE: 73.8% reduction in the 0.5 mg/mL group and 51.7% reduction in the 0.1mg/mL group compared to the control group. DIZE treatment resulted in significantly increased ACE2 activity in the retina (P < 0.001).
  
  
  CONCLUSIONS
  Endogenous ACE2 activation by DIZE has a preventive effect on LPS-induced ocular inflammation in the EIU mouse model. These results support the notions that RAS plays a role in modulating ocular immune response and that enhancing ACE2 provides a novel therapeutic strategy for uveitis.
authors.affiliation: Department of Ophthalmology, University of Florida, Gainesville, Florida, United States The First Affiliated Hospital of Chongqing Medical University, Chongqing Key Laboratory of Ophthalmology, Chongqing Eye Institute, Chongqing, China.
authors.forename: Yiguo
authors.initials: Y
authors.lastname: Qiu
authors.affiliation: Department of Ophthalmology, University of Florida, Gainesville, Florida, United States.
authors.forename: Pollob Kumar
authors.initials: PK
authors.lastname: Shil
authors.affiliation: Department of Ophthalmology, University of Florida, Gainesville, Florida, United States.
authors.forename: Ping
authors.initials: P
authors.lastname: Zhu
authors.affiliation: The First Affiliated Hospital of Chongqing Medical University, Chongqing Key Laboratory of Ophthalmology, Chongqing Eye Institute, Chongqing, China.
authors.forename: Hongxia
authors.initials: H
authors.lastname: Yang
authors.affiliation: Department of Ophthalmology, University of Florida, Gainesville, Florida, United States.
authors.forename: Amrisha
authors.initials: A
authors.lastname: Verma
authors.affiliation: The First Affiliated Hospital of Chongqing Medical University, Chongqing Key Laboratory of Ophthalmology, Chongqing Eye Institute, Chongqing, China.
authors.forename: Bo
authors.initials: B
authors.lastname: Lei
authors.affiliation: Department of Ophthalmology, University of Florida, Gainesville, Florida, United States.
authors.forename: Qiuhong
authors.initials: Q
authors.lastname: Li
mesh_terms: D000085962:Angiotensin-Converting Enzyme 2; D000818:Animals; D000869:Anterior Eye Segment / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D004133:Diminazene / Q000008:administration & dosage / Q000031:analogs & derivatives*; D004195:Disease Models, Animal; D004789:Enzyme Activation / Q000187:drug effects*; D005260:Female; D005500:Follow-Up Studies; D007364:Intercalating Agents / Q000008:administration & dosage; D051379:Mice; D008807:Mice, Inbred BALB C; D009883:Ophthalmic Solutions; D007703:Peptidyl-Dipeptidase A / Q000187:drug effects* / Q000378:metabolism; D012084:Renin-Angiotensin System / Q000187:drug effects* / Q000502:physiology; D014605:Uveitis / Q000139:chemically induced / Q000188:drug therapy* / Q000201:enzymology
chemical_list: D007364:Intercalating Agents; D009883:Ophthalmic Solutions; D007703:Peptidyl-Dipeptidase A; C000705328:Ace2 protein, mouse; D000085962:Angiotensin-Converting Enzyme 2; C003915:diminazene aceturate; D004133:Diminazene
keywords: EIU; angiotensin-converting enzyme 2; diminazene aceturate; renin angiotensin system; uveitis