title: Development of an ex vivo coculture system to model pulpal infection by Streptococcus anginosus group bacteria.
abstract: BACKGROUND
  Streptococcus anginosus group (SAG) bacteria are opportunistic pathogens and a major cause of pulpal infection and subsequent abscess formation. Understanding of the processes involved in SAG oral infections has been limited by the lack of an appropriate model system.
  
  
  METHODS
  Cocultures of SAG bacteria and mammalian tooth slices were maintained using a combination of Dulbecco modified eagle medium and brain-heart infusion broth at 60 rpm, 37°C, 5% CO(2) for 4, 8, or 24 hours before histologic examination or staining with acridine orange/ethidium bromide. Tooth slices were also incubated as described with SAG bacteria stained with fluorescein diacetate. Pulps were extirpated from infected and sterile cultured tooth slices, messenger RNA was extracted and converted to complementary DNA, and polymerase chain reaction were performed for genes encoding tumor necrosis factor α, interleukin 1β, and interleukin-6.
  
  
  RESULTS
  SAG bacteria were able to adhere directly to the central region of the pulpal matrix in small foci that were associated with a localized matrix breakdown. Acridine orange-ethidium bromide staining and cell counts indicated a decrease in mammalian cell viability with increasing incubation times in the presence of SAG bacteria. The increased expression of tumor necrosis factor α and interleukin 1β was detected in infected tooth slices.
  
  
  CONCLUSIONS
  A novel ex vivo model system has been developed that allows coculture of SAG bacteria with a 3-dimensional organotypic tooth slice. The model allows observation of bacterial growth patterns and subsequent responses from host tissues. Therefore, it may be of future use in testing the efficacy of both antimicrobial and anti-inflammatory treatments for use in endodontic therapy.
authors.affiliation: Tissue Engineering and Reparative Dentistry, Cardiff University, Cardiff, United Kingdom.
authors.forename: Jessica L
authors.initials: JL
authors.lastname: Roberts
authors.forename: Jean-Yves
authors.initials: JY
authors.lastname: Maillard
authors.forename: Rachel J
authors.initials: RJ
authors.lastname: Waddington
authors.forename: Stephen P
authors.initials: SP
authors.lastname: Denyer
authors.forename: Christopher D
authors.initials: CD
authors.lastname: Lynch
authors.forename: Alastair J
authors.initials: AJ
authors.lastname: Sloan
mesh_terms: D000165:Acridine Orange; D000818:Animals; D001422:Bacterial Adhesion / Q000502:physiology; D058491:Bacterial Load; D001431:Bacteriological Techniques; D016923:Cell Death / Q000502:physiology; D002470:Cell Survival / Q000502:physiology; D018920:Coculture Techniques; D003470:Culture Media; D003782:Dental Pulp / Q000382:microbiology* / Q000473:pathology; D003788:Dental Pulp Diseases / Q000382:microbiology* / Q000473:pathology; D003804:Dentin / Q000382:microbiology / Q000473:pathology; D004996:Ethidium; D005347:Fibroblasts / Q000473:pathology; D005452:Fluoresceins; D005456:Fluorescent Dyes; D007180:Incisor / Q000382:microbiology / Q000473:pathology; D053583:Interleukin-1beta / Q000032:analysis; D015850:Interleukin-6 / Q000032:analysis; D008297:Male; D009804:Odontoblasts / Q000473:pathology; D009924:Organ Culture Techniques; D051381:Rats; D017208:Rats, Wistar; D013290:Streptococcal Infections / Q000382:microbiology*; D034366:Streptococcus anginosus / Q000254:growth & development / Q000502:physiology*; D034365:Streptococcus constellatus / Q000254:growth & development / Q000502:physiology*; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis
chemical_list: D003470:Culture Media; D005452:Fluoresceins; D005456:Fluorescent Dyes; D053583:Interleukin-1beta; D015850:Interleukin-6; D014409:Tumor Necrosis Factor-alpha; D004996:Ethidium; D000165:Acridine Orange; C018506:diacetylfluorescein