Clopidogrel enhances periodontal repair in rats through decreased inflammation

Abstract

Aim: We hypothesized that platelet inactivation induced by drugs might interfere with periodontal repair in experimental periodontitis by suppressing the release of biological mediators from platelets at the site of injury.

Material and methods: Sixty rats were randomly assigned to six groups (n = 10) and ligatures were placed around lower first molars of three groups. The other three groups were used as negative controls. Ligatures were removed after 10 days of periodontitis induction and all groups were submitted to treatment with aspirin (Asp) (30 mg/kg), clopidogrel (Clop) (75 mg/kg) or NaCl 0.9% intra-gastrically once daily for 3 days. Periodontal tissue was assessed by the measurement of CXCL12, CXCL4, CCL5 and platelet-derived growth factor (PDGF) by enzyme-linked immunosorbent assay; histomorphometrical analysis of polymorphonuclear (PMN) infiltration, attachment loss, bone loss and osteoclast numbers and quantification of blood vessels by imunnohistochemistry.

Results: During periodontal repair and treatment with NaCl 0.9%, CCL5 was decreased and CXCL12 increased when compared with negative control groups. Asp and Clop did not affect CCL5 expression, decreased CXCL12 but only Clop decreased CXCL4 and PDGF content compared with saline-treated animals. Clop increased blood vessel number, reduced PMN count and decreased attachment and bone loss, also decreased osteoclast number in animals submitted or not to periodontal repair.

Conclusion: Systemic administration of Clop for 3 days improved the repair process associated with experimental periodontal disease, suggesting that it may have therapeutic value under situations where tissues undergo a transition from inflammation to repair.

Keywords: chemokines; clopidogrel; inflammation; innate immunity; periodontal repair.

FIGURE 1

Effect of systemic treatment with Asp or Clop on the gingival contents of CCL5 (panel A), CXCL4 (panel B), CXCL12 (panel C) in rats with ligature induced periodontal disease when stimulus is removed. Results were normalized to the total protein contents in each sample and expressed as pg/mg of protein. Values are expressed as mean±SD. * P<0.0001 vs Repair (Rep)+ NaCl, Rep+ Asp, Rep+ Clop; **P<0.001 vs Clop; #P<0.0001 vs Rep+ Asp, Rep+ Clop; ##P<0.001 vs Rep+ Asp and Rep+ Clop.

FIGURE 2

Effect of systemic treatment with Asp or Clop on the gingival contents of PDGF in rats with ligature induced periodontal disease when stimulus is removed. Results were normalized to the total protein contents in each sample and expressed as pg/mg of protein. Values are expressed as mean±SD. *P<0.001 vs Rep+ Clop.

FIGURE 3

Histological analysis of the alveolar bone crest/gingival margin region of right lower first molar stained with hematoxylin and eosin (H&E). Images are representative of the results observed in 5 animals in each group. 200X magnification.

FIGURE 4

Effect of systemic treatment with Asp or Clop on the PMN count (panel A); attachment loss (panel B); alveolar bone loss (panel C) and bone percentage (panel D) in animals submitted or not to experimental periodontal disease. The results are representative from 5 animals/group. Values are expressed as mean±SD. *P<0.0001 vs Rep+Clop; #P<0.001 vs Rep+Clop.

FIGURE 5

Immunohistochemistry sections of the alveolar bone crest of right lower first molar of rats stained with anti-tartrate-resistant acid phosphatase (TRAP) to measure osteoclast number. Images are representative of the results observed in 5 animals in each group. 200X magnification.

FIGURE 6

Effect of systemic treatment with Asp or Clop on the osteoclast number in rats with ligature induced periodontal disease when stimulus is removed. Values are expressed as mean±SD. *P<0.0001 vs Clop, Rep+Clop.

FIGURE 7

Effect of systemic treatment with Asp or Clop on the number of blood vessels in rats submitted or not to experimental periodontal repair. The results are representative from 5 animals/group. Values are expressed as mean±SD. *P<0.0001 vs Rep+Clop.