2. Refractory periodontitis
periodontal disease is a chronic infectious
disease of the supporting tissue of the teeth
Nowadays, a large majority of patients with
periodontitis respond well to conventional
therapies
However, a small percentage of patients
respond poorly of treatment
“ Refractory periodontitis ”
3. Characteristic of RP (Haffajee et al, 2004; Colombo et al,
2012)
Poor responder or refractory of treatment
AL and/or >3 sites
AL >2.5 mm from the baseline visit to any
monitoring visit 1 year post-therapy
Refractory periodontitis
Photo courtesy of: Dr. Darika
Saitawee
4. About 10–15% of US adults are ‘refractory’ to
therapy for chronic periodontitis
Recently, studies suggest that these patients
have investigation was to identify of this
disease
“ proper treatment “
Refractory periodontitis
5. Clinical and Laboratory
Antibody response
Real-Time Quantitative
Polymerase Chain
Reaction
Human Oral Microbe
Identification
Microarray
Diagnosis for refractory
periodontitis
http://www.bio-rad.com/en-us/applications-technologies/what-real-time-pcr-
qpcr
6. Clinical and laboratory
parameters
Colombo et al. 1999
investigation was to use baseline clinical and
laboratory parameters to distinguish subjects
refractory to conventional periodontal therapy
Baseline clinical
microbial
host parameters
7. 27 refractory subjects
poor response after both SRP and surgery with systemic
tetracycline
Clinical and laboratory
parameters
clinic
• Att gain & no site
with new att loss
• Gingival redness
• BOP
• Suppuration
• Supragingival
plaque
accumulation
• PD
• AL
microbial
• Level of 40
subgingival texa
(subging sample)
• Checker board
DNA-DNA
hybridization
Immune
• Serum Ab to 85
subging species
(IgG)
• Checker board
immunoblotting
11. selected by stepwise discriminant analysis:
namely, number of species exhibiting serum
antibody ±50 mg/ml
% of S. constellatus in plaque (similar previous study
Colombo 1998)
% of sites with attachment level ±6 mm
Clinical and laboratory
parameters
12. Clinical and laboratory
parameters
Discriminant analysis using these variables provided
Sensitivity 0.66
Specificity 0.92
Positive predictive value 0.80
Negative predictive values 0.85
13. The relatively high sensitivities, specificities
and predictive values of the clinical,
microbiologic and immunologic variables
these tests may distinguish a relatively
common form of refractory disease
Refractory periodontitis subjects could be
distinguished using a subset of clinical,
microbiological and immunological parameters
Clinical and laboratory
parameters
14. Antibody-based diagnostic
Levine et al. 2002
this study was to determine whether an
elevated IgG Ab response to lysine
decarboxylase, alone or with antibody to
bacterial Ag and baseline clinical
measurements predict the ‘refractory’
patients
Gingival microbiota were found to inhibit the
growth of cultured mammalian cells by
depleting them of “lysine”
15. Eikenella corrodens and Capnocytophaga spp.
Are major sources of the responsible enzyme,
lysine decarboxylase
Lysine decarboxylase Lysine
inhibit the turnover of the most coronally situated,
dentally attached (DAT) cells and JE basal cells
that lie remote from capillaries
cells stop proliferating causing the intermediate
layer of JE to become attenuated and weaken the
epithelial barrier
Antibody-based diagnostic
16. Lysine
decarboxylase
activity
Effect of DAT cell
High - Inhibit DAT cell turnover
Low - DAT cell can turnover
- Maintaining oral hygiene and
- Low levels of bacteria in the
sulcus or pocket (retard accumulation)
- Stop attachment loss
Antibody-based diagnostic
17. IgG antibody contents to a purified antigen
from
Actinomyces spp. (A-Ab)
Streptococcal d-alanyl glycerol lipoteichoic acid
(S-Ab)
related in ‘refractory’ patients
Antibody-based diagnostic
23. Subjects with advanced disease need for a
diagnostic test the immunoassay is
proposed for use in subjects with moderate
periodontitis (mean attachment loss level of
2–4mm)
HKL-Ab facilitated an accurate prediction of
therapeutic outcome in subjects with moderate
periodontitis
Antibody-based diagnostic
24. Real-Time Quantitative
Polymerase Chain Reaction
Marconcini et al. 2011
“Genetic risk factors” were proposed to influence
the natural history of periodontitis
The study use this technology evaluate the
expression levels of leader genes in the
leukocytes of refractory pt
Blood samples
25. PCR efficiencies were calculated with a relative
standard curve derived from a five cDNA dilution
series in triplicate
The standard curves were obtained using
1. glyceraldehyde-3-phosphate dehydrogenase
(GAPDH)
2. growth factor bound protein (GRB2)
3. casitas B-lineage lymphoma (CBL)
4. nuclear factorKB1 (NFKB1)
5. REL-A (gene for transcription factor p65)
Real-Time Quantitative
Polymerase Chain Reaction
26. Real-Time Quantitative
Polymerase Chain Reaction
Results : levels of CBL
and GRB2 were statically
significant in patients with
periodontitis compared to
healthy patients
characteristics of GRB2,
CBL and NFKB1 : a
specific role during
inflammation and process
of bone resorption
27. CBL oncogene part of a transforming retrovirus
GRB2 signal transduction pathwayThis gene has
never been associated with periodontitis
Molecular studies, our hypothesis
CBL played during the phase of bone resorption that can be
supposed to be the acute phase of the disease
GRB2 might play a central role during the inflammation
process that gave the onset at the periodontitis acute phase
Genes are important role in refractory chronic
periodontitis
develop a treatment plan
reduce risk factors and to focus of therapy
Real-Time Quantitative
Polymerase Chain Reaction
28. Human Oral Microbe
Identification Microarray
(HOMIM)
Colombo et al. 2012
investigation evaluated the post-therapy
changes on the subgingival microbiota
of periodontitis patients who were RP or
GR as measured by using the HOMIM
technique
Tx: SRP + OHI + MWF + ABT (Amoxy
500 mg+Metro 250 mg)
29. The HOMIM methodology used a total of 400 16S
rRNA-based, reverse-capture oligonucleotide probes
targeting >300 bacterial taxa
16S rRNA genes were PCR amplified from DNA
extracts
labeled 16S amplicons were hybridized overnight to
probes on the slides
After washing, the microarray slides were scanned
and crude data were extracted using software for
microarray image analysis
Human Oral Microbe
Identification Microarray
(HOMIM)
32. The majority of species evaluated decreased in
prevalence in both groups after treatment
Species that increased or persisted in high frequency in
RP but were significantly reduced in GR included
Human Oral Microbe
Identification Microarray
(HOMIM)
Bacteroidetes sp.
Porphyromonas
endodontalis
Porphyromonas
gingivalis
Prevotella spp.
Tannerella forsythia
Dialister spp.
Selenomonas spp.
Catonella morbi
Eubacterium spp.
Filifactor alocis
Parvimonas micra
Peptostreptococcus sp.
OT113
Fusobacterium sp. OT203
Pseudoramibacter
alactolyticus
Streptococcus intermedius
/Streptococcus
constellatus
Shuttlesworthia satelles
33. In contrast
Capnocytophaga sputigena, Cardiobacterium
hominis, Gemella haemolysans, Haemophilus
parainfluenzae, Kingella oralis, Lautropia
mirabilis, Neisseria elongata, Rothia
dentocariosa, Streptococcus australis, and
Veillonella spp.
associated with therapeutic success
Human Oral Microbe
Identification Microarray
(HOMIM)
34. GR patients
able to maintain low frequency and
proportions of these organisms
RP patients
rapidly colonized by these species
incapable of maintaining low levels of
putative periodontal pathogens attributable to
host impairment and/or colonization by a more
virulent periodontal microbiota
Human Oral Microbe
Identification Microarray
(HOMIM)
35. Haffajee et al. 2004
Clinical and microbiological changes
associated with the use of combined
antimicrobial therapies to treat ‘‘refractory’’
periodontitis
Combine therapies
SRP
periodontal surgery
locally delivered tetracycline at pockets ≥ 4 mm
systemically administered amoxicillin (500 mg, t.i.d. for 14
days) metronidazole (250 mg, t.i.d. for 14 days)
professional removal of supragingival plaque weekly
for 3 months
f/u every 3 months post-therapy for 2 years
Treatment of refractory
periodontitis
37. Combined antimicrobial
therapies to treat ‘‘refractory’’
periodontitis
members of the green, orange and red complexes as well as some of the
species in the other category were significantly elevated in the periodontitis
subjects compared with the refractory subjects
41. conventional periodontal therapies could be
successfully treated using a combination of
mechanical and antimicrobial therapies
combine therapies that were known to affect the
subgingival microbiota and provide a beneficial
clinical response (synergistic effects)
Mean PD reduction was 0.83 ± 0.13 mm
Mean attachment level gain was 0.44 ±0.12 mm
Combined antimicrobial
therapies to treat ‘‘refractory’’
periodontitis
42. SRP lower the biomass on the tooth surface
and PD
Local drug delivery directly diminish the
pathogen load in any residual pockets of ≥4 mm
Systemically administered amoxicillin plus
metronidazole rapidly lower the level of
pathogens throughout the oral cavity/poor
clinical access, and to attempt to control
pathogenic species that might have entered the
host’s tissues
Repeated professional supragingival plaque
Combined antimicrobial
therapies to treat ‘‘refractory’’
periodontitis
43. conclusion
The combined antibacterial therapy was
successful in controlling disease progression in
refractory periodontitis
Proposed reasons for ‘‘refractory’’ disease have
included differences in the
subgingival microbiota
host response
Environmental factors (smoking)
All of these hypotheses were presented with the
understanding that proper home care procedures
and treatment of refractory patients